Patent Application
20240261418
The present invention belongs to the technical field of pharmaceuticals, and particularly relates to an SOS1 degrader, a preparation method therefor and use thereof as a therapeutic agent preventing and/or treating cancer.
SOS1 (son of sevenless homolog 1) protein is a regulatory protein widely expressed in cells, and as a key protein in signal pathways, SOS1 plays an important role in the regulation in many signal transduction pathways in cells, such as being involved in regulation of Ras and Rac signal pathways. SOS1 consists of 1333 amino acids and comprises a proline-rich domain (PxxP) at its C-terminus. The domain can bind to growth factor receptor-bound protein 2 (Grb2) in the Ras pathway to form a complex of Grb2 and SOS1, thereby bringing SOS1 near the Ras protein on the cell membrane. SOS1 catalyzes the binding of Ras to GTP and promotes the activation of Ras, thereby activating multiple downstream signal pathways, such as Ras-Raf-Mek-Erk and Ras-PI3K-AKT-mTOR. PxxP can also bind to an SH3 (Src homology 3) domain of a protein such as E3B1 in a Rac pathway to form an EPS8-E3B1-SOS1 complex, and the EPS8-E3B1-SOS1 complex is linked to actin filaments through EPS8 to cause the conversion of GTP, so that Rac is activated, and then a signal pathway such as INK and MAPK is activated.
A mutant ras gene is considered to be the major oncogene with high incidence in human cancer. Studies have shown that Ras mutations occur in 20-30% of tumor patients, where 85% are KRas mutations, 12% are NRas mutations, and 3% are HRas mutations. However, due to the picomolar affinity of GTP for its binding site and the smooth surface of Ras proteins, which means other suitable binding pockets are absent, direct acting on Ras to inhibit its activity is considered to be extremely challenging.
Abnormal expression or mutation of SOS1 is also closely related to the occurrence of clinical diseases. Studies have shown that SOS1 mutations are present in NS patients and CFC patients. HGF1 is a rare autosomal dominant genetic disease whose cause is also associated with mutations in the PxxP domain of SOS1. In addition, abnormal expression or mutation of SOS1 is also related to the occurrence of cancer.
WO2018172250A1, WO2020173935A1, WO2019201848A1, WO2020180768A1, WO2020180770A1, WO2019122129A1 and EP3558979A1 disclose several classes of SOS1 inhibitors, but so far there are no reports on SOS1 degraders. Proteolysis targeting chimera (PROTAC) is a technology different from the traditional small molecule inhibitors. The traditional small molecule inhibitors usually need to act on the active site of a target protein to inhibit the activity of the target protein, while the PROTAC is a heterogeneous bifunctional molecule, one end of which is a small molecule inhibitor capable of recognizing the target protein, and the other end of which is an E3 ubiquitin ligase ligand capable of recognizing E3 ubiquitin ligase via a linking chain. Such a bifunctional molecule recognizes the target protein and the E3 ubiquitin ligase in vivo, draws the target protein and the E3 ubiquitin ligase closer to form a ternary complex, and degrades the target protein in vivo through a ubiquitin-proteasome pathway after ubiquitinating the target protein. Compared with the traditional small molecule inhibitors, in one aspect, the PROTAC only needs to draw the target protein and the E3 ubiquitin ligase closer to degrade the substrate, and the action mode enables the technology to be applied to some non-druggable targets; in other aspect, the PROTAC molecules can be released to continue to participate in the degradation process of the next protein after the target protein is degraded, so that the degradation effect with a catalytic effect can realize efficient degradation with less dosage of the PROTAC medicament; in yet another aspect, the traditional small molecule inhibitors easily generate drug resistance often because a point mutation occurs, so that the small molecule inhibitors lose the inhibition effect on the target point, while the PROTAC can directly degrade the target protein, so that the drug resistance generated by the point mutation can be avoided to a certain extent. Therefore, compared with the traditional small molecule inhibitors, the PROTAC technology has high advantages and feasibility in research and development of small molecule new drugs.
The present invention provides a compound of formula I, and/or a stereoisomer, an enantiomer, a diastereomer, a deuteride, a hydrate, a solvate, a prodrug and/or a pharmaceutically acceptable salt thereof
wherein:
The present invention provides a compound of formula I, and/or a stereoisomer, an enantiomer, a diastereomer, a deuteride, a hydrate, a solvate, a prodrug and/or a pharmaceutically acceptable salt thereof
S, L and E are as defined below.
In certain embodiments of the present invention, S is S7:
wherein in S7,
C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C4-C8 cycloalkenyl, 4- to 7-membered heterocycloalkyl (e.g.,
5- to 10-membered heterocycloalkenyl, spiroheterocycloalkyl, fused heterocycloalkyl, bridged heterocycloalkyl, phenyl, heteroaryl, C1-C6 haloalkyl, —COOH and —COORc; Rc is —C1-C6 alkyl, C3-C6 alkenyl, C3-C6 alkynyl, C3-C8 cycloalkyl or —C4-C8 cycloalkenyl; R2 is NS(O)(Rd)(Re), wherein Rd and Re are each independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C3-C8 cycloalkyl, C4-C8 cycloalkenyl, 6- to 10-membered aryl or 5- to 10-membered heteroaryl;
Preferably, in certain embodiments of the present invention, S is S7:
In certain embodiments of the present invention, in S7, R1 is hydrogen.
In certain embodiments of the present invention, in S7, R2 is hydrogen, C1-C4 alkyl, —O(C1-C4 alkyl), —NH2, —NH(C1-C4 alkyl), —N(C1-C4 alkyl)2 or halogen, and the C1-C4 alkyl and the —O(C1-C4 alkyl) are each independently optionally substituted with —O(C1-C4 alkyl).
In certain embodiments of the present invention, in S7, R2 is hydrogen, C1-C4 alkyl, —O(C1-C4 alkyl), —NH2, —NH(C1-C4 alkyl), —N(C1-C4 alkyl)2 or halogen.
In certain embodiments of the present invention, in S7, R2 is hydrogen, C1-C4 alkyl or —O(C1-C4 alkyl), and the —O(C1-C4 alkyl) is optionally substituted with —O(C1-C4 alkyl).
In certain embodiments of the present invention, in S7, R2 is
In certain embodiments of the present invention, in S7, R2 is —O(C1-C4 alkyl).
In certain embodiments of the present invention, in S7, R4 is hydrogen.
In certain embodiments of the present invention, in S7, R3 is hydrogen, halogen or C1-C6 alkyl, and the alkyl is optionally substituted with —NRkRl; Rk and Rl are each independently hydrogen or C1-C6 alkyl.
In certain embodiments of the present invention, in S7, A2 is aryl.
In certain embodiments of the present invention, in S7, Y is 0, 1, 2 or 3.
In certain embodiments of the present invention, in S7, L′ is a bond.
In certain embodiments of the present invention, in S7, R7 is C1-C6 alkyl.
In certain embodiments of the present invention, in S7, R8 is C1-C6 alkyl.
In certain embodiments of the present invention, in S7:
In certain embodiments of the present invention, in S7, R2 is hydrogen, methyl, —OCH3 or
In certain embodiments of the present invention, in S7, R2 is —O(C1-C4 alkyl), e.g., —OCH3.
In certain embodiments of the present invention, in S7, R3 is hydrogen, F, Cl, methyl or ethyl, and the methyl and the ethyl are substituted with —NRkRl; Rk is methyl or ethyl, and Rl is hydrogen, methyl or ethyl.
In certain embodiments of the present invention, in S7, R3 is hydrogen, Cl, —CH2NHCH3 or —CH2N(CH3)2.
In certain embodiments of the present invention, in S7, A2 is phenyl.
In certain embodiments of the present invention, in S7, R7 is methyl.
Preferably, in certain embodiments of the present invention, in S7, Rs is methyl.
Preferably, in certain embodiments of the present invention, S is S7′:
wherein R2, R3, R4, R7, R8, Y, A2 and L′ are as defined and described in S7.
In certain embodiments of the present invention, in S7 and S7 the fragment
is
In certain embodiments of the present invention, S is —O—S7′, wherein the structure and definition of S7′ are as described above.
Preferably, in certain embodiments of the present invention, S is S7″:
wherein R2, R3, R4, R7, Y, A2 and L′ are as defined and described in S7.
In certain embodiments of the present invention, S is —O—S7″, wherein the structure and definition of S7″ are as described above.
Preferably, in certain embodiments of the present invention, S is S7a:
wherein R1, R2, R3, R4, R7, R8 and Y are as defined and described in S7.
Preferably, in certain embodiments of the present invention, S is S7a′:
wherein R2, R3, R4, R7, R8 and Y are as defined and described in S7.
In certain embodiments of the present invention, in S7a and S7a′, the fragment
is
In certain embodiments of the present invention, S is —O—S7a′, wherein the structure and definition of S7a′ are as described above.
Preferably, in certain embodiments of the present invention, S is S7a″:
wherein R2, R3, R4, R7 and Y are as defined and described in S7.
In certain embodiments of the present invention, S is —O—S7a″, wherein the structure and definition of S7a″ are as described above.
In certain embodiments of the present invention, in S7a, S7a′ and S7a″, the fragment
is
and the definitions of R3a and R3b are independently as described in R3. In certain embodiments of the present invention, R3b is C1-C6 alkyl, and the alkyl is substituted with —NRkRl; Rk and Rl are each independently hydrogen or C1-C6 alkyl. In certain embodiments of the present invention, R3b is methyl or ethyl, the methyl and the ethyl are substituted with —NHRl or —NRkRl; Rk and Rl are each independently methyl or ethyl. In certain embodiments of the present invention, R3b is —CH2NHCH3 or —CH2N(CH3)2. In certain embodiments of the present invention, R3a is hydrogen or halogen, e.g., H or Cl.
Preferably, in certain embodiments of the present invention, S, S7, S7′, S7″, S7a, S7a′ and S7a″ are any of the following structures:
Preferably, in certain embodiments of the present invention, S, S7, S7, S7″, S7a, S7a′ and S7a″ are
Preferably, in certain embodiments of the present invention, S, S7, S7′, S7″, S7a, S7a′ and S7a″ are the following structure:
Preferably, in certain embodiments of the present invention, S, S7, S7′, S7″, S7a, S7a′ and S7a″ are the following structure:
In certain embodiments of the present invention, S is
In certain embodiments of the present invention, L is a bond.
In certain embodiments of the present invention, L is —(CH2)j—, wherein 1 or more of methylene groups in the —(CH2)j— are optionally replaced by a group selected from —NR3′—, —O—, —S—, —S(O)—, —S(O)NR3′—, —NR3′S(O)—, —S(O)2—, —S(O)2NR3′—, —NR3′S(O)2—, —NR4′S(O)2NR3′—, —CR1′R2′—, —C(O)—, —C(O)O—, —OC(O)—, —NR3′C(O)O—, —OC(O)NR3′—, —C(O)NR3′—, —NR3′C(O)—, —NR4′C(O)NR3′—, —P(O)—, —P(O)O—, —OP(O)—, —OP(O)O—, ethenylene, ethynylene, 3- to 12-membered cycloalkylene, 3- to 12-membered heterocycloalkylene containing 1 or more heteroatoms selected from N, O and S, 6- to 10-membered arylene and 5- to 10-membered heteroarylene, wherein the ethenylene, the cycloalkylene, the heterocycloalkylene, the arylene and the heteroarylene are each independently optionally substituted with 1 or more substituents selected from halogen, —OR3′, —NR3′R4′, oxo, nitro, cyano, C1-C6 alkyl, —S(C1-C6 alkyl), C3-C10 cycloalkyl, C3-C10 heterocycloalkyl, C6-C10 aryl, 5- to 10-membered heteroaryl, —C(O)R1′, —C(O)OR3′, —OC(O)R1′, —C(O)NR3′, —NR3′C(O)R1′, —S(O)R1′, —S(O)NR3′, —S(O)2R1′, —S(O)2NR3′, NR3′S(O)2R1′, —NR4′S(O)2NR3′, —OC(O)NR3′ and —NR4′C(O)NR3′, wherein the alkyl, the cycloalkyl, the heterocycloalkyl, the aryl and the heteroaryl are each independently optionally substituted with 1 or more substituents selected from halogen, —OH, —NR3′R4′, oxo, nitro, cyano, C1-C6 alkyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl, 6- to 10-membered aryl and 5- to 10-membered heteroaryl; R1′ and R2′ are each independently halogen, —OH, —NR3′R4′, C1-C6 alkyl, C1-C6 chloroalkyl, hydroxy C1-C6 alkyl, —O(C1-C6 alkyl), —NH(C1-C6 alkyl), —NH(C1-C6 alkyl)2, C3-C10 cycloalkyl, —O(C3-C10 cycloalkyl), —NH(C3-C10 cycloalkyl), C3-C10 heterocycloalkyl, —O(C3-C10 heterocycloalkyl), —NH(C3-C10 heterocycloalkyl), 6- to 10-membered aryl, —O(6- to 10-membered aryl), —NH(6- to 10-membered aryl), 5- to 10-membered heteroaryl, —O(5- to 10-membered heteroaryl) or —NH(5- to 10-membered heteroaryl); R3′ and R4′ are each independently hydrogen, deuterium, C1-C6 alkyl, C3-C10 cycloalkyl, C3-C10 heterocycloalkyl, 6- to 10-membered aryl or 5- to 10-membered heteroaryl; j is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24 or 25.
Preferably, in certain embodiments of the present invention, L is —(CH2)j—, wherein 1 or more of methylene groups in the —(CH2)j— are optionally replaced by a group selected from —NR3′—, —O—, —CR1′R2′—, —C(O)—, —S(O)—, —S(O)2—, —C(O)O—, —OC(O)—, —C(O)NR3′—, —NR3′C(O)—, —S(O)2NR3′—, —NR3′S(O)2—, ethenylene, ethynylene, phenyl, 8- to 10-membered bicyclic arylene, 3- to 7-membered saturated or partially unsaturated cycloalkylene, 5- to 11-membered saturated or partially unsaturated spirocycloalkylene, 5- to 11-membered saturated or partially unsaturated fused cycloalkylene, 8- to 10-membered bicyclic saturated or partially unsaturated cycloalkylene, 4- to 7-membered saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 5- to 11-membered saturated or partially unsaturated spiroheterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 5- to 11-membered saturated or partially unsaturated fused heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 8- to 10-membered bicyclic saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 5- to 6-membered heteroarylene having 1-4 heteroatoms independently selected from nitrogen, oxygen and sulfur, and 8- to 10-membered bicyclic heteroaryl having 1-5 heteroatoms selected from nitrogen, oxygen and sulfur, wherein the ethenylene, the ethynylene, the cycloalkylene, the heterocycloalkylene, the phenyl, the spiroheterocycloalkylene, the fused heterocycloalkylene, the spirocycloalkylene, the fused cycloalkylene and the heteroarylene are each independently optionally substituted with 1 or more substituents selected from halogen, oxo, —NR3′R4′, —OR3′, nitro, —CN, C1-C6 alkyl, C3-C10 cycloalkyl and C3-C10 heterocycloalkyl, wherein the alkyl, the cycloalkyl and the heterocycloalkyl are optionally substituted with 1 or more substituents selected from halogen, —OH, —NH2, —CN, C1-C4 alkyl and C3-C6 cycloalkyl; R1′ and R2′ are each independently halogen, —OH, —NH2, C1-C4 alkyl, C1-C4 chloroalkyl, C1-C4 hydroxyalkyl, —O(C1-C4 alkyl), —NH(C1-C4 alkyl), C3-C6 cycloalkyl, —O(C3-C6 cycloalkyl), —NH(C3-C6 cycloalkyl), C3-C6 heterocycloalkyl, or —O(C3-C6 heterocycloalkyl); R3′ and R4′ are each independently hydrogen, deuterium, C1-C4 alkyl, C3-C6 cycloalkyl, or C3-C6 heterocycloalkyl, and j is 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12.
Preferably, in certain embodiments of the present invention, L is —(CH2)j—, wherein 1 or more of methylene groups in the —(CH2)j— are optionally replaced by a group selected from —NR3′—, —O—, —CR1′R2′—, —C(O)—, —S(O)—, —S(O)2—, —C(O)O—, —OC(O)—, —C(O)NR3′—, —NR3′C(O)—, —S(O)2NR3′—, —NR3′S(O)2—, ethenylene, ethynylene, phenyl, 8- to 10-membered bicyclic arylene, 3- to 7-membered saturated or partially unsaturated monocyclic cycloalkylene, 5- to 11-membered saturated or partially unsaturated spirocycloalkylene, 5- to 11-membered saturated or partially unsaturated fused cycloalkylene, 8- to 10-membered bicyclic saturated or partially unsaturated cycloalkylene, 5- to 7-membered saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen and oxygen, 5- to 11-membered saturated or partially unsaturated spiroheterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen and oxygen, 5- to 11-membered saturated or partially unsaturated fused heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen and oxygen, 8- to 10-membered bicyclic saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen and oxygen, 5- to 6-membered heteroarylene having 1-4 heteroatoms independently selected from nitrogen and oxygen, and 8- to 10-membered bicyclic heteroaryl having 1-5 heteroatoms selected from nitrogen and oxygen, wherein the ethenylene, the ethynylene, the cycloalkylene, the heterocycloalkylene, the phenyl, the spiroheterocycloalkylene, the fused heterocycloalkylene, the fused cycloalkylene, the spirocycloalkylene and the heteroarylene are each independently optionally substituted with 1 or more substituents selected from halogen, oxo, —NR3′R4′, —OR3′, nitro, —CN, C1-C6 alkyl, C3-C6 cycloalkyl and C3-C6 heterocycloalkyl, wherein the alkyl, the cycloalkyl and the heterocycloalkyl are optionally substituted with 1 or more substituents selected from halogen, —OH, —NH2, —CN, C1-C4 alkyl and C3-C6 cycloalkyl; R1′ and R2′ are each independently halogen, —OH, —NH2, C1-C4 alkyl, C1-C4 chloroalkyl, C1-C4 hydroxyalkyl, —O(C1-C4 alkyl), —NH(C1-C4 alkyl), C3-C6 cycloalkyl, —O(C3-C6 cycloalkyl), —NH(C3-C6 cycloalkyl), C3-C6 heterocycloalkyl, or —O(C3-C6 heterocycloalkyl); R3′ and R4′ are each independently hydrogen, deuterium, C1-C4 alkyl, C3-C6 cycloalkyl, or C3-C6 heterocycloalkyl, and j is 2, 3, 4, 5, 6, 7, 8, 9 or 10.
Preferably, in certain embodiments of the present invention, L is —(CH2)j—, wherein 1, 2, 3, 4 or 5 methylene groups in the —(CH2)j— are optionally replaced by a group selected from —NH—, —NCH3—, —O—, —C(CH3)2—, —CHF—, —CHCF3—, —C(O)—, —C(O)O—, —OC(O)—, —C(O)NH—, —C(O)NCH3—, —NHC(O)—, —NCH3C(O)—, ethenylene, ethynylene, cyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene, oxiranylidene, oxetanylidene, oxolanylidene, oxanylidene, aziridinylidene, azetidinylidene, azolidinylidene, piperidinylidene, piperazinylidene, morpholinylidene, homomorpholinylidene, phenylene, pyrrolylidene, thienylidene, furylidene, imidazolylidene, pyrazolylidene, triazolylidene, tetrazolylidene, oxazolylidene, isoxazolylidene, thiazolylidene, isothiazolylidene, pyridinylidene, pyrimidinylidene, pyridazinylidene, pyrazinylidene,
wherein the group for the replacement is optionally substituted with 1 or more substituents selected from halogen, oxo, —NR3′R4′, —OR3′ and C1-C4 alkyl, wherein the alkyl is optionally substituted with 1 or more substituents selected from halogen, —OH and —NH2; R3′ and R4′ are each independently hydrogen, deuterium or C1-C4 alkyl, and j is 2, 3, 4, 5, 6, 7, 8, 9 or 10.
Preferably, in certain embodiments of the present invention, L is —(CH2)j—, wherein 1, 2, 3, 4 or 5 methylene groups in the —(CH2)j— are optionally replaced by a group selected from —NH—, —NCH3—, —O—, —C(CH3)2—, —CHF—, —CHCF3—, —C(O)—, —C(O)O—, —OC(O)—, —C(O)NH—, —C(O)NCH3—, —NHC(O)—, —NCH3C(O)—, cyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene, oxiranylidene, oxetanylidene, oxolanylidene, oxanylidene, aziridinylidene, azetidinylidene, azolidinylidene, piperidinylidene, piperazinylidene, morpholinylidene, homomorpholinylidene,
wherein the group for the replacement is optionally substituted with 1 or more substituents selected from F, Cl, oxo, —NR3′R4′, —OR3′ and C1-C4 alkyl, wherein the alkyl is optionally substituted with 1 or more substituents selected from halogen, —OH and —NH2; R3′ and R4′ are each independently hydrogen, deuterium, methyl, ethyl or propyl, and j is 3, 4, 5, 6, 7, 8, 9 or 10.
Preferably, in certain embodiments of the present invention, L is —(CH2)j—, wherein 1, 2, 3, 4 or 5 methylene groups in the —(CH2)j— are optionally replaced by a group selected from —O—, —NH—, —NCH3—, —C(O)—, —C(O)NH—, —NHC(O)—, —NCH3C(O)—, —C(O)NCH3—, cyclohexylene, aziridinylidene, azetidinylidene, azolidinylidene, piperidinylidene, piperazinylidene,
and j is 4, 5, 6, 7, 8, 9 or 10.
Preferably, in certain embodiments of the present invention, L is selected from —(CH2)j-1—C(O)—, wherein the methylene in the —(CH2)j-1—C(O)— is as defined in L described above, and is optionally replaced by 1 or more groups, and j is as defined in L described above.
Preferably, in certain embodiments of the present invention, L is selected from
In certain embodiments of the present invention, L is any one of the following structures:
Preferably, in certain embodiments of the present invention, L is LA,
wherein in LA
ring A is a bond, C3-C12 cycloalkylene (e.g.,
wherein end a is linked to S, and end b is linked to X′″) or 3- to 12-membered heterocycloalkylene containing 1-2 heteroatoms selected from N, O and S (e.g.,
wherein end a is linked to S, and end b is linked to X′″), and the cycloalkylene and heterocycloalkylene are optionally substituted with substituents selected from halogen, oxo, cyano, amino, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 hydroxyalkyl and —O—(C1-C6 alkyl);
ring B is a bond, C3-C12 cycloalkylene (e.g.,
wherein end c is linked to X′″, and end d is linked to L3) or 3- to 12-membered heterocycloalkylene containing 1-2 heteroatoms selected from N, O and S (e.g.,
wherein end c is linked to X′″, and end d is linked to L3), and the cycloalkylene and heterocycloalkylene are optionally substituted with substituents selected from halogen, oxo, cyano, amino, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 hydroxyalkyl and —O—(C1-C6 alkyl);
ring C is C3-C12 cycloalkylene (e.g.,
wherein end e is linked to L3, and end f is linked to X″) or 3- to 12-membered heterocycloalkylene containing 1-2 heteroatoms selected from N, O and S (e.g.,
wherein end e is linked to L3, and end f is linked to X″), and the cycloalkylene and heterocycloalkylene are optionally substituted with substituents selected from halogen, oxo, cyano, amino, hydroxy, C1-C6 alkyl, C1-C6 haloalkyl, C1-C6 hydroxyalkyl and —O—(C1-C6 alkyl);
In some embodiments, in LA:
In certain embodiments of the present invention, in LA, L3 is —(CH2)k, wherein one or two methylene groups in L3 are optionally replaced by a group selected from —O—, —NH—, —C≡C—, —N(C1-C6 alkyl)-, —N(C1-C6 haloalkyl)-, —N(C1-C6 hydroxyalkyl)- and —N(C3-C8 cycloalkyl)-, and k is 0, 1, 2, 3, 4, 5, 6 or 7.
Preferably, in certain embodiments of the present invention, in LA, ring A is a bond.
Preferably, in certain embodiments of the present invention, in LA, ring A is 3- to 7-membered saturated or partially unsaturated cycloalkylene, 4- to 11-membered saturated or partially unsaturated spirocycloalkylene, 4- to 11-membered saturated or partially unsaturated fused cycloalkylene, 8- to 10-membered bicyclic saturated or partially unsaturated cycloalkylene, 4- to 7-membered saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated spiroheterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated fused heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur or 8- to 10-membered bicyclic saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur.
Preferably, in certain embodiments of the present invention, in LA, ring A is 3- to 6-membered saturated cycloalkylene or 4- to 7-membered saturated monocyclic heterocycloalkylene having 1 or 2 heteroatoms independently selected from nitrogen.
Preferably, in certain embodiments of the present invention, in LA, ring A is cyclohexylene (e.g., cyclohex-1,4-diyl, e.g., trans cyclohex-1,4-diyl), piperidinylidene (e.g., piperidine-1,4-diyl) or piperazinylidene (e.g., piperazine-1,4-diyl).
In certain embodiments of the present invention, in LA, ring A is
wherein end a is linked to S, and end b is linked to X′″.
In certain embodiments of the present invention, in LA, ring A is
wherein end a is linked to S, and end b is linked to X′″.
In certain embodiments of the present invention, in LA, ring A is
wherein end a is linked to S, and end b is linked to X′″.
Preferably, in certain embodiments of the present invention, in LA, ring B is a bond.
Preferably, in certain embodiments of the present invention, in LA, ring B is 3- to 7-membered saturated or partially unsaturated cycloalkylene, 4- to 11-membered saturated or partially unsaturated spirocycloalkylene, 4- to 11-membered saturated or partially unsaturated fused cycloalkylene, 8- to 10-membered bicyclic saturated or partially unsaturated cycloalkylene, 4- to 7-membered saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated spiroheterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated fused heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur or 8- to 10-membered bicyclic saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur.
Preferably, in certain embodiments of the present invention, in LA, ring B is 4- to 7-membered saturated monocyclic heterocycloalkylene containing 1 or 2 nitrogen heteroatoms or 7- to 11-membered spiroheterocycloalkylene or fused heterocycloalkylene containing 1 or 2 nitrogen heteroatoms.
Preferably, in certain embodiments of the present invention, in LA, ring B is piperidinylidene (e.g., piperidine-1,4-diyl) or piperazinylidene (e.g., piperazine-1,4-diyl).
In certain embodiments of the present invention, in LA, ring B is
wherein end c is linked to X′″, and end d is linked to L3.
Preferably, in certain embodiments of the present invention, ring B is
wherein end c is linked to X′″, and end d is linked to L3.
Preferably, in certain embodiments of the present invention, in LA, ring C is 3- to 7-membered saturated or partially unsaturated cycloalkylene, 4- to 11-membered saturated or partially unsaturated spirocycloalkylene, 4- to 11-membered saturated or partially unsaturated fused cycloalkylene, 8- to 10-membered bicyclic saturated or partially unsaturated cycloalkylene, 4- to 7-membered saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated spiroheterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur, 4- to 11-membered saturated or partially unsaturated fused heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur or 8- to 10-membered bicyclic saturated or partially unsaturated heterocycloalkylene having 1-2 heteroatoms independently selected from nitrogen, oxygen and sulfur.
Preferably, in certain embodiments of the present invention, in LA, ring C is 4- to 7-membered saturated monocyclic heterocycloalkylene containing 1 or 2 nitrogen heteroatoms or 7- to 11-membered spiroheterocycloalkylene or fused heterocycloalkylene containing 1 or 2 nitrogen heteroatoms.
Preferably, in certain embodiments of the present invention, in LA, ring C is piperidinylidene (e.g., piperidine-1,4-diyl), piperazinylidene (e.g., piperazine-1,4-diyl),
In certain embodiments of the present invention, in LA, ring C is
wherein end e is linked to L3, and end f is linked to X″.
In certain embodiments of the present invention, in LA, ring C is
wherein end e is linked to L3, and end f is linked to X″.
Preferably, in certain embodiments of the present invention, in LA, ring A is 3- to 6-membered saturated cycloalkylene; ring B is 4- to 7-membered saturated monocyclic heterocycloalkylene containing 1 or 2 nitrogen heteroatoms; ring C is 4- to 7-membered saturated monocyclic heterocycloalkylene containing 1 or 2 nitrogen heteroatoms or 7- to 11-membered spiroheterocycloalkylene or fused heterocycloalkylene containing 1 or 2 nitrogen heteroatoms; X″ is a bond or —C(O)—; L3 is —(CH2)k, and k is 1, 2, 3, 4 or 5 (e.g., k is 1, 2, 3 or 4).
Preferably, in certain embodiments of the present invention, in LA, X″ is a bond or —C(O)—.
Preferably, in certain embodiments of the present invention, in LA, X″ is a bond.
Preferably, in certain embodiments of the present invention, in LA, X″ is —C(O)—.
Preferably, in certain embodiments of the present invention, in LA, X′″ is a bond or —C(O)—.
Preferably, in certain embodiments of the present invention, in LA, X′″ is a bond.
Preferably, in certain embodiments of the present invention, in LA, X′″ is —C(O)—.
Preferably, in certain embodiments of the present invention, in LA, k is 1, 2, 3, 4 or 5.
Preferably, in certain embodiments of the present invention, in LA, L3 is —(CH2)k, wherein k is 1, 2, 3, 4 or 5.
Preferably, in certain embodiments of the present invention, in LA, L3 is —(CH2)k, wherein one or two methylene groups in L3 are optionally replaced by —O—, —NH—, —C≡C— or —N(C1-C6 alkyl)- (e.g., —N(CH3)—), and k is 1, 2, 3, 4 or 5.
Preferably, in certain embodiments of the present invention, in LA, k is 1, 2, 3, or 4. Preferably, in certain embodiments of the present invention, in LA, L3 is —(CH2)k—, wherein one or two of CH2 contained in L3 are each independently optionally replaced by —O—, —NH— or —N(C1-C6 alkyl)- (e.g., —N(CH3)—), or one of —CH2CH2— contained in L3 is optionally replaced by —C≡C—; k is 1, 2, 3 or 4.
Preferably, in certain embodiments of the present invention, in LA, L3 is —(CH2)k—, wherein one methylene group in L3 is optionally replaced by a group, which is —O—, —NH— or —N(C1-C6 alkyl)- (e.g., —N(CH3)—); k is 1, 2, 3 or 4.
Preferably, in certain embodiments of the present invention, LA is LA-1:
wherein ring A, ring B, ring C, L3 and X″ are as defined and described in LA.
In certain embodiments of the present invention, LA is LA-2:
wherein ring A is
wherein end a is linked to S, and end b is linked to X′″; 1, 2, 3 or 4 hydrogen atoms in ring A are optionally substituted with F;
wherein end c is linked to X′″, and end d is linked to L3; 1, 2, 3 or 4 hydrogen atoms in ring B are optionally substituted with F;
wherein end e is linked to L3, and end f is linked to X″; 1, 2, 3 or 4 hydrogen atoms in ring C are optionally substituted with F;
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, ring A is
wherein end a is linked to S, and end b is linked to X′″.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, ring A is
wherein end a is linked to S, and end b is linked to X′″.
Preferably, in certain embodiments of the present invention, ring B is
wherein end c is linked to X′″, and end d is linked to L3.
In certain embodiments of the present invention, in LA, LA-1 and LA-2, L3 is —(CH2)k—, wherein one or two of CH2 contained in L3 are each independently optionally replaced by —O—, —NH— or —N(CH3)—; k is 1, 2, 3 or 4.
In certain embodiments of the present invention, in LA, LA-1 and LA-2, L3 is —(CH2)k—, wherein one or two of CH2 contained in L3 are each independently optionally replaced by —O—; k is 1, 2, 3 or 4.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, k is 3 or 4, preferably 4.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, L3 is —(CH2)k—, wherein one of CH2 contained in L3 is optionally replaced by —O—; k is 4.
In certain embodiments of the present invention, in LA, LA-1 and LA-2, L3 may be —CH2—, —(CH2)2—, —(CH2)3—, —(CH2)4—,
wherein end k″ is linked to ring B, and end k′ is linked to ring C.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, L3 may be —(CH2)4—,
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-2, ring C is,
wherein end e is linked to L3, and end f is linked to X″.
In certain embodiments of the present invention, LA-2 is any one of the following structures:
Preferably, in certain embodiments of the present invention, LA-2 is any one of the following structures:
In certain embodiments of the present invention, LA is LA-3:
a wherein end a is linked to S, and end b is linked to X′″; 1, 2, 3 or 4 hydrogen atoms in ring A are optionally substituted with F;
wherein end c is linked to X′″, and end d is linked to L3; 1, 2, 3 or 4 hydrogen atoms in ring B are optionally substituted with F;
wherein end e is linked to L3, and end f is linked to X″; 1, 2, 3 or 4 hydrogen atoms in ring C are optionally substituted with F;
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-3, ring A is
wherein end a is linked to S, and end b is linked to X′″.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-3, ring B is
wherein end c is linked to X′″, and end d is linked to L3.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-3, L3 is —(CH2)k—, wherein k is 1 or 2.
Preferably, in certain embodiments of the present invention, in LA, LA-1 and LA-3, ring C is
wherein end e is linked to L3, and end f is linked to X″.
In certain embodiments of the present invention, LA-3 is any one of the following structures:
Preferably, in certain embodiments of the present invention, LA-3 is any one of the following structures:
In certain embodiments of the present invention LA is LA-4:
a wherein end a is linked to S, and end b is linked to X′″; 1, 2, 3 or 4 hydrogen atoms in ring A are optionally substituted with F;
wherein end e is linked to L3, and end f is linked to X″; 1, 2, 3 or 4 hydrogen atoms in ring C are optionally substituted with F;
In certain embodiments of the present invention, in LA-4, ring A is
herein end a is linked to S, and end b is linked to X′″.
In certain embodiments of the present invention, in LA-4, ring A is
wherein end a is linked to S, and end b is linked to X′″.
In certain embodiments of the present invention, in LA-4, X′″ is —C(O)NCH3—.
In certain embodiments of the present invention, in LA-4, L3 is —(CH2)k—.
In certain embodiments of the present invention, in LA-4, k is 1, 2 or 3, e.g., 3.
In certain embodiments of the present invention, in LA-4, ring C is
wherein end e is linked to L3, and end f is linked to X″.
In certain embodiments of the present invention, in LA-4, ring C is
wherein end e is linked to L3, and end f is linked to X″.
In certain embodiments of the present invention, LA-4 is any one of the following structures:
Preferably, in certain embodiments of the present invention, LA is
Preferably, in certain embodiments of the present invention, E is a small molecule ligand of CRBN, VHL, XAIP or MDM2 E3 ubiquitin ligases.
Preferably, in certain embodiments of the present invention, E is E1, E1a, E1b or E1c
Preferably, in certain embodiments of the present invention, E is E2
wherein in E2,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E3:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E4:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E5:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E6, E6a, E6b, E6c, E6d, E6e, E6f, E6g, E6h or E6i:
Ring A, X, X1, X2, Y100, R1, R3, R3′, R4, R5, t, m and n are as defined in WO2017/007612 and US2018/0134684, which are incorporated herein by reference in their entirety.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E7, E7b, E7c, E7d or E7e:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E8, E8a, E8b, E8c, E8d, E8e, E8f, E8g, E8h, E8i, E8j, E8k, E8l, E8m, E8n, E8o, E8p, E8q, E8s, E8t, E8u, E8v, E8w, E8x, E8y, E8z, E8aa, E8bb or E8cc:
In one preferred embodiment of the compound of formula I, the stereoisomer, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E9a, E9b, E9c, E9d, E9e, E9f, E9g, E9h, E9i, E9j, E9k, E9l, E9m, E9n, E9o, E9p, E9q, E9r or E9s:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E10, E10a or E10b:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E11:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E12:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E13:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E14, E14a, E14b, E14c, E14d or E14e:
In some embodiments, in E14, E14a, E14b, E14c, E14d and E14e, Ar, R1, R2, R3, R4, R5, R6, R7, R8, A, L, x, y and are as defined and described in WO2017/161119 (incorporated herein by reference in its entirety).
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E15, E15a, E15b, E15c, E15d, E15e, E15f, E15g or E15h:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E16, E16a or E16b:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E17, E17a, E17b or E17c:
or R1*, wherein X is O or N, and when X is O, R2 is absent;
In some embodiments, in E17, E17a, E17b and E17c, R1, R4, R10, R11, R14, R16, W1, W2, X, and n″ are as defined in WO2018/237026, which is incorporated herein by reference in its entirety.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E18, E18a, E18b, E18c, E18d, E18e, E18f, E18g, E18h, E18i, E18j, E18k, E18l or E18m:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E19, E19a, E19b, E19c, E19d, E19e, E19f, E19g, E19h or E19i:
In some embodiments, in E19, E19a, E19b, E19c, E19d, E19e, E19f, E19g, E19h and E19i, A1, A2, A3, Z, G and R5 are as defined and described in WO2017/176958 (incorporated herein by reference in its entirety).
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E20, E20a, E20b, E20c, E20d, E20e or E20f:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E21:
In certain embodiments of the present invention, 1 or 2 of Q1, Q2, Q3, Q4 and Q5 in E21 are N, and the rest are each independently CR3″.
In certain embodiments of the present invention, Q1, Q2, Q3, Q4 and Q5 in E21 are each independently CR3″.
In certain embodiments of the present invention, X″ in E21 is N.
In certain embodiments of the present invention, X″ in E21 is C.
In certain embodiments of the present invention, Y″ in E21 is N.
In certain embodiments of the present invention, each R1″ in E21 is independently hydrogen, deuterium, —F, —Cl or C1-C6 alkyl, and the alkyl is optionally substituted with 1-3 halogens; preferably, R1″ is hydrogen.
In certain embodiments of the present invention, R2″ in E21 is hydrogen or C1-C6 alkyl, and the alkyl is optionally substituted with 1-3 halogens; preferably, R2″ is hydrogen.
In certain embodiments of the present invention, each R3″ in E21 is independently hydrogen, deuterium, halogen, —O(C1-C6 alkyl) or C1-C6 alkyl, and the alkyl is optionally substituted with 1-3 halogens; preferably, each R3″ is independently hydrogen, deuterium, F, Cl, methyl, methoxy, ethoxy, trifluoromethoxy, 2-hydroxypropan-2-yl or trifluoromethyl.
In certain embodiments of the present invention, m in E21 is 2.
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, in E21 and E21-1a to 1j, R3″ is hydrogen, halogen (e.g., F or Cl), C1-C6 alkyl or —O(C1-C6 alkyl) (e.g., —OCH3), and the C1-C6 alkyl and the —O(C1-C6 alkyl) are optionally substituted with 1-3 halogens (e.g., F).
In certain embodiments of the present invention, in E21 and E21-1a to 1j, R3″ is hydrogen, F, Cl, CF3, - OCF3 or —OCH3.
In certain preferred embodiments of the present invention, in E21-1h, R3″ is halogen (e.g., F or Cl) or —O(C1-C6 alkyl) (e.g., —OCH3).
In certain embodiments of the present invention, E21-1h is any one of the following structures:
In certain preferred embodiments of the present invention, E21-1h is any one of the following structures:
In certain embodiments of the present invention, E21-1i is the following structure:
In certain embodiments of the present invention, E21-1j is any one of the following structures:
In certain embodiments of the present invention, E21 is selected from
In certain embodiments of the present invention, E21 is selected from
In certain preferred embodiments of the present invention, E21 has a structure shown in the following formula,
In certain preferred embodiments of the present invention, E21 has a structure shown in the following formula,
In certain preferred embodiments of the present invention, E21 has a structure shown in the following formula,
In certain preferred embodiments of the present invention, E21 has a structure shown in the following formula,
In certain preferred embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 has a structure shown in the following formula,
In certain embodiments of the present invention, E21 is selected from
In certain embodiments of the present invention, E21 is selected from
Preferably, in certain embodiments of the present invention, E21 is any one of the following structures:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E22:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E23, E23a, E23b, E23c, E23d or E23e:
and the aryl, the heteroaryl and the heterocyclyl may be optionally substituted;
and the heterocycloalkyl, the alkoxy, the heteroaryl and the aryl may be optionally substituted;
In some embodiments, in E23, E23a, E23b, E23c, E23d and E23e, R1′, R2′, R3′, R5, R6, R7, R9, R10, R11, R14, R15, R16, R17, R23, R25, E, M, o, X, Y, Z1, Z2, Z3 and Z4 are as defined and described in US 2016/0272639, which is incorporated herein by reference in its entirety.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E23a:
or optionally substituted phenyl;
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E23c
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E23d:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E23d-1
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is the specific compound according to Example 1 to Example 208 in CN108601764 (incorporated herein by reference in its entirety).
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E24:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E25 or E25a:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E26, E26a or E26b,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E27, E27a, E27b, E27c, E27d, E27e, E27f or E27g:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E28, E28a, E28b, E28c, E28d, E28e, E28f, E28g, E28h or E28i, (MDM2)
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E29:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E30, E30a, E30b, E30c or E30d:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E31
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E32,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E33,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E34,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E35,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E36,
is selected from hydrogen, alkyl and aryl;
is nitrogen-containing heteroaryl.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E37,
is selected from H, alkyl and aryl;
is selected from CH2 and O; and
in and is nitrogen-containing heteroaryl.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E38,
is selected from H, alkyl and aryl;
is selected from CH2 and O; and
in and is nitrogen-containing heteroaryl; and
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E39,
is 5- or 6-membered nitrogen-containing heteroaryl, more preferably 5-membered nitrogen-containing heteroaryl, and most preferably thiazole; and
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E40,
R is selected from halogen (e.g., fluorine), cyano,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E41,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E42,
is 6-membered nitrogen-containing heteroaryl.
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E43,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E44, E44a or E44b,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E45,
is a 5-membered heterocycle selected from the following:
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is E46 or E46a,
In one preferred embodiment of the compound of formula I, the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention, E is:
Preferably, in certain embodiments of the present invention, the compound of formula I is
In certain embodiments, the deuterid has a structure formed by replacing 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 H with deuterium in a compound structure (for example, L, e.g., ring A, ring B, L3 and ring C in LA, LA-1, LA-2, LA-3 and LA-4). In certain embodiments, the deuterated LA structure is
The present invention provides a method for preparing the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof.
The present invention provides a pharmaceutical composition comprising a therapeutically effective amount of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, diluent, or excipient.
The present invention provides a method for degrading SOS1 protein, comprising contacting the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof with the SOS1 protein.
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof as a medicament for the treatment or prevention of an SOS1-mediated disease or disorder.
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof as a medicament for the treatment or prevention of a disease or disorder (cancer) caused by the interaction of SOS1 with Ras (e.g., KRAS) or SOS1 with Rac (e.g., KRAS).
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for the treatment or prevention of an SOS1-mediated disease or disorder (e.g., cancer).
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for the treatment or prevention of a disease or disorder (e.g., cancer) caused by the interaction of SOS1 with Ras (e.g., KRAS) or SOS1 with Rac (e.g., KRAS).
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for the treatment or prevention of cancer.
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for the treatment or prevention of pancreatic cancer, lung cancer, colorectal cancer, cholangiocarcinoma, multiple myeloma, melanoma, uterine cancer, endometrial cancer, thyroid cancer, acute myeloid leukemia, bladder cancer, urothelial cancer, gastric cancer, cervical cancer, head and neck squamous cell carcinoma, diffuse large B-cell lymphoma, esophageal cancer, chronic lymphocytic leukemia, hepatocellular carcinoma, breast cancer, ovarian cancer, prostate cancer, glioblastoma, renal cancer and sarcoma.
The present invention provides use of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof in preparing a medicament for the treatment or prevention of neurofibromatosis type 1 (NF1), Noonan syndrome (NS), Noonan syndrome with multiple lentigines (NSML), capillary malformation-arteriovenous malformation syndrome (CM-AVM), Costello syndrome (CS), cardio-facio-cutaneous syndrome (CFC), Legius syndrome and hereditary gingival fibromatosis.
The present invention provides a method for treating or preventing an SOS1-mediated disease or disorder, comprising administering to a patient in need thereof a therapeutically effective amount of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof.
The present invention provides a method for treating or preventing a disease or disorder (e.g., cancer) regulated by the interaction of SOS1 with Ras (e.g., KRAS) or SOS1 with Rac (e.g., KRAS), comprising administering to a patient in need thereof a therapeutically effective amount of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof.
In certain embodiments of the present invention, the cancer may be selected from
heart: sarcoma (angiosarcoma, fibrosarcoma, rhabdomyosarcoma and liposarcoma), myxoma, rhabdomyoma, fibroma, lipoma and teratoma;
lung: bronchial cancer (squamous cell carcinoma, undifferentiated small cell carcinoma, undifferentiated large cell carcinoma and adenocarcinoma), bronchioloalveolar carcinoma (bronchiolar carcinoma), bronchial adenoma, sarcoma, lymphoma, chondroma, hamartoma and mesothelioma;
gastrointestinal tract and esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma and lymphoma), stomach (cancer, lymphoma and leiomyosarcoma), pancreas (ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, carcinoid tumor and vipoma), small intestine (adenocarcinoma, lymphoma, carcinoid tumor, Kaposi's sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma and fibroma) and large intestine (adenocarcinoma, tubular adenoma, villous adenoma, hamartoma and leiomyoma);
genitourinary system: kidney (adenocarcinoma, Wilms' tumor, lymphoma and leukemia), bladder and urinary tract (squamous cell carcinoma, transitional cell carcinoma and adenocarcinoma), prostate (adenocarcinoma and sarcoma) and testis (seminoma, teratoma, embryonal carcinoma, teratoma, choriocarcinoma, sarcoma, interstitial cell carcinoma, fibroma, fibroadenoma, adenomatoid tumor and lipoma);
liver: liver cancer (hepatocellular carcinoma), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma and hemangioma;
biliary tract: gallbladder cancer, ampullary cancer and cholangiocarcinoma;
bone: osteosarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, Ewing's sarcoma, malignant lymphoma (reticulosarcoma), multiple myeloma, malignant giant cell tumor, chordoma, chondroma (osteochondral exostosis), benign chondroma, chondroblastoma, chondromyxoid fibroma, osteoid osteoma and giant cell tumor;
nervous system: skull (osteoma, hemangioma, granuloma, xanthoma and osteitis deformans), meninges (meningioma, meningosarcoma and glioma), brain (astrocytoma, medulloblastoma, glioma, ependymoma, germinal cell tumor (pinealoma), glioblastoma multiforme, oligodendroglioblastoma, neurilemmoma, retinoblastoma and congenital tumor), spinal neurofibroma, meningioma, glioma and sarcoma);
gynaecology: uterus (endometrial cancer (serous cystadenocarcinoma, mucinous cystadenocarcinoma and unclassified cancer), granulosa theca cell tumor, Sertoli-Leydig cell tumor, dysgerminoma and malignant teratoma), vulva (squamous cell carcinoma, intraepithelial carcinoma, adenocarcinoma, fibrosarcoma and melanoma), vagina (clear cell carcinoma, squamous cell carcinoma, botryoid sarcoma (embryonal rhabdomyosarcoma) and fallopian tube (cancer);
hematology: blood (myeloid leukemia (acute and chronic), acute lymphoblastic leukemia, chronic lymphocytic leukemia, myeloproliferative disease, multiple myeloma and myelodysplastic syndrome), Hodgkin's disease and non-Hodgkin's lymphoma (malignant lymphoma);
skin: malignant melanoma, basal cell carcinoma, squamous cell carcinoma, Kaposi's sarcoma, dysplastic nevus, lipoma, hemangioma, dermatofibroma, keloid and psoriasis; and adrenal gland: neuroblastoma.
In certain embodiments of the present invention, the cancer may be selected from pancreatic cancer, lung cancer, colorectal cancer, cholangiocarcinoma, multiple myeloma, melanoma, uterine cancer, endometrial cancer, thyroid cancer, acute myeloid leukemia, bladder cancer, urothelial cancer, gastric cancer, cervical cancer, head and neck squamous cell carcinoma, diffuse large B-cell lymphoma, esophageal cancer, chronic lymphocytic leukemia, hepatocellular carcinoma, breast cancer, ovarian cancer, prostate cancer, glioblastoma, renal cancer and sarcoma.
The present invention also provides use of the compound of formula I described above, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition described above in preparing a medicament for the treatment and/or prevention of a KRAS-mediated disease or disorder.
The present invention also provides a method for treating and/or preventing a KRAS-mediated disease or disorder, comprising administering to a patient in need thereof a therapeutically effective amount of the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof or the pharmaceutical composition thereof.
In certain embodiments of the present invention, the KRAS may be a KRAS mutant. The KRAS mutant is preferably one or more of KRAS G12C, KRAS G12D, KRAS G13D and KRAS G12V.
The present invention provides a method for preparing the compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof:
and ring B contains an NH group, the preparation method is
subjecting INT-A and INT-B to a reductive amination reaction to give the target compound, wherein the reductive amination reductant includes, but is not limited to, Pd/C, sodium borohydride, sodium cyanoborohydride, borane and sodium triacetoxyborohydride, wherein the
refers to ring B containing an NH group; R2, R3, R4, R5, R6 and R7 are as defined and described in S6b;
refers to L3 containing an aldehyde group; ring A, ring B, ring C and X″ are as defined in LA; E is as defined and described in the present invention, and E is preferably E7, E7a, E7b, E7c, E7d, E7e, E7a-1, E7a-2, E21, E21-1a, E21-1b, E21-1c, E21-1d, E21-1e, E21-1f, E21-1g, E21-1h, E21-1aa, E21-1bb, E21-1cc, E21-1dd, E21-1ee, E21-1ff, E21-1gg or E21-1hh.
When S is S6c
ring C contains an NH group, and X″ is C(O), the preparation method is
subjecting INT-C and INT-D to a substitution reaction under an alkaline condition to give the target compound, wherein the base includes, but is not limited to, triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate and sodium bicarbonate, wherein the
refers to ring C containing an NH group; P100 is pentafluorophenyl or p-nitrophenyl; R2, R3, R4, R5, R6 and R7 are as defined and described in S6b; ring A, ring B, ring C and L3 are as defined in LA; E is as defined and described in the present invention, and E is preferably E7, E7a, E7b, E7c, E7d, E7e, E7a-1, E7a-2, E21, E21-1a, E21-1b, E21-1c, E21-1d, E21-1e, E21-1f, E21-1g, E21-1h, E21-1aa, E21-1bb, E21-1cc, E21-1dd, E21-1ee, E21-1ff, E21-1gg or E21-1hh.
ring C contains an NH group, and X″ is C(O), the preparation method is
subjecting INT-E and INT-D to a substitution reaction under an alkaline condition to give the target compound, wherein the base includes, but is not limited to, triethylamine, N,N-diisopropylethylamine, potassium carbonate, sodium carbonate and sodium bicarbonate, wherein the
refers to ring C containing an NH group; P100 is pentafluorophenyl or p-nitrophenyl; R2, R3, R4, R5, R6 and R7 are as defined and described in S6d; ring A, ring B, ring C and L3 are as defined in LA; E is as defined and described in the present invention, and E is preferably E7, E7a, E7b, E7c, E7d, E7e, E7a-1, E7a-2, E21, E21-1a, E21-1b, E21-1c, E21-1d, E21-1e, E21-1f, E21-1g, E21-1h, E21-1aa, E21-1bb, E21-1cc, E21-1dd, E21-1ee, E21-1ff, E21-1gg or E21-1hh.
Detailed description: Unless otherwise stated, the following terms used in the specification and claims have the following meanings.
“Alkyl” refers to a saturated aliphatic hydrocarbon group including linear or branched alkyl; C1-C8 alkyl refers to alkyl containing 1-8 carbon atoms, e.g., methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, tert-butyl, sec-butyl, n-pentyl, 1,1-dimethylpropyl, 1,2-dimethylpropyl, 2,2-dimethylpropyl, 1-ethylpropyl, 2-methylbutyl, 3-methylbutyl, n-hexyl, 1-ethyl-2-methylpropyl, 1,1,2-trimethylpropyl, 1,1-dimethylbutyl, 1,2-dimethylbutyl, 2,2-dimethylbutyl, 1,3-dimethylbutyl, 2-ethylbutyl, 2-methylpentyl, 3-methylpentyl, 4-methylpentyl, 2,3-dimethylbutyl, n-heptyl, 2-methylhexyl, 3-methylhexyl, 4-methylhexyl, 5-methylhexyl, 2,3-dimethylpentyl, 2,4-dimethylpentyl, 2,2-dimethylpentyl, 3,3-dimethylpentyl, 2-ethylpentyl, 3-ethylpentyl, n-octyl, 2,3-dimethylhexyl, 2,4-dimethylhexyl, 2,5-dimethylhexyl, 2,2-dimethylhexyl, 3,3-dimethylhexyl, 4,4-dimethylhexyl, 2-ethylhexyl, 3-ethylhexyl, 4-ethylhexyl, 2-methyl-2-ethylpentyl, 2-methyl-3-ethylpentyl or various branched isomers thereof; preferably, C1-C6 alkyl; more preferably, C1-C4 alkyl. The alkyl may be substituted or unsubstituted. In some embodiments, the alkyl is C1, C2, C3, C4, C5, C6, C7 or C8 alkyl.
“Cycloalkyl” refers to a monocyclic or polycyclic hydrocarbon substituent that is saturated or partially unsaturated; “C3-C11 cycloalkyl” refers to cycloalkyl containing 3 to 11 carbon atoms; “C3-C8 cycloalkyl” refers to cycloalkyl containing 3 to 8 carbon atoms; “C5-C10 cycloalkyl” refers to cycloalkyl containing 5 to 10 carbon atoms.
Non-limiting examples of monocyclic cycloalkyl include cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cyclohexadienyl, cycloheptyl, cycloheptatrienyl, cyclooctyl and the like, preferably cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl, preferably C3-C8 cycloalkyl, more preferably C3-C6 cycloalkyl.
Polycyclic cycloalkyl includes spiro cycloalkyl, fused cycloalkyl and bridged cycloalkyl. “Spiro cycloalkyl” refers to a polycyclic group in which a carbon atom (called spiro atom) is shared among monocyclic rings, wherein the rings may contain one or more double bonds, but none of the rings has a fully conjugated π-electron system. According to the number of the spiro atoms shared among the rings, the spiro cycloalkyl may be monospiro cycloalkyl, bispiro cycloalkyl or polyspiro cycloalkyl, preferably 7- to 12-membered bispiro cycloalkyl. Non-limiting examples of spiro cycloalkyl include:
“Fused cycloalkyl” refers to an all-carbon polycyclic group in which each ring shares a pair of adjacent carbon atoms with the other rings in the system, wherein one or more of the rings may contain one or more double bonds, but none of the rings has a fully conjugated π-electron system. According to the number of the formed rings, the fused cycloalkyl may be bicyclic, tricyclic, tetracyclic or polycyclic, preferably bicyclic fused cycloalkyl. Non-limiting examples of fused cycloalkyl include:
“Bridged cycloalkyl” refers to an all-carbon polycyclic group in which any two rings share two carbon atoms that are not directly connected to each other, wherein the rings may contain one or more double bonds, but none of the rings has a fully conjugated π-electron system. According to the number of the formed rings, the bridged cycloalkyl may be bicyclic, tricyclic, tetracyclic or polycyclic. Non-limiting examples of bridged cycloalkyl include:
The cycloalkyl ring may be fused to an aryl, heteroaryl or heterocycloalkyl ring, wherein the ring connected to the parent structure is a cycloalkyl ring; non-limiting examples include indanyl, tetrahydronaphthyl, benzocycloheptyl, and the like. The cycloalkyl may be optionally substituted or unsubstituted.
In some embodiments, the cycloalkyl is C3, C4, C5, C6, C7, C8, C9, C10, C11 or C12 monocyclic or polycyclic (e.g., spiro, fused or bridged) cycloalkyl.
“Heterocycloalkyl” refers to a monocyclic or polycyclic hydrocarbon substituent that is saturated or partially unsaturated, wherein one or more (e.g., 2, 3, 4 or 5) of the ring atoms are selected from nitrogen, oxygen and S(O)r (wherein r is an integer of 0, 1 or 2), excluding ring portions of —O—O—, —O—S— or —S—S—, and the remaining ring atoms are carbon atoms. “3- to 11-membered heterocycloalkyl” refers to a cyclic group containing 3 to 11 ring atoms, “5- to 10-membered heterocycloalkyl” refers to a cyclic group containing 5 to 10 ring atoms, and “3- to 8-membered heterocycloalkyl” refers to a cyclic group containing 3 to 8 ring atoms, preferably “3- to 11-membered heterocycloalkyl” containing 1-2 heteroatoms selected from N, O and S, and more preferably “3- to 11-membered heterocycloalkyl” containing 1 or 2 N atoms.
Monocyclic heterocycloalkyl is preferably 3- to 8-membered monocyclic heterocycloalkyl containing 1-2 N heteroatoms; non-limiting examples of monocyclic heterocycloalkyl include pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl, homopiperazinyl and the like, preferably piperidinyl and piperazinyl.
Polycyclic heterocycloalkyl includes spiro heterocyclyl, fused heterocyclyl and bridged heterocycloalkyl. “Spiroheterocycloalkyl” refers to a polycyclic heterocycloalkyl group in which an atom (called spiro atom) is shared among monocyclic rings, wherein one or more of the ring atoms are selected from nitrogen, oxygen and S(O)r (wherein r is an integer of 0, 1 or 2), and the remaining ring atoms are carbon atoms. The rings may contain one or more double bonds, but none of the rings has a fully conjugated it-electron system. According to the number of the spiro atoms shared among the rings, the spiroheterocycloalkyl may be monospiroheterocycloalkyl, bispiroheterocycloalkyl or polyspiroheterocycloalkyl, preferably saturated “3- to 11-membered bispiroheterocycloalkyl” containing 1-2 heteroatoms selected from N, O and S, and more preferably saturated “7- to 11-membered bispiroheterocycloalkyl” containing 1 or 2 N atoms. Non-limiting examples of spiroheterocycloalkyl include:
“Fused heterocycloalkyl” refers to a polycyclic heterocycloalkyl group in which each ring shares a pair of adjacent atoms with the other rings in the system, wherein one or more of the rings may contain one or more double bonds, but none of the rings has a fully conjugated π-electron system, wherein one or more of the ring atoms are selected from nitrogen, oxygen and S(O)r (wherein r is an integer of 0, 1 or 2), and the remaining ring atoms are carbon atoms. According to the number of the formed rings, the fused heterocycloalkyl may be bicyclic, tricyclic, tetracyclic or polycyclic, preferably “3- to 11-membered bicyclic fused heterocycloalkyl” containing 1-3 heteroatoms selected from N, O and S, and more preferably saturated “3- to 11-membered bicyclic fused heterocycloalkyl” containing 1 or 2 N atoms. Non-limiting examples of fused heterocycloalkyl include:
“Bridged heterocycloalkyl” refers to a polycyclic heterocycloalkyl group in which any two rings share two atoms that are not directly connected to each other, wherein the rings may contain one or more double bonds, but none of the rings has a fully conjugated 7-electron system, wherein one or more of the ring atoms are selected from nitrogen, oxygen and S(O)r (wherein r is an integer of 0, 1 or 2), and the remaining ring atoms are carbon atoms. According to the number of the formed rings, the bridged heterocycloalkyl may be bicyclic, tricyclic, tetracyclic or polycyclic; non-limiting examples of bridged heterocycloalkyl include:
The heterocycloalkyl ring may be fused to an aryl, heteroaryl or cycloalkyl ring, wherein the ring connected to the parent structure is a heterocycloalkyl ring; non-limiting examples include:
and the heterocycloalkyl may be optionally substituted or unsubstituted.
In some embodiments, the heterocycloalkyl is 3-, 4-, 5-, 6-, 7-, 8-, 9-, 10-, 11- or 12-membered monocyclic or polycyclic (e.g., spiro, fused or bridged) heterocycloalkyl, wherein the number of the heteroatoms may be 1, 2, 3, 4 or 5, each heteroatom being independently nitrogen, oxygen or S(O)r (wherein r is an integer of 0, 1 or 2).
“Aryl” refers to an all-carbon monocyclic or fused polycyclic group (i.e., rings that share a pair of adjacent carbon atoms) and a polycyclic group having a conjugated π-electron system, and “6- to 10-membered aryl” refers to all-carbon aryl containing 6-10 carbons, such as phenyl and naphthyl, preferably phenyl. The aryl ring may be fused to a heteroaryl, heterocycloalkyl or cycloalkyl ring, wherein the ring connected to the parent structure is the aryl ring; non-limiting examples include:
and the aryl may be optionally substituted or unsubstituted. In some embodiments, the aryl is 6- to 10-membered aryl.
“Heteroaryl” refers to a heteroaromatic system containing 1 to 4 heteroatoms, and the heteroatoms include nitrogen, oxygen or S(O)r (where r is an integer of 0, 1 or 2); 5- to 6-membered heteroaryl refers to a heteroaromatic system containing 5-6 ring atoms, and 5- to 10-membered heteroaryl refers to a heteroaromatic system containing 5-10 ring atoms, preferably 5- to 6-membered heteroaryl, and more preferably 5- to 6-membered heteroaryl containing 1 or 2 N atoms; non-limiting examples include furyl, thienyl, pyridyl, pyrrolyl, N-alkylpyrrolyl, pyrimidinyl, pyrazinyl, pyrazole, imidazolyl, triazolyl, tetrazolyl and the like, preferably pyridyl. The heteroaryl ring may be fused to an aryl, heterocycloalkyl or cycloalkyl ring, wherein the ring connected to the parent structure is the heteroaryl ring; non-limiting examples include:
and the heteroaryl may be optionally substituted or unsubstituted. In some embodiments, the heteroaryl is 5-, 6-, 7-, 8-, 9- or 10-membered heteroaryl, wherein the number of the heteroatoms may be 1, 2, 3, 4 or 5, each heteroatom being independently nitrogen, oxygen or S.
“Alkenyl” refers to alkyl as defined above consisting of at least two carbon atoms and at least one carbon-carbon double bond, and “C2-8 alkenyl” refers to linear or branched alkenyl containing 2-8 carbons, including but not limited to ethenyl, 1-propenyl, 2-propenyl, 1-, 2- or 3-butenyl and the like, preferably “C2-6 alkenyl”, and more preferably “C2-4 alkenyl”. The alkenyl may be substituted or unsubstituted. In some embodiments, the alkenyl is C2, C3, C4, C5, C6, C7 or C8 alkenyl.
“Alkynyl” refers to alkyl as defined above consisting of at least two carbon atoms and at least one carbon-carbon triple bond, and “C2-8 alkynyl” refers to linear or branched alkynyl containing 2-8 carbons, including but not limited to ethynyl, 1-propynyl, 2-propynyl, 1-, 2- or 3-butynyl, preferably “C2-6 alkynyl”, and more preferably “C2-4 alkynyl”. The alkynyl may be substituted or unsubstituted. In some embodiments, the alkynyl is C2, C3, C4, C5, C6, C7 or C8 alkynyl.
“-ylene” refers to a divalent group. For example, alkylene refers to divalent alkyl, alkenylene refers to divalent alkenyl, alkynylene refers to divalent alkynyl, cycloalkylene refers to divalent cycloalkyl, heterocycloalkylene refers to divalent heterocycloalkyl, arylene refers to divalent aryl, and heteroarylene refers to divalent heteroaryl, wherein the alkyl, the alkenyl, the alkynyl, the cycloalkyl, the heterocycloalkyl, the aryl and the heteroaryl are as defined above, and the “-ylene” may be optionally substituted or unsubstituted.
“Haloalkyl” refers to alkyl optionally substituted with one or more fluorine, chlorine, bromine or iodine atoms, wherein the alkyl is as defined above, and non-limiting examples include difluoromethyl, dichloromethyl, dibromomethyl, trifluoromethyl, trichloromethyl, tribromomethyl and the like.
“Hydroxyalkyl” refers to alkyl optionally substituted with one or more —OH, wherein the alkyl is as defined above, and non-limiting examples include hydroxymethyl, hydroxyethyl, hydroxypropyl and hydroxyisopropyl.
“Alkoxy” refers to —O-alkyl, wherein the alkyl is as defined above, and non-limiting examples include methoxy, ethoxy, isopropoxy, tert-butoxy and the like.
“Cycloalkoxy” refers to —O-cycloalkyl, wherein the cycloalkyl is as defined above, and non-limiting examples include cyclopropaneoxy, cyclobutaneoxy, cyclopentanyloxy, cyclohexyloxy and the like.
“Heterocycloalkoxy” refers to —O-heterocycloalkyl, wherein the heterocycloalkyl is as defined above, and non-limiting examples include azetidinyloxy, azacyclopentyloxy, piperidinyloxy, piperazinyloxy, oxolanyloxy, oxanyloxy and the like.
“—C(O)C—C3 alkyl” refers to —C(O)—CH3, —C(O)—CH2CH3 and the like.
“Cyano” refers to —CN.
“Hydroxy” refers to —OH.
“Sulfonyl” refers to —S(O)2—.
“Carboxyl” or “carboxylic acid” refers to —COOH.
“Oxo” refers to a ═O group.
“Halogen” refers to fluorine, chlorine, bromine or iodine.
“Pd(PPh3)2Cl2” refers to (bis(triphenylphosphine)palladium(II) chloride).
“TEA” refers to triethylamine.
“EA or EtOAc” refers to ethyl acetate.
“THF” refers to tetrahydrofuran.
“NaBH4” refers to sodium borohydride.
“DCM” refers to dichloromethane.
“Tf2O” refers to trifluoromethanesulfonic anhydride.
“(Bpin)2” refers to bis(pinacolato)diboron.
“Pd(dppf)2Cl2” refers to [1,1′-bis(diphenylphosphino)ferrocene]palladium dichloride.
“KOAc” refers to potassium acetate.
“DMF” refers to N,N-dimethylformamide.
“NBS” refers to N-bromosuccinimide.
“EtOH” refers to ethanol.
“MeOH” refers to methanol.
“Pd/C” refers to palladium/carbon.
“Pd(OH)2/C” refers to palladium hydroxide/carbon.
“DMSO” refers to dimethyl sulfoxide.
“HATU” refers to 2-(7-azabenzotriazol-1-yl)-N,N,N′,N′-tetramethyluronium hexafluorophosphate.
“TFA” refers to trifluoroacetic acid.
“NMP” refers to N-methylpyrrolidone.
“IBX” refers to 2-iodoxybenzoic acid.
“DIEA” refers to N,N-diisopropylethylamine.
“STAB” refers to sodium triacetoxyborohydride.
“T3P” refers to 2,4,6-tripropyl-1,3,5,2,4,6-trioxatriphosphorinane-2,4,6-trioxide.
“NaOH” refers to sodium hydroxide.
“DMAP” refers to 4-dimethylaminopyridine.
“TPSCl” refers to 2,4,6-triisopropylbenzenesulfonyl chloride.
“DPBS” refers to Dulbecco's phosphate-buffered saline.
“Dess-Martin” refers to a Dess-Martin reagent.
“PBS” refers to phosphate-buffered saline.
“SDS-PAGE” refers to sodium dodecyl sulfate-polyacrylamide gel electrophoresis.
“PVDF” refers to polyvinylidene fluoride.
“PE” refers to petroleum ether.
“NaHCO3” refers to sodium bicarbonate.
“Na2SO4” refers to sodium sulfate.
“NH4Cl” refers to ammonium chloride.
“AcOH” refers to acetic acid.
“HCl” refers to hydrochloric acid.
“DCC” refers to N,N-dicyclohexylcarbodiimide.
“Ti(OEt)4” refers to tetraethyl titanate.
“L-Selectride” refers to lithium triisobutylborohydride.
“BOP” refers to benzotriazol-1-yloxytris(dimethylamino)phosphonium hexafluorophosphate.
“DBU” refers to 1,8-diazabicyclo[5.4.0]undec-7-ene.
“LiOH·H2O” refers to lithium hydroxide monohydrate.
“EDCI” refers to 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride.
“NIS” refers to N-iodosuccinimide.
“Ac2O” refers to acetic anhydride.
“HOAc” refers to acetic acid.
“Hantzsch-ester” refers to diethyl 1,4-dihydro-2,6-dimethyl-3,5-pyridinedicarboxylate.
“DMA” refers to dimethylacetamide.
“NiBr2(dtbpy)” refers to (4,4-di-tert-butylbipyridine)nickel dibromide.
“K2CO3” refers to potassium carbonate.
“9-BBN” refers to 9-borabicyclo[3.3.1]nonane.
“MTBE” refers to methyl tert-butyl ether.
“LiOH” refers to lithium hydroxide.
“DCE” refers to dichloroethane.
“NaBH3CN” refers to sodium cyanoborohydride.
“CBr4” refers to carbon tetrabromide.
“PPh3” refers to triphenylphosphine.
“n-BuLi” refers to n-butyllithium.
“NaH” refers to sodium hydride.
“i-PrOH” refers to isopropanol.
“ACN” refers to acetonitrile.
“NH4HCO3” refers to ammonium bicarbonate.
“m-CPBA” refers to m-chloroperoxybenzoic acid.
“KI” refers to potassium iodide.
“IPA” refers to isophthalic acid.
“N2H4—H2O” refers to hydrazine hydrate.
“NMI” refers to N-methylimidazole.
“TCFH” refers to chloro-N,N,N′,N′-tetramethylformamidinium hexafluorophosphate.
“Ruphos Pd G2” refers to chloro(2-dicyclohexylphosphino-2′,6′-di-isopropoxy-1,1′-biphenyl)(2-amino-1,1′-biphenyl-2-yl)palladium (II).
“K3PO4” refers to potassium phosphate.
“(Boc)2O” refers to di-tert-butyl dicarbonate.
“Prep-HPLC” refers to preparative high performance liquid chromatography.
“sat.” refers to a saturated solution.
“aq” refers to an aqueous solution.
In the chemical structure herein, “—” as a connecting bond represents a single bond, and “═” represents a double bond (in the case of an undefined configuration, it may be trans or cis).
“Optional” or “optionally” means that the event or circumstance subsequently described may, but does not necessarily, occur, and that the description includes instances where the event or circumstance occurs or does not occur. For example, “a heterocycloalkyl group optionally substituted with alkyl” means that the alkyl may be, but not necessarily, present, and that the description includes cases where the heterocycloalkyl group is or is not substituted with the alkyl.
“Substituted” means that one or more, preferably up to 5, more preferably 1-3 hydrogen atoms in the group are independently substituted with a corresponding number of substituents. It goes without saying that a substituent is only in its possible chemical position, and those skilled in the art will be able to determine (experimentally or theoretically) possible or impossible substitution without undue effort. For example, it may be unstable when amino or hydroxy having a free hydrogen is bound to a carbon atom having an unsaturated (such as olefin) bond.
As used herein, unless otherwise indicated, the term “optionally substituted” may be unsubstituted or substituted; when it is substituted, the substituent may be one or more (e.g., 2, 3, 4, 5 or 6) groups independently selected from alkyl, alkenyl, alkynyl, hydroxy, hydroxyalkyl, haloalkyl, alkoxy, amino, aminoalkyl, cyano, halogen, oxo, cycloalkyl, heterocycloalkyl, aryl and heteroaryl, and the alkyl, the alkenyl, the alkynyl, the hydroxy, the hydroxyalkyl, the haloalkyl, the alkoxy, the amino and the aminoalkyl are optionally substituted with one or more (e.g., 2, 3, 4, 5 or 6) cycloalkyl, heterocycloalkyl, aryl or heteroaryl; the cycloalkyl, the heterocycloalkyl, the aryl and the heteroaryl are optionally substituted with one or more groups selected from alkyl, alkenyl, alkynyl, hydroxy, hydroxyalkyl, haloalkyl, alkoxy, amino, aminoalkyl, cyano, halogen and oxo.
“Pharmaceutical composition” refers to a mixture containing one or more of the compounds or the physiologically/pharmaceutically acceptable salts or pro-drugs thereof described herein, and other chemical components, for example, physiologically/pharmaceutically acceptable carriers and excipients. The pharmaceutical composition is intended to promote the administration to an organism, so as to facilitate the absorption of the active ingredient, thereby exerting biological activity.
The present invention also provides a pharmaceutically acceptable salt of the compound of formula (I). The term “pharmaceutically acceptable salt” refers to a relatively non-toxic acid addition salt or base addition salt of the compound of the present invention. The acid addition salt is a salt formed from the compound of formula (I) of the present invention and a suitable inorganic acid or organic acid. These salts may be prepared during the final isolation and purification of the compound or by reacting the purified compound of formula (I) in its free base form with a suitable organic acid or inorganic acid. The representative acid addition salt includes hydrobromide, hydrochloride, sulfate, bisulfate, sulfite, acetate, oxalate, valerate, oleate, palmitate, stearate, laurate, borate, benzoate, lactate, phosphate, biphosphate, carbonate, bicarbonate, toluate, citrate, maleate, fumarate, succinate, tartrate, benzoate, methanesulfonate, p-toluenesulfonate, gluconate, lactobionate, lauryl sulfonate and the like. The base addition salt is a salt formed from the compound of formula (I) and a suitable inorganic base or organic base, including, for example, a salt formed with alkali metal, alkaline earth metal and quaternary ammonium cations, such as sodium salts, lithium salts, potassium salts, calcium salts, magnesium salts, tetramethyl quaternary ammonium salts, tetraethyl quaternary ammonium salts and the like; the amine salt includes salts formed with ammonia (NH3) and primary, secondary or tertiary amines, such as methylamine salts, dimethylamine salts, trimethylamine salts, triethylamine salts, ethylamine salts and the like.
The compound of the present invention or the pharmaceutically acceptable salt thereof may be administered to mammals including humans, and may be administered orally, rectally, parenterally (intravenously, intramuscularly or subcutaneously), topically (powders, ointments or drops) or intratumorally.
The compound of the present invention may be administered at a dose of about 0.05-300 mg/kg body weight/day, preferably 10-300 mg/kg body weight/day, and more preferably 10-200 mg/kg body weight/day.
The compound of the present invention or the pharmaceutically acceptable salt thereof may be formulated into a solid dosage form for oral administration, including but not limited to capsules, tablets, pills, pulvis, granules and the like. In these solid dosage forms, the compound of formula (I) of the present invention, as an active ingredient, is mixed with at least one conventional inert excipient (or carrier), such as sodium citrate or dicalcium phosphate, or with the following ingredients: (1) fillers or extenders, such as starch, lactose, sucrose, glucose, mannitol, silicic acid and the like; (2) binders, such as hydroxymethyl cellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose, acacia and the like; (3) humectants, such as glycerol and the like; (4) disintegrants, such as agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, sodium carbonate and the like; (5) solution retarders, such as paraffin and the like; (6) absorption accelerators, such as quaternary ammonium compounds and the like; (7) wetting agents, such as cetyl alcohol, glycerol monostearate and the like; (8) adsorbents, such as kaolin and the like; and (9) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate and the like, or mixtures thereof. Capsules, tablets and pills may further comprise buffers.
The solid dosage forms such as tablets, dragees, capsules, pills and granules may be coated or microencapsulated using coatings and shells such as enteric coatings and other materials well known in the art. They may comprise opacifying agents, and the active ingredient in such a composition may be released in a certain part of the digestive tract in a delayed manner. Examples of embedding components that can be used are polymeric substances and wax-based substances. If necessary, the active ingredient can also be in microcapsule form with one or more of the above-mentioned excipients.
The compound of the present invention or the pharmaceutically acceptable salt thereof may be formulated into a liquid dosage form for oral administration, including but not limited to pharmaceutically acceptable emulsions, solutions, suspensions, syrups, tinctures and the like. In addition to the compound of formula (I) or the pharmaceutically acceptable salt thereof as an active ingredient, the liquid dosage form may comprise inert diluents commonly used in the art, such as water and other solvents, solubilizers and emulsifiers, for example, ethanol, isopropanol, ethyl carbonate, ethyl acetate, propylene glycol, 1,3-butanediol, dimethylformamide, and oils, especially cottonseed oil, peanut oil, corn germ oil, olive oil, castor oil, sesame oil and the like, or mixtures of these substances. In addition to such inert diluents, the liquid dosage form in the present invention may further comprise conventional adjuvants, such as wetting agents, emulsifiers, suspending agents, sweeteners, flavoring agents, perfuming agents and the like.
The suspending agents include, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methylate, agar and the like, or mixtures of these substances.
The compound of the present invention or the pharmaceutically acceptable salt thereof may be formulated into a dosage form for parenteral injection, including but not limited to physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for redissolving into sterile injectable solutions or dispersions. Suitable carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.
The compound of the present invention or the pharmaceutically acceptable salt thereof may also be formulated into a dosage form for topical administration, including, for example, ointments, pulvis, suppositories, drops, sprays, inhalants and the like. The compound of formula (I) or the pharmaceutically acceptable salt thereof in the present invention as an active ingredient is mixed under sterile conditions with a physiologically acceptable carrier and optional preservatives, buffers or propellants that may be required if necessary.
The present invention also provides a pharmaceutical composition comprising the compound of formula (I) or the pharmaceutically acceptable salt thereof as an active ingredient, and a pharmaceutically acceptable carrier, excipient or diluent. In the preparation of the pharmaceutical composition, the compound of formula (I) or the pharmaceutically acceptable salt thereof in the present invention is typically mixed with the pharmaceutically acceptable carrier, excipient or diluent.
The composition in the present invention may be formulated into a conventional pharmaceutical formulation according to a conventional preparation method. For example, tablets, pills, capsules, pulvis, granules, emulsions, suspensions, dispersions, solutions, syrups, elixirs, ointments, drops, suppositories, inhalants, sprays and the like.
The compound or the pharmaceutically acceptable salt thereof described in the present invention may be administered alone or, if desired, in combination with other pharmaceutically acceptable therapeutic agents, such as other anti-tumor medicaments. The ingredients to be combined may be administered simultaneously or sequentially, and administered in a single formulation or in different formulations. The combination may include not only a combination of the compound in the present invention and one additional active agent, but also a combination of the compound in the present invention and two or more additional active agents.
In the present invention, other pharmaceutically acceptable therapeutic agents that may be used together with or in combination with the SOS1 degrading agent, the compound of formula (I), may be: EGFR and/or a mutant inhibitor thereof, ErbB2(Her2) and/or a mutant inhibitor thereof, ALK and/or a mutant inhibitor thereof, MEK and/or a mutant inhibitor thereof, Kras and/or a mutant inhibitor thereof, BCR-ABL and/or a mutant inhibitor thereof, FGFR1/FGFR2/FGFR3 and/or a mutant inhibitor thereof, ROS1 and/or a mutant inhibitor thereof, c-MET and/or a mutant inhibitor thereof, AXL and/or a mutant inhibitor thereof, NTRK1 and/or a mutant inhibitor thereof, RET and/or a mutant inhibitor thereof, taxane, a platinum-containing compound, an antimetabolite, a mitotic kinase inhibitor, an immunotherapeutic agent, an anti-angiogenic drug, a topoisomerase inhibitor, A-Raf/B-Raf/C-Raf and/or a mutant inhibitor thereof, ERK and/or a mutant inhibitor thereof, an apoptosis inhibitor, an mTOR inhibitor, an epigenetic modulator, an IGF1/2 and/or IGF1-R inhibitor, Ras GEF and/or a mutant inhibitor thereof or PI3K and/or a mutant inhibitor thereof.
In the present invention, other pharmaceutically acceptable therapeutic agents that may be used together with or in combination with the SOS1 degrading agent, the compound of formula (I), may be: afatinib, erlotinib, gefitinib, lapatinib, cetuximab, panitumumab, osimertinib, olmutinib, EGF-816, trastuzumab, pertuzumab, crizotinib, alectinib, entrectinib, brigatinib, trametinib, cobimetinib, binimetinib, selumetinib, refametinib, imatinib, dasatinib, nilotinib, nintedanib, crizotinib, lorlatinib, ceritinib, merestinib, paclitaxel, nab-paclitaxel, docetaxel, cisplatin, carboplatin, oxaliplatin, 5-fluorouracil, capecitabine, floxuridine, cytarabine, gemcitabine, a combination of trifluridine and tipiracil (=TAS102), palbociclib, ribociclib, abemaciclib, ipilimumab, nivolumab, pembrolizumab, atezolizumab, avelumab, durvalumab, pidilizumab, PDR-001 (=spartalizumab), bevacizumab, irinotecan, liposomal irinotecan, topotecan, ulixertinib, rapamycin, temsirolimus, everolimus, ridaforolimus, JQ-1, GSK 525762, OTX 015(=MK8628), CPI 0610, TEN-010 (=RO6870810), xentuzumab (antibody 60833 in WO2010/066868) or MEDI-573 (=dusigitumab).
According to the present invention, SOS1 kinase activity tests prove that the compound of formula I described in the present invention is able to effectively bind to the SOS1 target protein or have an inhibitory effect, and it is proved that the compound of formula I described herein is able to effectively and specifically degrade the SOS1 protein in NCI-H358 cells by means of Western Blot. The compound of formula I, and/or the stereoisomer, the enantiomer, the diastereomer, the deuteride, the hydrate, the solvate, the metabolite, the prodrug and/or the pharmaceutically acceptable salt thereof in the present invention can effectively degrade the SOS1 protein, thereby achieving the effect of preventing or treating a disease or disorder related to SOS1 or caused by the interaction of SOS1 with Ras or SOS1 with Rac.
The present invention will be further explained in detail below with reference to examples, which are not intended to limit the present invention, and the present invention is not merely limited to the contents of the examples. Starting materials in the examples of the present invention are known and commercially available, or may be synthesized by using or according to methods known in the art. Unless otherwise stated, experimental procedures without specific conditions indicated in the examples of the present invention are generally conducted according to conventional conditions, or according to conditions recommended by the manufacturers of the starting materials or commercial products.
A reaction mixture of 1-bromo-3-nitro-5-(trifluoromethyl)benzene (18 g, 66.7 mmol), tributyl(1-ethoxyvinyl)stannane (31 g, 86.7 mmol), Pd(PPh3)2Cl2 (4.7 g, 6.67 mmol), and TEA (13.5 g, 133 mmol) in a 1,4-dioxane (200 mL) solution was stirred at 80° C. under nitrogen atmosphere for reaction overnight. Potassium fluoride (saturated aqueous solution, 300 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with EA (200 mL×3). The organic phase was collected and concentrated under reduced pressure. The concentrate was dissolved in THF (180 mL), hydrochloric acid (100 mL, 6 M) was added thereto, and the mixture was stirred at room temperature for 3 h. Water (300 mL) was added to the mixture to quench the reaction, and the resulting mixture was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography to give the target product 1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl-1-one.
1H NMR (400 MHz, CDCl3) δ 8.95 (t, J=1.6 Hz, 1H), 8.69 (s, 1H), 8.54 (d, J=0.5 Hz, 1H), 2.76 (s, 3H).
A reaction mixture of 1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl-1-one (10.6 g, 45.5 mmol), (R)-2-methylpropyl-2-sulfinamide (8.3 g, 68.2 mmol), and tetraethyl titanate (25.9 g, 114 mmol) in THE (200 mL) solution was stirred at 60° C. for reaction for 2 h. Water (300 mL) was added to the reaction mixture to quench the reaction, and the solution was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give the target product (R,E)-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethylidene)propyl-2-sulfinamide.
1H NMR (400 MHz, DMSO-d6) δ 8.87 (s, 1H), 8.61 (s, 1H), 8.43 (s, 1H), 2.90 (s, 3H), 1.37 (s, 9H).
NaBH4 (1.28 g, 33.7 mmol) was added to a mixture of (R,E)-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethylidene)propyl-2-sulfinamide (10 g, 30.7 mmol) in a solution THE (100 mL) and water (20 mL). After the reaction mixture was stirred at −10° C. for reaction for 1 min, water (300 mL) was added to the reaction mixture to quench the reaction, and the solution was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give the target product (R)-2-methyl-N—((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)propyl-2-sulfinamide.
1H NMR (400 MHz, CDCl3) δ 8.45-8.42 (m, 2H), 7.96 (s, 1H), 4.76-4.68 (m, 1H), 3.56 (d, J=4.2 Hz, 1H), 1.62 (d, J=6.7 Hz, 3H), 1.26 (s, 9H).
(R)-2-Methyl-N—((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)propyl-2-sulfinamide (5.5 g, 16.3 mmol) was placed in a solution of hydrochloric acid (80 mL, 4 M) in dioxane, and the mixture was stirred at 25° C. for reaction for 2 h. The crude product was filtered and washed with petroleum ether (100 mL) to give the target product (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl-1-amine hydrochloride.
LC-MS: (ESI, m/z): [M+H]+=235.2.
Tf2O (9 g, 32.0 mmol) was added dropwise to a stirred mixture of ethyl 4-oxocyclohexane-1-formate (5 g, 32.0 mmol) and 2,6-dimethylpyridine (4.6 g, 42.9 mmol) DCM (60 mL) at 0° C. The reaction mixture was stirred at 0° C. for reaction for 1 h. The reaction mixture was then stirred at 25° C. for reaction for 15 min, and Tf2O (7 g, 25.6 mmol) was added. Then the reaction mixture was stirred at 25° C. overnight and then concentrated. Water (300 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give ethyl 4-(((trifluoromethyl)sulfonyl)oxy)cyclohex-3-ene-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 5.90 (d, J=3.9 Hz, 1H), 4.14-4.02 (m, 2H), 2.68-2.60 (m, 1H), 2.47-2.27 (m, 4H), 2.08-1.98 (m, 1H), 1.88-1.76 (m, 1H), 1.19 (t, J=7.1 Hz, 3H).
A mixture of ethyl 4-(((trifluoromethyl)sulfonyl)oxy)cyclohex-3-ene-1-carboxylate (4.7 g, 16.8 mmol), (Bpin)2 (6.4 g, 25.2 mmol), Pd(dppf)2Cl2 (1.2 g, 1.68 mmol), and KOAc (4.9 g, 50.4 mmol) in a 1,4-dioxane (80 mL) solution was stirred at 90° C. for reaction overnight. Water (300 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give ethyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 6.42 (d, J=2.2 Hz, 1H), 4.11-4.02 (m, 2H), 2.33-1.95 (m, 4H), 1.93-1.83 (m, 1H), 1.53-1.38 (m, 1H), 1.21-1.14 (m, 16H).
NBS (35 g, 197 mmol) was added to a stirred solution of 2-amino-4-methoxybenzoic acid (30 g, 179 mmol) in DMF (600 mL) at 0° C., and the reaction mixture was stirred at 25° C. for reaction for 1 h. Sodium thiosulfate (saturated aqueous solution, 500 mL) was added to quench the reaction liquid, and the resulting mixture was extracted with EA (500 mL×3). The organic phase was collected, washed with sodium chloride (saturated aqueous solution, 500 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated to give a crude product of 2-amino-5-bromo-4-methoxybenzoic acid, which was directly used in the next step without further purification.
LC-MS: (ESI, m/z): [M+H]+=246.0.
Ammonium acetate (53.2 g, 691 mmol) and trimethyl orthoacetate (83.0 g, 691 mmol) were added to a stirred solution of 2-amino-5-bromo-4-methoxybenzoic acid (17.0 g, 69.1 mmol) in methanol (170 mL). The resulting reaction mixture was stirred in a closed container at 120° C. for reaction for 24 h. The reaction mixture was poured into water (1 L), and the solid product was filtered. The crude product was washed with EA (1 L) to give the target product 6-bromo-7-methoxy-2-methylquinazolin-4(3H)-one. The crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=268.9.
A mixture of 6-bromo-7-methoxy-2-methylquinazolin-4(3H)-one (10 g, 37.2 mmol) in a phosphorus oxychloride (200 mL) solution was stirred at 105° C. overnight. The reaction mixture was concentrated. Sodium bicarbonate (saturated aqueous solution, 500 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (300 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product 6-bromo-4-chloro-7-methoxy-2-methylquinazoline.
LC-MS: (ESI, m/z): [M+H]+=287.0.
(R)-1-(3-Nitro-5-(trifluoromethyl))phenyl)ethyl-1-amine hydrochloride (1.7 g, 7.52 mmol) DIEA (1.6 g, 12.5 mmol) were added to a mixture of 6-bromo-4-chloro-7-methoxy-2-methylquinazoline (1.8 g, 6.27 mmol) in an EtOH (12 mL) solution. The reaction mixture was stirred in microwave at 100° C. for reaction for 5 h. Water (300 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (300 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give (R)-6-bromo-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)quinazolin-4-amine as a yellow oil.
LC-MS: (ESI, m/z): [M+H]+=484.9.
A reaction mixture of (R)-6-bromo-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)quinazolin-4-amine (2 g, 4.12 mmol), ethyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-yl)cyclohex-3-ene-1-carboxylate (2.31 g, 8.24 mmol), sodium carbonate (873 mg, 8.24 mmol), and Pd(dppf)2Cl2 (300 mg, 0.41 mmol) in DMF (24 mL) and water (3 mL) was stirred at 110° C. for reaction overnight. Water (300 mL) was added to quench the reaction.
The solution was extracted with EA (300 mL×3) and washed with sodium chloride (saturated aqueous solution, 500 mL×3), and the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give ethyl 4-(7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=559.7.
Pd/C (1.02 g, 0.6 w/w) was added to a mixture of ethyl 4-(7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohex-3-ene-1-carboxylate (1.7 g, 3.04 mmol) in an MeOH (60 mL) solution. The reaction mixture was stirred at 50° C. under hydrogen atmosphere for reaction for 4 days. The crude product was filtered to give ethyl (R)-4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=531.2.
Lithium hydroxide (203 mg, 8.46 mmol) was added to a mixture of ethyl (R)-4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (1.5 g, 2.82 mmol) in methanol (8 mL), THE (8 mL) and water (16 mL). The reaction mixture was stirred at 60° C. for reaction overnight. The crude product was purified by preparative HPLC to give (1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-formic acid.
LC-MS: (ESI, m/z): [M+H]+=503.2.
1H NMR (400 MHz, DMSO-d6) δ 8.13 (d, J=8.0 Hz, 1H), 8.06 (s, 1H), 6.99 (s, 1H), 6.88 (s, 1H), 6.86 (s, 1H), 6.69 (s, 1H), 5.56 (dd, J=13.6, 5.8 Hz, 3H), 3.88 (s, 3H), 2.92 (t, J=11.7 Hz, 1H), 2.35 (s, 3H), 2.24 (t, J=11.8 Hz, 1H), 2.05 (d, J=10.8 Hz, 2H), 1.87 (d, J=11.8 Hz, 2H), 1.66-1.40 (m, 7H).
tert-Butyl piperazine-1-formate (163 mg, 0.88 mmol), DIEA (169 mg, 1.31 mmol), and T3P (209 mg, 0.66 mmol) were added to a mixture of (1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-formic acid (220 mg, 0.44 mmol) in an EA (12 mL) solution. The reaction mixture was stirred at 0° C. for 3 h. Water (100 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (100 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give tert-butyl 4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=671.3.
1H NMR (400 MHz, DMSO-d6) δ 8.19-8.07 (m, 2H), 6.99 (s, 1H), 6.86 (d, J=10.5 Hz, 2H), 6.70 (s, 1H), 5.56 (dd, J=16.7, 9.3 Hz, 3H), 3.89 (s, 3H), 3.47 (m, 4H), 3.32 (m, 4H), 2.95 (s, 1H), 2.67 (s, 1H), 2.36 (s, 3H), 1.83 (d, J=14.5 Hz, 4H), 1.59 (t, J=22.3 Hz, 7H), 1.41 (s, 9H).
A mixture of tert-butyl 4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazine-1-carboxylate (220 mg, 0.33 mmol) in a solution of ethyl acetate in hydrochloric acid (3 M, 10 mL) was stirred at 25° C. for reaction for 2 h. The reactant was concentrated to give ((1R,4R)-4-(4-(((R)-1-(3-amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(piperazin-1-yl)methanone, and the crude product could be used in the next step without further purification.
LC-MS: (ESI, m/z): [M+H]+=571.2.
3-Aminopiperidine-2,6-dione hydrochloride (5.0 g, 0.03 mol) and potassium acetate (8.8 g, 0.09 mol) were added to a solution of 4-fluorophthalic acid (5.52 g, 0.03 mol) in acetic acid (50 mL). The reaction mixture was stirred at 120° C. for reaction overnight and concentrated under reduced pressure, and the concentrate was diluted with water (100 mL). The resulting mixture was stirred at room temperature for 30 min and filtered. The solid was washed with water (50 mL×2) and dried under reduced pressure to give the product.
LC-MS: (ESI, m/z): [M+H]+=277.1
A mixture of 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindoline-1,3-dione (500 mg, 1.81 mmol), 2-(piperidin-4-yl)ethyl-1-ol (280 mg, 2.17 mmol), DIEA (701 mg, 5.43 mmol) NMP (5 mL) was reacted in a microwave reactor at 140° C. for 5 h. The reaction mixture was diluted with water (50 mL), and the resulting mixture was extracted with ethyl acetate (50 mL×2). The organic phase was collected, washed with water (100 mL×2) and saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the product.
LC-MS: (ESI, m/z): [M+H]+=386.1.
A mixture of 2-(2,6-dioxopiperidin-3-yl)-5-(4-(2-hydroxyethyl)piperidin-1-yl)isoindoline-1,3-dione (300 mg, 0.78 mmol) and IBX (436 mg, 1.56 mmol) in acetonitrile (6 mL) was stirred at 80° C. for reaction for 2 h. The reaction mixture was diluted with water (30 mL), and the resulting mixture was extracted with ethyl acetate (30 mL×2).
The organic phase was collected, washed with water (50 mL×2) and saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the product.
LC-MS: (ESI, m/z): [M+H]+=384.1.
1H NMR (400 MHz, DMSO-d6) δ 11.07 (s, 1H), 9.69 (t, J=1.6 Hz, 1H), 7.65 (d, J=8.6 Hz, 1H), 7.31 (d, J=2.0 Hz, 1H), 7.23 (dd, J=8.6, 2.2 Hz, 1H), 5.06 (dd, J=12.9, 5.4 Hz, 1H), 4.03 (dd, J=10.3, 2.9 Hz, 2H), 3.04-2.82 (m, 3H), 2.65-2.52 (m, 2H), 2.41 (dd, J=6.7, 1.6 Hz, 2H), 2.17-1.97 (m, 2H), 1.73 (d, J=11.1 Hz, 2H), 1.2-1.18 (m, 2H).
3-Amino-4-chlorobenzoic acid (5.0 g, 2.93 mmol) was suspended in acrylic acid (8.05 mL, 117 mmol), and the suspension was stirred at 100° C. for reaction for 3 h. Then the reaction liquid was cooled to room temperature while stirring. Acetic acid (33 mL) was added, and the stirred suspension was heated at 100° C. for 10 min. Urea (11.00 g, 183 mmol) was added, and the reaction liquid was stirred at 120° C. for reaction overnight. The reaction liquid was added to a mixture of ice water and concentrated hydrochloric acid (37%), and the mixture was stirred. The resulting suspension was stored in a refrigerator at 5° C. overnight and filtered, and the solid was washed with water and dried to give a solid. The solid was triturated in a hydrochloric acid solution (0.05 M), filtered, washed with methyl tert-butyl ether, and dried under reduced pressure at 40° C. to give the target product.
LC-MS: (ESI, m/z): [M+H]+=269.0
A mixture of 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoic acid (2.0 g, 7.58 mmol), 2,3,4,5,6-pentafluorophenol (1.67 g, 9.09 mmol), and N,N-dicyclohexylcarbodiimide (1.87 g, 9.09 mmol) in an N,N-dimethylformamide (20 mL) solution was stirred at room temperature for reaction for 3 h. The reaction liquid was poured into water (200 mL), and the resulting solution was stirred for 0.5 h and extracted with ethyl acetate (200 mL×3). The organic phase was collected, washed with water (500 mL×2) and saturated brine (300 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=435.0
Pd(OH)2/C (200 mg) was added to a mixture of benzyl 9-(2-hydroxyethyl)-3-azaspiro[5.5]undecane-3-carboxylate (1.0 g, 3 mmol) in EA (10 mL) solution, and the reaction mixture was stirred at room temperature under hydrogen atmosphere for reaction for 16 h. The mixture was filtered, and the filtrate was concentrated to give the target product.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (1.0 g, 2.5 mmol) DIEA (0.5 mL) were added to a mixture of 2-(3-azaspiro[5.5]undecan-9-yl)ethan-1-ol (500 mg, 2.5 mmol) in DMSO (5 mL) solution. Water (30 mL) was added to quench the reaction, and the resulting mixture was extracted with EA (10 mL×3). The organic phase was collected, washed with water (10 mL×2) and saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by column chromatography to give the target compound.
LC-MS: (ESI, m/z): [M+H]+=448.1
IBX (873 mg, 3.1 mmol) was added to a mixture of 1-(2-chloro-5-(9-(2-hydroxyethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (700 mg, 1.56 mmol) in THF (12 mL) solution. The reaction mixture was stirred at 80° C. for reaction for 2 h. Water (30 mL) was added to quench the reaction, and the solution was extracted with EA (10 mL×3). The organic phase was collected, washed with water (10 mL×2) and saturated brine (10 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by column chromatography to give the target compound.
LC-MS: (ESI, m/z): [M+H]+=446.1
2-Bromo-1,1-dimethoxyethane (3.63 g, 21.51 mmol) was added to a mixture of tert-butyl piperazine-1-carboxylate (2.0 g, 10.75 mmol), potassium carbonate (4.45 g, 32.26 mmol), and potassium iodide (892 mg, 5.38 mmol) in acetone (20 mL), and the mixture was stirred at 80° C. for reaction overnight. The reaction mixture was concentrated under reduced pressure. The concentrate was dissolved in ethyl acetate (50 mL) and washed with water (50 mL×2) and saturated brine (50 mL). The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the desired product.
LC-MS: (ESI, m/z): [M+H]+=275.0.
A mixture of tert-butyl 4-(2,2-dimethoxyethyl)piperazine-1-carboxylate (2.2 g, 8.03 mmol) in a hydrochloric acid/dioxane (4 M, 10 mL) solution was stirred at room temperature overnight. The mixture was concentrated under reduced pressure to give the target product (1.5 g, crude product). The crude product was directly used in the next step.
A mixture of 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindoline-1,3-dione (1.48 g, 5.36 mmol), 1-(2,2-dimethoxyethyl)piperazine (1.4 g, 8.05 mmol), DIEA (4.15 g, 36.16 mmol) in NMP (10 mL) solution was reacted in a microwave reactor at 140° C. for 5 h. The mixture was diluted with water (50 mL), and the resulting mixture was extracted with ethyl acetate (50 mL×2). The organic phase was collected, washed with water (100 mL×2) and saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=431.1.
A mixture of 5-(4-(2,2-dimethoxyethyl)piperazin-1-yl)-2-(2,6-dioxopiperidin-3-yl)isoindoline-1,3-dione (1.8 g, 4.19 mmol) in a trifluoroacetic acid/dichloromethane (5 mL/5 mL) solution was stirred at room temperature for reaction for 60 h. The mixture was concentrated under reduced pressure, water was added, and the pH was adjusted to 8 with sodium bicarbonate (aqueous solution). The mixture was extracted with ethyl acetate (100 mL×2). The organic phase was collected, washed with water (100 mL×2) and saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=285.0.
HATU (1.66 g, 4.37 mmol), tert-butyl 3,9-diazaspiro[5.5]undecyl-3-carboxylate (0.96 g, 3.78 mmol), and N-methylmorpholine (0.8 g, 7.92 mmol) were added to a solution of 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoic acid (1.0 g, 3.78 mmol) in N,N-dimethylformamide (10 mL) at room temperature under nitrogen atmosphere. After the reaction mixture was stirred for reaction for 2 h, water (50 mL) was added to the reaction mixture to quench the reaction, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phase was collected, washed with saturated brine (50 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=505.2
A reaction liquid of tert-butyl 9-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3,9-diazaspiro[5.5]undecane-3-carboxylate (1.56 g, 3.1 mmol) in a trifluoroacetic acid/dichloromethane (2.5 mL/5 mL) solution was stirred at room temperature for reaction for 4 h. The reaction liquid was concentrated under reduced pressure to give a crude product of the target compound. The crude product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=405.1
2-Bromo-1,1-dimethoxyethane (46 mg, 0.3 mmol) was added to a mixture of 1-(2-chloro-5-(3,9-diazaspiro[5.5]undecyl-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (200 mg, 0.25 mmol), potassium carbonate (155 mg, 1.25 mmol), and potassium iodide (19 mg, 0.11 mmol) in an acetone (4 mL) solution. The reaction liquid was stirred at 80° C. for reaction for 3 h and then concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to give the target compound.
LC-MS: (ESI, m/z): [M+H]+=493.3.
A mixture of 1-(2-chloro-5-(9-(2,2-dimethoxyethyl)-3,9-diazaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (121 mg, 0.246 mmol) in a trifluoroacetic acid/dichloromethane (1 mL/2 mL) solution was stirred at room temperature for reaction for 4 h. The reaction liquid was concentrated under reduced pressure. The concentrate was purified by silica gel column chromatography to give the target compound.
LC-MS: (ESI, m/z): [M+H]+=447.1.
A mixture of 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindoline-1,3-dione (500 mg, 1.81 mmol), 3-(piperidin-4-yl)ethyl-1-ol (310 mg, 2.17 mmol), DIEA (701 mg, 5.43 mmol) NMP (5 mL) was reacted in a microwave reactor at 140° C. for 5 h. The reaction mixture was diluted with water (50 mL), and the resulting mixture was extracted with ethyl acetate (50 mL×2). The organic phase was collected, washed with water (100 mL×2) and saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=400.2.
A mixture of 2-(2,6-dioxopiperidin-3-yl)-5-(4-(3-hydroxypropyl)piperidin-1-yl)isoindoline-1,3-dione (310 mg, 0.78 mmol) and IBX (436 mg, 1.56 mmol) in acetonitrile (6 mL) was stirred at 80° C. for reaction for 2 h. The reaction mixture was diluted with water (30 mL), and the resulting mixture was extracted with ethyl acetate (30 mL×2). The organic phase was collected, washed with water (50 mL×2) and saturated brine (30 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by silica gel column chromatography to give the product.
LC-MS: (ESI, m/z): [M+H]+=398.2.
NBS (19.6 g, 110.0 mmol) was added to a stirred solution of 2-amino-4-methoxybenzoic acid (16.7 g, 100.0 mmol) DMF (300 mL) at 0° C. The reaction mixture was stirred at 25° C. for reaction for 1 h, and water (300 mL) was added to quench the reaction. The mixture was extracted with EA (300 mL×3), and the organic phase was collected, washed with sodium chloride (a saturated aqueous solution, 300 mL×3), dried over anhydrous ammonium sulfate, and concentrated to give the target crude product. The crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=247.4.
Ammonium acetate (61.0 g, 793 mmol) and trimethyl orthoacetate (95.1 g, 793 mmol) were added to a mixture of 2-amino-5-bromo-4-methoxybenzoic acid (19.5g, 79.3 mmol) in a MeOH (195 mL) solution. The reaction mixture was stirred in a closed container at 120° C. for reaction for 24 h. The reaction mixture was poured into water (500 mL) and the solid product was filtered. The crude product was washed with EA to give the target crude product, and the crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=271.0.
tert-Butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborane-2-yl)-3,6-dihydropyridin-1(2H)-carboxylate (1.73 g, 6.18 mmol), sodium carbonate (982 mg, 9.27 mmol), and Pd(dppf)2Cl2 (226 mg, 0.319 mmol) were added to a mixture of 6-bromo-7-methoxy-2-methylquinazolin-4-ol (1.5 g, 3.09 mmol) in a mixed solution of DMF (30 mL) and water (4 mL). The reaction mixture was stirred at 110° C. for reaction overnight. The reaction was quenched with water (300 mL), and the mixture was extracted with EA (200 mL×3). The organic phase was collected, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=372.2.
Pd/C (300 mg) was added to a solution of tert-butyl 4-(4-hydroxy-7-methoxyquinazolin-6-yl)-3,6-dihydropyridin-1(2H)-carboxylate (500 mg, 0.894 mmol) in MeOH (20 mL). The reaction mixture was stirred at 50° C. under H2 atmosphere for reaction for 2 days. The reaction liquid was filtered through celite, and the filtrate was concentrated under reduced pressure to give the target product tert-butyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)piperidine-1-carboxylate. The crude product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=374.2.
TEA (0.3 mL, 2.4 mmol) was added to a solution of tert-butyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)piperidine-1-carboxylate (250.0 mg, 0.76 mmol), TPSCl (295.0 mg, 1.0 mmol), DMAP (13.0 mg, 0.1 mmol) DCM (5.0 mL). The reaction mixture was stirred at room temperature for reaction for 12 h. The reaction liquid was diluted with DCM and washed with a saturated NaHCO3 solution. The organic phase was dried over Na2SO4, filtered, and concentrated. The resulting crude product was purified by column chromatography (the eluent system was EA/PE) to give the product tert-butyl 4-(7-methoxy-2-methyl-4-(((2,4,6-triisopropylphenyl)sulfonyl)oxy)quinazolin-6-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=640.0.
(R)-1-(3-Nitro-5-(trifluoromethyl)phenyl)ethan-1-amine hydrochloride (203 mg, 0.75 mmol) TEA (0.8 mL) were added to a solution of tert-butyl 4-(7-methoxy-2-methyl-4-(((2,4,6-triisopropylphenyl)sulfonyl)oxy)quinazolin-6-yl)piperidine-1-carboxylate (400 mg, 0.03 mmol) DMSO (8 mL). The reaction mixture was stirred at 90° C. for reaction overnight. Water was added to quench the reaction, and the resulting mixture was extracted with EA. The organic phase was dried over Na2SO4, filtered, and concentrated. The resulting crude product was purified by normal phase column chromatography to give the target product tert-butyl (R)-4-(7-methoxy-2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=590.3.
TFA was added to a solution of tert-butyl (R)-4-(7-methoxy-2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidine-1-carboxylate (220 mg, 0.328 mmol) in DCM. The reaction mixture was stirred at 25° C. for reaction for 2 h. then concentrated to give the target product (R)-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-yl)quinazolin-4-amine. The resulting crude product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=490.2
NH4Cl (135.6 mg, 2.5 mmol) and Fe powder (141.5 mg, 2.5 mmol) were added to a solution of (R)-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-yl)quinazolin-4-amine (124 mg, 0.25 mmol) in EtOH/H2O (3 mL/1 mL). The reaction mixture was stirred at 70° C. under N2 atmosphere for reaction for 2 h. The reaction liquid was filtered and concentrated to give a crude product. The crude product was purified by preparative HPLC to give the target product (R)—N-(1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)-7-methoxy-2-methyl-6-(piperidin-4-yl)quinazolin-4-amine.
LC-MS: (ESI, m/z): [M+H]+=460.2.
1H NMR (400 MHz, DMSO-d6) δ 8.32 (s, 1H), 8.09 (s, 1H), 7.03 (s, 1H), 6.88 (d, J=11.1 Hz, 1H), 6.70 (s, 1H), 5.57 (d, J=6.9 Hz, 1H), 3.89 (s, 3H), 3.39-3.25 (m, 2H), 3.22-3.13 (m, 1H), 3.07-2.95 (m, 2H), 2.36 (s, 3H), 2.03-1.85 (m, 4H), 1.56 (d, J=7.0 Hz, 3H).
Methanesulfonyl chloride (7.4 g, 64.9 mmol) dissolved in a DCM (20.0 mL) solution was added dropwise to a mixture of tert-butyl 4-hydroxypiperidine-1-carboxylate (10.0 g, 49.7 mmol) and TEA (15.0 g, 149.1 mmol) in a DCM (100 mL) solution at 0° C. The reaction mixture was stirred at room temperature for reaction overnight, then water (200 mL) was added, and the resulting mixture was extracted with DCM (200 mL×2). The organic phase was collected, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The concentrate was purified by column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+H]+=280.1.
Acetamidine hydrochloride (19.0 g, 0.2 mol) and sodium acetate (16.4 g, 0.2 mol) were added to a mixture of 2-amino-4,5-dimethoxybenzoic acid (20.0 g, 0.1 mol) in a 2-methoxyethanol (120.0 mL) solution. The reaction mixture was stirred at 140° C. for reaction overnight. The reaction mixture was poured into water (250 mL), and the resulting mixture was stirred at room temperature for 10 min and then filtered. The filter cake was washed with water, and the solid was dried under vacuum to give the target product.
LC-MS: (ESI, m/z): [M+H]+=221.2.
A reaction mixture of 6,7-dimethoxy-2-methylquinazolin-4-ol (9.1 g, 41.4 mmol) and DL-methionine (10.5 g, 70.4 mmol) in methanesulfonic acid (50 mL) was stirred at 80° C. for reaction overnight. Ice water was added to quench the reaction, and the resulting mixture was basified with a sodium hydroxide (2 N) solution. The precipitate was filtered off to give the target crude product.
LC-MS: (ESI, m/z): [M+H]+=207.2.
A reaction mixture of 7-methoxy-2-methylquinazolin-4,6-diol (1.4 g, 6.8 mmol), tert-butyl 4-((methylsulfonyl)oxy)piperidine-1-carboxylate (2.2 g, 8.2 mmol), and potassium carbonate (1.8 g, 13.6 mmol) in an NMP (10.0 mL) solution was stirred at 100° C. for reaction overnight. Then water (50.0 mL) was added, and the resulting mixture was extracted with EA (50 mL×3). The organic phase was collected, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The concentrate was purified by reversed-phase column chromatography (acetonitrile:water=0-100%) to give the target product.
LC-MS: (ESI, m/z): [M+H]+=390.2.
A reaction mixture of tert-butyl 4-((4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)oxy)piperidine-1-carboxylate (800.0 mg, 2.0 mmol), 2,4,6-triisopropylbenzenesulfonyl chloride (748.0 mg, 2.4 mmol), DMAP (125.0 mg, 0.4 mmol), and TEA (623.0 mg, 6.0 mmol) in a DCM (10.0 mL) solution was stirred at room temperature for reaction overnight. The reaction mixture was concentrated, and the concentrate was purified by column chromatography to give the target product.
LC-MS: (ESI, m/z): [M+Na]+=678.1.
A reaction mixture of tert-butyl 4-((7-methoxy-2-methyl-4-(((2,4,6-triisopropylphenyl)sulfonyl)oxy)quinazolin-6-yl)oxy)piperidine-1-carboxylate (0.8 g, 1.2 mmol), (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-amine hydrochloride (0.43 g, 1.8 mmol), and TEA (1.1 g, 10.8 mmol) in a DMSO (15.0 mL) solution was stirred at 90° C. for reaction overnight. Then water (50.0 mL) was added, and the resulting mixture was extracted with EA (50 mL×3). The organic phase was collected, washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated. The concentrate was purified by reversed-phase column chromatography (acetonitrile-water=0-100%) to give the target product.
LC-MS: (ESI, m/z): [M+H]+=606.0.
A mixture of tert-butyl (R)-4-((7-methoxy-2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)oxy)piperidine-1-carboxylate (140 mg, 0.23 mmol) in a solution of DCM (5.0 mL) and TFA (1.0 mL) was stirred at room temperature for reaction for 2.0 h. The reactant was concentrated under reduced pressure to give (R)-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-oxy)quinazolin-4-amine.
LC-MS: (ESI, m/z): [M+H]+=506.1.
A reaction mixture of (R)-7-methoxy-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-oxy)quinazolin-4-amine (100.0 mg, 0.19 mmol) and Pd/C (20.0 mg) in a methanol (5.0 mL) solution was stirred at room temperature under hydrogen atmosphere for reaction overnight. The reaction liquid was filtered, and the filtrate was concentrated. The concentrate was purified by preparative HPLC to give the target product.
LC-MS: (ESI, m/z): [M+H]+=476.3.
1H NMR (400 MHz, DMSO-d6) δ 7.91 (d, J=7.9 Hz, 1H), 7.78 (s, 1H), 7.04 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.69 (s, 1H), 5.66-5.46 (m, 3H), 4.54-5.48 (m, 1H), 3.87 (s, 3H), 3.01-2.93 (m, 2H), 2.64-2.53 (m, 2H), 2.35 (s, 3H), 1.93-1.89 (m, 2H), 1.58-1.44 (m, 5H).
Acrylic acid (23.62 g, 328.1 mmol) was added to a solution of 4-aminobenzoic acid (15.0 g, 109.4 mmol) in AcOH/H2O (40 mL/200 mL), and the resulting mixture was stirred at 110° C. for 12 h. The reaction liquid was cooled and filtered to give the target product 4-((2-carboxyethyl)amino)benzoic acid.
LC-MS: (ESI, m/z): [M+H]+=210.0.
Urea (114.83 g, 1913.8 mmol) was added to a solution of 4-((2-carboxyethyl)amino)benzoic acid (20.0 g, 95.69 mmol) in AcOH (240 mL), and the resulting mixture was stirred at 110° C. for 24 h. After the reaction liquid was cooled, an HCl solution was added to adjust the pH to 1-2. A solid was precipitated, and the mixture was filtered to give the target product 4-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoic acid.
LC-MS: (ESI, m/z): [M+H]+=233.1.
Pentafluorophenol (12.97 g, 70.51 mmol) and DCC (14.52 g, 70.51 mmol) were added to a solution of 4-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoic acid (15.0 g, 64.1 mmol) in DMF (200 mL), and the resulting mixture was stirred at room temperature overnight and extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (300 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=3:7) to give the target product pentafluorophenyl 4-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate.
1H NMR (400 MHz, DMSO-d6) δ 10.59 (s, 1H), 8.20 (d, J=8.7 Hz, 2H), 7.64 (d, J=8.7 Hz, 2H), 3.94 (t, J=6.6 Hz, 2H), 2.76 (t, J=6.6 Hz, 2H).
A reaction liquid of 1-bromo-3-nitro-5-(trifluoromethyl)benzene (50.0 g, 185.1 mmol), Pd(PPh3)2Cl2 (12.9 g, 18.5 mmol), TEA (37.4 g, 370.2 mmol), and tributyl (1-ethoxyvinyl)stannane (86.6 g, 240.6 mmol) in dioxane (300 mL) was stirred at 80° C. for 16 h. Water (900 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with EA (500 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous Na2SO4, and concentrated under reduced pressure. The residue was stirred in a mixed solution of 6 N HCl (200 mL) and THF (200 mL) for 30 min, and then water (500 mL) was added. The resulting mixture was extracted with EA (500 mL×2). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=0-1:9) to give 1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-one.
1H NMR (400 MHz, CDCl3) δ 8.95 (s, 1H), 8.69 (s, 1H), 8.53 (s, 1H), 2.75 (s, 3H).
A mixture of 1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-one (1.5 g, 6.4 mmol) and Pd/C (400 mg) in THF (30 mL) was stirred at 60° C. for 16 h. The reaction liquid was filtered, and the filtrate was concentrated under reduced pressure to give 1-(3-amino-5-(trifluoromethyl)phenyl)ethan-1-one. The resulting product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=203.0.
A mixture of 1-(3-amino-5-(trifluoromethyl)phenyl)ethan-1-one (1.1 g, 5.4 mmol) and (Boc)2O (5.9 g, 27.0 mmol) in dioxane (20 mL) was stirred at 80° C. for 16 h. The reaction liquid was directly concentrated under reduced pressure, and the resulting crude product was purified by column chromatography (EA:PE=0-3:17) to give tert-butyl (3-acetyl-5-(trifluoromethyl)phenyl)carbamate.
LC-MS: (ESI, m/z): [M+H]+=303.0.
1H NMR (400 MHz, DMSO-d6) δ 8.26 (s, 1H), 8.18 (d, J=6.5 Hz, 1H), 8.05 (s, 1H), 2.67 (d, J=11.4 Hz, 3H), 1.41 (d, J=11.7 Hz, 9H).
A mixture of tert-butyl (3-acetyl-5-(trifluoromethyl)phenyl)carbamate (1.0 g, 3.3 mmol), Ti(OEt)4 (1.8 g, 6.6 mmol), diethylene glycol dimethyl ether (2.3 g, 17.5 mmol), and (S)-propane-2-sulfinamide (1.2 g, 9.9 mmol) in THF (10 mL) was stirred at 70° C. for 12 h. Water (50 mL) was added to the reaction liquid, and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=0-1:4) to give tert-butyl (S,E)-(3-(1-((tert-butylsulfinyl)imino)ethyl)-5-(trifluoromethyl)phenyl)carbamate.
LC-MS: (ESI, m/z): [M+H]+=407.0.
L-Selectride (1.0 mol/L/THF, 3.6 mL) was added to a solution of tert-butyl (S,E)-(3-(1-((tert-butylsulfinyl<sulfinyl>)imino)ethyl)-5-(trifluoromethyl)phenyl)carbamate (1.0 g, 2.4 mmol) in THF (20 mL) at −78° C., and the reaction liquid was stirred at −78° C. for 50 min. Water (50 mL) was added to the reaction liquid to quench the reaction, and then the resulting mixture was extracted with EA (50 mL×2). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=0-1:1) to give tert-butyl (3-((R)-1-(((S)-tert-butylsulfinyl)amino)ethyl)-5-(trifluoromethyl)phenyl)carbamate.
LC-MS: (ESI, m/z): [M+H]+=409.0.
1H NMR (400 MHz, DMSO-d6) δ 9.71 (s, 1H), 7.74 (d, J=5.1 Hz, 2H), 7.39 (s, 1H), 4.15 (q, J=6.7 Hz, 1H), 1.48 (s, 15H), 1.31 (d, J=6.7 Hz, 3H), 1.11 (d, J=2.1 Hz, 3H).
tert-Butyl (3-((R)-1-(((S)-tert-butylsulfinyl)amino)ethyl)-5-(trifluoromethyl)phenyl)carbamate (1.0 g, 2.4 mmol) was stirred in 1 N HCl/EtOAc (20 mL) at 0° C. for 3 h. The reaction liquid was rapidly filtered through a chromatographic column, washed with EtOAc, and then washed with a mixed solvent of DCM/MeOH (5/1). The filtrate was basified with 7 N NH3/MeOH, and the organic phase was washed with water, dried over anhydrous Na2SO4, filtered, and concentrated to give tert-butyl (R)-(3-(1-aminoethyl)-5-(trifluoromethyl)phenyl)carbamate.
LC-MS: (ESI, m/z): [M+H]+=305.1.
1H NMR (400 MHz, DMSO-d6) δ 9.63 (s, 1H), 7.71 (s, 1H), 7.69 (s, 1H), 7.35 (s, 1H), 4.02-3.99 (m, 1H), 1.48 (s, 9H), 1.23 (d, J=6.6 Hz, 3H).
BOP (574 mg, 1.3 mmol) and DBU (380 mg, 2.5 mmol) were added to a solution of methyl (1R,4R)-4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (330 mg, 1.0 mmol) in DMF (5 mL) while stirring.
After stirring for 15 min, tert-butyl (R)-(3-(1-aminoethyl)-5-(trifluoromethyl)phenyl)carbamate hydrochloride (396 mg, 1.3 mmol) was added, and the reaction liquid was stirred at 70° C. for another 12 h. Water (10 mL) was added to the reaction liquid to quench the reaction, and then the resulting mixture was extracted with EA (30 mL×3). The organic phase was dried over anhydrous Na2SO4 and then concentrated under reduced pressure. The resulting crude product was purified by column chromatography (EA:PE=0-1) to give methyl (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=617.4.
1H NMR (400 MHz, DMSO-d6) δ 9.69 (s, 1H), 8.12 (s, 1H), 7.77 (d, J=25.2 Hz, 2H), 7.41 (s, 1H), 7.01 (s, 1H), 5.71-5.55 (m, 1H), 3.90 (s, 3H), 3.62 (s, 3H), 2.96 (d, J=11.8 Hz, 1H), 2.39 (d, J=10.4 Hz, 4H), 2.06 (d, J=12.0 Hz, 2H), 1.89 (d, J=14.0 Hz, 2H), 1.64-1.50 (m, 7H), 1.46 (s, 9H).
LiOH—H2O (48 mg, 1.2 mmol) was added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (250 mg, 0.4 mmol) in THF/H2O (10 mL/5 mL), and the reaction liquid was stirred at 40° C. overnight. The reaction liquid was adjusted to pH with 10% citric acid to be weakly acidic, and then the resulting mixture was extracted with EtOAc (30 mL×3). The organic phases were combined, washed with saturated brine, and concentrated under reduced pressure to give (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=603.1.
1H NMR (400 MHz, DMSO-d6) δ 9.67 (s, 1H), 8.20 (d, J=7.4 Hz, 1H), 8.07 (s, 1H), 7.77 (d, J=24.0 Hz, 2H), 7.41 (s, 1H), 7.00 (s, 1H), 5.68-5.52 (m, 1H), 3.89 (s, 3H), 2.95 (d, J=11.3 Hz, 1H), 2.40-2.23 (m, 4H), 2.06 (d, J=12.0 Hz, 2H), 1.88 (d, J=12.0 Hz, 2H), 1.67-1.41 (m, 16H).
BOP (871 mg, 1.97 mmol) and DBU (577 mg, 3.80 mmol) were added to a solution of methyl (1R,4R)-4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (500 mg, 1.52 mmol) in DMF (10 mL) while stirring. After the resulting mixture was stirred for another 15 min, (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-amine (461 mg, 1.97 mmol) was added. The reaction liquid was then stirred at 70° C. overnight. Water (50 mL) was added to the reaction liquid, and the resulting mixture was extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by flash column chromatography (EA:PE=0-1) to give methyl (1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=547.1.
1H NMR (400 MHz, DMSO-d6) δ 8.62 (s, 1H), 8.34 (d, J=6.0 Hz, 3H), 8.04 (s, 1H), 7.01 (s, 1H), 5.68 (p, J=6.9 Hz, 1H), 3.89 (s, 3H), 3.64 (s, 3H), 2.94 (t, J=11.4 Hz, 1H), 2.48-2.37 (m, 1H), 2.34 (s, 3H), 2.08 (d, J=12.0 Hz, 2H), 1.97-1.86 (m, 2H), 1.69 (d, J=7.1 Hz, 3H), 1.65-1.47 (m, 4H).
LiOH—H2O (240 mg, 8.0 mmol) was added to a solution of methyl (1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylate (300 mg, 0.55 mmol) in THF/H2O (15 mL, 1:2). The reaction liquid was then stirred at 50° C. overnight. Water (50 mL) was added to the reaction liquid to quench the reaction, and then the resulting mixture was extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give (1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=533.3.
1H NMR (400 MHz, DMSO-d6) δ 12.09 (s, 1H), 8.89 (s, 1H), 8.63 (s, 1H), 8.36 (d, J=10.6 Hz, 2H), 8.14 (s, 1H), 7.02 (s, 1H), 5.89-5.64 (m, 1H), 3.92 (s, 3H), 2.95 (t, J=11.6 Hz, 1H), 2.41 (s, 3H), 2.37-2.26 (m, 1H), 2.08 (d, J=10.7 Hz, 2H), 1.96-1.85 (m, 2H), 1.72 (d, J=7.1 Hz, 3H), 1.68-1.44 (m, 4H).
EDCI (34.7 g, 118.2 mmol) and DMAP (1.3 g, 10.7 mmol) were added to a solution of (1R,4R)-4-(methoxycarbonyl)cyclohexane-1-carboxylic acid (20 g, 107.4 mmol) and 2-hydroxyisoindoline-1,3-dione (20 g, 107.4 mmol) in DCM (300 mL). Then the reaction liquid was stirred at room temperature for 2 h. Brine and DCM were added to the reaction liquid. The organic phase was isolated, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (DCM) to give 1-(1,3-dicarbonylisoindolin-2-yl) 4-methyl (1R,4R)-cyclohexane-1,4-dicarboxylate.
LC-MS: (ESI, m/z): [M+Na]+=354.1.
NIS (133 g, 613 mmol) was added to a solution of 2-amino-4-methoxybenzonitrile (86.3 g, 582 mmol) in DMF (1000 mL). Then the reaction liquid was stirred at room temperature for 2 h. Brine and DCM were added to the reaction liquid. The organic phase was isolated, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure to give 2-amino-5-iodo-4-methoxybenzonitrile.
LC-MS: (ESI, m/z): [M+Na]+=296.9.
Ac2O (71.3 g, 700 mmol) was added to a solution of 2-amino-5-iodo-4-methoxybenzonitrile (160 g, 580 mmol) in HOAc (1600 mL). Then the reaction liquid was stirred at room temperature overnight, filtered, and then concentrated to give N-(2-cyano-4-iodo-5-methoxyphenyl)acetamide.
LC-MS: (ESI, m/z): [M+H]+=317.0.
NiBr2(dtbpy) (2.31 g, 4.75 mmol) was added to a solution of N-(2-cyano-4-iodo-5-methoxyphenyl)acetamide (15.0 g, 47.45 mmol), 1-(1,3-dicarbonylisoindolin-2-yl) 4-methyl (1R,4R)-cyclohexane-1,4-dicarboxylate (31.40 g, 94.90 mmol) and Hantzsch-ester (24.00 g, 94.90 mmol) in DMA (237 mL, 0.2 M). The reaction liquid was purged with N2 for 5 min and irradiated with two Kessil PR160-purple LED lamps (30 W High Luminous DEX 2100 LED, λmax=390 nm) for 16 h. The reaction temperature was controlled at 30-40° C. After the reaction was completed, the reaction liquid was poured into water (1 L), and then the resulting mixture was extracted with EtOAc (150 mL×3). The organic phases were combined, washed with saturated brine (400 mL×3), dried over anhydrous Na2SO4, and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (MeOH:DCM=1:99). The eluate containing the product with a concentration value of 80 mL was collected and filtered. The filtrate was concentrated to give a mixture with a cis-to-trans ratio of 1:1, and the filter cake was a mixture with a cis-to-trans ratio of 15:1. The mixture with the cis-to-trans ratio of 15:1 was dissolved in DMA (85 mL), and a 0.5M aqueous K2CO3 solution (200 mL) was added. The resulting mixture was stirred at room temperature for 2 h and filtered to give a mixture with a cis-to-trans ratio of 20:1. The mixture was recrystallized with acetonitrile (180 mL) to give the target product (with a cis-to-trans ratio of 97:2).
LC-MS: (ESI, m/z): [M+H]+=331.1.
1H NMR (400 MHz, CDCl3) δ 8.04 (s, 1H), 7.55 (s, 1H), 7.29 (s, 1H), 3.89 (s, 3H), 3.69 (s, 3H), 2.90-2.84 (m, 1H), 2.35-2.26 (m, 1H), 2.25 (s, 3H), 2.11-2.08 (m, 2H), 1.92-1.89 (m, 2H), 1.63-1.56 (m, 2H), 1.39-1.33 (m, 2H).
Methyl (1R,4R)-4-(4-acetylamino-5-cyano-2-methoxyphenyl)cyclohexane-1-carboxylate (3 g, 9.08 mmol) was added to i-PrOH (80 mL), and the resulting mixture was purged with HCl gas at room temperature for 1 h. The reaction liquid was filtered, and the solid was dried to give methyl (1R,4R)-4-(7-methoxy-2-methyl-4-carbonyl-3,4-dihydroquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=331.1.
1H NMR (400 MHz, DMSO-d6) δ 7.87 (s, 1H), 7.23 (s, 1H), 3.95 (s, 3H), 3.61 (s, 3H), 2.98-2.87 (m, 1H), 2.58 (s, 3H), 2.47-2.39 (m, 1H), 2.06-1.98 (m, 2H), 1.88-1.80 (m, 2H), 1.59-1.42 (m, 4H).
A reaction liquid of 1-(4-bromothiophen-2-yl)ethan-1-one (10.0 g, 48.80 mmol), (R)-2-methylpropane-2-sulfinamide (8.90 g, 73.20 mmol), and Ti(OEt)4 (27.8 g, 122.00 mmol) in THF (70 mL) was stirred at 80° C. for 3 h. Water (200 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with EA (100 mL×3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (DCM:MeOH=100:1) to give (R,Z)—N-(1-(4-bromothiophen-2-yl)ethylidene)-2-methylpropane-2-sulfinamide.
1H NMR (400 MHz, DMSO-d6) δ 7.96 (d, J=1.2 Hz, 1H), 7.85 (d, J=1.2 Hz, 1H), 2.68 (s, 3H), 1.19 (s, 9H).
9-BBN/THF (0.5 M, 200 mL) was added to a solution of (R,Z)—N-(1-(4-bromothiophen-2-yl)ethylidene)-2-methylpropane-2-sulfinamide (7.0 g, 22.7 mmol) in THE (10 mL). The reaction liquid was stirred at 25° C. for 0.5 h. MeOH (10 mL) was added to the reaction liquid to quench the reaction, then water (100 mL) was added, and the resulting mixture was extracted with EA (100 mL×3). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (DCM:MeOH=100:1) to give (R)—N—((R)-1-(4-bromothiophen-2-yl)ethyl)-2-methylpropane-2-sulfinamide.
1H NMR (400 MHz, DMSO-d6) δ 7.54 (d, J=1.2 Hz, 1H), 7.08 (s, 1H), 5.93 (d, J=7.2 Hz, 1H), 4.64-4.58 (m, 1H), 1.48 (d, J=6.8 Hz, 3H), 1.12 (s, 9H).
HCl (12 M, 454 mmol) was added to a solution of (R)—N—((R)-1-(4-bromothiophen-2-yl)ethyl)-2-methylpropane-2-sulfinamide (6.0 g, 19.34 mmol) in 1,4-dioxane (60 mL). The reaction liquid was then stirred at 25° C. for 2 h. The reaction liquid was directly concentrated. The resulting crude product was stirred in an MTBE (100 mL) solution, filtered, and dried to give (R)-1-(4-bromothiophen-2-yl)ethan-1-amine hydrochloride.
1H NMR (400 MHz, DMSO-d6) δ 8.62 (s, 3H), 7.71 (s, 1H), 7.32 (s, 1H), 4.71-4.67 (m, 1H), 1.57 (d, J=6.8 Hz, 3H).
DBU (3.45 g, 22.67 mmol) was added to a solution of methyl (1R,4R)-4-(7-methoxy-2-methyl-4-carbonyl-3,4-dihydroquinazolin-6-yl)cyclohexane-1-carboxylate (3.00 g, 9.07 mmol) and BOP (5.21 g, 11.79 mmol) in DMF (35 mL) at room temperature. After stirring for 15 min, a solution of (R)-1-(4-bromothiophen-2-yl)ethan-1-amine hydrochloride (2.86 g, 11.79 mmol) in DMF (35 mL) was added dropwise. Then the reaction liquid was stirred at room temperature for 20 min, and then heated to 70° C. and stirred for another 8 h. The reaction liquid was purified by reversed-phase column chromatography to give methyl (1R,4R)-4-(4-(((R)-1-(4-bromothiophene-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=518.0, 520.0.
LiOH (0.81 g, 19.30 mmol) was added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-bromothiophen-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (2.00 g, 3.86 mmol) in THF/H2O (20.0 mL/20.0 mL). The reaction liquid was stirred at 25° C. for 16 h. acidified with a 2 N HCl solution, and a precipitate was formed. The mixture was filtered, and the filter cake was dried under vacuum to give (1R,4R)-4-(4-(((R)-1-(4-bromothiophene-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=504.0, 506.0.
Benzyltrimethylammonium hydroxide (6.03 g, 36.057 mmol) and acrylonitrile (5.727 g, 79.32 mmol) were added to a solution of methyl 4-(2-methoxy-2-oxoethane)benzoate (15.0 g, 72.12 mmol) intoluene (200 mL). The resulting mixture was stirred at 30° C. for 16 h and extracted with ethyl acetate (100 mL×5). The organic phase was washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=3:7) to give the target product methyl 4-(4-cyano-1-methoxy-1-oxobutan-2-yl)benzoate.
1H NMR (400 MHz, DMSO-d6) δ 7.95 (d, J=4.0 Hz, 2H), 7.46 (d, J=6.0 Hz, 2H), 3.90-3.80 (m, 4H), 3.62 (s, 3H), 2.48-2.38 (m, 2H), 2.36-2.26 (m, 1H), 2.10-1.96 (m, 1H).
Methyl 4-(4-cyano-1-methoxy-1-oxobutan-2-yl)benzoate (8.5 g, 32.56 mmol) was dissolved in acetic acid (30 mL), and sulfuric acid (15 mL) was added. The resulting mixture was stirred at 110° C. for 12 h and extracted with dichloromethane/methanol (10/1, 150 mL×3). The organic phase was washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:10) to give 4-(2,6-dioxopiperidin-3-yl)benzoate.
1H NMR (400 MHz, DMSO-d6) δ 12.89 (s, 1H), 10.89 (s, 1H), 7.91 (d, J=4.0 Hz, 2H), 7.36 (d, J=4.0 Hz, 2H), 4.02-3.92 (m, 1H), 2.72-2.63 (m, 1H), 2.56-2.45 (m, 1H), 2.30-2.20 (m, 1H), 2.10-2.00 (m, 1H).
4-(2,6-Dioxopiperidin-3-yl)benzoic acid (1.3 g, 5.579 mmol) was dissolved in DMF (100 mL), and pentafluorophenol (1.129 g, 6.14 mmol) and DCC (1.265 g, 6.14 mmol) were added. The resulting mixture was stirred at room temperature overnight and extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=3:7) to give pentafluorophenyl 4-(2,6-dioxopiperidin-3-yl)benzoate.
1H NMR (400 MHz, DMSO-d6) δ 10.95 (s, 1H), 8.17 (d, J=4.0 Hz, 2H), 7.56 (d, J=4.0 Hz, 2H), 4.14-4.08 (m, 1H), 2.79-2.69 (m, 1H), 2.66-2.55 (m, 1H), 2.30-2.23 (m, 1H), 2.15-2.05 (m, 1H).
3-Amino-4-methoxybenzoic acid (5.0 g, 2.93 mmol) was suspended in acrylic acid (8.05 mL, 117 mmol), and the suspension was stirred at 100° C. for reaction for 3 h. Then the reaction liquid was cooled to room temperature while stirring. Acetic acid (33 mL) was added, and the stirred suspension was heated at 100° C. for 10 min. Urea (11.00 g, 183 mmol) was added, and the reaction liquid was stirred at 120° C. for reaction overnight. The reaction liquid was added to a mixture of ice water and concentrated hydrochloric acid (37%), and the mixture was stirred. The resulting suspension was stored in a refrigerator at 5° C. overnight and filtered, and the solid was washed with water and dried to give a solid. The solid was triturated in a hydrochloric acid solution (0.05 M), filtered, washed with methyl tert-butyl ether, and dried under reduced pressure at 40° C. to give the target product.
1H NMR (400 MHz, DMSO-d6) δ 12.70 (s, 1H), 10.34 (s, 1H), 7.92 (dd, J=8.6, 2.2 Hz, 1H), 7.83 (d, J=2.2 Hz, 1H), 7.21 (d, J=8.8 Hz, 1H), 3.94-3.82 (m, 3H), 3.60 (t, J=6.7 Hz, 2H), 2.69 (s, 2H).
A mixture of 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoic acid (2.0 g, 7.58 mmol), 2,3,4,5,6-pentafluorophenol (1.67 g, 9.09 mmol), and N,N-dicyclohexylcarbodiimide (1.87 g, 9.09 mmol) in an N,N-dimethylformamide (20 mL) solution was stirred at room temperature for reaction for 3 h. The reaction liquid was poured into water (200 mL), and the resulting solution was stirred for 0.5 h and extracted with ethyl acetate (200 mL×3). The organic phase was collected, washed with water (500 mL×2) and saturated brine (300 mL), dried over anhydrous sodium sulfate, filtered, and concentrated. The concentrate was purified by column chromatography to give the target product pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidine-1(2H)-yl)-4-methoxybenzoate.
LC-MS: (ESI, m/z):[M+H]+=431.1.
Intermediate 19 was prepared by reference to the preparation method of intermediate 18.
LC-MS: (ESI, m/z):[M+H]+=419.0.
Intermediate 19 was prepared by reference to the preparation method of intermediate 18.
LC-MS: (ESI, m/z):[M+H]+=401.0.
K3PO4 (255 mg, 1.2 mmol), (PPh3)2PdCl2 (34 mg, 0.048 mmol), and (2-formylphenyl) boronic acid (87 mg, 0.58 mmol) were added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-bromothiophen-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (250 mg, 0.48 mmol) in 1,4-dioxane/H2O (1.5 mL/0.5 mL).
The reaction liquid was stirred at 100° C. overnight. The reaction liquid was concentrated and then purified by column chromatography (PE/EA=2/1 to 1/2) to give methyl (1R,4R)-4-(4-(((R)-1-(4-(2-formylphenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=544.1.
MeOH (3 mL), DCE (3 mL), and AcOH (5 drops) were added to a mixture of methyl (1R,4R)-4-(4-(((R)-1-(4-(2-formylphenyl)thiophen-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (200 mg, 0.36 mmol) and dimethylamine hydrochloride (60 mg, 0.74 mmol). The above solution was stirred at room temperature for 1 h, and NaBH3CN (46 mg, 0.74 mmol) was added. Then the reaction liquid was stirred at room temperature for another 3 h. DCM and water were added to the reaction liquid. The organic phase was isolated, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC to give methyl (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=573.3
1H NMR (400 MHz, CDCl3) δ 7.44-7.34 (m, 3H), 7.33-7.28 (m, 4H), 7.12 (s, 1H), 6.11-6.02 (m, 1H), 5.69-5.60 (m, 1H), 3.92 (s, 3H), 3.69 (s, 3H), 3.38 (s, 2H), 3.04-2.98 (m, 1H), 2.63 (s, 3H), 2.44-2.33 (m, 1H), 2.24-2.18 (m, 6H), 2.16-2.07 (m, 2H), 2.05-1.95 (m, 2H), 1.82-1.79 (d, J=6.8 Hz, 3H), 1.67-1.60 (m, 4H).
PPh3 (22.13 g, 84.48 mmol) was added to a solution of CBr4 (14.0 g, 42.24 mmol) in DCM (200 mL) while stirring at 0° C., and the resulting mixture was stirred at 0° C. for another 30 min, followed by the addition of tert-butyl 4-formylpiperidine-1-carboxylate (6.0 g, 28.16 mmol). The reaction liquid was stirred at room temperature overnight, diluted with EA (200 mL), and filtered. The filtrate was concentrated under reduced pressure. The resulting crude product was purified by flash column chromatography (EA:PE=0-3:7) to give tert-butyl 4-(2,2-dibromovinyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M-tBu]+=313.9.
n-BuLi (13.6 mL, 1.6 M/n-hexane) was added to a solution of tert-butyl 4-(2,2-dibromovinyl)piperidine-1-carboxylate (4.0 g, 10.86 mmol) in THF (60 mL) at −78° C. The resulting mixture was stirred at −78° C. for 1 h, and then paraformaldehyde (977 mg, 32.60 mmol) was added. The reaction liquid was stirred at room temperature overnight. NH4Cl (100 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with EA (100 mL×3). The organic phases were combined, washed with saturated brine (300 mL), dried over anhydrous Na2SO4, filtered, and concentrated. The resulting crude product was purified by flash column chromatography (EA:PE=0-4:1) to give tert-butyl 4-(3-hydroxyprop-1-yn-1-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M-Boc]+=140.2.
1H NMR (400 MHz, DMSO-d6) δ 5.04 (t, J=5.9 Hz, 1H), 4.04 (dd, J=5.8, 2.0 Hz, 2H), 3.65-3.55 (m, 2H), 3.04 (t, J=10.1 Hz, 2H), 2.60 (ddd, J=8.8, 7.0, 3.5 Hz, 1H), 1.77-1.64 (m, 2H), 1.43-1.31 (m, 11H).
CBr4 (1.94 g, 5.83 mmol) was added to a solution of tert-butyl 4-(3-hydroxyprop-1-yn-1-yl)piperidine-1-carboxylate (700 mg, 2.92 mmol) and PPh3 (1.94 g, 5.83 mmol) in DCM (10 mL) while stirring at 0° C. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with EA (10 mL) and then filtered.
The filtrate was concentrated under reduced pressure. The resulting crude product was purified by flash column chromatography (EA:PE=0-3:7) to give tert-butyl 4-(3-bromoprop-1-yn-1-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M-tBu]+=246.0.
1H NMR (400 MHz, DMSO-d6) δ 4.24 (d, J=2.0 Hz, 2H), 3.64-3.54 (m, 2H), 3.13-3.00 (m, 2H), 2.75-2.65 (m, 1H), 1.78-1.68 (m, 2H), 1.44-1.33 (m, 11H).
NaH (60%, 1.2 g, 30.58 mmol) was added to a solution of tert-butyl 4-(2-hydroxyethyl)piperidine-1-carboxylate (1.0 g, 4.36 mmol) in THF (20 mL) at 0° C., and the resulting mixture was stirred at 0° C. for 20 min. After warming to room temperature, 4-(bromomethyl)pyridine hydrogen bromide (1.3 g, 5.13 mmol) was added, and the reaction mixture was stirred at room temperature for reaction overnight. Water (30 mL) was added to the reaction liquid, and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with brine (30 mL), dried over Na2SO4, and concentrated under reduced pressure. The concentrate was purified by flash column chromatography (EA:PE=0-4:1) to give the target product tert-butyl 4-(2-(pyridin-4-ylmethoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=321.1.
1H NMR (400 MHz, CDCl3) δ 8.57 (d, J=5.9 Hz, 2H), 7.24 (d, J=5.7 Hz, 2H), 4.51 (s, 2H), 4.09 (d, J=15.9 Hz, 2H), 3.55 (t, J=6.1 Hz, 2H), 2.70 (t, J=12.2 Hz, 2H), 1.66 (d, J=13.9 Hz, 2H), 1.58 (m, 2H), 1.45 (s, 9H), 1.26 (m, 1H), 1.13 (d, J=8.4 Hz, 2H).
A mixture of tert-butyl 4-(2-(pyridin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (1.9 g, 5.9 mmol) and Pd/C (570 mg) in i-PrOH/H2O (30 mL/35 mL) was stirred at 75° C. overnight. The reaction liquid was filtered through celite, and the filtrate was concentrated to give the target product tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate. The resulting product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=327.2.
2-(Piperidin-4-yl)ethan-1-ol (467 mg, 3.6 mmol) and DIEA (1.4 g, 10.8 mmol) were added to a solution of 2-(2,6-dioxopiperidin-3-yl)-4-fluoroisoindoline-1,3-dione (1.0 g, 3.6 mmol) in DMF (10 mL). The reaction liquid was stirred at 100° C. overnight. Water (100 mL) was added to the reaction liquid to quench the reaction, and then the resulting mixture was extracted with EA (20 mL×3). The organic phases were combined, washed with saturated brine (60 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by flash column chromatography (EA:PE=0-1:1) to give 2-(2,6-dioxopiperidin-3-yl)-4-(4-(2-hydroxyethyl)piperidin-1-yl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M+H]+=386.1.
IBX (1.7 g, 6.0 mmol) was added to a solution of 2-(2,6-dioxopiperidin-3-yl)-4-(4-(2-hydroxyethyl)piperidin-1-yl)isoindoline-1,3-dione (1.1 g, 3.0 mmol) in ACN (20 mL) while stirring. The reaction liquid was stirred at 80° C. for 2 h. filtered, and the filtrate was concentrated. The resulting crude product was purified by flash column chromatography (EA:PE=0-1:1) to give 2-(1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)piperidin-4-yl)acetaldehyde.
LC-MS: (ESI, m/z): [M+H]+=384.0.
1H NMR (400 MHz, DMSO-d6) δ 11.08 (s, 1H), 9.72 (s, 1H), 7.77-7.58 (m, 1H), 7.34 (dd, J=7.7, 4.9 Hz, 2H), 5.09 (dd, J=12.9, 5.4 Hz, 1H), 3.68 (d, J=11.8 Hz, 2H), 2.97-2.82 (m, 3H), 2.64-2.53 (m, 2H), 2.45 (d, J=6.6 Hz, 2H), 2.13-1.99 (m, 3H), 1.82-1.72 (m, 2H), 1.50-1.30 (m, 2H).
TFA (4.0 mL) was added to tert-butyl 4-(3-hydroxypropyl)piperidine-1-carboxylate (930.0 mg, 3.8 mmol) in DCM (8.00 mL). The reaction liquid was stirred at room temperature overnight. The reaction liquid was directly concentrated to give 3-(piperidin-4-yl)propan-1-ol. The resulting crude product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=144.2.
2-(2,6-Dioxopiperidin-3-yl)-4-fluoroisoindoline-1,3-dione (1.0 g, 3.8 mmol) and DIEA (1.5 g, 11.4 mmol) were added to a solution of 3-(piperidin-4-yl)propan-1-ol (730 mg, TFA salt, 3.8 mmol) in DMF (10 mL) while stirring.
The reaction liquid was stirred at 100° C. overnight. Water (100 mL) was added to the reaction liquid to quench the reaction, and then the resulting mixture was extracted with EA (20 mL×3). The organic phase was washed with saturated brine (60 mL), dried over anhydrous Na2SO4, filtered, and concentrated. The resulting crude product was purified by flash column chromatography (EA:PE=0-1:1) to give 2-(2,6-dioxopiperidin-3-yl)-4-(4-(3-hydroxypropyl)piperidin-1-yl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M+H]+=400.0.
IBX (982 mg, 3.5 mmol) was added to a solution of 2-(2,6-dioxopiperidin-3-yl)-4-(4-(3-hydroxyethyl)piperidin-1-yl)isoindoline-1,3-dione (700 mg, 1.7 mmol) in ACN (20 mL) while stirring. The reaction liquid was stirred at 80° C. for 2 h. filtered, and the filtrate was concentrated. The resulting crude product was purified by flash column chromatography (EA:PE=0-1:1) to give 3-(1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindolin-4-yl)piperidin-4-yl)propionaldehyde.
LC-MS: (ESI, m/z): [M+H]+=398.0.
1H NMR (400 MHz, DMSO-d6) δ 11.08 (s, 1H), 9.71 (t, J=1.4 Hz, 1H), 7.74-7.60 (m, 1H), 7.33 (dd, J=7.7, 3.4 Hz, 2H), 5.09 (dd, J=12.9, 5.4 Hz, 1H), 3.69 (d, J=11.9 Hz, 2H), 2.93-2.79 (m, 3H), 2.64-2.46 (m, 4H), 2.07-2.00 (m, 1H), 1.80-1.72 (m, 2H), 1.54 (dd, J=14.0, 7.1 Hz, 2H), 1.40-1.27 (m, 3H) Intermediate 26: benzyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate
tert-Butyl 9-formyl-3-azaspiro[5.5]undecane-3-carboxylate (1.0 g, 3.56 mmol) and (9H-fluoren-9-yl)methylpiperazine-1-carboxylate (1.35 g, 3.91 mmol) were dissolved in THF (20 mL), and STAB (2.26 g, 10.68 mmol) was added. The resulting mixture was stirred at room temperature overnight, and water (100 mL) was added, followed by extraction with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA/PE=3:2) to give tert-butyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=574.2.
tert-Butyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (1.1 g, 1.92 mmol) was dissolved in DCM (10 mL), and TFA (2 mL) was added. The resulting mixture was stirred at room temperature for 1 h and concentrated under reduced pressure to give (9H-fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-carboxylate, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=474.1.
(9H-Fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-carboxylate (1.1 g, 2.33 mmol) was dissolved in ACN (20 mL), and a saturated NaHCO3 solution (20 mL) was added, followed by the addition of CbzCl (0.60 g, 3.50 mmol) while stirring. The resulting mixture was reacted at room temperature overnight and then concentrated under reduced pressure to remove acetonitrile. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA/PE=11:9) to give benzyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=608.1.
Benzyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (0.97 g, 1.60 mmol) was dissolved in CAN (10 mL), and piperidine (2 mL) was added. The resulting mixture was stirred at room temperature for 3 h and then concentrated under reduced pressure to remove the solvent. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH/DCM=3:17) to give benzyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=386.1.
2,6-Dimethyl-1λ2-piperazine (51.4 g, 480.37 mmol) and Tf2O (90.38 g, 320.50 mmol) were added to a solution of methyl 4-oxocyclohexane-1-carboxylate (50.0 g, 320.51 mmol) in dichloromethane (500 mL). The resulting mixture was stirred at 0° C. for 1 h, and then Tf2O (72.3 g, 256.38 mmol) was added. The mixture was stirred at room temperature overnight, and the organic phase was washed with water, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was subjected to column chromatography (PE:EA=5:1) to give methyl 4-(((trifluoromethoxy)sulfonyl)oxy)cyclohex-3-ene-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 5.90 (s, 1H), 3.63 (s, 3H), 2.70-2.60 (m, 1H), 2.47-2.28 (m, 4H), 2.11-1.98 (m, 1H), 1.81 (tt, J=8.9, 6.0 Hz, 1H).
(Bpin)2 (39.1 g, 153.94 mmol), Pd(dppf)2Cl2(7.5 g, 10.24 mmol), and potassium acetate (30.4 g, 310.2 mmol) were added to a solution of methyl 4-(((trifluoromethoxy)sulfonyl)oxy)cyclohex-3-ene-1-carboxylate (30.0 g, 104.17 mmol) in dioxane (500 mL), and the resulting mixture was stirred at 90° C. overnight. The reaction liquid was diluted with water (2 L), and the resulting mixture was extracted with ethyl acetate (500 mL×3). The organic phase was washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=0:100) to give methyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate.
1H NMR (400 MHz, CDCl3) δ 6.53 (s, 1H), 3.71-3.64 (m, 3H), 2.61-2.46 (m, 1H), 2.42-1.95 (m, 6H), 1.28-1.19 (m, 12H).
Methyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate (25.0 g, 93.98 mmol), Pd(dppf)2Cl2 (5.3 g, 7.24 mmol), and Na2CO3 (15.3 g, 144.34 mmol) were added to a solution of 6-bromo-7-methoxy-2-methylquinazolin-4-ol (19.4 g, 72.12 mmol) in DMF/H2O (300 mL/30 mL), and the resulting mixture was stirred at 110° C. overnight. The reaction liquid was diluted with water (1 L), and the resulting mixture was extracted with ethyl acetate (500 mL×3). The organic phase was washed with saturated brine (500 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=0:100) to give the product methyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=329.0.
Methyl (1R,4R)-4-(4-(((R)-1-(4-(2-formylphenyl)thiophen-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (180 mg, 0.33 mmol) and a methylamine solution (0.33 mL, 0.66 mmol, 2 M in THF) were added to MeOH/DCE (2 mL/2 mL), and then a few drops of AcOH (4 drops) were added dropwise. The mixed solution was stirred at room temperature for 1 h, and then NaBH3CN (42 mg, 0.66 mmol) was added. The resulting mixture was stirred for reaction for another 3 h. DCM and water were added to the reaction liquid, and the organic phase was isolated, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by Prep-HPLC to give methyl (1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thiophen-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=559.4.
1H NMR (400 MHz, CDCl3) δ 7.42-7.39 (m, 1H), 7.35-7.26 (m, 5H), 7.24-7.20 (m, 1H), 7.10 (s, 1H), 6.08-6.01 (m, 1H), 5.65-5.60 (m, 1H), 3.92 (s, 3H), 3.73-3.69 (m, 5H), 3.04-2.98 (m, 1H), 2.63 (s, 3H), 2.44-2.36 (m, 4H), 2.14-1.99 (m, 4H), 1.80 (d, J=6.8 Hz, 3H), 1.70-1.53 (m, 4H).
Pd(dppf)Cl2 (100 mg, 0.1 mmol) and KOAc (450 mg, 4.6 mmol) were added to a solution of 2-bromo-3-chlorobenzaldehyde (500 mg, 2.3 mmol) and 4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bis(1,3,2-dioxaborolane) (700 mg, 2.7 mmol) in 1,4-dioxane (5 mL), and the mixture was purged 3 times with N2 and stirred at 100° C. for 2 h. The reaction liquid was poured into water and extracted with EA (30 mL×3), and the organic phase was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=0-1:4) to give 3-chloro-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzaldehyde.
1H NMR (400 MHz, CDCl3) δ 9.93 (s, 1H), 7.69 (dd, J=7.4, 0.7 Hz, 1H), 7.61-7.52 (m, 1H), 7.47 (t, J=7.7 Hz, 1H), 1.48 (s, 12H).
Pd(dppf)Cl2 (30 mg, 0.004 mmol) and K2CO3 (110 mg, 0.6 mmol) were added to a solution of 3-chloro-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzaldehyde (127 mg, 0.48 mmol) and methyl (1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl-1-carboxylate (200 mg, 0.4 mmol) in DMF/H2O (3 mL/0.5 mL), and the reaction liquid was purged 3 times with N2 and stirred at 100° C. for 2 h. The reaction liquid was poured into water and extracted with EA (20 mL×2), and the organic phase was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=0-3:2) to give methyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-formylphenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=578.2.
NaBH(OAc)3 (254 mg, 1.2 mmol) was added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-formylphenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (200 mg, 0.4 mmol) and dimethylamine (0.8 mL, 2 mmol) in dichloroethane (5 mL), and the mixture was stirred at 80° C. for 1 h. The reaction liquid was poured into water and extracted with DCM (20 mL×2), and the organic phase was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (DCM:MeOH=19:1) to give methyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=607.3.
LiOH (36 mg, 1.5 mmol) was added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (230 mg, 0.38 mmol)) in MeOH/H2O (1 mL), and the mixture was stirred at 60° C. for 1 h. HCl (1 mol/L, aq) was poured into the reaction liquid to adjust the pH to 3-4, and then the pH was adjusted to 8-9 with NaHCO3 (aq). The mixture was extracted with EA (30 mL×2), and the organic phase was dried over Na2SO4, filtered and concentrated under reduced pressure to give (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=593.3.
A solution of tert-butyl 9-((4-(((9H-fluorenyl-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (500 mg, 0.87 mmol) in dioxane hydrochloride (4 M, 3 mL) was stirred at room temperature for 1 h. The reaction liquid was concentrated under reduced pressure to give (9H-fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate. The resulting product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=474.2.
(9H-fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (400 mg, 0.84 mmol) was dissolved in DMSO (4 mL), and then pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (367 mg, 0.84 mmol) and DIEA (325 mg, 2.5 mmol) were added to the solution. The reaction liquid was stirred at room temperature for 16 h. The reaction liquid was poured into water, extracted with EA (20 mL×3), dried over anhydrous Na2SO4 and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (DCM:MeOH=19:1) to give (9H-fluoren-9-yl)methyl 4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=724.3.
Piperidine (587 mg, 6.9 mmol) was added to a solution of (9H-fluoren-9-yl)methyl 4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (500 mg, 0.69 mmol) in DCM (5 mL). The reaction liquid was stirred at room temperature for 3 h. The reaction liquid was poured into water, extracted with DCM (20 mL×3), dried over anhydrous Na2SO4 and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (DCM:MeOH=19:1) to give 1-(2-chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=502.4.
(1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (70 mg, 0.12 mmol) and 1-(2-chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (60 mg, 0.12 mmol) were dissolved in DMF (3 mL), and HATU (53 mg, 0.14 mmol) and DIEA (50 mg, 0.36 mmol) were added to the solution. The reaction liquid was stirred at 20° C. for 16 h. The reaction liquid was poured into water, extracted with EA (20 mL×3), dried over anhydrous Na2SO4 and concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC to give 1-(2-chloro-5-(9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1076.1.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.32 (d, J=8.2 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.55 (d, J=1.8 Hz, 1H), 7.46-7.29 (m, 4H), 7.24 (d, J=1.3 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 5.96 (t, J=7.4 Hz, 1H), 3.89 (s, 3H), 3.79-3.72 (m, 1H), 3.67-3.51 (m, 3H), 3.46 (s, 4H), 3.18-3.07 (m, 2H), 2.94 (s, 1H), 2.79-2.56 (m, 4H), 2.42 (s, 3H), 2.29 (d, J=26.9 Hz, 4H), 2.12 (s, 2H), 2.01 (s, 6H), 1.90-1.25 (m, 21H), 1.07 (s, 4H).
NMI (533 mg, 6.50 mmol) and TCFH (910 mg, 3.25 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (656 mg, 1.30 mmol) and tert-butyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate (550 mg, 1.56 mmol) in DMF (20 mL). The reaction mixture was stirred at 30° C. for 2 h. After the reaction was completed, the reaction liquid was poured into water (50 mL) and extracted with EA (100 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (MeOH:DCM=0-1:9) to give the target product tert-butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=837.4, 839.4.
TFA (4 mL) was added to a solution of tert-butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (1.1 g, 1.31 mmol) in DCM (12 mL) at room temperature, and the reaction liquid was stirred at room temperature for 2 h. The reaction liquid was concentrated under reduced pressure to give a crude product of (4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone. The resulting crude product was directly used in the next step without being purified.
LC-MS: (ESI, m/z): [M+H]+=737.1, 739.1.
K3PO4 (0.63 g, 2.98 mmol), Ruphos Pd G2 (120 mg, 0.15 mmol) and (2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)boronic acid (590 mg, 2.23 mmol) were added to a mixture of (4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone (1.1 g, 1.49 mmol) in 1,4-dioxane/water (25 mL/5 mL).
The reaction liquid was stirred at 100° C. for 16 h. The reaction liquid was filtered through celite, the filtrate was concentrated under reduced pressure, and the resulting crude product was purified by reversed-phase column chromatography (ACN:H2O=0-100%) to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=878.2.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (95 mg, 0.22 mol) and tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carboxylate (160 mg, 0.18 mmol) were dissolved in DMSO (3 mL), and then DIEA (71 mg, 0.55 mmol) was added. The reaction liquid was stirred at room temperature for 1 h. After the reaction was completed, the reaction liquid was poured into water (50 mL) and the mixture was extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carbamate. The resulting crude product was directly used in the next step without being further purified.
LC-MS: (ESI, m/z): [1/2M+H]+=565.7.
TFA (2 mL) was added to a solution of tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carbamate (250 mg, 0.22 mmol) in DCM (6 mL), and then the mixture was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC to give 1-(2-chloro-5-(9-((4-((1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1028.5.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.28 (d, J=8.2 Hz, 1H), 8.03 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.8 Hz, 1H), 7.48-7.41 (m, 2H), 7.38 (dd, J=8.2, 1.9 Hz, 1H), 7.32-7.25 (m, 4H), 7.01 (s, 1H), 6.08-5.90 (m, 1H), 3.89 (s, 3H), 3.81-3.42 (m, 10H), 2.98-2.89 (m, 1H), 2.77-2.71 (m, 2H), 2.68-2.55 (m, 1H), 2.43 (s, 3H), 2.34-2.21 (m, 7H), 2.15-2.05 (m, 3H), 2.02-1.97 (m, 1H), 1.87-1.49 (m, 18H), 1.26-0.96 (m, 7H). MS
A solution of 1,3-di(piperidin-4-yl)propane (2.0 g, 9.5 mmol) and (Boc)2O (415 mg, 1.9 mmol) in DCM (30 mL) was stirred at 0° C. for 2 h. The reaction liquid was directly concentrated and the resulting crude product was purified by column chromatography (DCM:MeOH=15:1) to give tert-butyl 4-(3-(piperidin-4-yl)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=311.2.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (270 mg, 0.62 mmol) and tert-butyl 4-(3-(piperidin-4-yl)propyl)piperidine-1-carboxylate (230 mg, 0.74 mmol) were dissolved in DMSO (10 mL), and then DIEA (240 mg, 1.86 mmol) was added. The reaction liquid was stirred at 30° C. for 1 h. After the reaction was completed, the reaction liquid was diluted with water (50 mL) and the mixture was extracted with EA (100 mL×3). The organic phases were combined, washed with saturated brine (100 mL), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. EA (50 mL) was added to the resulting crude product, and the mixture was stirred and filtered to give tert-butyl 4-(3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=461.2.
A solution of tert-butyl 4-(3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)piperidine-1-carboxylate (281 mg, 0.50 mmol) in TFA/DCM (2 mL/4 mL) was stirred at 25° C. for 1 h. The reaction liquid was directly concentrated to give 1-(2-chloro-5-(4-(3-(piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione. The resulting crude product was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=461.1.
K3PO4 (1.0 g, 4.83 mmol), Ruphos Pd G2 (156 mg, 0.2 mmol) and (2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)boronic acid (615 mg, 2.32 mmol) were added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (1.0 g, 1.93 mmol) in 1,4-dioxane/H2O (20.0 mL/4.0 mL). The reaction liquid was stirred at 100° C. for 16 h. The reaction liquid was filtered through celite, and the filtrate was concentrated under reduced pressure.
The resulting crude product was purified by reversed-phase column chromatography (acetonitrile:H2O=0-100%) to give methyl (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=659.2.
LiOH (210 mg, 5.0 mmol) was added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (645 mg, 1.0 mmol) in THF/H2O (10.0 mL/10.0 mL). The reaction liquid was stirred at 25° C. for 16 h. The pH of the reaction liquid was adjusted with 2 N HCl, and a precipitate was produced. The mixture was filtered and dried to give (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
1H NMR (400 MHz, CD3OD) δ 8.20 (s, 1H), 7.29-7.03 (m, 7H), 6.20-6.14 (m, 1H), 4.47 (s, 2H), 4.02 (s, 3H), 3.38 (s, 2H), 2.68-2.65 (m, 6H), 2.34 (s, 1H), 2.12 (s, 2H), 1.98-1.96 (m, 2H), 1.85 (d, J=6.8 Hz, 3H), 1.63-1.58 (m, 4H), 1.44-1.28 (m, 10H).
LC-MS: (ESI, m/z): [M+H]+=645.2.
NMI (168 mg, 2.05 mmol) and TCFH (289 mg, 1.03 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (213 mg, 0.33 mmol) and 1-(2-chloro-5-(4-(3-(piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (231 mg, 0.50 mmol) in DMF (5 mL). The reaction liquid was stirred at 30° C. for 16 h. After the reaction was completed, water (25 mL) was added to the reaction liquid and the mixture was extracted with EA (100 mL×3). The organic phases were combined (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (MeOH:DCM=8%) to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-(3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carbamate.
LC-MS: (ESI, m/z): [M+H]+=1087.3.
TFA (2.0 mL) was added to a solution of tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-(3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carbamate (218 mg, 0.20 mmol) in DCM (4.0 mL) at 25° C., and then the mixture was stirred at 25° C. for 1 h. The reaction liquid was directly concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC to give 1-(2-chloro-5-(4-(3-(1-((1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
1H NMR (400 MHz, CD3OD) δ 8.01 (s, 1H), 7.63-7.62 (m, 1H), 7.43-7.41 (m, 2H), 7.41-7.23 (m, 4H), 7.23 (s, 1H), 7.14 (s, 1H), 7.01 (s, 1H), 6.09-6.07 (m, 1H), 4.59-4.52 (m, 2H), 4.10-4.06 (m, 3H), 3.96 (s, 3H), 3.80-3.76 (m, 3H), 3.32-3.30 (m, 3H), 3.30-2.84 (m, 4H), 2.84-2.53 (m, 1H), 2.52 (s, 3H), 2.44 (s, 3H), 2.03-1.57 (m, 17H), 1.38-1.12 (m, 10H).
LC-MS: (ESI, m/z): [M+H]+=987.4.
LiOH (210 mg, 5.0 mmol) was added to a solution of methyl (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (570 mg, 1.0 mmol) in THF/H2O (10.0 mL/10.0 mL). The reaction liquid was stirred at 25° C. for 16 h. The reaction liquid was acidified with 2 N HCl, a precipitate was produced, the mixture was filtered, and the filtrate was concentrated under reduced pressure to give (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=559.2.
HATU (173 mg, 0.456 mmol) and DIEA (118 mg, 0.912 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (170 mg, 0.30 mmol) and tert-butyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate (128 mg, 0.365 mmol) in DMF (5 mL). The reaction liquid was stirred at 30° C. for 2 h. After the reaction was completed, water (25 mL) was added to the reaction liquid and the mixture was extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting crude product was purified by column chromatography (MeOH:DCM=0-1:9) to give tert-butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [1/2(M-100)+H]+=396.8.
TFA (2.0 mL) was added to a solution of tert-butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (300 mg, 0.336 mmol) in DCM (6 mL) at room temperature, and the mixture was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure to give (4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone.
LC-MS: (ESI, m/z): [M+H]+=792.3.
DIEA (130 mg, 1.01 mmol) was added to a solution of pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (175 mg, 0.40 mmol) and (4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone in DMSO (5 mL). Then the reaction liquid was stirred at room temperature for 1 h. After the reaction was completed, the reaction liquid was diluted with water (50 mL) and the mixture was extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC to give 1-(2-chloro-5-(9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.28 (d, J=8.2 Hz, 1H), 8.03 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.7 Hz, 1H), 7.48 (d, J=1.2 Hz, 1H), 7.47-7.26 (m, 6H), 7.01 (s, 1H), 6.09-5.77 (m, 1H), 3.89 (s, 3H), 3.78-3.73 (m, 1H), 3.65-3.39 (m, 7H), 3.29-3.21 (m, 4H), 2.96-2.92 (m, 1H), 2.78-2.56 (m, 3H), 2.43 (s, 3H), 2.28 (d, J=22.1 Hz, 4H), 2.12-1.96 (m, 8H), 1.87-1.41 (m, 19H), 2.26-1.13 (m, 5H).
LC-MS: (ESI, m/z): [M+H]+=1042.5.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (407 mg, 0.092 mmol) and DIEA (356 mg, 2.76 mmol) were added to a solution of tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (300 mg, 0.92 mmol) in DMSO (3 mL), and the reaction liquid was stirred at 25° C. for 16 h. The reaction liquid was poured into water and then extracted with EA (30 mL×2). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The resulting crude product was purified by silica gel column chromatography (EA:PE=0-1:1) to give tert-butyl 4-(2-((1-(4-chloro-3-(2,4-dicarbonyltetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=577.2.
A solution of tert-butyl 4-(2-((1-(4-chloro-3-(2,4-dicarbonyltetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (440 mg, 0.76 mmol) in HCl/dioxane (3 mL) was stirred at room temperature for 1 h. The reaction liquid was directly concentrated under reduced pressure to give 1-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=477.3.
1-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (57 mg, 0.12 mmol), HATU (53 mg, 0.14 mmol) and DIEA (50 mg, 0.36 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (70 mg, 0.12 mmol) in DMF (3 mL). The reaction liquid was stirred at 20° C. for 16 h. The reaction liquid was poured into water and then extracted with EA (20 mL×2). The organic phases were combined, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure.
The resulting crude product was purified by Prep-HPLC to give 1-(2-chloro-5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M−H]−=1049.1.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.31 (d, J=8.2 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.9 Hz, 1H), 7.46-7.32 (m, 4H), 7.24 (d, J=1.3 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.01-5.90 (m, 1H), 4.53-4.33 (m, 2H), 3.96-3.85 (m, 4H), 3.80-3.71 (m, 1H), 3.65-3.54 (m, 2H), 3.40 (t, J=6.5 Hz, 2H), 3.23 (d, J=6.3 Hz, 2H), 3.15-3.10 (m, 2H), 3.02-2.88 (m, 3H), 2.80-2.60 (m, 4H), 2.42 (s, 3H), 2.01 (s, 6H), 1.89-1.25 (m, 20H), 1.20-0.89 (m, 4H).
(R)-4-(4-((1-(3-Amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (220 mg, 0.44 mmol) was dissolved with ethyl acetate (12 mL) and cooled to 0° C., and N-Boc piperazine (163 mg, 0.88 mmol), DIEA (169 mg, 1.31 mmol), and T3P (209 mg, 0.66 mmol) were separately added. The reaction liquid was stirred at 0° C. for 3 h. Water (30 mL) was added to quench the reaction, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed once with saturated brine (20 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure, and the crude product was purified by column chromatography (PE:EA=10:1) to give tert-butyl (R)-4-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=671.3.
tert-Butyl (R)-4-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazine-1-carboxylate (220 mg, 0.33 mmol) was added to a solution of hydrogen chloride in ethyl acetate (3 M/EA, 10 mL). The reaction liquid was stirred at room temperature for 2 h and directly concentrated under reduced pressure to give (R)-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(piperazin-1-yl)methanone, which was directly used in the next step without purification.
(R)-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(piperazin-1-yl)methanone (115 mg, 0.165 mmol) was dissolved in tetrahydrofuran (5 mL), 2-(3-(4-chloro-3-(2,4-oxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)acetaldehyde (39 mg, 0.09 mmol) and sodium triacetoxyborohydride (55 mg, 0.27 mmol) were added sequentially, and the reaction liquid was stirred at room temperature for 2 h, quenched with water (10 mL), extracted with dichloromethane (30 mL×3), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was subjected to preparative reversed-phase chromatography (first purified with a formic acid system, followed by an ammonium bicarbonate system) to give 1-(5-(9-(2-(4-((1S,4S)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1000.1.
δ 10.50 (s, 1H), 8.31 (d, J=7.9 Hz, 1H), 7.95 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.9 Hz, 1H), 7.38 (dd, J=8.2, 1.8 Hz, 1H), 6.98 (s, 1H), 6.88 (s, 1H), 6.84 (s, 1H), 6.69 (s, 1H), 5.66-5.42 (m, 3H), 3.87 (s, 3H), 3.75-3.50 (m, 5H), 3.50-3.40 (m, 5H), 3.00-2.70 (m, 5H), 2.39-2.23 (m, 7H), 2.00-1.84 (m, 4H), 1.75-1.63 (m, 6H), 1.59-1.18 (m, 13H), 1.13-0.96 (m, 4H).
Acetaldehyde (40% aqueous solution, 16 mg, 0.142 mmol) was added to a solution of 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione (70 mg, 0.071 mmol) in THE (3 ml). The mixture was stirred at room temperature for 30 min.
Sodium triacetoxyborohydride (75 mg, 0.355 mmol) was added, and the resulting mixture was stirred at room temperature overnight. The reaction liquid was diluted with water (15 mL), extracted with DCM (20 mL×3), dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure, and the resulting crude product was purified by prep-HPLC to give 1-(2-chloro-5-(9-((4-((1R,4R)-4-(4-(((R)-1-(3-(ethylamino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1014.3.
1H NMR (400 MHz, DMSO-d6) δ 10.52 (s, 1H), 8.13 (s, 1H), 8.07 (s, 1H), 7.64 (d, J=8.3 Hz, 1H), 7.55 (s, 1H), 7.39 (d, J=8.0 Hz, 1H), 7.00 (s, 1H), 6.90 (d, J=6.5 Hz, 2H), 6.64 (s, 1H), 6.05 (s, 1H), 5.60 (t, J=6.4 Hz, 1H), 3.89 (s, 3H), 3.75-3.40 (m, 8H), 3.11-3.00 (m, 2H), 3.00-2.90 (m, 1H), 2.87-2.70 (m, 2H), 2.70-2.60 (m, 1H), 2.40-2.25 (m, 7H), 2.18-2.07 (m, 2H), 1.90-1.75 (m, 4H), 1.75-1.20 (m, 18H), 1.18-0.96 (m, 7H).
5-Amino-2-chloropyridine-4-carboxylic acid (14.0 g, 81.13 mmol) was dissolved with DCM (200 mL), and HOBt (13.2 g, 97.67 mmol) and EDCI (18.7 g, 97.67 mmol) were added. The reaction liquid was stirred at room temperature for 1 h. NH3·H2O (200 mL) was added, and the resulting mixture was stirred for 2 h, filtered, and concentrated under reduced pressure to give the crude product 5-amino-2-chloropyridine-4-carboxamide.
LC-MS: (ESI, m/z): [M+H]+=172.0.
5-Amino-2-chloropyridine-4-carboxamide (5.0 g, 29.24 mmol) was dissolved with dioxane/H2O (200 mL/20 mL), and Cs2CO3 (47.51 g, 146.18 mmol), Pd(dppf)Cl2 (2.67 g, 3.65 mmol), and methyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate (10.1 g, 38.1 mmol) were separately added. The reaction liquid was heated to 100° C. under nitrogen atmosphere, stirred overnight, diluted with water (500 mL), extracted with ethyl acetate (600 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure, and the crude product was purified by column chromatography (PE:EA=1:3) to give methyl 4-(5-amino-4-carbamoylpyridin-2-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=276.0.
Methyl 4-(5-amino-4-carbamoylpyridin-2-yl)cyclohex-3-ene-1-carboxylate (4.0 g, 14.55 mmol) was added to a solution of trimethyl orthoacetate (17.46 g, 145.5 mmol) in methanol (100 ml). The reaction liquid was stirred at 120° C. overnight in a sealed container, cooled, and concentrated under reduced pressure to give the crude product methyl 4-(4-hydroxy-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=300.0.
Methyl 4-(4-hydroxy-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohex-3-ene-1-carboxylate (800 mg, 3.33 mmol) and Pd(OH)2 (320 mg) were added to a solution of methanol (50 mL), and the reaction liquid was purged 3 times with hydrogen, warmed to 70° C. and stirred overnight, filtered and concentrated under reduced pressure. The crude product was dissolved in a solution of methanol (40 mL). Sodium methoxide (1 mL, 5.4 mmol/L) was added, and the mixture was stirred at 70° C. overnight. The pH was adjusted to 3-4 with H2SO4, and the mixture was stirred overnight at 50° C., concentrated under reduced pressure and purified by preparative reversed-phase chromatography to give methyl (1R,4R)-4-(4-hydroxy-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=302.1.
2,4,6-Triisopropylphenylsulfonyl chloride (1.0 g, 3.3 mmol), DMAP (20.3 mg, 0.165 mmol), and TEA (503 mg, 4.98 mmol) were added to a solution of methyl (1R,4R)-4-(4-hydroxy-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate (500 mg, 1.66 mmol) in dichloromethane (30 mL). The reaction liquid was stirred at room temperature overnight. The organic phase was washed with water (50 mL) and saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA=1:3) to give methyl (1R,4R)-4-(2-methyl-4-(((2,4,6-triisopropylphenyl)sulfonyl)oxy)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=568.2.
Methyl (1R,4R)-4-(2-methyl-4-(((2,4,6-triisopropylphenyl)sulfonyl)oxy)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate (800 mg, 1.41 mmol) and TEA (546.1 mg, 4.23 mmol) were added to a solution of 1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl-1-amine (462 mg, 1.97 mmol) in DMSO (20 mL). The reaction liquid was stirred at room temperature overnight. The crude product was purified by reversed-phase column chromatography (ACN/H2O=2/1) to give methyl (1R,4R)-4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=518.4.
Methyl (1R,4R)-4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylate (400 mg, 0.774 mmol) was dissolved in a solution of THF/H2O (40 mL/10 mL), and LiOH (93 mg, 3.87 mmol) was added. The reaction liquid was heated to 50° C. and stirred overnight. HCl (1 M) was added to adjust the pH to neutrality, and the resulting mixture was dissolved with water (50 mL), extracted with ethyl acetate (100 mL), and concentrated under reduced pressure to give the crude product (1R,4R)-4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=504.2.
(1R,4R)-4-(2-Methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylic acid (180 mg, 0.357 mmol), HATU (203.5 mg, 0.536 mmol), and DIEA (138.2 mg, 1.07 mmol) were added to a solution of 1-(2-methoxy-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione in DMF (6 mL). The reaction liquid was stirred at room temperature overnight, dissolved with water (50 mL), extracted with dichloromethane (30 mL×3), and concentrated under reduced pressure. The crude product was purified by reversed-phase column chromatography (ACN/H2O=1/2) to give 1-(2-methoxy-5-(4-((2-(1-((1R,4R)-4-(2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1l-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=958.5.
1-(2-Methoxy-5-(4-((2-(1-((1R,4R)-4-(2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (300 mg, 0.313 mmol) was dissolved in ethanol (20 mL), and Raney-Ni (aq., 1 mL) and hydrazine hydrate (0.5 mL) were added. The reaction liquid was stirred at room temperature for 1 h, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-methoxyphenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=928.5.
1H NMR (400 MHz, DMSO-d6) δ 10.33 (s, 1H), 8.91 (s, 1H), 8.58 (d, J=7.9 Hz, 1H), 8.10 (s, 1H), 7.36 (dd, J=8.4, 1.9 Hz, 1H), 7.32 (d, J=1.9 Hz, 1H), 7.15 (d, J=8.5 Hz, 1H), 6.89 (s, 1H), 6.85 (s, 1H), 6.71 (s, 1H), 5.57-5.51 (m, 3H), 4.45-4.33 (m, 1H), 4.00-3.90 (m, 1H), 3.84 (s, 3H), 3.59 (t, J=6.6 Hz, 2H), 3.41 (t, J=6.2 Hz, 2H), 3.24 (d, J=6.2 Hz, 2H), 3.05-2.95 (m, 2H), 2.83-2.70 (m, 2H), 2.70-2.65 (m, 3H), 2.43 (s, 3H), 2.05-1.96 (m, 2H), 1.88-1.40 (m, 20H), 1.23-0.89 (m, 4H).
A mixture of tert-butyl 4-((1-((benzyloxy)carbonyl)piperidin-4-yl)methyl)piperazine-1-carboxylate (500 mg, 1.20 mmol) and Pd/C (100 mg) in MeOH (10 mL) was stirred at room temperature for 2.0 h under H2 atmosphere. The reaction liquid was filtered and concentrated under reduced pressure to give tert-butyl 4-(piperidin-4-ylmethyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=284.1.
A mixture of 2-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenoxy)acetic acid (240 mg, 0.805 mmol), tert-butyl 4-(piperidin-4-ylmethyl)piperazine-1-carboxylate (251 mg, 0.886 mmol) HATU (612 mg, 1.61 mmol), DIEA (312 mg, 2.42 mmol) DMF (5 mL) was stirred at room temperature for 3 h. Water (20 mL) was added to quenchthe reaction, and the resulting mixture was extracted with DCM (20 mL×2). The organic phase was isolated, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH: DCM=0-1:24) to give tert-butyl 4-((1-(2-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenoxy)acetyl)piperidin-4-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=564.2.
A mixture of tert-butyl 4-((1-(2-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenoxy)acetyl)piperidin-4-yl)methyl)piperazine-1-carboxylate (350 mg, 0.622 mmol) TFA/DCM (10 mL/3 mL) was stirred at room temperature for 2 h. The reaction liquid was directly concentrated to give 1-(2-chloro-5-(2-oxo-2-(4-(piperazine-1-ylmethyl)piperidine-1-yl)ethoxy)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=464.2.
(1R,4R)-4-(4-(((R)-1-(3-((tert-Butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (405.6 mg, 0.674 mmol) was added to a solution of 1-(2-chloro-5-(2-oxo-2-(4-(piperazin-1-ylmethyl)piperidin-1-yl)ethoxy)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (260 mg, 0.562 mmol), HATU (426 mg, 1.12 mmol), and DIEA (217 mg, 3.36 mmol) in DMF (3 mL). Then the reaction liquid was stirred at room temperature for 1 h. Water (20 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with DCM (20 mL×2). The organic phase was dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=0-2:23) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-((1-(2-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenoxy)acetyl)piperidin-4-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1048.6.
A solution of tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-((1-(2-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenoxy)acetyl)piperidin-4-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (140 mg, 0.13 mmol) in DCM/TFA (3 mL/1 mL) was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure. The resulting crude product was purified by Prep-HPLC (acetonitrile/0.05% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-(2-(4-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)piperidin-1-yl)-2-oxoethoxy)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=948.4.
1H NMR (400 MHz, DMSO-d6) δ 10.47 (s, 1H), 8.24-7.87 (m, 2H), 7.45 (d, J=8.9 Hz, 1H), 7.11 (d, J=2.9 Hz, 1H), 6.99 (s, 1H), 6.93 (dd, J=9.0, 3.0 Hz, 1H), 6.86 (d, J=10.9 Hz, 2H), 6.70 (s, 1H), 5.67-5.40 (m, 3H), 4.93-4.63 (m, 2H), 4.40-4.24 (m, 1H), 3.89 (s, 3H), 3.82-3.65 (m, 2H), 3.63-3.42 (m, 4H), 3.09-2.89 (m, 2H), 2.77-2.55 (m, 5H), 2.41-2.25 (m, 7H), 2.12 (d, J=6.5 Hz, 2H), 1.9-1.48 (m, 14H), 1.15-0.88 (m, 2H).
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (2.0 g, 4.61 mmol) was dissolved in anhydrous tetrahydrofuran (40 mL) and cooled to −60° C. under nitrogen atmosphere, and a solution of lithium aluminum hydride (9.22 mL, 1.0 mmol/L) in tetrahydrofuran was added dropwise to the above solution while stirring. After the addition, the reaction liquid was stirred at −60° C. for 1 h. A saturated aqueous ammonium chloride solution (20 mL) was added dropwise to the reaction liquid, and the resulting mixture was filtered. The mother liquor was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give 1-(2-chloro-5-(hydroxymethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=255.0
1-(2-Chloro-5-(hydroxymethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (450 mg, 1.76 mmol) was dissolved with acetonitrile (20 mL), and IBX (988 mg, 3.53 mmol) was added. The reaction liquid was heated to 80° C. and stirred for 1 h. The reaction liquid was filtered, the filter cake was washed with methanol (20 ml×3), and the filtrate was concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzaldehyde.
LC-MS: (ESI, m/z): [M+H]+=253.0.
1H NMR (400 MHz, DMSO-d6) δ 10.57 (s, 1H), 10.01 (s, 1H), 8.06 (d, J=1.8 Hz, 1H), 7.93 (dd, J=8.2, 1.9 Hz, 1H), 7.84 (d, J=8.2 Hz, 1H), 3.80 (dd, J=16.6, 9.3 Hz, 1H), 3.64 (dt, J=12.1, 6.1 Hz, 1H), 2.77 (dd, J=12.5, 6.4 Hz, 2H).
tert-Butyl 4-(3-hydroxypropyl)piperidine-1-carboxylate (500 mg, 2.05 mmol) was dissolved in acetonitrile (20 mL), and IBX (1.15 g, 4.11 mmol) was added. The reaction liquid was heated to 70° C. and stirred for 1 h. The reaction liquid was filtered, the filter cake was washed with methanol (20 mL×3), and the filtrate was concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:1) to give tert-butyl 4-(3-oxopropyl)piperidine-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 9.68 (t, J=1.5 Hz, 1H), 3.91 (d, J=12.0 Hz, 2H), 2.65 (m, 2H), 2.46 (td, J=7.5, 1.5 Hz, 2H), 1.61 (d, J=13.0 Hz, 2H), 1.46 (2H), 1.42-1.33 (m, 10H), 0.94 (2H).
tert-Butyl 4-(3-oxopropyl)piperidine-1-carboxylate (300 mg, 1.2 mmol) and benzyl 4-(methylamino)piperidine-1-carboxylate (308 mg, 1.2 mmol) were dissolved with tetrahydrofuran (20 mL), and sodium triacetoxyborohydride (1.2 g, 6.0 mmol) was added while stirring. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was extracted with dichloromethane (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give benzyl 4-((3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=474.3.
Benzyl 4-((3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate (400 mg, 0.84 mmol) was dissolved in EA (20 mL), and Pd(OH)2/C (200 mg, 0.28 mmol) was added. The reaction liquid was heated to 70° C. under hydrogen atmosphere and stirred overnight. The reaction liquid was cooled to room temperature and filtered, and the filtrate was concentrated under reduced pressure to give tert-butyl 4-(3-(methyl(piperidin-4-yl)amino)propyl)piperidine-1-carboxylate. The crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=340.3.
tert-Butyl 4-(3-(methyl(piperidin-4-yl)amino)propyl)piperidine-1-carboxylate (280 mg, 0.82 mmol) and 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzaldehyde (311 mg, 1.23 mmol) were dissolved with tetrahydrofuran (20 mL), and sodium triacetoxyborohydride (869 mg, 4.1 mmol) was added. The reaction liquid was heated to 50° C. and stirred for 2 h, diluted with water (50 mL), and extracted with dichloromethane (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenyl)piperidin-4-yl)(methyl)amino)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=576.2.
tert-Butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenyl)piperidin-4-yl)(methyl)amino)propyl)piperidine-1-carboxylate (180 mg, 0.31 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h and directly concentrated under reduced pressure to give 1-(2-chloro-5-((4-(methyl(3-(piperidin-4-yl)propyl)amino)-1-yl)methyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=476.3.
1-(2-Chloro-5-((4-(methyl(3-(piperidin-4-yl)propyl)amino)-1-yl)methyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (180 mg, 0.31 mmol) was dissolved in DMF (10 mL), and (1R,4R)-4-(4-(((R)-1-(3-(tert-butoxycarbonyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (150 mg, 0.24 mmol), HATU (118 mg, 0.31 mmol), and N,N-diisopropylethylamine (93 mg, 0.72 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenyl)piperidin-4-yl)(methyl)amino)propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1060.5.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)phenyl)piperidin-4-yl)(methyl)amino)propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (200 mg, 0.19 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The resulting oil was dissolved with a mixed solution (20 mL) of 10% methanol/dichloromethane. 5% aqueous sodium bicarbonate solution (20 mL) was added, and the resulting mixture was stirred for 1 h. The resulting organic phase was isolated, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude product, which was purified by Pre-HPLC (CH3CN/0.08% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-((4-((3-(1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidin-1-yl)methyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=960.5.
1H NMR (400 MHz, DMSO-d6) δ 10.45 (s, 1H), 8.16-8.02 (m, 2H), 7.50 (d, J=8.2 Hz, 1H), 7.38 (s, 1H), 7.29 (d, J=8.3 Hz, 1H), 6.99 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.64-5.47 (m, 3H), 4.45-4.35 (m, 1H), 3.96-3.85 (m, 4H), 3.76-3.54 (m, 2H), 3.47-3.40 (m, 2H), 3.08-2.90 (m, 2H), 2.89-2.77 (m, 2H), 2.77-2.60 (m, 3H), 2.35 (s, 6H), 2.15 (s, 3H), 1.98-1.40 (m, 23H), 1.24-1.18 (m, 2H), 1.08-0.82 (m, 2H).
1,3-Dioxo-1,3-dihydroisobenzofuran-5-carboxylic acid (1.0 g, 5.21 mmol) was dissolved withAcOH (10 mL), and potassium acetate (945 mg, 5.73 mmol) and 3-aminopiperidine-2,6-dione hydrochloride (945 mg, 5.73 mmol) were added. The reaction liquid was heated to 90° C. and stirred overnight, and concentrated under reduced pressure.
Water (20 mL) was added to the crude product, and the resulting mixture was stirred for 10 min and filtered. The filter cake was washed with water (10 mL×3) and dried under vacuum to give the crude product 2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=303.0.
2-(2,6-Dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carboxylic acid (1.4 g, 4.64 mmol) was dissolved with DMF (20 mL), and DCC (1.43 g, 6.96 mmol) and 2,3,4,5,6-pentafluorophenol (1.30 g, 6.96 mmol) were added. The reaction liquid was stirred at room temperature overnight, diluted with water (150 mL), extracted with ethyl acetate (100 mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:1) to give pentafluorophenyl 2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 11.16 (s, 1H), 8.41 (dd, J=7.8, 1.3 Hz, 1H), 8.28 (s, 1H), 8.06 (d, J=7.8 Hz, 1H), 5.20 (dd, J=12.8, 5.4 Hz, 1H), 2.66-2.47 (m, 5H).
Pentafluorophenyl 2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carboxylate (480 mg, 1.02 mmol) was added to a solution of tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (400 mg, 1.22 mmol) in DMSO (10 mL). The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), extracted with ethyl acetate (30 mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:10) to give tert-butyl 4-(2-((1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=611.3.
tert-Butyl 4-(2-((1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (250 mg, 0.41 mmol) was dissolved with DCM (5 mL), and trifluoroacetic acid (1 mL) was added. The reaction liquid was stirred at room temperature overnight and concentrated under reduced pressure to give the crude product 2-(2,6-dioxopiperidin-3-yl)-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M+H]+=511.3.
2-(2,6-Dioxopiperidin-3-yl)-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)isoindoline-1,3-dione (200 mg, 0.40 mmol), HATU (130 mg, 0.35 mmol) and DIEA (89 mg, 0.69 mmol) were added to a solution of (1S,4S)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (140 mg, 0.23 mmol) in DMF (10 mL), and the reaction liquid was stirred at room temperature for 2 h, diluted with water (50 mL), stirred for 10 min and filtered. The solid was washed with water (5 mL×2) to give the crude product of tert-butyl (3-((1R)-1-((6-((1S,4S)-4-(4-(2-((1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1095.5.
tert-Butyl (3-((1R)-1-((6-((1S,4S)-4-(4-(2-((1-(2-(2,6-dioxopiperidin-3-yl)-1,3-dioxoisoindoline-5-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (250 mg, 0.23 mmol) was dissolved with DCM (5 mL), and trifluoroacetic acid (1 mL) was added. The reaction liquid was stirred at room temperature overnight and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 5-(4-((2-(1-((1S,4S)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(2,6-dioxopiperidin-3-yl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M+H]+=995.4.
1H NMR (400 MHz, DMSO-d6) δ 11.13 (s, 1H), 8.15 (s, 1H), 8.11 (d, J=8.1 Hz, 1H), 8.06 (s, 1H), 7.98 (d, J=7.6 Hz, 1H), 7.88-7.82 (m, 2H), 6.99 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.59-5.50 (m, 3H), 5.18 (dd, J=12.8, 5.4 Hz, 1H), 4.47-4.39 (m, 2H), 3.95-3.85 (m, 4H), 3.41 (t, J=6.4 Hz, 2H), 3.24 (d, J=5.3 Hz, 2H), 3.12-2.77 (m, 5H), 2.70-2.55 (m, 4H), 2.36 (s, 3H), 2.09-1.40 (m, 21H), 1.22-0.90 (m, 4H).
Trifluoroacetic acid (2.34 g, 20.5 mmol) was added dropwise to benzylamine (2.19 g, 20.5 mmol), and a solution of CD2O in D2O (20%, 1.5 g of CD2O was dissolved in 6 mL of D2O) was added. The reaction liquid was sonicated for 10 min and stirred at 20° C. for 1 h until the solution became clear. Allyltrimethylsilane (2.56 g, 22.5 mmol) was added, and the resulting mixture was heated to 40° C. and stirred overnight, diluted with water (10 mL), and adjusted to above pH 9 with solid potassium carbonate. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9, 1% ammonia water was added to the eluent) to give 1-phenylpiperidin-2,2,6,6-d4-4-ol.
LC-MS: (ESI, m/z): [M+H]+=196.3.
1-Phenylpiperidine-2,2,6,6-d4-4-ol (470 mg, 2.4 mmol) was dissolved in methanol (30 mL), and Pd/C (100 mg) and Pd(OH)2/C (100 mg) were added. The reaction liquid was stirred at 70° C. overnight under hydrogen atmosphere, filtered, and concentrated under reduced pressure to give piperidine-2,2,6,6-d4-4-ol.
LC-MS: (ESI, m/z): [M+H]+=106.4.
Piperidine-2,2,6,6-d4-4-ol (215 mg, 2.05 mmol) was dissolved with DCM (10 mL), and (Boc)2O (491 mg, 2.26 mmol), TEA (620 mg, 6.15 mmol), and DMAP (25 mg, 0.21 mmol) were added. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with water (50 mL) and extracted with dichloromethane (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=1:1) to give tert-butyl 4-hydroxypiperidine-1-carboxylate-2,2,6,6-d4.
LC-MS: (ESI, m/z): [M+Na]+=228.
tert-Butyl 4-hydroxypiperidine-1-carboxylate-2,2,6,6-d4 (200 mg, 0.98 mmol) was dissolved with THF (20 mL), and NaH (60%, 235 mg, 5.88 mmol) was added at 0° C. The reaction liquid was stirred for 2 h. Bromopicoline hydrobromide (296 mg, 1.18 mmol) was added, and the reaction liquid was heated to 70° C. and stirred overnight.
The reaction liquid was diluted with water (50 mL) and extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:19) to give tert-butyl 4-(pyridin-4-ylmethoxy)piperidine-1-carboxylate-2,2,6,6-d4.
LC-MS: (ESI, m/z): [M+H]+=297.2.
tert-Butyl 4-(pyridin-4-ylmethoxy)piperidine-1-carboxylate-2,2,6,6-d4 (150 mg, 0.51 mmol) was dissolved with a mixed solution of isopropanol (6 mL) and water (7 mL), and Pd(OH)2/C (45 mg) was added. The reaction liquid was stirred at 70° C. overnight under hydrogen atmosphere. The reaction liquid was filtered and concentrated under reduced pressure to give tert-butyl 4-(piperidine-4-ylmethoxy)piperidine-1-carboxylate-2,2,6,6-d4.
LC-MS: (ESI, m/z): [M+H]+=303.2.
tert-Butyl 4-(piperidin-4-ylmethoxy)piperidine-1-carboxylate-2,2,6,6-d4 (130 mg, 0.43 mmol) was dissolved with DMSO (5 mL), and DIEA (133 mg, 1.04 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (150 mg, 0.35 mmol) were added. The reaction liquid was stirred at room temperature for 2 h.
Water (30 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give tert-butyl 4-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)piperidine-1-carboxylate-2,2,6,6-d4.
LC-MS: (ESI, m/z): [M+Na]+=575.3.
tert-Butyl 4-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)piperidine-1-carboxylate-2,2,6,6-d4 (130 mg, 0.24 mmol) was dissolved with DCM (5 mL), and trifluoroacetic acid (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure to give 1-(2-chloro-5-(4-(((piperidin-4-yl-2,2,6,6-d4)oxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=453.2.
1-(2-Chloro-5-(4-(((piperidin-4-yl-2,2,6,6-d4)oxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (130 mg, 0.29 mmol) was dissolved with DMSO (5 mL), and (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (130 mg, 0.22 mmol), HATU (110 mg, 0.29 mmol), and DIEA (142 mg, 1.10 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (30 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)piperidine-1-carbonyl-2,2,6,6-d4)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1037.5.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)piperidine-1-carbonyl-2,2,6,6-d4)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (130 mg, 0.13 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The crude product was dissolved with DCM (20 mL), and a saturated aqueous sodium bicarbonate solution (20 mL) was added to adjust the pH to neutrality. The resulting mixture was stirred for 10 min, extracted with MeOH/DCM (v:v=1:9, 20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude was purified by preparative reversed-phase chromatography to give 1-(5-(4-(((1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl-2,2,6,6-d4)oxy)methyl)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=937.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.14-8.02 (m, 2H), 7.64 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.9 Hz, 1H), 7.38 (dd, J=8.2, 1.9 Hz, 1H), 6.99 (s, 1H), 6.86 (d, J=10.5 Hz, 2H), 6.70 (s, 1H), 5.64-5.46 (m, 3H), 4.51-4.39 (m, 1H), 3.88 (s, 3H), 3.75-3.42 (m, 4H), 3.12-3.01 (m, 1H), 2.95 (t, J=11.2 Hz, 1H), 2.83-2.57 (m, 5H), 2.35 (s, 3H), 1.95-1.50 (m, 17H), 1.45-1.27 (m, 2H), 1.25-1.07 (m, 2H).
DIEA (675 mg, 3.9 mmol) was added to a solution of tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (430 mg, 1.3 mmol) and 2-(2,6-dioxopiperidin-3-yl)-5-fluoroisoindoline-1,3-dione (718 mg, 2.6 mmol) in DMF (3 mL). The reaction mixture was heated to 110° C. and stirred overnight. The reaction liquid was poured into water, and the resulting mixture was extracted with EA (30 mL×2). The organic phase was dried over anhydrous Na2SO4 and concentrated under reduced pressure. The resulting crude product was purified by automatic column chromatography (EA/PE) to give tert-butyl 4-(2-((1-(2-(2,4-dioxocyclohexyl)-1,3-dioxoisoindolin-5-yl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=583.4.
A solution of tert-butyl 4-(2-((1-(2-(2,4-dioxocyclohexyl)-1,3-dioxoisoindolin-5-yl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (210 mg, 0.36 mmol) in HCl/1,4-dioxane (4 M, 5 mL) was stirred at room temperature for 1 h. The reaction liquid was directly concentrated to give 2-(2,6-dioxopiperidine-3-yl)-5-(4-((2-(piperidine-4-yl)ethoxy)methyl)piperidine-1-yl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M+H]+=483.3.
HATU (77 mg, 0.20 mmol) and DIEA (65 mg, 0.51 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (100 mg, 0.17 mmol) and 2-(2,6-dioxopiperidin-3-yl)-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidin-1-yl)isoindoline-1,3-dione (87 mg, 0.17 mmol) in DMF (5 mL). The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was poured into water, and the resulting mixture was extracted with EA (20 mL×3), dried over anhydrous Na2SO4, and concentrated under reduced pressure. The resulting crude product was purified by prep-HPLC to give 5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidin-1-yl)-2-(2,4-dioxocyclohexyl)isoindoline-1,3-dione.
LC-MS: (ESI, m/z): [M−H]−=1057.4.
1H NMR (400 MHz, DMSO-d6) δ 11.07 (s, 1H), 8.31 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.64 (d, J=8.5 Hz, 1H), 7.49-7.38 (m, 2H), 7.36-7.29 (m, 2H), 7.23 (dd, J=9.8, 1.7 Hz, 2H), 7.02 (s, 1H), 6.96 (s, 1H), 6.03-5.87 (m, 1H), 5.10-5.00 (m, 1H), 4.42-4.31 (m, 1H), 4.10-4.00 (m, 2H), 3.97-3.85 (m, 4H), 3.41 (t, J=6.3 Hz, 2H), 3.23 (d, J=6.2 Hz, 2H), 3.18-3.07 (m, 2H), 3.05-2.81 (m, 5H), 2.71-2.52 (m, 3H), 2.42 (s, 3H), 2.01 (s, 6H), 1.89-1.44 (m, 20H), 1.26-0.86 (m, 4H).
3-(Methylamino)propan-1-ol (5.0 g, 56.17 mmol) and benzyl chloroformate (10.5 g, 61.79 mmol) were dissolved in 1,4-dioxane (40 mL), and a sodium hydroxide solution (2 mol/L, 48 mL) was added. The reaction liquid was stirred at 50° C. for 0.5 h and directly concentrated under reduced pressure to give the crude product benzyl (3-hydroxypropyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=224.3.
Benzyl (3-hydroxypropyl)(methyl)carboxylate (4.6 g, 20 mmol) was dissolved in tetrahydrofuran (40 mL), and triphenylphosphine (7.8 g, 30 mmol) and carbon tetrabromide (10 g, 30 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h, diluted with water (200 mL), extracted with ethyl acetate (200 mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:1) to give benzyl (3-bromopropyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=288.1.
4-Bromopyridin-2(1H)-one (790 mg, 4.5 mmol) was dissolved with anhydrous DMF (5 mL) and cooled to 0° C., and NaH (218 mg, 5.5 mmol) was added. The reaction liquid was stirred for 0.5 h. Then Benzyl (3-bromopropyl)(methyl)carboxylate (1.3 g, 4.5 mmol) was added at room temperature, and the resulting mixture was stirred for 12 h, diluted with water (100 mL), and extracted with ethyl acetate (150 mL×4). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:2) to give benzyl (3-(4-bromo-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): (ESI, m/z): [M+H]+=379.0.
Benzyl (3-(4-bromo-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate (1.2 g, 3.2 mmol) and 4,4,4′,4′,5,5,5′,5′-octamethyl-2,2′-bis(1,3,2-dioxaborolane) (979 mg, 3.8 mmol) were dissolved in anhydrous 1,4-dioxane, Pd(dppf)Cl2 (120 mg) and potassium carbonate (880 mg, 6.47 mmol) were added, and then the mixture was stirred overnight at 100° C. under nitrogen atmosphere, diluted with water (100 mL) and extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:1) to give the product benzyl (3-(2-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-1(2H)-yl)propyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=427.4.
6-Bromo-2,7-dimethylquinazolin-4-ol (450 mg, 1.77 mmol) and benzyl methyl(3-(2-oxo-4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridin-1(2H)-yl)propyl)carboxylate (980 mg, 2.3 mmol) were dissolved in DMF/water solution (10/1, 4 mL), and potassium carbonate (480 mg, 3.5 mmol) and Pd(dppf)Cl2 (90 mg) were added. The reaction liquid was purged with nitrogen, heated to 100° C. under nitrogen atmosphere, stirred for 2 h, diluted with water (50 mL), and extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (DCM:MEOH=10:1) to give the product benzyl (3-(4-(4-hydroxy-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): (ESI, m/z): [M+H]+=473.3.
Benzyl (3-(4-(4-hydroxy-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate (480 mg, 1.02 mmol) was dissolved with DMF (4 mL), and DBU (388 mg, 2.55 mmol) and BOP (583 mg, 1.32 mmol) were added. The reaction liquid was stirred at room temperature for 0.5 h. tert-Butyl (R)-(3-(1-aminoethyl)-5-(trifluoromethyl)phenyl)carboxylate (403 mg, 1.32 mmol) was added, and the reaction liquid was heated to 70° C. and stirred overnight, diluted with water (50 mL), and extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:1) to give the product benzyl (R)-(3-(4-(4-((1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=759.4.
Benzyl (R)-(3-(4-(4-((1-3-((tert-butoxycarbonyl)amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)(methyl)carboxylate (300 mg, 0.4 mmol) was dissolved room temperature for 2 h under hydrogen atmosphere, filtered, and concentrated to give the crude product tert-butyl (R)-(3-1-((2,7-dimethyl-6-(1-(3-(methylamino)propyl)2-oxo-1,2-dihydropyridin-4-yl)quinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=625.5.
tert-butyl (R)-9-(((3-(4-(4-((1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)methyl)amino)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=890.5.
tert-Butyl (R)-9-(((3-(4-(4-((1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)methyl)amino)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (100 mg, 0.11 mmol) was added to a solution of hydrogen chloride in 1,4-dioxane (4 M, 3 mL). The mixture was reacted at room temperature for 2 h and directly concentrated under reduced pressure to give the crude product of (R)-1-(3-(((3-azaspiro[5.5]undecan-9-yl)methyl)(methyl)amino)propyl)-4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)pyridin-2(1H)-one.
LC-MS: (ESI, m/z): [M+H]+=690.5.
(R)-1-(3-(((3-Azaspiro[5.5]undecan-9-yl)methyl)(methyl)amino)propyl)-4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)pyridin-2(1H)-one (80 mg, 0.12 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (51 mg, 0.12 mmol) were dissolved in DMSO (3 mL), and DIEA (60 mg, 0.48 mmol) was added. The reaction liquid was stirred at room temperature for 2 h, diluted with water (30 mL), extracted with ethyl acetate (30 mL×3), washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by Prep-HPLC to give the product (R)-1-(5-(9-(((3-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)-2-oxopyridin-1(2H)-yl)propyl)(methyl)amino)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M−H]−=940.5.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.36 (d, J=8.2 Hz, 1H), 8.29 (s, 1H), 7.73 (d, J=6.6 Hz, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (s, 1H), 7.50 (s, 1H), 7.38 (d, J=8.3 Hz, 1H), 6.88 (s, 1H), 6.84 (s, 1H), 6.69 (s, 1H), 6.42 (s, 1H), 6.36 (d, J=7.0 Hz, 1H), 5.60-5.48 (m, 3H), 4.00-3.90 (m, 2H), 3.80-3.50 (m, 5H), 3.29-3.20 (m, 3H), 2.77-2.70 (m, 2H), 2.44-2.26 (m, 7H), 2.20-2.06 (m, 4H), 1.87-1.78 (m, 2H), 1.74-1.65 (m, 2H), 1.62-1.20 (m, 10H), 1.16-0.92 (m, 4H).
2-Methoxyethane-1-ol (365 mg, 4.8 mmol) was added to a three-necked flask and dissolved with THF (10 mL). The reaction liquid was purged three times with nitrogen. t-BuOK (1 M/THF, 4.2 mL, 4.2 mmol) was added at 0° C., and the resulting mixture was reacted at 0° C. for 1 h. A solution of methyl 4-fluoro-2-nitrobenzoate (600 mg, 3.0 mmol) in THF (10 mL) was added dropwise, and the resulting mixture was successively reacted at 0° C. for 2 h. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=2:3) to give methyl 4-(2-methoxyethoxy)-2-nitrobenzoate.
1H NMR (400 MHz, DMSO-d6) δ 7.87 (d, J=8.7 Hz, 1H), 7.57 (d, J=2.5 Hz, 1H), 7.34 (dd, J=8.7, 2.5 Hz, 1H), 4.29-4.24 (m, 2H), 3.81 (s, 3H), 3.70-3.66 (m, 2H), 3.31 (s, 3H).
Methyl 4-(2-methoxyethoxy)-2-nitrobenzoate (850 mg, 3.3 mmol) was dissolved in EtOH (20 mL), and Raney-Ni (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h under H2 atmosphere, filtered, and concentrated under reduced pressure to give methyl 2-amino-4-(2-methoxyethoxy)benzoate.
LC-MS: (ESI, m/z): [M+H]+=226.1.
Methyl 2-amino-4-(2-methoxyethoxy)benzoate (680 mg, 3.0 mmol) was dissolved with DMF (10 mL), and NBS (484 mg, 2.7 mmol) was added at 0° C. The reaction liquid was reacted at 0° C. for 1 h. Water (100 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (EA:PE=2:3) to give methyl 2-amino-5-bromo-4-(2-methoxyethoxy)benzoate.
LC-MS: (ESI, m/z): [M+H]+=304.0.
Methyl 2-amino-5-bromo-4-(2-methoxyethoxy)benzoate (710 mg, 2.3 mmol) was dissolved in THF/H2O (15 mL, 2:1), and LiOH (570 mg, 23.0 mmol) was added. The reaction liquid was stirred at 50° C. overnight. Water (20 mL) was added, and the resulting mixture was adjusted to pH 6 with 1 M hydrochloric acid, extracted with a mixed solution of MeOH/DCM (1/10, 30 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 2-amino-5-bromo-4-(2-methoxyethoxy)benzoic acid.
LC-MS: (ESI, m/z): [M+3]+=292.0.
2-Amino-5-bromo-4-(2-methoxyethoxy)benzoic acid (665 mg, 2.3 mmol) was added to a sealed container and dissolved with MeOH (20 mL), and ammonium acetate (2.66 g, 34.5 mmol) and trimethyl orthoacetate (4.14 g, 34.5 mmol) were added. The reaction liquid was reacted at 120° C. overnight and concentrated under reduced pressure to remove methanol. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:4) to give 6-bromo-7-(2-methoxyethoxy)-2-methylquinazolin-4-ol.
LC-MS: (ESI, m/z): [M+H]+=312.9.
6-Bromo-7-(2-methoxyethoxy)-2-methylquinazolin-4-ol (210 mg, 0.67 mmol) was dissolved with a mixed solution of DMF/H2O (11 mL, 10:1), and methyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-ene-1-carboxylate (233 mg, 0.88 mmol), Na2CO3 (143 mg, 1.34 mmol), and Pd(dppf)Cl2 (25 mg, 0.03 mmol) were added. The reaction liquid was reacted at 110° C. overnight under nitrogen atmosphere. Water (100 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (50 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give methyl 4-(4-hydroxy-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=373.1.
Methyl 4-(4-hydroxy-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (500 mg, 1.3 mmol) was added to an autoclave and dissolved with MeOH (30 mL), and Pd(OH)2/C (150 mg, 20%) was added.
The reaction liquid was reacted at 70° C. for 48 h under hydrogen atmosphere (2 MPa), filtered, and concentrated under reduced pressure. The crude product was isolated by preparative reversed-phase chromatography to give methyl (1R,4R)-4-(4-hydroxy-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=375.2.
Methyl (1R,4R)-4-(4-hydroxy-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (80 mg, 0.21 mmol) was dissolved in DMF (5 mL), and BOP (123 mg, 0.28 mmol) and DBU (81 mg, 0.53 mmol) were added. The reaction liquid was stirred at room temperature for 0.5 h. tert-Butyl (R)-(3-(1-aminoethyl)-5-(trifluoromethyl)phenyl)carboxylate (70 mg, 0.23 mmol) was added, and the resulting mixture was reacted at 70° C. overnight in an oil bath. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reversed-phase chromatography to give methyl (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=661.2.
Methyl (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (66 mg, 0.10 mmol) was dissolved in a mixed solution of THF/H2O (15 mL, 2:1), and LiOH (48 mg, 2.0 mmol) was added. The reaction liquid was stirred at 50° C. overnight. Water (20 mL) was added, and the resulting mixture was adjusted to pH 6 with concentrated hydrochloric acid, extracted with MeOH/DCM (10%, 30 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reversed-phase chromatography to give (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=647.4.
(1R,4R)-4-(4-(((R)-1-(3-((tert-Butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (50 mg, 0.08 mmol) was dissolved in DMF (5 mL), and 1-(2-chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (48 mg, 0.10 mmol), DIEA (52 mg, 0.40 mmol), and pyBOP (50 mg, 0.10 mmol) were added. The reaction liquid was reacted at room temperature for 1 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-(2-methoxyethoxy)-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1130.4.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-(2-methoxyethoxy)-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (80 mg, 0.07 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 1 h and concentrated under reduced pressure. The residue was dissolved with MeOH/DCM (1:9, 20 mL), and a saturated aqueous sodium bicarbonate solution (20 mL) was added to adjust the pH to neutrality. The resulting mixture was stirred for 10 min, extracted with MeOH/DCM (1:9, 20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-(2-methoxyethoxy)-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1030.5.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.12 (d, J=7.7 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.55 (d, J=1.8 Hz, 1H), 7.39 (d, J=8.2 Hz, 1H), 6.98 (s, 1H), 6.87 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.61-5.48 (m, 3H), 4.22-4.17 (m, 2H), 3.75 (d, J=4.6 Hz, 3H), 3.65-3.52 (m, 3H), 3.51-3.41 (m, 4H), 3.36 (s, 3H), 3.02-2.88 (m, 1H), 2.79-2.71 (m, 2H), 2.68-2.60 (m, 1H), 2.29-2.38 (m, 5H), 2.28-2.21 (m, 2H), 2.16-2.06 (m, 2H), 1.95-1.85 (m, 2H), 1.84-1.80 (m, 2H), 1.76-1.19 (m, 18H), 1.17-0.95 (m, 4H).
(1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (100 mg, 0.17 mmol) was dissolved in DMF (8 mL), and benzyl 4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate (72 mg, 0.2 mmol), HATU (90 mg, 0.23 mmol) and DIEA (64 mg, 0.5 mmol) were added separately. The reaction liquid was stirred at room temperature for 5 h. Water (50 mL) was added to the reaction liquid, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give benzyl 4-((2-(1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=945.5.
Benzyl 4-((2-(1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate (100 mg, 0.11 mmol) was dissolved with ethyl acetate (8 mL), and Pd(OH)2/C (50 mg, 0.14 mmol) was added. The reaction liquid was heated to 70° C. under hydrogen atmosphere and stirred overnight. The reaction liquid was cooled to room temperature and filtered, and the filtrate was concentrated under reduced pressure to give tert-butyl (3-((R)-1-((7-methoxy-2-methyl-6-((1R,4R)-4-(4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)quinolinazol-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=811.5.
tert-Butyl (3-((R)-1-((7-methoxy-2-methyl-6-((1R,4R)-4-(4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)quinolinozol-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (80 mg, 0.098 mmol) was dissolved with DMSO (8 mL), and 1-(2-fluoro-5-(pentafluorobenzoyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (40 mg, 0.098 mmol) and DIEA (40 mg, 0.28 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added to the reaction liquid, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-(2-((1-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1045.3.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-(2-((1-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (50 mg, 0.047 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure. The crude product was purified by Prep-HPLC to give 1-(5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-fluorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=945.4.
1H NMR (400 MHz, MeOD) δ 10.52 (s, 1H), 8.19-7.99 (m, 2H), 7.49 (d, J=7.9 Hz, 1H), 7.37 (d, J=8.3 Hz, 2H), 6.99 (s, 1H), 6.87 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.65-5.47 (m, 3H), 4.50-4.33 (m, 2H), 3.96-3.85 (m, 4H), 3.74 (t, J=6.7 Hz, 2H), 3.41 (t, J=6.4 Hz, 2H), 3.23 (d, J=6.2 Hz, 2H), 3.08-2.89 (m, 3H), 2.72 (t, J=6.7 Hz, 2H), 2.69-2.60 (m, 1H), 2.40-2.30 (m, 4H), 1.99-1.37 (m, 22H), 1.16-0.94 (m, 4H).
tert-Butyl 4-(piperidin-4-ylmethoxy)piperidine-1-carboxylate-2,2,6,6-d4 (150 mg, 0.50 mmol) was dissolved with DMF (5 mL), and (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (150 mg, 0.25 mmol), HATU (123 mg, 0.32 mmol), and DIEA (161 mg, 1.25 mmol) were added. The reaction liquid was stirred at room temperature for 2 h.
Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give tert-butyl 4-((1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)piperidine-1-carboxylate-2,2,6,6-d4.
LC-MS: (ESI, m/z): [M+H]+=887.4.
tert-Butyl 4-((1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)piperidine-1-carboxylate-2,2,6,6-d4 (200 mg, 0.23 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure to give ((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(((piperidin-4-yl-2,2,6,6-d4)oxy)methyl)piperidin-1-yl)methanone, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=687.4.
((1R,4R)-4-(4-(((R)-1-(3-Amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(((piperidin-4-yl-2,2,6,6-d4)oxy)methyl)piperidin-1-yl)methanone (200 mg, 0.29 mmol) was dissolved in DMSO (5 mL), and DIEA (149 mg, 1.15 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (100 mg, 0.23 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(4-((1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)piperidine-1-carbonyl-2,2,6,6-d4)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): (ESI, m/z): [M+H]+=937.5.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.13 (s, 1H), 8.09 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.57 (d, J=1.9 Hz, 1H), 7.40 (dd, J=8.2, 2.0 Hz, 1H), 6.99 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.62-5.50 (m, 3H), 4.42 (d, J=11.5 Hz, 1H), 4.0-3.92 (m, 4H), 3.74 (s, 1H), 3.67-3.47 (m, 2H), 3.29 (d, J=4.5 Hz, 2H), 3.09-2.87 (m, 2H), 2.76-2.73 (m, 2H), 2.67-2.63 (m, 1H), 2.36 (s, 3H), 1.94-1.50 (m, 17H), 1.50-1.38 (m, 2H), 1.16-0.91 (m, 2H).
(R)-1-(4-bromo-2-yl)ethan-1-amine (607 mg, 3 mmol) was dissolved in DMF (5 mL), BOP (574.6 mg, 1.3 mmol) and DBU (456.0 mg, 3.0 mmol) were added, and the reaction liquid was stirred at room temperature for 0.5 h. Then methyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (328 mg, 1 mmol) was added to the reaction liquid, and the mixture was warmed to 70° C. and reacted for 16 h. The reaction liquid was diluted with water (50 mL) and extracted with ethyl acetate (30 mL×3). The upper organic phases were combined, washed with saturated brine, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give methyl 4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=516.1.
3-Chloro-2-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)benzaldehyde (206 mg, 0.77 mmol) and methyl 4-(4-(((R)-1-(4-bromothien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (400 mg, 0.77 mmol) were dissolved in DMF/H2O (3 mL/0.3 mL), and Pd(dppf)Cl2 (100 mg) and K2CO3 (214 mg, 1.55 mmol) were added. The reaction liquid was purged with nitrogen and stirred at 100° C. for 2 h under nitrogen atmosphere. After the reaction was completed, the reaction liquid was diluted with water (50 mL) and extracted with ethyl acetate (30 mL×3). The organic phases were combined and washed with saturated brine, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give methyl 4-(4-(((R)-1-(4-(2-chloro-6-benzaldehyde)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=576.3.
Methyl 4-(4-(((R)-1-(4-(2-chloro-6-benzaldehyde)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (130 mg, 0.22 mmol) was dissolved in 1,2-dichloroethane (5 mL), and sodium triacetoxyborohydride (143 mg, 0.68 mmol) was added. The reaction liquid was stirred at 80° C. for 1 h. The reaction liquid was diluted with water (50 mL) and extracted with dichloromethane (30 mL×3). The organic phases were combined, dried over anhydrous Na2SO4, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:19) to give methyl 4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=605.3.
Methyl 4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (170 mg, 0.28 mmol) was dissolved with MeOH/H2O (2 mL/0.2 mL), and LiOH (52 mg, 1.3 mmol) was added. The reaction liquid was heated to 60° C. and stirred for 1 h.
The reaction liquid was cooled, the pH was adjusted to 3-4 with diluted hydrochloric acid (1.0 mol/L), and a NaHCO3 saturated solution was added to adjust the pH to 7-8. The mixture was extracted with ethyl acetate (30 mL×3), and the organic phases were combined, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give 4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=591.3.
4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid (125 mg, 0.22 mmol) and 3-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)piperidine-2,6-dione (108 mg, 0.22 mmol) were dissolved with DMF (3 mL), and HATU (96 mg, 0.25 mmol) and DIEA (82 mg, 0.64 mmol) were added. The reaction liquid was stirred at room temperature for 1 h. The reaction liquid was diluted with water (50 mL) and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine, dried over anhydrous Na2SO4, filtered, and concentrated under reduced pressure. The crude product was purified by Prep-HPLC to give the product 3-(2-chloro-5-(4-((2-(1-(4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidin-1-carbonyl)phenyl)piperidin-2,6-dione.
LC-MS: (ESI, m/z): [M+H]+=1049.3.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.41 (s, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (s, 1H), 7.48-7.30 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.01-5.89 (m, 1H), 5.83 (s, 1H), 4.47-4.37 (m, 1H), 3.99-3.91 (m, 1H), 3.86 (s, 3H), 3.74 (s, 1H), 3.68-3.52 (m, 2H), 3.44-3.37 (m, 2H), 3.26-3.18 (m, 3H), 3.18-2.96 (m, 3H), 2.90-2.82 (m, 1H), 2.81-2.70 (m, 2H), 2.43 (s, 3H), 2.39-2.29 (m, 2H), 2.25-2.16 (m, 1H), 2.03 (s, 6H), 1.89-1.55 (m, 10H), 1.49-1.40 (m, 2H), 1.30-1.20 (m, 2H), 1.20-0.95 (m, 4H).
CbzCl (5.40 mL, 38.40 mmol) was added to a solution of 3-(piperidin-4-yl)propan-1-ol (5.00 g, 34.94 mmol) in THF (20 mL) at 0° C. The reaction liquid was reacted for 1 h. After the reaction was completed, a saturated aqueous NaHCO3 solution (50 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with brine (100 mL), dried over sodium sulfate, and filtered, and the filtrate was concentrated. The crude product was purified by silica gel column chromatography (MeOH:DCM=1:100-1:10) to give benzyl 4-(3-hydroxypropyl)piperidine-1-carboxylate as a pale yellow oil.
LC-MS: (ESI, m/z): [M+H]+=278.2.
Dess-Martin (5.50 g, 13.0 mmol) was added to a solution of benzyl 4-(3-hydroxypropyl)piperidine-1-carboxylate (3.00 g, 10.8 mmol) in DCM (60 mL) at 0° C. for reaction for 1 h. After the reaction was completed, saturated Na2S2O3 (50 mL) and saturated NaHCO3 (50 mL) were added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with DCM (80 mL×3). The organic phases were combined, washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated by rotary evaporation. The crude product was purified by silica gel chromatography (EA:PE=3:17-3:7) to give benzyl 4-(3-oxopropyl)piperidine-1-carboxylate.
1H NMR (400 MHz, CDCl3) δ 9.78 (s, 1H), 7.38-7.28 (m, 5H), 5.12 (s, 2H), 4.28-4.02 (m, 2H), 2.90-2.65 (m, 2H), 2.51-2.42 (m, 2H), 1.74-1.54 (m, 4H), 1.51-1.37 (m, 1H), 1.21-1.03 (m, 2H).
Methylamine (846 mg, 27.24 mmol) and AcOH (0.1 mL) were added to a solution of benzyl 4-(3-oxopropyl)piperidine-1-carboxylate (1.50 g, 5.45 mmol) in THE (10 mL), the mixture was reacted at 30° C. for 4 h, then NaBH3CN (684 mg, 10.90 mmol) was added, and the mixture was reacted for another 16 h. After the reaction was completed, saturated NaHCO3 (50 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated by rotary evaporation. The crude product was purified by silica gel chromatography (NH3/MeOH:DCM=1:9-2:8) to give benzyl 4-(3-(methylamino)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=291.7.
(Boc)2O (1.2 mL, 5.10 mmol) was added to a solution of benzyl 4-(3-(methylamino)propyl)piperidine-1-carboxylate (295 mg, 1.02 mol) in DCM (3 mL). The reaction liquid was reacted at room temperature for 16 h. After the reaction was completed, water (50 mL) was added to quench the reaction liquid, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phases were combined, washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated by rotary evaporation. The crude product was purified by silica gel chromatography (NH3/MeOH:DCM=1:19-1:9) to give benzyl 4-(3-((tert-butoxycarbonyl)(methyl)amino)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M-100+H]+=291.2.
Pd/C (80 mg) was added to a solution of benzyl 4-(3-((tert-butoxycarbonyl)(methyl)amino)propyl)piperidine-1-carboxylate (291 mg, 0.75 mmol) in MeOH (5 mL). The reaction liquid was purged three times with hydrogen and reacted at room temperature for 16 h. The reaction liquid was concentrated by rotary evaporation to give the crude product tert-butyl methyl(3-piperidin-4-yl)propyl)carboxylate, which was used in the next step without further purification.
LC-MS: (ESI, m/z): [M+H]+=257.2.
DIEA (232 mg, 1.8 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (381 mg, 0.88 mmol) were added to a solution of tert-butyl methyl(3-(piperidin-4-yl)propyl)carbamate (150 mg, 0.59 mmol) DMSO (2 mL). The reaction liquid was reacted at room temperature for 16 h. After the reaction was completed, water (50 mL) was added to quench the reaction liquid, and the resulting mixture was extracted with ethyl acetate (50 mL×3). The organic phases were combined, washed with brine (100 mL), dried over sodium sulfate, filtered, and concentrated by rotary evaporation. The crude product was purified by silica gel chromatography (NH3/MeOH:DCM=1:19-1:9) to give the target compound tert-butyl (3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+Na]+=529.1.
The compound tert-butyl (3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)(methyl)carboxylate (256 mg, 0.50 mmol) was dissolved DCM (2 mL), TFA (1 mL) was added at room temperature. The reaction liquid was reacted for 0.5 h. The reaction liquid was concentrated to give the crude product 1-(2-chloro-5-(4-(3-(methylamino)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was used in the next step without further purification.
LC-MS: (ESI, m/z): [M+H]+=407.0.
1-(2-Chloro-5-(4-(3-(methylamino)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (216 mg, 0.53 mmol), TCFH (249 mg, 0.89 mmol), and NMI (146 mg, 1.78 mmol) were added to a solution of the compound (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (229 mg, 0.36 mmol) in DMF (3 mL). The reaction liquid was reacted at room temperature for 16 h. After the reaction was completed, water (20 mL) was added to the reaction liquid to quench the reaction, and the resulting mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined, washed with brine (50 mL), dried over sodium sulfate, filtered, and concentrated under vacuum. The crude product was purified by silica gel chromatography (NH3/MeOH:DCM=1:19-1:9) to give the target compound tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-((3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)(methyl)carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1033.2.
TFA (1 mL) was added to a solution of the compound tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-((3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)(methyl)carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)phenyl)(methyl)carboxylate (107 mg, 0.10 mmol) in DCM (2 mL) at room temperature. The reaction liquid was reacted for 0.5 h. The reaction liquid was concentrated to give the crude product, which was purified by high performance liquid chromatography to give the target compound (1R,4R)—N-(3-(1-(4-chloro-3-(2,4-dioxotetrahydropyrimidine-1(2H)-yl)benzoyl)piperidin-4-yl)propyl)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)-N-methylcyclohexane-1-carboxamide.
LC-MS: (ESI, m/z): [M+H]+=933.4.
1H NMR (400 MHz, CD3OD) δ 7.99 (s, 1H), 7.66-7.58 (m, 1H), 7.54-7.48 (m, 1H), 7.43-7.36 (m, 2H), 7.33-7.26 (m, 3H), 7.18 (d, J=1.2 Hz, 1H), 7.15-7.11 (m, 1H), 7.01 (m, 1H), 6.08-6.01 (m, 1H), 4.65-4.53 (m, 1H), 3.95 (s, 3H), 3.81-3.69 (m, 5H), 3.48-3.34 (m, 2H), 3.16-3.04 (m, 4H), 2.95-2.68 (m, 5H), 2.52 (s, 3H), 2.22 (s, 3H), 2.00-1.57 (m, 16H), 1.35-1.11 (m, 4H).
The compound benzyl 4-(2-hydroxyethyl)piperidine-1-carboxylate (400 mg, 1.52 mmol) was dissolved THF (5 mL), and triethylamine (230 mg, 2.28 mmol) was added. The reaction liquid was cooled to 0° C., and methanesulfonyl chloride (208 mg, 1.82 mmol) was added dropwise. After the addition, the reaction liquid was heated to room temperature and reacted for 2 h. An aqueous ammonium chloride solution was added to quench the reaction, and the resulting mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined and washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated to give an intermediate. This intermediate was dissolved in tetrahydrofuran, and the resulting solution was added to a solution of potassium tert-butoxide (340 mg, 3.04 mmol) and the compound tert-butyl 4-(hydroxymethyl)piperidine-1-carboxylate (652 mg, 3.04 mmol) in THF (6 mL). The reaction liquid was reacted at 30° C. for 16 h. The reaction liquid was diluted with water and extracted with ethyl acetate (20 ml×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, concentrated, and purified by column chromatography (PE:EA=5:1) to give the target product benzyl 4-(2-((1-(tert-butoxycarbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+Na]+=483.2.
The compound benzyl 4-(2-((1-(tert-butoxycarbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate was dissolved in methanol (180 mg, 0.39 mmol), and palladium on carbon (70 mg) was added. The reaction liquid was purged three times under hydrogen (1 atmosphere pressure) atmosphere, reacted at room temperature for 16 h, and filtered through celite. The filtrate was concentrated to give the crude product tert-butyl 4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate, which was directly used in the next step.
The compound tert-butyl 4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate (114 mg, 0.35 mmol), TcFH (162 mg, 0.58 mmol), and NMI (94 mg, 1.15 mmol) were added to a solution of the compound (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (130 mg, 0.23 mmol) in DMF (2 mL). The reaction liquid was reacted at room temperature for 16 h. After the reaction was completed, water (20 mL) was added to quench the reaction liquid, and the resulting mixture was extracted with ethyl acetate (20 mL×3). The organic phases were combined, washed with brine (50 mL), dried over sodium sulfate, filtered, and concentrated under vacuum. The crude product was purified by column chromatography (NH3/MeOH:DCM=1:19-1:9) to give the target product tert-butyl 4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=867.3.
The compound tert-butyl 4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carboxylate (40 mg, 0.05 mmol) was dissolved in DCM (2 mL) at room temperature, and TFA (1 mL) was added. The reaction liquid was reacted at room temperature for 0.5 h. The reaction liquid was concentrated to give the crude product ((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(2-(piperidin-4-ylmethoxy)ethyl)piperidin-1-yl)methanone, which was directly used in the next step without further purification.
LC-MS: (ESI, m/z): [M+H]+=767.3.
DIEA (103 mg, 0.80 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (42 mg, 0.10 mmol) were added to a solution of ((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(2-(piperidin-4-ylmethoxy)ethyl)piperidin-1-yl)methanone (50 mg, 0.07 mmol) in DMSO (2 mL). The reaction liquid was reacted at room temperature for 16 h. After the reaction was completed, water (10 mL) was added to quench the reaction liquid, and the resulting mixture was extracted with ethyl acetate (10 mL×3). The organic phases were combined, washed with brine (10 mL), dried over sodium sulfate, filtered, and concentrated by rotary evaporation. The crude product was purified by preparative chromatography to give 1-(2-chloro-5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl))))phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1017.3.
1H NMR (400 MHz, CD3OD) δ 7.99 (s, 1H), 7.65-7.59 (m, 1H), 7.52 (s, 1H), 7.44-7.26 (m, 5H), 7.24-7.16 (m, 2H), 7.01 (s, 1H), 6.11-6.01 (m, 1H), 4.66-4.46 (m, 2H), 4.09-3.98 (m, 1H), 3.95 (s, 3H), 3.81-3.67 (m, 3H), 3.53-3.40 (m, 4H), 3.16-3.00 (m, 3H), 2.93-2.57 (m, 5H), 2.52 (s, 3H), 2.23-2.05 (m, 7H), 1.95-1.57 (m, 15H), 1.56-1.51 (m, 2H), 1.34-1.00 (m, 7H).
NaH (214 mg, 5.33 mmol, 60%) was added DMF (15 mL) at 0° C., and then ethyl 2-(diethoxyphosphoryl)acetate (1.20 g, 5.33 mmol) was added. The reaction liquid was reacted at 0° C. for 30 min. tert-Butyl 9-formyl-3-azaspiro[5.5]undecane-3-carboxylate (1.00 g, 3.55 mmol) was added, and the resulting mixture was stirred at room temperature for 3 h. After the reaction was completed, the reaction liquid was cooled to 0° C., diluted with an aqueous NH4Cl solution (100 mL), and extracted with ethyl acetate (100 mL×3). The combined organic phase was washed with brine (100 mL), dried over sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (PE:EA=0-1:4) to give tert-butyl (E)-9-(3-ethoxy-3-oxoprop-1-en-1-yl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−56]+=296.2.
A solution of tert-butyl (E)-9-(3-ethoxy-3-oxoprop-1-en-1-yl)-3-azaspiro[5.5]undecane-3-carboxylate (1.1 g, 3.13 mmol) in EtOH (20 mL) was added to Pd/C (150 mg) at room temperature under H2 atmosphere. The reaction liquid was stirred overnight. The mixture was filtered and concentrated to give tert-butyl 9-(3-ethoxy-3-oxopropyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−56]+=298.2.
1H NMR (400 MHz, CDCl3) δ 4.12 (q, J=7.2 Hz, 2H), 3.40-3.30 (m, 4H), 2.30 (t, J=7.6 Hz, 2H), 1.71-1.62 (m, 2H), 1.58-1.52 (m, 5H), 1.47-1.42 (m, 10H), 1.31-1.22 (m, 6H), 1.13-1.02 (m, 4H).
tert-Butyl 9-(3-ethoxy-3-oxopropyl)-3-azaspiro[5.5]undecane-3-carboxylate (1 g, 2.83 mmol) was dissolved in THF (25 mL), LAH (3.4 mmol, 3.4 mL, 1 M/THF) was added dropwise at 0° C. The reaction liquid was reacted at 0° C. for 1 h. Then Na2SO4-10H2O was added to quench the reaction, and the resulting mixture was stirred for 20 min or longer. The mixture was filtered, and the filtrate was concentrated to give tert-butyl 9-(3-hydroxypropyl)-3-azaspiro[5.5]undecane-3-carboxylate, which was directly used in the next step.
1H NMR (400 MHz, CDCl3) δ 3.63 (t, J=7.2 Hz, 2H), 3.42-3.29 (m, 4H), 1.69-1.53 (m, 6H), 1.46-1.42 (m, 10H), 1.32-1.22 (m, 6H), 1.12-1.03 (m, 4H).
Dess-Martin oxidant (1.37 g, 3.3 mmol) was added to a solution of tert-butyl 9-(3-hydroxypropyl)-3-azaspiro[5.5]undecane-3-carboxylate (850 mg, 2.7 mmol) DCM (25 mL) in batches. The mixture was stirred at room temperature for 1.5 h. An aqueous Na2SO3 solution (50 mL) and an aqueous NaHCO3 solution (50 mL) were added to quench the reaction, and the resulting mixture was extracted with DCM (100 mL L×3). The combined organic phase was dried over anhydrous sodium sulfate, and filtered, and the filtrate was concentrated and purified by silica gel column chromatography (PE:EtOAc=20:1-2:1) to give tert-butyl 9-(3-oxopropyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−100]+=210.2.
tert-Butyl 9-(3-oxopropyl)-3-azaspiro[5.5]undecane-3-carboxylate (400 mg, 1.29 mmol) and 1-(4-methoxyphenyl)-N-methylmethanamine (0.39 g, 2.58 mmol) were dissolved DCM/AcOH (25 mL/0.5 mL). The reaction liquid was stirred at room temperature for 1 h. Then NaBH3 CN (162 mg, 2.58 mmol) was added, and the resulting mixture was reacted at room temperature for 16 h. The reaction liquid was diluted with an aqueous NH4Cl solution (50 mL) and extracted with DCM (50 mL×3). The combined organic phase was dried over sodium sulfate, and filtered, and the filtrate was concentrated and purified by column chromatography (DCM:MeOH=200:1-5:1) to give tert-butyl 9-(3-((4-methoxybenzyl)(methyl)amino)propyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=445.4.
tert-Butyl 9-(3-((4-methoxybenzyl)(methyl)amino)propyl)-3-azaspiro[5.5]undecane-3-carboxylate (400 mg, 0.89 mmol) was dissolved DCM (4.5 mL), TFA (1.5 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated to give N-(4-methoxybenzyl)-N-methyl-3-(3-azaspiro[5.5]undecan-9-yl)propan-1-amine, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=345.3.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (470 mg, 1.08 mmol) DIEA (350 mg, 2.7 mmol) were added to a solution of N-(4-methoxybenzyl)-N-methyl-3-(3-azaspiro[5.5]undecan-9-yl)propan-1-amine (400 mg, 0.9 mmol) DMSO (2 mL). The reaction liquid was stirred at room temperature for 2 h. Water (10 mL) was added, and the resulting mixture was extracted with EtOAc (20 mL×2). The organic phases were combined, washed with saturated brine (50 mL×3), dried over anhydrous sodium sulfate, and filtered, and the filtrate was concentrated and purified by column chromatography (DCM:MeOH=200:1-5:1) to give 1-(2-chloro-5-(9-(3-((4-methoxybenzyl)(methyl)amino)propyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=595.2.
1-(2-Chloro-5-(9-(3-((4-methoxybenzyl)(methyl)amino)propyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (130 mg, 0.218 mmol) DIEA (14 mg, 0.11 mmol) were dissolved DCM (3 mL), and 2-chloroethyl chloroformate (47 mg, 0.33 mmol) was added dropwise at 0° C. The mixture was stirred at room temperature for 2 h. Then MeOH (3 mL) was added, and the resulting mixture was reacted at 65° C. for 1 h. The mixture was concentrated and purified by column chromatography (DCM:MeOH=200:1-5:1) to give 1-(2-chloro-5-(9-(3-(methylamino)propyl)-3-)azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=475.2.
1-(2-Chloro-5-(9-(3-(methylamino)propyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (100 mg, 0.21 mmol) was dissolved in DMSO (2 mL), and pentafluorophenyl (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (187 mg, 0.23 mmol) and DIEA (82 mg, 0.63 mmol) were added.
The reaction liquid was reacted at room temperature for 2 h. After the reaction was completed, water (2 mL) was added, and the resulting mixture was extracted with ethyl acetate and water. The combined organic phase was dried over sodium sulfate, filtered, concentrated, and purified by column chromatography (DCM:MeOH=100:1-5:1) to give the target compound tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-((3-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidine))-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)propyl)(methyl)carbamoyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)aminocarboxylate.
LC-MS: (ESI, m/z): [1/2(M-100)+H]+=501.2.
The compound tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-((3-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidine))-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)propyl)(methyl)carbamoyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)aminocarboxylate (170 mg, crude product) was dissolved in DCM (4.5 mL), and TFA (1.5 mL) was added. The mixture was stirred at room temperature for 2 h, concentrated, and purified by high performance liquid chromatography to give (1R,4R)—N-(3-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)propyl)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)-N-methylcyclohexane-1-carboxamide.
LC-MS: (ESI, m/z): [M+H]+=1001.4.
1H NMR (400 MHz, CD3OD) δ 8.02-7.96 (m, 1H), 7.65-7.58 (m, 1H), 7.54-7.48 (m, 1H), 7.42-7.36 (m, 2H), 7.32-7.26 (m, 3H), 7.20-7.10 (m, 2H), 7.01 (s, 1H), 6.06 (q, J=6.8 Hz, 1H), 3.95 (s, 3H), 3.80-3.63 (m, 6H), 3.48-3.34 (m, 4H), 3.16-2.99 (m, 3H), 2.93-2.82 (m, 3H), 2.76-2.62 (m, 1H), 2.52 (s, 3H), 2.22 (s, 3H), 2.03-1.85 (m, 4H), 1.80-1.42 (m, 16H), 1.37-1.15 (m, 8H).
Ethyl 2-(diethoxyphosphoryl)acetate (1.26 g, 5.61 mmol) was added to a solution of NaH (230 mg, 5.61 mmol, 60%) DMF (15 mL) while stirring. The mixture was stirred at 0° C. for 30 min. Then tert-butyl 9-oxo-3-azaspiro[5.5]undecane-3-carboxylate (1 g, 3.74 mmol) was added, and the resulting mixture was stirred at room temperature for 3 h. The mixture was diluted with an aqueous NH4Cl solution (100 mL) and extracted with ethyl acetate (100 mL×3). The combined organic phase was washed with brine (100 mL), dried over sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (EA:PE=0-1:4) to give tert-butyl 9-(2-ethoxy-2-oxoethylidene)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=338.3.
tert-Butyl 9-(2-ethoxy-2-oxoethylidene)-3-azaspiro[5.5]undecane-3-carboxylate (1.1 g, 3.26 mmol) was dissolved in EtOH (20 mL), and the reaction liquid was stirred overnight under H2 atmosphere. The mixture was filtered and concentrated to give tert-butyl 9-(2-ethoxy-2-oxoethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−56]+=284.2.
LAH (3.6 mmol, 3.6 mL, 1 M/THF) was added to a solution of tert-butyl 9-(2-ethoxy-2-oxoethyl)3-azaspiro[5.5]undecane-3-carboxylate (1 g, 3 mmol) in THF (25 mL) while stirring. The reaction liquid was reacted at 0° C. to room temperature for 1 h. Na2SO4·10H2O was added to the mixture at room temperature, and the resulting mixture was stirred for 1 h or longer. The mixture was filtered and concentrated to give the target compound tert-butyl 9-(2-hydroxyethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−56]+=242.2.
Dess-Martin oxidant (1.42 g, 3.6 mmol) was added to a solution of tert-butyl 9-(2-hydroxyethyl)-3-azaspiro[5.5]undecane-3-carboxylate (850 mg, 2.5 mmol) DCM (25 mL) while stirring. The mixture was stirred at room temperature for 1.5 h. The mixture was diluted with an aqueous Na2SO3 solution (50 mL) and an aqueous NaHCO3 solution (50 mL) and extracted with DCM (100 mL×3). The combined organic phase was dried over sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (EA:PE=0-3:2) to give tert-butyl 9-(2-oxoethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H−56]+=240.2.
1-(4-Methoxybenzyl)piperazine (0.79 g, 3.81 mmol) was added to a solution of tert-butyl 9-(2-oxoethyl)-3-azaspiro[5.5]undecane-3-carboxylate (750 mg, 2.54 mmol) in DCM/HAc (25 mL/0.5 mL) while stirring. The mixture was stirred at room temperature for 1 h. Then NaBH3 CN (320 mg, 5.08 mmol) was added. The mixture was diluted with an aqueous NH4Cl solution (50 mL) and extracted with DCM (50 mL×3). The combined organic phase was dried over sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (DCM:MeOH=0-1:4) to give tert-butyl 9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=486.4.
TFA (0.5 mL) was added dropwise to a solution of tert-butyl 9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate (100 mg, 0.21 mmol) DCM (1.5 mL) while stirring. The mixture was stirred at room temperature for 2 h, and the reaction liquid was concentrated under reduced pressure to give 9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane.
LC-MS: (ESI, m/z): [M+H]+=386.4.
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (99 mg, 0.23 mmol) DIEA (80 mg, 0.62 mmol) were added to a solution of 9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane (100 mg, crude product) DMSO (2 mL) while stirring. The mixture was stirred at room temperature for 2 h and extracted with EA and water. The combined organic phase was dried over sodium sulfate, filtered, concentrated under reduced pressure, and purified by column chromatography (DCM:MeOH=0-1:4) to give 1-(2-chloro-5-(9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=636.4.
1-(2-Chloro-5-(9-(2-(4-(4-methoxybenzyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (90 mg, 0.14 mmol) was dissolved in DCM (2 mL), and 2-chloroethyl chloroformate (30 mg, 0.22 mmol) and DIEA (9 mg, 0.071 mmol) were added. The mixture was stirred at 0° C. to room temperature for 2 h. Then MeOH (3 mL) was added at 65° C. and the mixture was successively stirred for 1 h. The mixture was concentrated and purified by column chromatography (DCM:MeOH=0-1:4) to give 1-(2-chloro-5-(9-(2-(piperazin-1-yl)ethyl)))-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=516.3.
1-(2-Chloro-5-(9-(2-(piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (63 mg, 0.12 mmol) was dissolved in DMSO (2 mL), and pentafluorophenyl (1R,4R)-4-(4-(((R)-1-(4-(2-(((tert-butoxycarbonyl)(methyl)amino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (109 mg, 0.13 mmol) and DIEA (48 mg, 0.36 mmol) were added.
The mixture was stirred at room temperature for 2 h and extracted with EA and water. The combined organic phase was dried over sodium sulfate, filtered, and concentrated to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-(2-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)ethyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)aminocarboxylate.
LC-MS: (ESI, m/z): [(M-56)/2+H]+=544.3.
tert-Butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-(2-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)ethyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carbamate (160 mg, crude product) was dissolved in DCM (4.5 mL), and TFA (1.5 mL) was added. The mixture was stirred at room temperature for 2 h, concentrated, and purified by preparative chromatography to give 1-(2-chloro-5-(9-(2-(4-((1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-))-4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1042.4.
1H NMR (400 MHz, CD3OD) δ 7.98 (s, 1H), 7.68-7.56 (m, 1H), 7.57-7.48 (m, 1H), 7.45-7.35 (m, 2H), 7.34-7.24 (m, 3H), 7.22-7.10 (m, 2H), 7.01 (s, 1H), 6.06 (q, J=6.9 Hz, 1H), 3.95 (s, 3H), 3.83-3.73 (m, 2H), 3.74-3.68 (m, 3H), 3.66-3.55 (m, 4H), 3.45-3.34 (m, 2H), 3.12-3.01 (m, 1H), 2.93-2.76 (m, 2H), 2.76-2.66 (m, 1H), 2.55-2.39 (m, 8H), 2.21 (s, 3H), 2.03-1.86 (m, 4H), 1.81-1.75 (m, 4H), 1.70-1.57 (m, 6H), 1.45-1.16 (m, 14H).
Benzyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate (400 mg, 1.04 mmol) was dissolved with DCM (10 mL), and (Boc)2O (272 mg, 1.25 mmol), DMAP (13 mg, 0.1 mmol), and TEA (315 mg, 3.12 mmol) were separately added. The reaction liquid was stirred at room temperature overnight and directly concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:1) to give benzyl 9-((4-(tert-butoxycarbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=486.3.
Benzyl 9-((4-(tert-butoxycarbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (300 mg, 0.62 mmol) was dissolved with ethyl acetate (10 mL), and Pd(OH)2/C (100 mg) was added. The reaction liquid was stirred at 70° C. for 3 h, cooled, and filtered. The solid was washed three times with ethyl acetate (10 mL×3) and concentrated under reduced pressure to give the crude product tert-butyl 4-((3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=352.2.
tert-Butyl 4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (200 mg, 0.57 mmol) was dissolved with DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (247 mg, 0.57 mmol) and DIEA (220 mg, 1.71 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:10) to give tert-butyl 4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=602.2.
tert-Butyl 4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (350 mg, 0.58 mmol) was dissolved with DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The residue was diluted with water (20 mL), and a saturated sodium bicarbonate solution was slowly added dropwise to adjust the reaction liquid to neutrality. The reaction liquid was extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give the crude product 1-(2-chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecan-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=502.3.
1-(2-Chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (125 mg, 0.25 mmol) was dissolved with DMF (10 mL), and 4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid (140 mg, 0.23 mmol), HATU (114 mg, 0.30 mmol), and DIEA (129 mg, 1.00 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added, and the solid was precipitated, collected by filtration, washed 3 times with water, dissolved with dichloromethane, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:10) to give tert-butyl (3-((1R)-1-((6-(4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexan-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1095.5.
tert-Butyl (3-((1R)-1-((6-(4-(4-((3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexan-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (100 mg, 0.09 mmol) was dissolved with DCM (5 mL), and TFA (1 mL) was added. Then the reaction liquid was stirred at room temperature overnight, concentrated under reduced pressure, and diluted with water (20 mL). A sodium bicarbonate solution was slowly added dropwise to adjust the reaction liquid to neutrality. The reaction liquid was extracted with dichloromethane (25 mL×4), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(9-((4-(4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=984.3.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.18 (s, 1H), 8.06 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.55 (d, J=1.7 Hz, 1H), 7.39 (d, J=8.3 Hz, 1H), 6.99 (s, 1H), 6.89 (s, 1H), 6.85 (s, 1H), 6.69 (s, 1H), 5.83 (s, 1H), 5.59-5.47 (m, 3H), 3.85 (s, 3H), 3.80-3.70 (m, 1H), 3.67-3.45 (m, 7H), 2.94-2.82 (m, 1H), 2.77-2.71 (m, 2H), 2.42-2.10 (m, 13H), 1.89-1.81 (m, 1H), 1.73-1.25 (m, 15H), 1.18-0.94 (m, 4H).
tert-Butyl 9-aldhyde-3-azaspiro[5.5]undecane-3-carboxylate (200 mg, 0.71 mmol) was dissolved in a tetrahydrofuran solution (4 mL), and (9H-fluoren-9-yl)methyl piperazine-1-carboxylate (219 mg, 0.71 mmol) and sodium triacetoxyborohydride (449 mg, 2.13 mmol) were added. The reaction liquid was stirred at room temperature overnight, diluted with water (40 mL), extracted with ethyl acetate (80 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:1) to give tert-butyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=574.4.
tert-Butyl 9-((4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (400 mg, 0.7 mmol) was dissolved in a solution of hydrogen chloride in 1,4-dioxane (4 mmol/L, 5 mL).
The reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give (9H-fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=474.5.
(9H-Fluoren-9-yl)methyl 4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (340 mg, 0.72 mmol) was dissolved with DMSO (4 mL), and pentafluorophenyl 4-(2,6-dioxopiperidin-3-yl)benzoate (287 mg, 0.72 mmol) and DIEA (371 mg, 2.87 mmol) were added. The reaction liquid was stirred at room temperature overnight, diluted with water (40 mL), and extracted with ethyl acetate (40 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:2) to give (9H-fluoren-9-yl)methyl 4-((3-(4-(2,6-dioxopiperidin-3-yl)benzoyl)-3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=689.5.
(9H-Fluoren-9-yl)methyl 4-((3-(4-(2,6-dioxopiperidin-3-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carboxylate (250 mg, 0.36 mmol) was dissolved with dichloromethane (4 mL), and piperidine (0.25 mL) was added. The reaction liquid was stirred at room temperature for 3 h and directly concentrated under reduced pressure to give 3-(4-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecan-3-carbonyl)phenyl)piperidine-2,6-dione.
LC-MS: (ESI, m/z): [M+H]+=467.4.
3-(4-(9-(Piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)piperidine-2,6-dione (200 mg, 0.43 mmol) was dissolved with DMF (3 mL), and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (254 mg, 0.43 mmol), HATU (196 mg, 0.52 mmol), and DIEA (221 mg, 1.7 mmol) were separately added. The reaction liquid was stirred at room temperature for 3 h, diluted with water (40 mL), and extracted with ethyl acetate (40 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by Prep-HPLC to give 3-(4-(9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)piperidine-2,6-dione.
LC-MS: (ESI, m/z): [M+H]+=1041.6.
1H NMR (400 MHz, DMSO-d6) δ 10.86 (s, 1H), 8.31 (d, J=8.5 Hz, 1H), 8.05 (s, 1H), 7.45-7.40 (s, 2H), 7.36-7.32 (s, 3H), 7.28 (d, J=8.1 Hz, 2H), 7.24 (d, J=1.2 Hz, 1H), 7.01 (s, 1H), 6.96 (s, 1H), 5.99-5.94 (m, 1H), 3.94-3.87 (m, 4H), 3.57 (s, 2H), 3.46 (s, 5H), 3.12 (s, 2H), 2.94 (s, 1H), 2.74-2.59 (m, 4H), 2.42 (s, 3H), 2.23 (d, J=13.2 Hz, 5H), 2.17-2.04 (m, 4H), 2.01 (s, 7H), 1.88-1.76 (m, 5H), 1.71 (d, J=6.9 Hz, 4H), 1.68 (s, 1H), 1.64-1.48 (m, 8H), 1.23 (s, 2H), 1.07 (s, 3H).
Pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (140 mg, 0.32 mmol) was dissolved with DMSO (5 mL), and tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperazine-1-carboxylate (116 mg, 0.35 mmol) and DIEA (124 mg, 0.96 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give tert-butyl 4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=578.2.
tert-Butyl 4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperazine-1-carboxylate (170 mg, 0.29 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure to give 1-(2-chloro-5-(4-((2-(piperazin-1-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=478.2.
1-(2-Chloro-5-(4-((2-(piperazin-1-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (170 mg, 0.36 mmol) was dissolved in DMF (5 mL), and 4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid (150 mg, 0.25 mmol), HATU (124 mg, 0.32 mmol), and DIEA (161 mg, 1.25 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure.
The crude product was purified by column chromatography (MeOH:DCM=1:9) to give tert-butyl (3-((1R)-1-((6-(4-(4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperazine-1-carbonyl)cyclohex-1-en-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1060.4.
tert-Butyl (3-((1R)-1-((6-(4-(4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperazine-1-carbonyl)cyclohex-1-en-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (140 mg, 0.13 mmol) was dissolved in DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The residue was dissolved with MeOH/DCM (1/9, 20 mL), and a saturated aqueous sodium bicarbonate solution (20 mL) was added to adjust the pH to neutrality. The resulting mixture was stirred for 10 min, extracted with MeOH/DCM (1/9, 20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(4-((2-(4-(4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-en-1-carbonyl)piperazin-1-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=960.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.18 (d, J=7.8 Hz, 1H), 8.06 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=2.0 Hz, 1H), 7.38 (dd, J=8.2, 2.0 Hz, 1H), 6.99 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.69 (s, 1H), 5.83 (s, 1H), 5.55-5.45 (m, 3H), 4.50-4.36 (m, 1H), 3.85 (s, 3H), 3.75 (d, J=7.2 Hz, 1H), 3.67-3.37 (m, 9H), 3.06 (s, 1H), 2.87 (s, 1H), 2.81-2.69 (m, 3H), 2.47-2.16 (m, 13H), 1.89-1.41 (m, 9H), 1.20-1.13 (m, 2H).
tert-Butyl 4-(3-hydroxypropyl)piperidine-1-carboxylate (3.6 g, 14.81 mmol) was dissolved with dichloromethane (20 mL), and TsCl (3.4 g, 17.80 mmol), DMAP (180 mg, 1.48 mmol), and TEA (4.5 g, 44.43 mmol) were separately added. The reaction liquid was stirred at room temperature overnight and directly concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=5:1) to give tert-butyl 4-(3-(p-toluenesulfonyl)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=398.1.
Benzyl 4-hydroxypiperidine-1-carboxylate (4.4 g, 18.51 mmol) was dissolved with anhydrous DMF (10 mL) and cooled to 0° C., and NaH (0.6 g, 24.68 mmol) was added. The reaction liquid was stirred for 0.5 h, and then stirred at room temperature for 1 h. tert-Butyl 4-(3-(p-toluenesulfonyl)propyl)piperidine-1-carboxylate (4.9 g, 12.34 mmol) was added, and the reaction liquid was stirred at room temperature overnight. Water (100 mL) was added, and the resulting mixture was extracted with ethyl acetate (100 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=5:1) to give benzyl 4-(3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propoxy)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=461.2.
Benzyl 4-(3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propoxy)piperidine-1-carboxylate (540 mg, 1.17 mmol) was dissolved with ethyl acetate (10 mL), and Pd(OH)2/C (20%, 170 mg) was added. The reaction liquid was heated to 70° C., stirred for 3 h, cooled, and filtered. The solid was washed three times with ethyl acetate (20 mL×3) and concentrated under reduced pressure to give the crude product tert-butyl 4-(3-(piperidine-4-oxy)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=327.3.
tert-Butyl 4-(3-(piperidine-4-oxy)propyl)piperidine-1-carboxylate (150 mg, 0.46 mmol) was dissolved with DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (200 mg, 0.46 mmol) and DIEA (178 mg, 1.38 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h.
Water (50 mL) was added, and the resulting mixture was extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:10) to give tert-butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=577.3.
tert-Butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperidine-1-carboxylate (220 mg, 0.58 mmol) was dissolved with DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature overnight and concentrated under reduced pressure.
The residue was diluted with water (10 mL), and a saturated sodium bicarbonate solution was slowly added dropwise to adjust the reaction liquid to neutrality. The resulting mixture was extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give the crude product 1-(2-chloro-5-(4-(3-(piperidin-4-yl)propoxy)piperidin-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=477.2.
1-(2-Chloro-5-(4-(3-(piperidin-4-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (71 mg, 0.14 mmol), HATU (63 mg, 0.17 mmol), and DIEA (54 mg, 0.42 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (80 mg, 0.14 mmol) in DMF (10 mL). The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added, and the resulting mixture was extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(2-chloro-5-(4-(3-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1051.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.31 (d, J=8.4 Hz, 1H), 8.06 (s, 1H), 7.64 (d, J=8.2 Hz, 1H), 7.57 (s, 1H), 7.42 (d, J=8.5 Hz, 2H), 7.36 (d, J=7.8 Hz, 1H), 7.24 (s, 1H), 7.02 (s, 1H), 6.97 (s, 1H), 5.97 (s, 1H), 4.45-4.35 (m, 1H), 3.98-3.84 (m, 5H), 3.82-3.71 (m, 1H), 3.67-3.57 (m, 1H), 3.61-3.50 (m, 2H), 3.41 (t, J=6.3 Hz, 3H), 3.18-2.90 (m, 6H), 2.78-2.70 (m, 1H), 2.70-2.60 (m, 1H), 2.43 (s, 3H), 2.02 (s, 6H), 1.90-1.40 (m, 20H), 1.30-1.21 (m, 2H), 1.08-0.87 (m, 2H).
4-(4-((1-(1,1-Dioxido-2,3-dihydrobenzo[b]thiophen-4-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (70 mg, 0.14 mmol) was dissolved with DMF (3 mL), and 1-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (73.3 mg, 0.15 mmol), HATU (79.8 mg, 0.21 mmol), and DIEA (91 mg, 0.7 mmol) were separately added. The reaction liquid was stirred at room temperature for 1 h, diluted with water (40 mL), and extracted with ethyl acetate (40 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(1,1-dioxide-2,3-dihydrobenzo[b]thiophen-4-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=968.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.27 (d, J=7.0 Hz, 1H), 8.06 (s, 1H), 7.80 (d, J=7.6 Hz, 1H), 7.67-7.57 (m, 2H), 7.56-7.48 (m, 2H), 7.38 (d, J=8.4 Hz, 1H), 6.98 (s, 1H), 5.59 (t, J=7.4 Hz, 1H), 4.39 (d, J=11.8 Hz, 2H), 3.91 (s, 1H), 3.87 (s, 3H), 3.81-3.70 (m, 2H), 3.67-3.59 (m, 3H), 3.54-3.46 (m, 1H), 3.42 (t, J=6.4 Hz, 2H), 3.29 (s, 1H), 3.24 (d, J=6.4 Hz, 2H), 3.11-2.90 (m, 3H), 2.82-2.62 (m, 4H), 2.32 (s, 3H), 2.12-1.23 (m, 20H), 1.20-0.90 (m, 4H).
tert-Butyl 4-(4-(piperazin-1-yl)butyl)piperidine-1-carboxylate (100 mg, 0.31 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (133 mg, 0.31 mmol) were dissolved DMSO (3 mL), DIEA (198 mg, 1.54 mmol) was added while stirring. The reaction liquid was stirred at room temperature for 1 h.
After the reaction was completed, the reaction liquid was poured into water (30 mL), and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (DCM:MeOH=10:1) to give tert-butyl 4-(4-(4-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperazin-1-yl)butyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=576.3.
tert-Butyl 4-(4-(4-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperazin-1-yl)butyl)piperidine-1-carboxylate (112 mg, 0.19 mmol) was dissolved in DCM (2 mL), 0.2 mL of TFA was added, and the reaction liquid was reacted at room temperature for 1 h and directly concentrated under reduced pressure to give 1-(2-chloro-5-(4-(4-(piperidin-4-yl)butyl)piperazine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=476.3
1-(2-Chloro-5-(4-(4-(piperidin-4-yl)butyl)piperazine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (165 mg, 0.195 mmol) and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (115 mg, 0.195 mmol) were dissolved in DMF (3 mL), and HATU (86 mg, 0.227 mmol) and DIEA (75 mg, 0.584 mmol) were added while stirring. The reaction liquid was reacted at room temperature for 1 h. After the reaction was completed, the reaction liquid was poured into water (50 mL), and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-(4-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)butyl)piperazine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1050.7.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.30 (d, J=8.3 Hz, 1H), 8.05 (s, 1H), 7.64 (d, J=8.2 Hz, 1H), 7.56 (d, J=2.0 Hz, 1H), 7.46-7.30 (m, 4H), 7.24 (d, J=1.4 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 5.96 (dd, J=14.4, 7.2 Hz, 1H), 4.40 (d, J=11.5 Hz, 1H), 3.89 (s, 4H), 3.78-3.71 (m, 1H), 3.62 (d, J=6.0 Hz, 3H), 3.11 (d, J=15.8 Hz, 2H), 3.01-2.92 (m, 2H), 2.76-2.63 (m, 2H), 2.68-2.58 (m, 2H), 2.42 (s, 6H), 2.34-2.18 (m, 4H), 2.01 (s, 6H), 1.85 (d, J=9.6 Hz, 2H), 1.79 (d, J=11.5 Hz, 2H), 1.72 (d, J=7.0 Hz, 4H), 1.65 (d, J=12.4 Hz, 2H), 1.60-1.50 (m, 3H), 1.42 (d, J=7.4 Hz, 3H), 1.34-1.14 (m, 5H), 1.02-0.82 (m, 2H).
tert-Butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (100 mg, 0.31 mmol) and pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-(trifluoromethyl)benzoate (133 mg, 0.31 mmol) were dissolved in DMSO (3 mL), and DIEA (198 mg, 1.54 mmol) was added while stirring. The reaction liquid was reacted at room temperature for 1 h. After the reaction was completed, the reaction liquid was diluted with water (30 mL) and extracted with ethyl acetate (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was isolated by column chromatography to give tert-butyl 4-(2-((1-(3-(2,4-dioxotetrahydropyrimidine-1(2H)-yl)-4-(trifluoromethyl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+Na]+=633.2.
tert-Butyl 4-(2-((1-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-(trifluoromethyl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (112 mg, 0.19 mmol) was dissolved in dichloromethane (2 mL), and trifluoroacetic acid (0.8 mL) was added. The reaction liquid was reacted at room temperature for 1 h. After the reaction was completed, the reaction liquid was concentrated under reduced pressure to give 1-(5-(4-((2-(piperidine-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was directly used in the next step without purification.
1-(5-(4-((2-(Piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (TFA salt, 165 mg, 0.195 mmol) and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (115.3 mg, 0.195 mmol) were dissolved in DMF (3 mL), and HATU (86.3 mg, 0.227 mmol) and DIEA (75.4 mg, 0.584 mmol) were added while stirring. The reaction liquid was reacted at room temperature for 1 h. After the reaction was completed, the reaction liquid was diluted with water (30 mL) and extracted with ethyl acetate (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1085.5.
1H NMR (400 MHz, DMSO-d6) δ 10.53 (s, 1H), 8.31 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.87 (d, J=8.1 Hz, 1H), 7.70 (s, 1H), 7.58 (d, J=8.0 Hz, 1H), 7.43 (t, J=8.0 Hz, 2H), 7.37-7.31 (m, 1H), 7.24 (d, J=1.3 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.00-5.92 (m, 1H), 4.53-4.31 (m, 2H), 3.94-3.82 (m, 5H), 3.52-3.37 (m, 4H), 3.23 (d, J=6.1 Hz, 2H), 3.12 (d, J=2.1 Hz, 2H), 3.07-2.90 (m, 3H), 2.85-2.74 (m, 1H), 2.72-2.58 (m, 3H), 2.42 (s, 3H), 2.01 (s, 6H), 1.89-1.50 (m, 18H), 1.48-1.40 (m, 2H), 1.21-0.91 (m, 4H).
Pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-(trifluoromethoxy)benzoate (150 mg, 0.31 mmol) and tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (100 mg, 0.31 mmol) were dissolved in DMSO (3 mL), and DIEA (198 mg, 1.54 mmol) was added while stirring. The reaction liquid was stirred at room temperature for 1 h. After the reaction was completed, the reaction liquid was poured into water (30 mL), and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (DCM:MeOH=10:1) to give tert-butyl 4-(2-((1-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-(trifluoromethoxy)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+Na]+=649.3.
tert-Butyl 4-(2-((1-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-(trifluoromethoxy)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (110 mg, 0.17 mmol) was dissolved in DCM (2 mL), TFA (0.2 mL) was added, and the reaction liquid was reacted at room temperature for 1 h and directly concentrated under reduced pressure to give 1-(5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethoxy)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=527.3
1-(5-(4-((2-(Piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (90 mg, 0.17 mmol) and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (100 mg, 0.17 mmol) were dissolved in DMF (3 mL), and HATU (84 mg, 0.22 mmol) and DIEA (66 mg, 0.51 mmol) were added while stirring. The reaction liquid was reacted at room temperature for 1 h. After the reaction was completed, the reaction liquid was poured into water (50 mL), and the resulting mixture was extracted with EA (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by prep-HPLC to give 1-(5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethoxy)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1101.5.
1H NMR (400 MHz, DMSO-d6) δ 10.54 (s, 1H), 8.31 (d, J=8.2 Hz, 1H), 8.05 (s, 1H), 7.59 (d, J=2.0 Hz, 1H), 7.52 (dd, J=8.4, 1.4 Hz, 1H), 7.49-7.30 (m, 4H), 7.24 (d, J=1.3 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.02-5.91 (m, 1H), 4.52-4.33 (m, 2H), 3.98-3.90 (m, 4H), 3.80-3.65 (m, 2H), 3.61-3.51 (m, 1H), 3.41 (t, J=6.3 Hz, 2H), 3.23 (d, J=6.2 Hz, 2H), 3.15-2.89 (m, 5H), 2.82-2.60 (m, 4H), 2.42 (s, 3H), 2.01 (s, 6H), 1.89-1.40 (m, 20H), 1.20-0.90 (m, 4H).
tert-Butyl 9-formyl-3-azaspiro[5.5]undecane-3-carboxylate (300 mg, 1.06 mmol) and benzyl piperazine-1-carboxylate (235 mg, 1.06 mmol) were dissolved with tetrahydrofuran (10 mL), and sodium triacetoxyborohydride (674 mg, 3.18 mmol) was added while stirring. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was extracted with dichloromethane (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 9-((4-((benzyloxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=486.2.
tert-Butyl 9-((4-((benzyloxy)carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (500 mg, 1.02 mmol) was dissolved in ethyl acetate (15 mL), and Pd(OH)2/C (250 mg, 0.35 mmol) was added. The reaction liquid was heated to 70° C. under hydrogen atmosphere and stirred overnight. The reaction liquid was cooled to room temperature and filtered, and the filtrate was concentrated under reduced pressure to give tert-butyl 9-(piperazine-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate. The crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=352.2.
tert-Butyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate (130 mg, 0.36 mmol) was dissolved in DMF (10 mL), and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (200 mg, 0.33 mmol), HATU (188 mg, 0.49 mmol), and DIEA (127 mg, 0.99 mmol) were added. The mixture was stirred at room temperature for 1 h. Water (100 mL) was added to the reaction liquid, and the resulting mixture was extracted with ethyl acetate (3×30 mL).
The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=926.4.
tert-Butyl 9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (230 mg, 0.25 mmol) was dissolved in DCM (3 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure to give (4-((3-azaspiro[5.5]undecane-9-yl)methyl)piperazine-1-yl)((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone.
LC-MS: (ESI, m/z): [M+H]+=826.5.
(4-((3-Azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl) ((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone (110 mg, 0.12 mmol) was dissolved in DMSO (10 mL), and pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoate (57 mg, 0.13 mmol) and DIEA (46 mg, 0.36 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added to the reaction liquid, and the resulting mixture was extracted with dichloromethane (30 mL×3). The organic phase was washed with a saturated aqueous sodium chloride solution (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude product. The crude product was purified by pre-HPLC (CH3CN/0.08% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-methoxyphenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1072.5.
1H NMR (400 MHz, DMSO-d6) δ 10.32 (s, 1H), 8.32 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.43 (t, J=8.1 Hz, 2H), 7.38-7.33 (m, 2H), 7.32 (d, J=2.0 Hz, 1H), 7.24 (s, 1H), 7.15 (d, J=8.6 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.00-5.92 (m, 1H), 3.89 (s, 3H), 3.84 (s, 3H), 3.59 (t, J=6.6 Hz, 2H), 3.49-3.37 (m, 4H), 3.17-3.09 (m, 2H), 2.94 (t, J=12.0 Hz, 1H), 2.68 (t, J=6.6 Hz, 2H), 2.65-2.43 (m, 2H), 2.42 (s, 3H), 2.33-2.26 (m, 4H), 2.16-2.08 (m, 2H), 2.02 (s, 6H), 1.83-1.46 (m, 21H), 1.35-1.24 (m, 2H), 1.16-0.90 (m, 4H).
(4-((3-Azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl) ((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)methanone (110 mg, 0.12 mmol) was dissolved in DMSO (5 mL), and pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoate (54 mg, 0.13 mmol) and DIEA (46 mg, 0.36 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added to the reaction liquid, and the resulting mixture was extracted with dichloromethane (30 mL×3). The organic phase was washed with a saturated aqueous sodium chloride solution (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude product. The crude product was purified by pre-HPLC (CH3CN/0.08% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-fluorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1060.5.
1H NMR (400 MHz, DMSO-d6) δ 10.52 (s, 1H), 8.32 (d, J=8.3 Hz, 1H), 8.05 (s, 1H), 7.50 (d, J=7.9 Hz, 1H), 7.38 (m, 5H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.04-5.88 (m, 1H), 3.89 (s, 3H), 3.75 (t, J=6.6 Hz, 2H), 3.57 (s, 2H), 3.46 (s, 4H), 3.30-3.26 (m, 1H), 3.11 (d, J=15.1 Hz, 2H), 2.94 (m, 1H), 2.73 (t, J=6.6 Hz, 2H), 2.63 (m, 1H), 2.42 (s, 3H), 2.32 (s, 2H), 2.29 (s, 2H), 2.11 (s, 2H), 2.01 (s, 6H), 1.90-1.39 (m, 19H), 1.34 (m, 2H), 1.07 (m, 4H).
tert-Butyl 4-(3-(piperidine-4-oxy)propyl)piperidine-1-carboxylate (105 mg, 0.32 mmol), HATU (145 mg, 0.38 mmol) and DIEA (123 mg, 0.96 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (190 mg, 0.32 mmol) in DMF (5 mL). The mixture was stirred at room temperature for 2 h, diluted with water (50 mL), extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product of tert-butyl 4-(3-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)oxy)propyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=901.4.
tert-Butyl 4-(3-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)oxy)propyl)piperidine-1-carboxylate (170 mg, 0.19 mmol) was dissolved with DCM (5 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and directly concentrated under reduced pressure to give the crude product ((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(3-(piperidin-4-yl)propoxy)piperidin-1-yl)methanone.
LC-MS: (ESI, m/z): [M+H]+=801.4.
((1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(3-(piperidin-4-yl)propoxy)piperidin-1-yl)methanone (150 mg, 0.19 mmol) was dissolved with DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (97 mg, 0.23 mmol) and DIEA (72 mg, 0.56 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h, diluted with water (50 mL), extracted with ethyl acetate (25 mL×4), washed with saturated brine, drying over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(2-chloro-5-(4-(3-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)oxy)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1051.4.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.30 (d, J=8.2 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.7 Hz, 1H), 7.48-7.30 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.03-5.87 (m, 1H), 4.53-4.33 (m, 1H), 3.93-3.81 (m, 4H), 3.79-3.45 (m, 5H), 3.41 (t, J=6.2 Hz, 2H), 3.27-3.20 (m, 1H), 3.18-2.99 (m, 4H), 2.99-2.89 (m, 1H), 2.80-2.60 (m, 4H), 2.42 (s, 3H), 2.01 (s, 6H), 1.88-1.23 (m, 22H), 1.10-1.00 (m, 2H).
The compound tert-butyl 9-(2-(4-(((9H-fluoren-9-yl)methoxy)carbonyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate (70 mg, 0.12 mmol) was dissolved in a solution of hydrogen chloride in 1,4-dioxane (4 M, 7 mL). The reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give (9H-fluoren-9-yl)methyl 4-(2-(3-azaspiro[5.5]undecane-9-yl)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=488.5.
(9H-Fluoren-9-yl)methyl 4-(2-(3-azaspiro[5.5]undecan-9-yl)ethyl)piperidine-1-carboxylate (70 mg, 0.12 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (52 mg, 0.12 mmol) were dissolved with DMSO (4 mL), and DIPEA (46 mg, 0.36 mmol) was added. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with water (40 mL), extracted with ethyl acetate (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (EA in PE=0-4:1) to give (9H-fluoren-9-yl)methyl 4-(2-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)ethyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=738.5.
(9H-Fluoren-9-yl)methyl 4-(2-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)ethyl)piperazine-1-carboxylate (70 mg, 0.09 mmol) was dissolved with DCM (2 mL), and piperidine (0.3 mL) was added. The reaction liquid was stirred at room temperature for 3 h, diluted with water (20 mL), extracted with dichloromethane (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (DCM:MeOH=95:5) to give 1-(2-chloro-5-(9-(2-(piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+Na]+=516.3.
1-(2-Chloro-5-(9-(2-(piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (30 mg, 0.06 mmol) and (1R,4R)-4-(4-(((R)-1-(4-(2-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (34 mg, 0.06 mmol) were dissolved with DMF (3 mL), and HATU (27 mg, 0.07 mmol) and DIEA (23 mg, 0.18 mmol) were added. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with water (30 mL), extracted with ethyl acetate (30 mL×3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(9-(2-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)ethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1090.5.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.30 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (s, 1H), 7.47-7.28 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.00-5.92 (m, 1H), 3.89 (s, 3H), 3.80-3.71 (m, 1H), 3.66-3.42 (m, 8H), 3.12 (s, 2H), 3.00-2.90 (m, 1H), 2.78-2.71 (m, 2H), 2.65-2.57 (m, 2H), 2.42 (s, 3H), 2.36-2.23 (m, 6H), 2.01 (s, 6H), 1.89-1.75 (m, 4H), 1.72-1.29 (m, 18H), 1.16-1.00 (m, 4H).
3-(Piperidin-4-yl)propan-1-ol (700 mg, 4.9 mmol) was dissolved in 1,4-dioxane (7 mL), and Boc anhydride (2.1 g, 9.8 mmol) and DMAP (30 mg, 0.24 mmol) were added. The reaction liquid was heated to 80° C. overnight. After the reaction was completed, the reaction liquid was poured into water (60 mL), and the resulting mixture was extracted with ethyl acetate (30 mL×3) and dried over anhydrous sodium sulfate. The organic phase was concentrated by rotary evaporation. The crude product was purified by column chromatography (EA:PE=0-3:7) to give tert-butyl 4-(3-hydroxypropyl)piperidine-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 4.34 (t, J=5.1 Hz, 1H), 3.91 (d, J=12.3 Hz, 2H), 3.37 (dd, J=11.9, 6.3 Hz, 2H), 2.66 (s, 2H), 1.61 (d, J=12.3 Hz, 2H), 1.50-1.28 (m, 12H), 1.20 (dd, J=15.2, 6.9 Hz, 2H), 0.93 (m, 2H).
tert-Butyl 4-(3-hydroxypropyl)piperidine-1-carboxylate (300 mg, 1.2 mmol) was dissolved in acetonitrile (5 mL), and IBX (567 mg, 2.4 mmol) was added. The reaction liquid was heated to 70° C., reacted for 2 h, and filtered, and the filter cake was washed with ethyl acetate (10 mL×2). The organic phase was concentrated by rotary evaporation.
The crude product was purified by column chromatography (EA:PE=0-1:4) to give tert-butyl 4-(3-propionaldehyde)piperidine-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 9.68 (t, J=1.5 Hz, 1H), 3.91 (d, J=12.0 Hz, 2H), 2.65 (m, 2H), 2.46 (td, J=7.5, 1.5 Hz, 2H), 1.61 (d, J=13.0 Hz, 2H), 1.46 (dd, J=14.5, 7.3 Hz, 2H), 1.42-1.33 (m, 10H), 0.94 (m, 2H).
tert-Butyl 4-(3-propionaldehyde)piperidine-1-carboxylate (200 mg, 0.82 mmol) and benzyl 4-(methylamino)piperidine-1-carboxylate (205 mg, 0.82 mmol) were dissolved in tetrahydrofuran (4 mL), and sodium triacetoxyborohydride (525 mg, 2.48 mmol) was added. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with water (40 mL), extracted with ethyl acetate (30 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (EA:PE=0-3:7) to give benzyl 4-((3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate as a white oil.
LC-MS: (ESI, m/z): [M+H]+=474.3.
Benzyl 4-((3-(1-(tert-butoxycarbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate (90 mg, 0.19 mmol) was dissolved in DCM (5 mL), and trifluoroacetic acid (0.5 mL) was added. The reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give benzyl 4-(methyl(3-(piperidin-4-yl)propyl)amino)piperidine-1-carboxylate as an oil.
LC-MS: (ESI, m/z): [M+H]+=374.1.
(1R,4R)-4-(4-(((R)-1-(3-((tert-Butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (110 mg, 0.18 mmol) and benzyl 4-(methyl(3-(piperidin-4-yl)propyl)amino)piperidine-1-carboxylate (trifluoroacetate salt, 90 mg, 0.18 mmol) were dissolved in DMF (3 mL), and HATU (83 mg, 0.22 mmol) and DIEA (70 mg, 0.54 mmol) were added. The reaction liquid was stirred at room temperature overnight, diluted with water (30 mL), extracted with ethyl acetate (30 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (MeOH:DCM=0-1:9) to give benzyl 4-((3-(1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=958.2.
Benzyl 4-((3-(1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carboxylate (100 mg, 0.1 mmol) was dissolved in MeOH (5 mL), and Pd/C (20 mg) was added. The reaction liquid was purged 3 times with hydrogen and stirred at room temperature for 1 h under hydrogen atmosphere. The reaction liquid was filtered, the filter cake was washed with methanol (5 mL×3), and the filtrate was concentrated under reduced pressure to give tert-butyl (3-((R)-1-((7-methoxy-2-methyl-6-((1R,4R)-4-(4-(3-(methyl(piperidin-4-yl)amino)propyl)piperidine-1-carbonyl)cyclohexyl)quinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=824.1.
The compound tert-butyl (3-((R)-1-((7-methoxy-2-methyl-6-((1R,4R)-4-(4-(3-(methyl(piperidin-4-yl)amino)propyl)piperidine-1-carbonyl)cyclohexyl)quinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (75 mg, 0.09 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (39 mg, 0.09 mmol) were dissolved in DMSO (2 mL), and DIEA (35 mg, 0.27 mmol) was added. The reaction liquid was stirred at room temperature overnight. The reaction liquid was diluted with water (20 mL), extracted with ethyl acetate (30 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (MeOH:DCM=0-5:95) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)(methyl)amino)propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1074.3.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)(methyl)amino) propyl)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (40 mg, 0.04 mmol) was dissolved in dichloromethane (3 mL), and trifluoroacetic acid (0.3 mL) was added. The reaction liquid was stirred at room temperature for 1 h, directly concentrated under reduced pressure, adjusted to pH 8-9 with a saturated aqueous sodium bicarbonate solution, extracted with dichloromethane (20 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by prep-HPLC to give 1-(5-(4-((3-(1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)(methyl)amino)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=974.3.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.15-8.02 (m, 2H), 7.63 (d, J=8.2 Hz, 1H), 7.56 (d, J=1.9 Hz, 1H), 7.40 (d, J=8.2 Hz, 1H), 6.99 (s, 1H), 6.86 (d, J=10.8 Hz, 2H), 6.70 (s, 1H), 5.64-5.47 (m, 3H), 4.54-4.48 (m, 2H), 3.96-3.88 (m, 4H), 3.88-3.80 (m, 1H), 3.80-3.74 (m, 2H), 3.05-2.95 (m, 3H), 2.91-2.85 (m, 2H), 2.40-2.30 (m, 6H), 2.15 (s, 3H), 1.91-1.42 (m, 23H), 1.25-1.15 (m, 2H), 1.06-0.84 (m, 2H).
6-Bromo-7-methoxy-2-methylquinazolin-4-ol (6.2 g, 23.22 mmol) was added to a mixed solution of Pd(dppf)Cl2 (850 mg, 1.16 mmol), sodium carbonate (4.9 g, 46.44 mmol), and tert-butyl 4-(4,4,5,5-tetramethyl-1,3-dioxolan-2-yl)cyclohex-3-ene-1-carboxylate (8.1 g, 30.19 mmol) in DMF/H2O (80 mL/8 mL). The reaction liquid was stirred at room temperature overnight. After the reaction was completed, the reaction liquid was diluted with water (800 mL), extracted with ethyl acetate (200 mL×5), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:100) to give methyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=329.0.
Methyl 4-(4-hydroxy-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (600 mg, 1.82 mmol) was added to a solution of DBU (820 mg, 5.37 mmol) and BOP (1.19 g, 2.69 mmol) in DMF (10 mL). The reaction liquid was stirred at room temperature for 30 min. tert-Butyl (R)-(3-(1-aminoethyl)-5-(trifluoromethyl)phenyl)carboxylate (600 mg, 1.97 mmol) was added, and the reaction liquid was stirred at 70° C. overnight. The reaction liquid was diluted with water (100 mL), and the solid was precipitated, collected by filtration, washed three times with water, dissolved with dichloromethane (100 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:1) to give methyl 4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=615.2.
Methyl 4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (1.1 g, 1.79 mmol) was added to a solution of lithium hydroxide (430 mg, 17.9 mmol) in THF/1-H2O (20 mL/10 mL). The reaction liquid was stirred at 50° C. overnight under nitrogen atmosphere. 10% citric acid was slowly added dropwise in an ice bath, and the solution was adjusted to weak acidity or neutrality, extracted with ethyl acetate (100 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give the crude product 4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=601.2.
4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylic acid (130 mg, 0.22 mmol), HATU (100 mg, 0.26 mmol) and DIEA (85 mg, 0.66 mmol) were added to a solution of 1-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (140 mg, 0.23 mmol) in DMF (10 mL). The mixture was stirred at room temperature for 2 h, water (100 mL) was added, a solid was precipitated, and the mixture was filtered. The solid was washed 3 times with water, dissolved in dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product of tert-butyl (3-((1R)-1-((6-(4-(4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohex-1-en-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1059.5.
tert-Butyl (3-((1R)-1-((6-(4-(4-(2-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)cyclohex-1-en-1-yl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (190 mg, 0.18 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (1 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(4-((2-(1-(4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohex-3-ene-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=959.2.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.19 (s, 1H), 8.07 (s, 1H), 7.63 (d, J=8.3 Hz, 1H), 7.54 (d, J=1.8 Hz, 1H), 7.38 (dd, J=8.2, 1.8 Hz, 1H), 6.99 (s, 1H), 6.89 (s, 1H), 6.85 (s, 1H), 6.69 (s, 1H), 5.83 (s, 1H), 5.61-5.43 (m, 3H), 4.51-4.37 (m, 2H), 4.00-3.90 (m, 1H), 3.85 (s, 3H), 3.80-3.70 (m, 1H), 3.67-3.52 (m, 2H), 3.41 (t, J=6.3 Hz, 2H), 3.24 (d, J=6.4 Hz, 2H), 3.12-2.97 (m, 2H), 2.92-2.70 (m, 4H), 2.42-2.16 (m, 7H), 1.90-1.56 (m, 8H), 1.56-1.41 (m, 5H), 1.21-0.90 (m, 4H).
1-(2-Chloro-5-(4-(3-(piperazin-1-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (90 mg, 0.13 mmol), HATU (59 mg, 0.16 mmol) and DIEA (50 mg, 0.39 mmol) were added to a solution of (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (80 mg, 0.16 mmol) in DMF (5 mL). The mixture was stirred at room temperature for 2 h, water (50 mL) was added, a solid was precipitated, and the mixture was filtered. The solid was washed 3 times with water, dissolved in dichloromethane (20 mL), dried over anhydrous sodium sulfate and concentrated under reduced pressure to give a crude product of tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2-oxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquizolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1062.5.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-(3-((1-(4-chloro-3-(2-oxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (130 mg, 0.12 mmol) was dissolved in dichloromethane (5 mL), and trifluoroacetic acid (1 mL) was added. Then the reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(4-(3-(4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)propyl)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=962.5.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.10 (d, J=8.2 Hz, 1H), 8.06 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.57 (s, 1H), 7.47-7.34 (m, 1H), 6.99 (s, 1H), 6.88 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.64-5.46 (m, 3H), 4.00-3.82 (m, 4H), 3.80-3.67 (m, 1H), 3.66-3.57 (m, 1H), 3.56-3.39 (m, 8H), 3.22-3.10 (m, 1H), 3.00-2.89 (m, 1H), 2.80-2.55 (m, 3H), 2.40-2.25 (m, 9H), 1.94-1.35 (m, 18H).
1-(5-(4-((2-(Piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (120 mg, 0.24 mmol) and (1R,4R)-4-(4-(((R)-1-(1,1-dioxido-2,3-dihydrobenzo[b]thiophen-4-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (120 mg, 0.24 mmol) were dissolved with DMF (3 mL), and HATU (108 mg, 0.28 mmol) and DIEA (90 mg, 0.71 mmol) were added. The reaction liquid was stirred at room temperature for 1 h. The reaction liquid was diluted with water (30 mL) and extracted with ethyl acetate (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by prep-HPLC to give 1-(5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(1,1-dioxide-2,3-dihydrobenzo[b]thiophen-4-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)-2-(trifluoromethyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1002.4.
1H NMR (400 MHz, DMSO-d6) δ 10.53 (s, 1H), 8.27 (d, J=7.2 Hz, 1H), 8.06 (s, 1H), 7.87 (d, J=8.2 Hz, 1H), 7.80 (d, J=7.1 Hz, 1H), 7.70 (s, 1H), 7.59 (t, J=6.6 Hz, 2H), 7.53 (t, J=7.6 Hz, 1H), 6.98 (s, 1H), 5.58 (t, J=7.0 Hz, 1H), 4.57-4.33 (m, 2H), 3.99-3.84 (m, 5H), 3.81-3.70 (m, 1H), 3.68-3.60 (m, 2H), 3.52-3.40 (m, 5H), 3.24 (d, J=6.2 Hz, 2H), 3.14-2.89 (m, 3H), 2.85-2.65 (m, 4H), 2.32 (s, 3H), 1.92-1.40 (m, 20H), 1.23-0.89 (m, 4H).
1-(2-Chloro-5-(4-((2-(piperazin-1-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (150 mg, crude product) was dissolved in DMF (5 mL), and (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (94 mg, 0.22 mmol), HATU (107 mg, 0.28 mmol), and DIEA (145 mg, 1.1 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure.
The crude product was purified by preparative reversed-phase chromatography to give 1-(2-chloro-5-(4-((2-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1052.4.
1H NMR (400 MHz, DMSO-d6) δ 10.48 (s, 1H), 8.29 (d, J=8.3 Hz, 1H), 8.04 (s, 1H), 7.62 (d, J=8.1 Hz, 1H), 7.53 (d, J=1.6 Hz, 1H), 7.45-7.31 (m, 4H), 7.23 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 5.96 (t, J=7.1 Hz, 1H), 4.52-4.28 (m, 1H), 3.89 (s, 3H), 3.80-3.40 (m, 10H), 3.26 (d, J=6.3 Hz, 2H), 3.12 (s, 2H), 2.97-2.88 (m, 1H), 2.79-2.59 (m, 4H), 2.45-2.39 (m, 5H), 2.38-2.33 (m, 2H), 2.01 (s, 6H), 1.94-1.46 (m, 16H), 1.13 (d, J=10.9 Hz, 2H).
tert-Butyl 4-(3-((1-((benzyloxy)carbonyl)piperidin-4-yl)oxy)propyl)piperazine-1-carboxylate (400 mg, 0.87 mmol) and Pd/C (10%, 100 mg) were dissolved with ethyl acetate (10 mL). The reaction liquid was heated to 70° C., stirred for 3 h, and filtered, and the solid was washed three times with ethyl acetate (10 mL×3) and concentrated under reduced pressure to give tert-butyl 4-(3-(piperidin-4-yloxy)propyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=328.3.
tert-Butyl 4-(3-(piperidin-4-yloxy)propyl)piperazine-1-carboxylate (230 mg, 0.70 mmol) was dissolved with DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (360 mg, 0.82 mmol) and DIEA (271 mg, 2.10 mmol) were separately added. The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:10) to give tert-butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperazine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=578.3.
tert-Butyl 4-(3-((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)oxy)propyl)piperazine-1-carboxylate (220 mg, 0.47 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (1 mL) was added. The reaction liquid was stirred at room temperature overnight. A saturated aqueous sodium bicarbonate solution was slowly added dropwise to the reaction liquid to adjust the pH to 7-8. The resulting mixture was extracted with a mixed solution (25 mL×4) of dichloromethane and isopropanol, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give 1-(2-chloro-5-(4-(3-(piperazin-1-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=478.2.
1-(2-Chloro-5-(4-(3-(piperazin-1-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (90 mg, 0.19 mmol), HATU (87 mg, 0.23 mmol), and DIEA (74 mg, 0.57 mmol) were separately added to a solution of (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (106 mg, 0.19 mmol) in DMF (10 mL). The reaction liquid was stirred at room temperature for 2 h and diluted with water (100 mL), and the solid was precipitated, collected by filtration, washed 3 times with water, dissolved with dichloromethane, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(2-chloro-5-(4-(3-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)propoxy)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1052.4.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.32 (d, J=7.7 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.3 Hz, 1H), 7.57 (s, 1H), 7.47-7.30 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.97 (s, 1H), 5.97 (t, J=7.2 Hz, 1H), 4.00-3.84 (s, 4H), 3.80-3.71 (m, 1H), 3.66-3.57 (m, 1H), 3.49-3.35 (m, 8H), 3.20-3.07 (m, 3H), 3.00-2.84 (m, 1H), 2.78-2.58 (m, 3H), 2.42 (s, 3H), 2.40-2.26 (m, 6H), 2.01 (s, 6H), 1.92-1.75 (m, 6H), 1.75-1.38 (m, 12H).
The compound methyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-formylphenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (450 mg, 0.78 mmol) and methylamine (2 M in THF, 7.8 mmol) were dissolved in dichloroethane (10 mL), and sodium triacetoxyborohydride (496 mg, 2.34 mmol) was added. The reaction liquid was heated to 80° C. and stirred for 1 h. The reaction liquid was directly concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:10, flow rate: 40 mL/min) to give tert-butyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]=593.3.
tert-Butyl (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylate (300 mg, 0.5 mmol) and lithium hydroxide (60 mg, 2.5 mmol) were dissolved with a mixed solution (MeOH/H2O=10/1, 5 mL). The reaction liquid was heated to 70° C. and stirred for 2 h. The reaction liquid was cooled, adjusted to pH 3-4 with 1 M HCl, adjusted to pH 7-8 with a saturated aqueous sodium bicarbonate solution, and extracted with ethyl acetate (100 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reversed-phase chromatography (ACN:H2O=1:1) to give (1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=579.2.
(1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (80 mg, 0.14 mmol) was dissolved with DMF (5 mL), and HATU (65 mg, 0.17 mmol), 1-(2-chloro-5-(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (80 mg, 0.17 mmol), and DIEA (54 mg, 0.42 mmol) were added. The reaction liquid was stirred at room temperature for 1 h, diluted with water (20 mL), and extracted with ethyl acetate (20 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-((2-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)ethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1037.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.32 (d, J=8.3 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.7 Hz, 1H), 7.45 (d, J=7.5 Hz, 1H), 7.42-7.30 (m, 3H), 7.28 (s, 1H), 7.02 (s, 1H), 6.98 (s, 1H), 6.07-5.86 (m, 1H), 4.38 (d, J=12.6 Hz, 2H), 3.89 (s, 4H), 3.73 (d, J=7.1 Hz, 1H), 3.66-3.51 (m, 2H), 3.44-3.36 (m, 4H), 3.23 (d, J=6.2 Hz, 2H), 3.11-2.88 (m, 3H), 2.78-2.59 (m, 4H), 2.42 (s, 3H), 2.12 (s, 3H), 1.92-1.39 (m, 21H), 1.18-0.87 (m, 4H).
(1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (80 mg, 0.13 mmol), HATU (62 mg, 0.16 mmol) and DIEA (50 mg, 0.39 mmol) were added to a solution of tert-butyl 4-(2-(piperidin-4-ylmethoxy)ethyl)piperidine-1-carboxylate (43 mg, 0.13 mmol) in DMF (5 mL). The mixture was stirred at room temperature for 1 h and diluted with water (50 mL), a solid was precipitated, and the mixture was filtered. The solid was washed 3 times with water, dissolved in dichloromethane, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give tert-butyl 4-(2-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=901.4.
tert-Butyl 4-(2-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carboxylate (80 mg, 0.09 mmol) was added to a solution of hydrogen chloride in dioxane (4 M, 2 mL), and the reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give a crude product of ((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidin-1-yl)methanone.
LC-MS: (ESI, m/z): [M+H]+=801.3.
((1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-((2-(piperidin-4-yl)ethoxy)methyl)piperidin-1-yl)methanone (70 mg, 0.09 mmol) was added to a solution of pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (39 mg, 0.08 mmol) and DIEA (35 mg, 0.27 mmol) in DMSO (5 mL). The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), extracted with ethyl acetate (25 mL×4), washed with saturated brine, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(2-chloro-5-(4-(2-((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)ethyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1051.5.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.30 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.62 (d, J=8.2 Hz, 1H), 7.53 (d, J=2.0 Hz, 1H), 7.45-7.31 (m, 4H), 7.24 (d, J=1.3 Hz, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.03-5.89 (m, 1H), 4.48-4.32 (m, 2H), 3.97-3.85 (m, 4H), 3.79-3.68 (m, 1H), 3.66-3.52 (m, 2H), 3.40 (t, J=6.4 Hz, 2H), 3.21 (d, J=6.1 Hz, 2H), 3.16-2.90 (m, 5H), 2.82-2.52 (m, 7H), 2.42 (s, 3H), 2.01 (s, 6H), 1.88-1.43 (m, 20H), 1.19-0.90 (m, 4H).
(1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (120 mg, 0.20 mmol), tert-butyl 4-(4-(piperazin-1-yl)butyl)piperidine-1-carboxylate (66 mg, 0.20 mmol), and HATU (91 mg, 0.24 mmol) were dissolved in DMF (3 mL), and DIEA (78 mg, 0.61 mmol) was added. The reaction liquid was stirred at room temperature for 1 h, diluted with water (30 mL), and extracted with ethyl acetate (30 mL×3). The organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography (DCM:MeOH=10:1) to give tert-butyl 4-(4-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)butyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=900.6.
tert-Butyl 4-(4-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)butyl)piperidine-1-carboxylate (149 mg, 0.17 mmol) was dissolved with a solution of hydrogen chloride in 1,4-dioxane (4 M, 5 mL). The reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give ((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(4-(piperidin-4-yl)butyl)piperazin-1-yl)methanone.
LC-MS: (ESI, m/z): [M+H]+=800.4.
((1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-(4-(piperidin-4-yl)butyl)piperazin-1-yl)methanone (158 mg, 0.11 mmol) and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (47 mg, 0.11 mmol) were dissolved in DMSO (2 mL), and DIEA (42 mg, 0.32 mmol) was added while stirring. The reaction liquid was stirred at room temperature for 1 h. The reaction liquid was diluted with water (30 mL) and extracted with ethyl acetate (30 mL×3). The upper organic phases were combined, washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-(4-(4-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)butyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1050.5.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.30 (d, J=7.7 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.3 Hz, 1H), 7.54 (s, 1H), 7.47-7.31 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.01-5.90 (m, 1H), 4.50-4.42 (m, 1H), 3.89 (s, 3H), 3.80-3.68 (m, 1H), 3.66-3.52 (m, 2H), 3.50-3.40 (m, 4H), 3.18-3.10 (m, 2H), 3.02-2.90 (m, 2H), 2.80-2.65 (m, 4H), 2.42 (s, 3H), 2.38-2.20 (m, 6H), 2.01 (s, 6H), 1.82-1.42 (m, 18H), 1.30-1.22 (m, 4H), 1.13-0.96 (m, 2H).
6-Bromo-2,7-dimethylquinazolin-4-ol (900 mg, 3.56 mmol) was dissolved with DMF/H2O (22 mL, 10/1), and methyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxopentan-2-yl)cyclohex-3-ene-1-carboxylate (1135 mg, 4.27 mmol), sodium carbonate (755 mg, 7.12 mmol), and Pd(dppf)Cl2 (264 mg, 0.36 mmol) were added. The reaction liquid was stirred at 110° C. overnight under nitrogen atmosphere. Water (100 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (100 mL×3). The organic phase was washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9) to give methyl 4-(4-hydroxy-2,7-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=313.1.
Methyl 4-(4-hydroxy-2,7-methylquinazolin-6-yl)cyclohex-3-ene-1-carboxylate (380 mg, 1.22 mmol) was added to an autoclave (100 mL) and dissolved with MeOH (30 mL), and Pd(OH)2/C (115 mg, 30%) was added. The reaction liquid was reacted at 70° C. for 48 h under hydrogen atmosphere (2 MPa), filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give methyl (1R,4R)-4-(4-hydroxy-2,7-dimethylquinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=315.2.
1H NMR (400 MHz, DMSO) δ 12.09 (s, 1H), 8.15 (s, 1H), 7.56 (s, 1H), 3.92 (s, 3H), 2.95 (t, J=11.6 Hz, 1H), 2.43 (s, 3H), 2.36 (s, 3H), 2.35-2.24 (m, 1H), 2.06 (d, J=10.7 Hz, 2H), 1.90-1.84 (m, 2H), 1.67-1.40 (m, 4H).
Methyl (1R,4R)-4-(4-hydroxy-2,7-dimethylquinazolin-6-yl)cyclohexane-1-carboxylate (250 mg, 0.80 mmol) was dissolved in DMF (5 mL), and BOP (506 mg, 1.19 mmol) and DBU (303 mg, 1.99 mmol) were added. The reaction liquid was stirred at room temperature for 0.5 h. (R)-1-(3-Nitro-5-(trifluoromethyl)phenyl)ethan-1-amine (224 mg, 0.96 mmol) was added, and the resulting mixture was heated to 70° C. and stirred overnight. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reversed-phase chromatography to give methyl (1R,4R)-4-(2,7-dimethyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=531.3.
Methyl (1R,4R)-4-(2,7-dimethyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylate (210 mg, 0.40 mmol) was dissolved in THF/H2O (15 mL, 2:1), and lithium hydroxide (190 mg, 8.00 mmol) was added. The reaction liquid was stirred at 50° C. overnight. Water (20 mL) was added, and 1 M hydrochloric acid was added to adjust the pH to 6. The resulting mixture was extracted with MeOH/DCM (1/9, 30 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give (1R,4R)-4-(2,7-dimethyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylic acid, which was directly used in the next step.
LC-MS: (ESI, m/z): [M+H]+=517.4.
(1R,4R)-4-(2,7-Dimethyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carboxylic acid (150 mg, 0.29 mmol) was dissolved in ethanol (10 mL), and Raney-Ni (0.5 mL) and N2H4-H2O (85%, 0.5 mL) were added. The reaction liquid was reacted at room temperature for 0.5 h, filtered, and concentrated under reduced pressure to give (1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)cyclohexane-1-carboxylic acid.
LC-MS: (ESI, m/z): [M+H]+=487.2.
(1R,4R)-4-(4-(((R)-1-(3-Amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)cyclohexane-1-carboxylic acid (70 mg, 0.14 mmol) was dissolved in DMF (5 mL), and 1-(2-chloro-5-(9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (84 mg, 0.17 mmol), DIEA (55 mg, 0.42 mmol), and pyBOP (88 mg, 0.17 mmol) were added. The reaction liquid was reacted at room temperature for 1 h. Water (50 mL) was added, and the resulting mixture was stirred for 5 min and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2,7-dimethylquinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=970.4.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.19 (d, J=8.0 Hz, 1H), 8.08 (s, 1H), 7.64 (d, J=8.2 Hz, 1H), 7.55 (d, J=1.9 Hz, 1H), 7.45-7.31 (m, 2H), 6.87 (d, J=11.3 Hz, 2H), 6.70 (s, 1H), 5.67-5.45 (m, 3H), 3.82-3.70 (m, 1H), 3.67-3.42 (m, 8H), 2.87-2.64 (m, 5H), 2.43 (s, 3H), 2.39-2.22 (m, 7H), 2.18-2.09 (m, 2H), 1.90-1.78 (m, 4H), 1.73-1.24 (m, 14H), 1.18-0.94 (m, 4H).
Pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoate (118 mg, 0.27 mmol) and (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylic acid (300 mg, 0.34 mmol) were dissolved in DMSO (6 mL), and DIEA (132 mg, 1.02 mmol) was added. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was diluted with water (50 mL) and extracted with EA (50 mL×3). The organic phases were combined, washed with brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH(NH3)/DCM, 0%-20%) to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M/2+H]+=563.5.
TFA (1 mL) was added to a solution of tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-methoxybenzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylate (72 mg, 0.064 mmol) in DCM (3 mL) at room temperature while stirring. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-methoxy-5-(9-((4-((1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1024.4.
1H NMR (400 MHz, CD3OD) δ 7.99 (s, 1H), 7.46-7.36 (m, 3H), 7.34-7.27 (m, 3H), 7.22-7.12 (m, 3H), 7.01 (s, 1H), 6.10-6.02 (m, 1H), 3.95-3.91 (m, 6H), 3.75-3.45 (m, 12H), 3.12-3.01 (m, 1H), 2.80 (t, J=6.7 Hz, 2H), 2.75-2.66 (m, 1H), 2.52 (s, 3H), 2.44 (m, 2H), 2.38 (m, 2H), 2.25-2.16 (m, 5H), 2.00-1.84 (m, 4H), 1.79 (d, J=7.0 Hz, 4H), 1.71-1.54 (m, 9H), 1.37-1.12 (m, 7H).
Pentafluorophenyl 3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoate (133 mg, 0.32 mmol) and tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylate (350 mg, 0.39 mmol) were dissolved in DMSO (6 mL), and then DIEA (132 mg, 1.02 mmol) was added. The reaction liquid was stirred at room temperature for 2 h. After the reaction was completed, the reaction liquid was diluted with water (50 mL) and extracted with EA (50 mL×3). The organic phases were combined, washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH(NH3)/DCM, 0%-20%) to give tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylate.
LC-MS: (ESI, m/z): [M/2+H]+=556.9.
TFA (1 mL) was added to a solution of tert-butyl (2-(5-((R)-1-((6-((1R,4R)-4-(4-((3-(3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)-4-fluorobenzoyl)-3-azaspiro[5.5]undecan-9-yl)methyl)piperazine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)thien-3-yl)benzyl)(methyl)carboxylate (100 mg, 0.09 mmol) in DCM (3 mL) at room temperature. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was directly concentrated. The crude product was purified by prep-HPLC to give 1-(2-fluoro-5-(9-((4-((1R,4R)-4-(7-methoxy-2-methyl-4-(((R)-1-(4-(2-((methylamino)methyl)phenyl)thien-2-yl)ethyl)amino)quinazolin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1012.3.
1H NMR (400 MHz, CD3OD) δ 7.99 (s, 1H), 7.52 (dd, J=7.1, 1.9 Hz, 1H), 7.46-7.39 (m, 2H), 7.36-7.27 (m, 4H), 7.21-7.17 (m, 1H), 7.15-7.11 (m, 1H), 7.01 (s, 1H), 6.08-6.03 (m, 1H), 3.95 (s, 3H), 3.83 (t, J=6.7 Hz, 2H), 3.77-3.55 (m, 8H), 3.50-3.37 (m, 2H), 3.10-3.00 (m, 1H), 2.83 (t, J=6.7 Hz, 2H), 2.76-2.65 (m, 1H), 2.52 (s, 3H), 2.44 (b, 2H), 2.38 (b, 2H), 2.25-2.16 (m, 5H), 2.00-1.83 (m, 4H), 1.82-1.75 (m, 4H), 1.69-1.46 (m, 9H), 1.36-1.06 (m, 7H).
1-(tert-Butyl)-4-ethylpiperidine-1,4-dicarboxylate (500 mg, 1.93 mmol) was dissolved in ethanol (20 mL), and NaBD4 (325 mg, 7.75 mmol) was added at 0° C. The reaction liquid was stirred for 2 h. Water (30 mL) was added to the reaction liquid, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:1) to give tert-butyl 4-(hydroxymethyl-d2)piperidine-1-carboxylate.
1H NMR (400 MHz, DMSO-d6) δ 4.42 (s, 1H), 3.93 (d, J=11.9 Hz, 2H), 2.67 (s, 2H), 1.62 (d, J=12.9 Hz, 2H), 1.55-1.44 (m, 1H), 1.39 (s, 9H), 0.96 (m, 2H).
tert-Butyl 4-(hydroxymethyl-d2)piperidine-1-carboxylate (250 mg, 1.14 mmol) was dissolved in anhydrous tetrahydrofuran (10 mL). The reaction liquid was cooled to 0° C. under nitrogen atmosphere. NaH (364 mg, 9.12 mmol) was added, and the resulting mixture was stirred for 3 h. 4-(Bromomethyl)pyridine hydrobromide (576 mg, 2.28 mmol) was added, and the resulting mixture was stirred at room temperature overnight. Water (20 mL) was added to the reaction liquid, and the resulting mixture was extracted with ethyl acetate (20 mL×3). The organic phase was washed with saturated brine (60 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 4-((pyridin-4-ylmethoxy)methyl-d2)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=309.1
1H NMR (400 MHz, DMSO-d6) δ 8.53 (dd, J=4.5, 1.5 Hz, 2H), 7.31 (d, J=5.8 Hz, 2H), 4.51 (s, 2H), 3.94 (d, J=12.0 Hz, 2H), 2.70 (s, 2H), 1.76 (m, 1H), 1.67 (d, J=12.9 Hz, 2H), 1.39 (s, 9H), 1.06 (m, 2H).
tert-Butyl 4-((pyridin-4-ylmethoxy)methyl-d2)piperidine-1-carboxylate (180 mg, 0.58 mmol) was dissolved in isopropanol (10 mL) and water (10 mL), and Pd/C (60 mg, 0.058 mmol) was added. The reaction liquid was heated to 70° C. under hydrogen atmosphere and stirred overnight. The reaction liquid was cooled to room temperature and filtered, and the filtrate was concentrated under reduced pressure to give tert-butyl 4-((piperidine-4-ylmethoxyl)methyl-d2)piperidine-1-carboxylate. The crude product was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=315.2.
tert-Butyl 4-((piperidin-4-ylmethoxy)methyl-d2)piperidine-1-carboxylate (130 mg, 0.41 mmol) was dissolved in DMF (10 mL), and (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (150 mg, 0.25 mmol), HATU (123 mg, 0.32 mmol), and DIEA (96 mg, 0.74 mmol) were added. The reaction liquid was stirred at room temperature for 1 h. The reaction liquid was diluted with water (100 mL) and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (50 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 4-(((1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=899.5.
tert-Butyl 4-(((1-((1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carboxylate (200 mg, 0.22 mmol) was dissolved in DCM (5 mL), and TFA (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h. The reaction liquid was directly concentrated under reduced pressure to give ((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-((piperidin-4-ylmethoxy-d2)methyl)piperidin-1-yl)methanone.
LC-MS: (ESI, m/z): [M+H]+=699.4
((1R,4R)-4-(4-(((R)-1-(3-Amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexyl)(4-((piperidin-4-ylmethoxy-d2)methyl)piperidin-1-yl)methanone (crude product, 200 mg, 0.22 mmol) was dissolved in DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (95 mg, 0.22 mmol) and DIEA (86 mg, 0.66 mmol) were added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added to the reaction liquid, and the resulting mixture was extracted with dichloromethane (30 mL×3). The organic phase was concentrated under reduced pressure to give the crude product.
The crude product was purified by pre-HPLC (CH3CN/0.08% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-(4-(((1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=949.4.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 8.15-8.02 (m, 2H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.8 Hz, 1H), 7.38 (dd, J=8.2, 1.9 Hz, 1H), 6.99 (s, 1H), 6.87 (s, 1H), 6.85 (s, 1H), 6.70 (s, 1H), 5.63-5.48 (m, 3H), 4.51-4.36 (m, 2H), 3.99-3.86 (m, 4H), 3.78-3.71 (m, 1H), 3.65-3.53 (m, 2H), 3.23 (d, J=6.1 Hz, 2H), 3.09-2.90 (m, 3H), 2.81-2.62 (m, 4H), 2.35 (s, 3H), 1.91-1.52 (m, 18H), 1.18-0.92 (m, 4H).
tert-Butyl 4-((piperidin-4-ylmethoxy)methyl-d2)piperidine-1-carboxylate (180 mg, 0.57 mmol) was dissolved with dimethylsulfoxide (5 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (247 mg, 0.57 mmol) and N,N-diisopropylethylamine (220 mg, 1.71 mmol) were added. The reaction liquid was stirred at room temperature for 2 h, diluted with water (50 mL). The organic phase was extracted with dichloromethane (30 mL×3), washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl 4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=565.2.
tert-Butyl 4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carboxylate (220 mg, 0.39 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h and directly concentrated under reduced pressure to give 1-(2-chloro-5-(4-((piperidin-4-ylmethoxy-d2)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione, which was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=465.3.
1-(2-Chloro-5-(4-((piperidin-4-ylmethoxy-d2)methyl)piperidine-1-carbonyl) phenyl)dihydropyrimidine-2,4(1H,3H)-dione (220 mg, 0.39 mmol) was dissolved with DMF (10 mL), and (1R,4R)-4-(4-(((R)-1-(3-((tert-butoxycarbonyl)amino)-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (150 mg, 0.24 mmol), HATU (118 mg, 0.31 mmol), and N,N-diisopropylethylamine (93 mg, 0.72 mmol) were separately added. The reaction liquid was stirred at room temperature for 2 h. Water (100 mL) was added, and the resulting mixture was extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (100 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (methanol:dichloromethane=1:9) to give tert-butyl (3-((R)-1-((6-((1R,4R)-4-(4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate.
LC-MS: (ESI, m/z): [M+H]+=1049.3.
tert-Butyl (3-((R)-1-((6-((1R,4R)-4-(4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl-d2)piperidine-1-carbonyl)cyclohexyl)-7-methoxy-2-methylquinazolin-4-yl)amino)ethyl)-5-(trifluoromethyl)phenyl)carboxylate (200 mg, 0.19 mmol) was dissolved with dichloromethane (5 mL), and trifluoroacetic acid (2 mL) was added. The reaction liquid was stirred at room temperature for 2 h and concentrated under reduced pressure. The resulting crude product was dissolved with a mixed solution (20 mL) of 10% methanol/dichloromethane. Then 5% aqueous sodium bicarbonate solution (20 mL) was added, and the resulting mixture was stirred for 1 h and left to stand for separation. The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to give the crude product, which was purified by pre-HPLC (CH3CN/0.08% aqueous NH4HCO3 solution, 5%-95%) to give 1-(5-(4-(((1-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy-d2)methyl)piperidine-1-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=949.2.
1H NMR (400 MHz, DMSO-d6) δ 10.49 (s, 1H), 8.13 (d, J=7.7 Hz, 1H), 8.06 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=2.0 Hz, 1H), 7.37 (dd, J=8.2, 2.0 Hz, 1H), 6.99 (s, 1H), 6.86 (d, J=10.1 Hz, 2H), 6.70 (s, 1H), 5.57 (dd, J=17.7, 10.2 Hz, 3H), 4.42 (d, J=12.0 Hz, 2H), 3.92 (m, 4H), 3.81-3.51 (m, 3H), 3.24 (d, J=6.2 Hz, 2H), 3.09-2.88 (m, 3H), 2.78-2.60 (m, 4H), 2.36 (s, 3H), 1.90-1.52 (m, 18H), 1.19-0.92 (m, 4H).
2-Amino-5-bromobenzamide (4.0 g, 14.55 mmol) was added to a solution of trimethyl orthoacetate (17.46 g, 145.5 mmol) in methanol (100 mL). The reaction liquid was stirred at 120° C. overnight in a sealed container, cooled, and concentrated under reduced pressure to give the crude product 6-bromo-2-methylquinazolin-4-ol, which was directly used in the next step without purification.
LC-MS: (ESI, m/z): [M+H]+=238.9.
6-Bromo-2-methylquinazolin-4-ol (5.00 g, 16.67 mmol) was dissolved with DMF/H2O (200 mL/20 mL), and potassium carbonate (5.80 g, 42.03 mmol), Pd(dppf)Cl2 (1.54 g, 2.10 mmol), and tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,6-dihydropyridine-1(2H)-carboxylate (7.79 g, 25.21 mmol) were separately added. The reaction liquid was stirred at 110° C. overnight under nitrogen atmosphere, diluted with water (500 mL), and extracted with ethyl acetate (200 mL×3). The organic phase was washed with saturated brine (200 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=1:2) to give tert-butyl 4-(4-hydroxy-2-methylquinazolin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=342.1.
tert-Butyl 4-(4-hydroxy-2-methylquinazolin-6-yl)-3,6-dihydropyridine-1(2H)-carboxylate (2.5 g, 14.55 mmol) was added to a solution of Pd(OH)2 (800 mg, 20%) in methanol (100 mL). The reaction liquid was stirred at 70° C. overnight in an autoclave under H2 atmosphere (10 atm), cooled, filtered, and concentrated under reduced pressure to give the crude product tert-butyl 4-(4-hydroxy-2-methylquinazolin-6-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=344.1.
BOP (3.30 g, 7.47 mmol), DBU (1.90 g, 12.5 mmol), and (R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethan-1-amine (1.53 g, 6.54 mmol) were separately added to a solution of tert-butyl 4-(4-hydroxy-2-methylquinazolin-6-yl)piperidine-1-carboxylate (1.73 g, 5.03 mmol) in DMF (100 mL). The reaction liquid was stirred at 70° C. overnight, diluted with water (300 mL), and extracted with ethyl acetate (100 mL). The organic phase was washed with saturated brine (150 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by reversed-phase column chromatography (H2O:ACN=1:2) to give tert-butyl (R)-4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=560.4.
tert-Butyl (R)-4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidine-1-carboxylate (300 mg, 0.537 mmol) was added to TFA (2 mL) in dichloromethane (20 mL). The reaction liquid was stirred at room temperature for 1 h and concentrated under reduced pressure to give the crude product (R)-2-methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-yl)quinazolin-4-amine.
LC-MS: (ESI, m/z): [M+H]+=460.4.
(R)-2-Methyl-N-(1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)-6-(piperidin-4-yl)quinazolin-4-amine (250 mg, 0.543 mmol) and potassium carbonate (300 mg, 2.17 mmol) were added to a solution of tert-butyl 2-bromoacetate (138 mg, 0.708 mmol) in THE (20 mL). The reaction liquid was stirred at 70° C. overnight, diluted with water (50 mL), extracted with ethyl acetate (50 mL×3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=5:1) to give tert-butyl (R)-2-(4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetate.
LC-MS: (ESI, m/z): [M+H]+=574.3.
tert-Butyl (R)-2-(4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetate (300 mg, 0.524 mmol) was added to HCl/1,4-dioxane (4 M, 20 mL), the mixture was stirred overnight at room temperature, and the reaction liquid was directly concentrated under reduced pressure to give a crude product of (R)-2-(4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetic acid.
LC-MS: (ESI, m/z): [M+H]+=518.1.
(R)-2-(4-(2-Methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetic acid (167 mg, 0.323 mmol), HATU (154 mg, 0.405 mmol), and DIEA (105 mg, 0.814 mmol) were added to a solution of tert-butyl 9-(2-aminoethyl)-3-azaspiro[5.5]undecane-3-carboxylate (80 mg, 0.27 mmol) in DMF (20 mL). The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), and extracted with ethyl acetate (50 mL×3). The crude product was purified by reversed-phase column chromatography (ACN:H2O=1:3) to give tert-butyl (R)-9-(2-(2-(4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetamido)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=796.4.
tert-Butyl (R)-9-(2-(2-(4-(2-methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)quinazolin-6-yl)piperidin-1-yl)acetamido)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate (70 mg, 0.088 mmol) was dissolved with ethanol (10 mL), and Raney-Ni (0.3 mL, water suspension) and hydrazine hydrate monohydrate (0.2 mL) were separately added. The reaction liquid was stirred at room temperature for 1 h, filtered, and concentrated under reduced pressure to give the crude product tert-butyl (R)-9-(2-(2-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylquinazolin-6-yl)piperidin-1-yl)acetamido)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=766.6.
tert-Butyl (R)-9-(2-(2-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylquinazolin-6-yl)piperidin-1-yl)acetamido)ethyl)-3-azaspiro[5.5]undecane-3-carboxylate (65 mg, 0.085 mmol) was dissolved with DCM (10 mL), and TFA (1 mL) was added. The reaction liquid was stirred at room temperature for 1 h and directly concentrated under reduced pressure to give the crude product (R)—N-(2-(3-azaspiro[5.5]undecane-9-yl)ethyl)-2-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylquinazolin-6-yl)piperidin-1-yl)acetamide.
LC-MS: (ESI, m/z): [M+H]+=666.4.
(R)—N-(2-(3-Azaspiro[5.5]undecan-9-yl)ethyl)-2-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylquinazolin-6-yl)piperidin-1-yl)acetamide (50 mg, 0.075 mmol) was dissolved with DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (40 mg, 0.092 mmol) and DIEA (30 mg, 0.232 mmol) were separately added. The reaction liquid was stirred at room temperature overnight, diluted with water (20 mL), and extracted with dichloromethane (30 mL×3). The organic phase was concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give (R)-2-(4-(4-((1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylquinazolin-6-yl)piperidin-1-yl)-N-(2-(3-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)-3-azaspiro[5.5]undecan-9-yl)ethyl)acetamide.
LC-MS: (ESI, m/z): [M+H]+=916.5.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.27 (d, J=7.9 Hz, 1H), 8.21 (s, 1H), 7.70-7.58 (m, 3H), 7.52 (d, J=8.7 Hz, 2H), 7.37 (dd, J=8.2, 1.8 Hz, 1H), 6.90 (s, 1H), 6.86 (s, 1H), 6.70 (s, 1H), 5.64-5.43 (m, 3H), 3.79-3.69 (m, 1H), 3.59-3.45 (m, 3H), 3.28-3.20 (m, 2H), 3.17-3.07 (m, 2H), 2.98-2.86 (m, 4H), 2.77-2.70 (m, 2H), 2.69-2.56 (m, 1H), 2.38 (s, 3H), 2.26-2.14 (m, 2H), 1.92-1.77 (m, 4H), 1.74-1.63 (m, 2H), 1.56-1.20 (m, 12H), 1.14-0.94 (m, 4H).
(1R,4R)-4-(2-Methyl-4-((1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carboxylic acid (170 mg, 0.337 mmol), HATU (192 mg, 0.505 mmol), and DIEA (130 mg, 1.01 mmol) were added to a solution of benzyl 9-(piperazin-1-ylmethyl)-3-azaspiro[5.5]undecane-3-carboxylate in DMF (30 mL). The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), and extracted with dichloromethane (50 mL×3). The crude product was purified by reversed-phase column chromatography (ACN:H2O=1:2) to give benzyl 9-((4-((1R,4R)-4-(2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=871.6.
Benzyl 9-((4-((1R,4R)-4-(2-methyl-4-(((R)-1-(3-nitro-5-(trifluoromethyl)phenyl)ethyl)amino)pyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (240 mg, 0.276 mmol) was dissolved in ethanol (20 mL), and Raney-Ni (water suspension, 0.3 mL) and hydrazine hydrate monohydrate (85%, 0.5 mL) were added. The reaction liquid was stirred at room temperature for 1 h, filtered, and concentrated under reduced pressure to give the crude product benzyl 9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=841.6.
Benzyl 9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carboxylate (220 mg, 0.262 mmol) was dissolved in ethyl acetate (20 mL), and Pd(OH)2 (44 mg, 20%) was added. The reaction liquid was stirred overnight at 70° C. under H2 atmosphere, filtered and concentrated under reduced pressure to give a crude product of (4-((3-azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexyl)methanone.
LC-MS: (ESI, m/z): [M+H]+=707.4.
(4-((3-Azaspiro[5.5]undecan-9-yl)methyl)piperazin-1-yl)((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexyl)methanone (200 mg, 0.283 mmol) was dissolved with DMSO (20 mL), and perfluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (147 mg, 0.338 mmol), and DIEA (110 mg, 0.853 mmol) were separately added. The reaction liquid was stirred at room temperature overnight, diluted with water (50 mL), and extracted with dichloromethane (50 mL×3). The organic phase was concentrated under reduced pressure. The crude product was purified by preparative reversed-phase chromatography to give 1-(5-(9-((4-((1R,4R)-4-(4-(((R)-1-(3-amino-5-(trifluoromethyl)phenyl)ethyl)amino)-2-methylpyrido[3,4-d]pyrimidin-6-yl)cyclohexane-1-carbonyl)piperazin-1-yl)methyl)-3-azaspiro[5.5]undecane-3-carbonyl)-2-chlorophenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=957.4.
1H NMR (400 MHz, DMSO-d6) δ10.51 (s, 1H), 8.91 (s, 1H), 8.58 (d, J=7.9 Hz, 1H), 8.19 (s, 1H), 8.10 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.55 (d, J=1.9 Hz, 1H), 7.39 (dd, J=8.2, 1.9 Hz, 1H), 6.89 (s, 1H), 6.85 (s, 1H), 6.71 (s, 1H), 5.61-5.47 (m, 3H), 3.78-3.72 (m, 1H), 3.64-3.56 (m, 3H), 3.54-3.41 (m, 6H), 3.30-3.24 (m, 2H), 2.82-2.71 (m, 3H), 2.70-2.61 (m, 1H), 2.43 (s, 3H), 2.36-2.21 (m, 4H), 2.17-2.07 (m, 2H), 2.04-1.98 (m, 2H), 1.85-1.71 (m, 2H), 1.77-1.31 (m, 18H), 1.16-0.90 (m, 4H).
1,3-Di(piperidin-4-yl)propane (193.5 mg, 0.92 mmol) DIEA (178 mg, 1.38 mmol) were dissolved DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (200 mg, 0.46 mmol) was added. The reaction liquid was reacted at room temperature for 30 min. The reaction liquid was diluted with water (30 mL) and extracted with ethyl acetate (30 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (MeOH:DCM=1:9, flow rate: 40 mL/min) to give 1-(2-chloro-5-(4-(3-(piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=461.2.
(1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (100 mg, 0.17 mmol), 1-(2-chloro-5-(4-(3-(piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (80 mg, 0.17 mmol), and DIEA (65.8 mg, 0.51 mmol) were dissolved with DMF (5 mL), and HATU (77.5 mg, 0.20 mmol) was finally added. The reaction liquid was stirred at room temperature for 30 min, diluted with water (20 mL), and extracted with ethyl acetate (20 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-(3-(1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)propyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1035.5.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.31 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.6 Hz, 1H), 7.43-7.35 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.05-5.83 (m, 1H), 4.52-4.35 (m, 2H), 3.95-3.80 (m, 4H), 3.80-3.69 (m, 1H), 3.67-3.50 (m, 2H), 3.20-3.08 (m, 2H), 3.06-2.87 (m, 3H), 2.83-2.70 (m, 3H), 2.68-2.57 (m, 1H), 2.42 (s, 3H), 2.01 (s, 6H), 1.92-1.39 (m, 18H), 1.36-1.15 (m, 6H), 1.13-0.82 (m, 4H).
tert-Butyl 4-(hydroxymethyl)piperidine-1-carboxylate (1.0 g, 4.65 mmol) was dissolved in a THF (40 mL) solution, and NaH (744 mg, 18.6 mmol, 60%) was added at 0° C. The reaction liquid was stirred for 2 h. Then 4-(bromomethyl)pyridine (795 mg, 4.65 mmol) was added, and the reaction liquid was stirred at room temperature for 2 h, diluted with water (100 mL), extracted with ethyl acetate (100 mL×2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The crude product was purified by column chromatography (ethyl acetate:petroleum ether=1:0) to give tert-butyl 4-((pyridin-4-ylmethoxy)methyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+Na]+=329.1.
tert-Butyl 4-((pyridin-4-ylmethoxy)methyl)piperidine-1-carboxylate (300 mg, 0.98 mmol) was dissolved in a solution of i-PrOH/H2O (6:7, 26 mL), and Pd(OH)/C (100 mg, 20%) was added under hydrogen atmosphere. The reaction liquid was stirred at 75° C. overnight and filtered through celite, and the filtrate was directly concentrated under reduced pressure to give tert-butyl 4-((piperidin-4-ylmethoxy)methyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+H]+=313.2.
tert-Butyl 4-((piperidin-4-ylmethoxy)methyl)piperidine-1-carboxylate (120 mg, 0.38 mmol) and DIEA (147 mg, 1.14 mmol) were added to DMSO (10 mL), and pentafluorophenyl 4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoate (165 mg, 0.38 mmol) was added. The reaction liquid was stirred at room temperature for 30 min, diluted with water (40 mL), and extracted with ethyl acetate (40 mL×3). The organic phase was washed with saturated brine (40 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by column chromatography (PE:EA=10:1) to give tert-butyl 4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl)piperidine-1-carboxylate.
LC-MS: (ESI, m/z): [M+NH4]+=580.2.
tert-Butyl 4-(((1-(4-chloro-3-(2,4-dioxotetrahydropyrimidin-1(2H)-yl)benzoyl)piperidin-4-yl)methoxy)methyl)piperidine-1-carboxylate (80 mg, 0.14 mmol) was dissolved with dichloromethane (5 mL), and a solution of hydrogen chloride in dioxane (4 M, 2 mL) was added. The reaction liquid was stirred at room temperature for 3 h and directly concentrated under reduced pressure to give 1-(2-chloro-5-(4-((piperidin-4-ylmethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=463.1.
(1R,4R)-4-(4-(((R)-1-(4-(2-Chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carboxylic acid (65 mg, 0.11 mmol) was dissolved with DMF (5 mL), and HATU (50 mg, 0.13 mmol), DIEA (50 mg, 0.39 mmol), and 1-(2-chloro-5-(4-((piperidin-4-ylmethoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione (60 mg, 0.13 mmol) were separately added. The reaction liquid was stirred at room temperature for 30 min, diluted with water (20 mL), and extracted with ethyl acetate (20 mL×3). The organic phase was washed with saturated brine (20 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The crude product was purified by prep-HPLC to give 1-(2-chloro-5-(4-(((1-((1R,4R)-4-(4-(((R)-1-(4-(2-chloro-6-((dimethylamino)methyl)phenyl)thien-2-yl)ethyl)amino)-7-methoxy-2-methylquinazolin-6-yl)cyclohexane-1-carbonyl)piperidin-4-yl)methoxy)methyl)piperidine-1-carbonyl)phenyl)dihydropyrimidine-2,4(1H,3H)-dione.
LC-MS: (ESI, m/z): [M+H]+=1037.4.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 8.31 (d, J=8.1 Hz, 1H), 8.05 (s, 1H), 7.63 (d, J=8.2 Hz, 1H), 7.54 (d, J=1.8 Hz, 1H), 7.47-7.29 (m, 4H), 7.24 (s, 1H), 7.02 (s, 1H), 6.96 (s, 1H), 6.02-5.90 (m, 1H), 4.52-4.32 (m, 2H), 3.98-3.91 (m, 1H), 3.89 (s, 3H), 3.80-3.69 (m, 1H), 3.66-3.54 (m, 2H), 3.28-3.17 (m, 4H), 3.15-3.09 (m, 2H), 3.09-2.89 (m, 3H), 2.84-2.69 (m, 3H), 2.69-2.60 (m, 2H), 2.42 (s, 3H), 2.01 (s, 6H), 1.91-1.46 (m, 17H), 1.20-0.89 (m, 4H).
The following method was used to determine the extent of inhibition of KRAS-G12C/SOS1 binding by the preferred compounds of the present invention under in vitro conditions. In this method, the competitive inhibition of KRAS-G12C/SOS1 binding by the preferred compounds was assayed by the homogeneous time-resolved fluorescence (HTRF) technology using the KRAS-G12C/SOS1 binding assay kit (Cisbio, 63ADK000CB16PEG).
The experimental procedures were as follows (refer to the kit instructions for details): the compounds of the present invention were each firstly dissolved in DMSO at a concentration of 20 mM, and then diluted in an equal gradient with a buffer in the kit, so that the final concentrations of the test compounds in the reaction system were in a range of 10000 nM-0.04 nM, and the final concentration of DMSO was 0.5%. 2 μL of the compounds were each incubated with 4 μL of 1×Tag1-SOS1, 4 μL of 1×Tag2-KRAS-G12C (containing 10 μM guanosine triphosphate, GTP) at 25° C. for 15 min. Then 5 μL of 1×Anti-Tag1-Tb3+ and 5 μL of 1×Anti-Tag2-XL665 were added to the reaction system, and the mixture was incubated on ice for 3 h. After the incubation, the fluorescence intensity of each well was measured at the excitation wavelength of 337 nm, the fluorescence intensity of each well was read at the emission wavelengths of 620 nm and 665 nm on a microplate reader EnVision (PerkinElmer, 2105) in an HTRF mode, and the Ratio value was calculated by the formula Ratio=(665 nm/620 nm)×104. The inhibition rates of the compounds at each concentration were calculated by comparing the fluorescence intensity ratio with that of the control group, and then nonlinear curve fitting was performed by GraphPad Prism 8 in the logarithmic concentration-inhibition rate to obtain the IC50 values of the compounds, as shown in Table 1.
The degradation of SOS1 by the compounds was investigated on KRAS G12C mutant human non-small cell lung cancer cells NCI-H358 (ATCC or Stem Cell Bank, Chinese Academy of Sciences), KRAS G12D mutant human lung cancer cells A-427 (ATCC), KRAS G12V mutant human colorectal adenocarcinoma cells SW-620 (ATCC), and KRAS G13D mutant human colorectal adenocarcinoma epithelial cells DLD-1 (ATCC). The specific procedures were as follows:
0.95 mL of cells were seeded into each well of a 24-well cell culture plate to achieve a cell density of 5×105 cells/well; the cell plate was incubated in an incubator at 37° C. with 5% CO2 overnight. Then 50 μL of the compound solution after dilution was added to each of the wells with the corresponding cells, so that the final concentration of the compound was in a range of 0.03-3000 nM. The final concentration of DMSO was 0.25%.
After dosing, the cell plate was incubated in an incubator at 37° C. with 5% CO2 for 24 h. The cell culture medium in the 24-well plate was removed, and the plate was washed twice with 1×PBS. 180 μL of RIPA (strong) lysis buffer (Beyotime, P0013B) supplemented with 1 mM phenylmethylsulfonyl fluoride, a protease inhibitor mixture (Beyotime, P1008), and a protease phosphatase inhibitor mixture (Beyotime, P1045) was added to lyse adherent NCI-H358 cells at the bottom of the cell culture plate. After the plate was left to stand on ice for 30 min, the protein lysate in each well was transferred to a 1.5 mL centrifuge tube and centrifuged at 15000 g for 20 min at 4° C. The cell lysate supernatant after centrifugation was cryopreserved at −80° C. for testing and was used for determining the degradation level of the SOS1 protein by a WB method. The concentration of total protein in the cell lysate supernatant was determined using the BCA Protein Quantification Kit (Tiangen, PA115-02).
The concentration of total protein in the cell lysate was determined by BCA. The total protein was adjusted to 0.5 μg/μL with PBS and 5×SDS- PAGE protein loading buffer (Beyotime, P001L), incubated in a water bath at 100° C. for 15 min, incubated in an ice bath on ice for 5 min, centrifuged at 14000 g for 1 min at 4° C., and mixed well as a loading sample for WB. Protein electrophoresis was performed on a precast gel (KeyGEN, KGMG010W15) with a loading volume of 10 μL (total protein: 5 μg) using a Tris-MOPS-SDS running solution (Adamas, P1598253) at a constant voltage of 120 V for 55 min. After the electrophoresis, the protein on the gel strip was transferred to a PVDF membrane at a constant current of 250 mA for 65 min. After the membrane transfer, the membrane was incubated in 1× QuickBlock blocking buffer (Beyotime, P0235) for 30 min at room temperature. After the blocking, the PVDF membrane was incubated with an SOS1 primary antibody (Abcam, ab140621) at 4° C. overnight, washed with a TBST buffer (2.4 g of Tris, 8.8 g of NaCl, 1.5 mL of Tween 20, with pH adjusted to 7.4, with the volume brought to 1 L) for 30 min (10 min/time), incubated with a secondary antibody (Abcam, ab205718) at room temperature for 2 h, washed with a TBST buffer for 30 min (10 min/time), and finally incubated with Clarity Western ECL Substrate (BIO-RAD, 170-5061) for 5 min for luminescence and color development, and a chemiluminescence imaging system (Clinx, ChemiScope 6200 Touch) was used for color development and protein mapping. The protein map was subjected to gray value analysis by the chemiluminescence imaging system from Clinx. The gray correction value for each sample was calculated by the formula: gray correction value=(gray value of target protein/gray value of corresponding internal reference)×103. Then the degradation rate was calculated by comparing the gray correction value with the gray correction value of the control group. Then nonlinear curve fitting was performed by GraphPad Prism 8 in the logarithmic concentration-inhibition rate to obtain the DC50 and Dmax values of the compounds.
The inhibitory effect of the compounds on the proliferation of 3D cells was investigated on KRAS G12C mutant human non-small cell lung cancer cells NCI-H358 (ATCC or Stem Cell Bank, Chinese Academy of Sciences), KRAS G12D mutant human metastatic pancreatic adenocarcinoma cells AsPC-1 (ATCC), KRAS G12V mutant human lung adenocarcinoma cells NCI-H441 (ATCC), and KRAS G13D mutant human colorectal adenocarcinoma epithelial cells DLD-1 (ATCC). The specific procedures were as follows:
200 nL of compound solution diluted in a gradient was added to corresponding microwells of a 384-well low-adsorption microwell plate (Corning, 3657) using an Echo sonic pipetting system (Labcyte, Echo 550), so that the final concentration of the test compound in the reaction system was in a range of 20000 nM-0.008 nM. Then 40 μL of cells at a certain density were added to the corresponding microplate. In addition to test wells for the test compounds, both a DMSO control well and a medium control well were set, with the DMSO control well containing DMSO and cells and the medium well containing only medium. After sample loading, the plate cover was applied, and the 384-well low-adsorption microplate was incubated in an incubator at 37° C. with 5% carbon dioxide for 7 days. After 7 days, the microplate was taken out, and 40 μL of CTG reagent (Promega, G9683) was added to each well. The plate was incubated at room temperature for 30 min and read on a microplate reader EnVision using the chemiluminescence program. The inhibition rates of the compounds at each concentration were calculated by the formula for the percentage inhibition of cell proliferation: inhibition rate %=(mean value of DMSO control group−single concentration reading value of compound)/(mean value of DMSO control group−mean value of medium control group)×100, and then nonlinear curve fitting was performed by GraphPad Prism 8 in the logarithmic concentration-inhibition rate to obtain the IC50 values of the compounds.
In Tables 1 and 2:
Experimental conclusion: the compounds of the present invention were able to effectively bind to the SOS1 target protein or have an inhibitory effect, and the compounds described herein were able to effectively and specifically degrade the SOS1 protein.
| Number | Date | Country | Kind |
|---|---|---|---|
| 202110443693.1 | Apr 2021 | CN | national |
| 202210201700.1 | Mar 2022 | CN | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/CN2022/088561 | 4/22/2022 | WO |
