Patent Application
20250019385
This application is filed as a U.S. Utility application and claims priority to EP Application No. EP23175994.5 filed May 30, 2023, and EP23175999.4 filed May 30, 2023, each of which are incorporated herein by reference in their entireties.
The present invention relates to annulated 2-amino-3-cyano thiophenes and derivatives of formula (I)
wherein R1a, R1b, R2a, R2b, Z, R3 to R5, A, B, p, q, V and W have the meanings given in the claims and specification, their use as inhibitors of KRAS, pharmaceutical compositions and preparations containing such compounds and their use as medicaments/medical uses, especially as agents for treatment and/or prevention of oncological diseases, e.g. cancer.
V-Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) is a small GTPase of the Ras family of proteins that exists in cells in either GTP-bound or GDP-bound states (McCormick et al., J. Mol. Med. (Berl)., 2016, 94(3):253-8; Nimnual et al., Sci. STKE., 2002, 2002(145):pe36). Binding of GTPase activating proteins (GAPs) such as NF1 increases the GTPase activity of Ras family proteins. The binding of guanine nucleotide exchange factors (GEFs) such as SOS1 (Son of Sevenless 1) promotes release of GDP from Ras family proteins, enabling GTP binding (Chardin et al., Science, 1993, 260(5112):1338-43). When in the GTP-bound state, Ras family proteins are active and engage effector proteins including C-RAF and phosphoinositide 3-kinase (PI3K) to promote the RAF/mitogen or extracellular signal-regulated kinases (MEK/ERK) pathway, PI3K/AKT/mammalian target of rapamycin (mTOR) pathway and RalGDS (Ral guanine nucleotide dissociation stimulator) pathway (McCormick et al., J. Mol. Med. (Berl)., 2016, 94(3):253-8; Rodriguez-Viciana et al., Cancer Cell. 2005, 7(3):205-6). These pathways affect diverse cellular processes such as proliferation, survival, metabolism, motility, angiogenesis, immunity and growth (Young et al., Adv. Cancer Res., 2009, 102:1-17; Rodriguez-Viciana et al., Cancer Cell. 2005, 7(3):205-6).
Cancer-associated mutations in Ras family proteins suppress their intrinsic and GAP-induced GTPase activity leading to an increased population of GTP-bound/active mutant Ras family proteins (McCormick et al., Expert Opin. Ther. Targets., 2015, 19(4):451-4; Hunter et al., Mol. Cancer Res., 2015, 13(9):1325-35). This in turn leads to persistent activation of effector pathways (e.g. RAF/MEK/ERK, PI3K/AKT/mTOR, RalGDS pathways) downstream of mutant Ras family proteins. KRAS mutations (e.g. amino acids G12, G13, Q61, A146) are found in a variety of human cancers including lung cancer, colorectal cancer and pancreatic cancer (Cox et al., Nat. Rev. Drug Discov., 2014, 13(11):828-51). Alterations (e.g. mutation, over-expression, gene amplification) in Ras family proteins/Ras genes have also been described as a resistance mechanism against cancer drugs such as the EGFR antibodies cetuximab and panitumumab (Leto et al., J. Mol. Med. (Berl). 2014 July; 92(7):709-22) and the EGFR tyrosine kinase inhibitor osimertinib/AZD9291 (Ortiz-Cuaran et al., Clin. Cancer Res., 2016, 22(19):4837-47; Eberlein et al., Cancer Res., 2015, 7 5(12):2489-500).
In a subset of tumor indications such as gastric cancer, gastroesophageal junction cancer and esophageal cancer prominent amplification of the wildtype (WT) KRAS proto-oncogene acts as a driver alteration and renders tumor models bearing this genotype addicted to KRAS in vitro and in vivo (Wong et al. Nat Med., 2018, 24(7):968-977). In contrast, non-amplified KRAS WT cell lines are KRAS independent, unless they carry secondary alterations in genes indirectly causing activation of KRAS (Meyers et al., Nat Genet., 2017, 49:1779-1784). Based on these data, a therapeutic window is expected for a KRAS targeting agent with a KRAS WT targeting activity.
Genetic alterations affecting e.g. codon 12 of KRAS substitute the glycine residue naturally occurring at this position for different amino acids such as aspartic acid (the G12D mutation or KRAS G12D), cysteine (the G12C mutation or KRAS G12C), valine (the G12V mutation or KRAS G12V) among others. Similarly, mutations within codons 13, 61 and 146 of KRAS are commonly found in the KRAS gene. Altogether KRAS mutations are detectable in 35% of lung, 45% of colorectal and up to 90% of pancreatic cancers (Herdeis et al., Curr Opin Struct Biol., 2021, 71:136-147).
In summary, binders/inhibitors of wildtype or mutated KRAS (e.g., G12D, G12V and G12C) are expected to deliver anti-cancer efficacy.
Thus, there is the need to develop new compounds efficacious in the treatment of cancers mediated by KRAS, especially KRAS mutated in position 12 or 13 and/or in wildtype amplified KRAS mediated cancer, which also possess desirable pharmacological properties, including but not limited to: metabolic stability, plasma protein binding, solubility, and permeability.
The present invention relates to annulated 2-amino-3-cyano thiophenes and derivatives of formula (I)
wherein R1a, R1b, R2a, R2b, Z, R3 to R5, A, B, p, q, V and W have the meanings given in the claims and specification, their use as inhibitors of KRAS, pharmaceutical compositions and preparations containing such compounds and their use as medicaments/medical uses, especially as agents for treatment and/or prevention of oncological diseases, e.g. cancer.
Surprisingly, the compounds described herein have been found to possess anti-tumour activity, being useful in inhibiting the uncontrolled cellular proliferation which arises from malignant diseases. It is believed that this anti-tumor activity is, inter alia, derived from inhibition of KRAS mutated in position 12 or 13, preferably G12D, G12V or G13D mutant KRAS, or inhibition of WT KRAS, especially KRAS WT amplified. Advantageously, the compounds can be selective for certain KRAS mutants, preferably KRAS G12V, or can be effective against a panel of KRAS mutants including KRAS wildtype amplified.
In addition, the compounds of the invention advantageously possess desirable pharmacological properties, including but not limited to metabolic stability, plasma protein binding, solubility and permeability.
It has now been found that, surprisingly, compounds of formula (I)
wherein
Surprisingly, the compounds described herein have been found to possess anti-tumour activity, being useful in inhibiting the uncontrolled cellular proliferation which arises from malignant diseases. It is believed that this anti-tumor activity is, inter alia, derived from inhibition of KRAS mutated in position 12 or 13, preferably G12D, G12V or G13D mutant KRAS, or inhibition of WT KRAS, especially KRAS WT amplified. Advantageously, the compounds can be selective for certain KRAS mutants, preferably KRAS G12V, or can be effective against a panel of KRAS mutants including KRAS wildtype amplified.
In addition, the compounds of the invention advantageously possess desirable pharmacological properties, including but not limited to metabolic stability, plasma protein binding, solubility, and permeability.
Thus, in a first aspect, the present invention relates to a compound of formula (I)
wherein
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein R1a and R1b are both independently selected from the group consisting of hydrogen and C1-4alkyl.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein R2a and R2b are both independently selected from the group consisting of hydrogen and halogen.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein R1a and R1b are both independently selected from the group consisting of hydrogen and methyl.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein R2a and R2b are both independently selected from the group consisting of hydrogen and fluorine.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein R1a, R1b, R2a and R2b are hydrogen.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein n is 0.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein n is 1; and each R6a and R6b is independently selected from the group consisting of hydrogen, C1-4alkyl, C1-4haloalkyl, C1-4alkoxy, C1-4haloalkoxy, halogen, —NH2, —NH(C1-4alkyl), —N(C1-4alkyl)2, C3-5cycloalkyl and 3-5 membered heterocyclyl.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein Z is —CH2—.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein n is 2;
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein Z is —S—.
In another aspect, the invention relates to the compound of the formula (I), or a salt thereof, wherein p is 0.
In another aspect, the present invention relates to a compound of the formula (I*) or a salt thereof
wherein
In another aspect the present invention relates to a compound of formula (Ia) or a salt thereof
wherein
In another aspect the present invention relates to a compound of formula (Ib) or a salt thereof
wherein
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is a ring selected from the group consisting of imidazole, pyrazole, oxazole, isoxazole, thiazole, isothiazole and triazole.
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is a ring selected from the group consisting of pyrrole, furan, thiophene, imidazole, pyrazole, isoxazole, isothiazole and triazole.
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is selected from the group consisting of
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is isoxazole or isothiazole.
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is selected from
In another aspect, the invention relates to the compound of the invention, or a salt thereof, wherein ring A is
In another aspect the invention relates to a compound of formula (Ic) or a salt thereof
wherein
In another aspect the invention relates to a compound of formula (Id) or a salt thereof
wherein
In another aspect the invention relates to a compound of formula (If) or a salt thereof
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic),
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein at least one of V and W is nitrogen.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein V and W are —CH═.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein R5 is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein R5 is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein R5 is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a C3-13alicycle.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a C3-13alicycle and q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a C3-13alicycle, q is 1 to 3 each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a C6-10arene.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a C6-10arene and q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a C6-10arene, q is 1 to 3 and R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-6 membered heteroarene and q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-6 membered heteroarene, q is 1 to 3 and R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene and q is 2.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene, q is 2 and R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene, q is 2 and R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-6 membered heteroarene, q is 2 and each R3 is R8;
In another aspect, the invention relates to the compound of the formula Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-6 membered heteroarene, q is 2 and each R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene and q is 1.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene, q is 1 and R3 is selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-6 membered heteroarene, q is 1 and R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-6 membered heteroarene, q is 1 and R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 3-13 membered heterocycle.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-7 membered heterocycle.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 3-13 membered heterocycle and q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 3-13 membered heterocycle, q is 1 to 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 1 to 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 2 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 2 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 1 and R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 1 and R3 is R7; and R7 is the bivalent substituent ═O.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 3-13 membered heterocycle, q is 1 and R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-7 membered heterocycle and q is 1 to 3.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-7 membered heterocycle, q is 1 to 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 1 to 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 3 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 2 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 2 and each R3 is independently selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 1 and R3 is selected from the group consisting of R7 and R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 1 and R3 is R7; and R7 is the bivalent substituent ═O.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-7 membered heterocycle, q is 1 and R3 is R8;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
wherein each of these groups is bound to ring B at any ring position by removal of a hydrogen atom and is optionally and independently substituted with one or more, identical or different R9 and/or R10;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
wherein each of these groups is bound to ring B at any ring position by removal of a hydrogen atom.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
wherein each of these groups is optionally and independently substituted with one or more, identical or different R9 and/or R10;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
wherein each of these groups is optionally and independently substituted with one or more, identical or different R9 and/or R10;
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
In another aspect, the invention relates to the compound of the formula (Ic), (Id), (Ie) or (If), or a salt thereof, wherein each R8 is independently selected from the group consisting of C1-6alkyl, C1-6alkoxy,
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B is a 5-13 membered heterocycle.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B has the substructure
wherein the substructure is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B has the substructure
wherein the substructure is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein R13 is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein Ring B has the substructure
wherein
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein R13 is selected from the group consisting of
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one oxygen.
In another aspect, the invention relates to the compound of the formula (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one oxygen.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing two oxygen.
In another aspect, the invention relates to the compound of the formula (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing two oxygen.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one nitrogen.
In another aspect, the invention relates to the compound of the formula (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one nitrogen.
In another aspect, the invention relates to the compound of the formula (I), (I*), (Ia), (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one oxygen and at least one nitrogen.
In another aspect, the invention relates to the compound of the formula (Ib), (Ic), (Id), (Ie) or (If), or a salt thereof, wherein ring B is a 5-11 membered heterocycle containing at least one oxygen and at least one nitrogen.
Preferred embodiments of the invention are example compounds I-1 to I-58 and compounds II-1 to II-20 and any subset thereof.
Preferred embodiments of the invention are example compounds I-1 to I-43 and compounds II-1 to II-11 and any subset thereof.
Preferred embodiments of the invention are example compounds I-44 to I-58 and compounds II-12 to II-20 and any subset thereof.
It is to be understood that any two or more aspects and/or preferred embodiments of formula (I)—or subformulas thereof—may be combined in any way leading to a chemically stable structure to obtain further aspects and/or preferred embodiments of formula (I)—or subformulas thereof.
The present invention further relates to hydrates, solvates, polymorphs, metabolites, derivatives, stereoisomers and prodrugs of a compound of formula (I), (including all embodiments thereof).
The present invention further relates to a hydrate of a compound of formula (I) (including all embodiments thereof).
The present invention further relates to a solvate of a compound of formula (I) (including all embodiments thereof).
Compounds of formula (I) (including all embodiments thereof) which e.g. bear ester groups are potential prodrugs the ester being cleaved under physiological conditions and are also part of the invention.
The present invention further relates to a pharmaceutically acceptable salt of a compound of formula (I) (including all embodiments thereof).
The present invention further relates to a pharmaceutically acceptable salt of a compound of formula (I) (including all embodiments thereof) with anorganic or organic acids or bases.
A further object of the invention is a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof—and one or more pharmaceutically acceptable excipient(s).
In one aspect, said pharmaceutical composition optionally comprises one or more other pharmacologically active substance(s). Said one or more other pharmacologically active substance(s) may be the pharmacologically active substances or combination partners as defined herein.
Suitable pharmaceutical compositions for administering the compounds of formula (I) according to the invention will be apparent to those with ordinary skill in the art and include for example tablets, pills, capsules, suppositories, lozenges, troches, solutions, suspensions—particularly solutions, suspensions or other mixtures for injection (s.c., i.v., i.m.) and infusion (injectables)—elixirs, syrups, sachets, emulsions, inhalatives or dispersible powders. The content of the compounds of formula (I) should be in the range from 0.1 to 90 wt.-%, preferably 0.5 to 50 wt.-% of the composition as a whole, i.e. in amounts which are sufficient to achieve the dosage range specified below. The doses specified may, if necessary, be given several times a day.
Suitable tablets may be obtained, for example, by mixing the compounds of formula (I) with known pharmaceutically acceptable excipients, for example inert diluents, carriers, disintegrants, adjuvants, surfactants, binders and/or lubricants. The tablets may also comprise several layers.
Coated tablets may be prepared accordingly by coating cores produced analogously to the tablets with excipients normally used for tablet coatings, for example collidone or shellac, gum arabic, talc, titanium dioxide or sugar. To achieve delayed release or prevent incompatibilities the core may also consist of a number of layers. Similarly, the tablet coating may consist of a number of layers to achieve delayed release, possibly using the excipients mentioned above for the tablets.
Syrups or elixirs containing one or more compounds of formula (I) or combinations with one or more other pharmaceutically active substance(s) may additionally contain excipients like a sweetener such as saccharine, cyclamate, glycerol or sugar and a flavour enhancer, e.g. a flavouring such as vanillin or orange extract. They may also contain excipients like suspension adjuvants or thickeners such as sodium carboxymethyl cellulose, wetting agents such as, for example, condensation products of fatty alcohols with ethylene oxide, or preservatives such as p-hydroxybenzoates.
Solutions for injection and infusion are prepared in the usual way, e.g. with the addition of excipients like isotonic agents, preservatives such as p-hydroxybenzoates, or stabilisers such as alkali metal salts of ethylenediamine tetra acetic acid, optionally using emulsifiers and/or dispersants, whilst if water is used as the diluent, for example, organic solvents may optionally be used as solvating agents or dissolving aids, and transferred into injection vials or ampoules or infusion bottles.
Capsules containing one or more compounds of formula (I) or combinations with one or more other pharmaceutically active substance(s) may for example be prepared by mixing the compounds/active substance(s) with inert excipients such as lactose or sorbitol and packing them into gelatine capsules.
Suitable suppositories may be made for example by mixing with excipients provided for this purpose such as neutral fats or polyethylene glycol or the derivatives thereof.
Excipients which may be used include, for example, water, pharmaceutically acceptable organic solvents such as paraffins (e.g. petroleum fractions), vegetable oils (e.g. groundnut or sesame oil), mono- or polyfunctional alcohols (e.g. ethanol or glycerol), carriers such as e.g. natural mineral powders (e.g. kaolins, clays, talc, chalk), synthetic mineral powders (e.g. highly dispersed silicic acid and silicates), sugars (e.g. cane sugar, lactose and glucose), emulsifiers (e.g. lignin, spent sulphite liquors, methylcellulose, starch and polyvinylpyrrolidone) and lubricants (e.g. magnesium stearate, talc, stearic acid and sodium lauryl sulfate).
The pharmaceutical compositions are administered by the usual methods, preferably by oral or transdermal route, most preferably by oral route. For oral administration the tablets may of course contain, apart from the above-mentioned excipients, additional excipients such as sodium citrate, calcium carbonate and dicalcium phosphate together with various excipients such as starch, preferably potato starch, gelatine and the like. Moreover, lubricants such as magnesium stearate, sodium lauryl sulfate and talc may be used at the same time for the tabletting process. In the case of aqueous suspensions, the active substances may be combined with various flavour enhancers or colourings in addition to the excipients mentioned above.
For parenteral use, solutions of the active substances with suitable liquid excipients may be used.
The dosage range of the compounds of formula (I) applicable per day is usually from 1 mg to 2000 mg, preferably from 250 to 1250 mg.
However, it may sometimes be necessary to depart from the amounts specified, depending on the body weight, age, the route of administration, severity of the disease, the individual response to the drug, the nature of its formulation and the time or interval over which the drug is administered (continuous or intermittent treatment with one or multiple doses per day). Thus, in some cases it may be sufficient to use less than the minimum dose given above, whereas in other cases the upper limit may have to be exceeded. When administering large amounts, it may be advisable to divide them up into a number of smaller doses spread over the day.
Thus, in a further aspect the invention relates to a pharmaceutical composition comprising at least one (preferably one) compound of formula (I) or a pharmaceutically acceptable salt thereof—and one or more pharmaceutically acceptable excipient(s).
The compounds of formula (I) or the pharmaceutically acceptable salts thereof—and the pharmaceutical compositions comprising such compound and salts may also be co-administered with other pharmacologically active substances, e.g. with other anti-neoplastic compounds (e.g. chemotherapy), i.e. used in combination (see combination treatment further below).
The elements of such combinations may be administered (whether dependently or independently) by methods customary to the skilled person and as they are used in monotherapy, e.g. by oral, enterical, parenteral (e.g., intramuscular, intraperitoneal, intravenous, transdermal or subcutaneous injection, or implant), nasal, vaginal, rectal, or topical routes of administration and may be formulated, alone or together, in suitable dosage unit formulations containing conventional non-toxic pharmaceutically acceptable excipients appropriate for each route of administration.
The combinations may be administered at therapeutically effective single or divided daily doses. The active components of the combinations may be administered in such doses which are therapeutically effective in monotherapy, or in such doses which are lower than the doses used in monotherapy, but when combined result in a desired (joint) therapeutically effective amount.
However, when the combined use of the two or more active substances or principles leads to a synergistic effect, it may also be possible to reduce the amount of one, more or all of the substances or principles to be administered, while still achieving the desired therapeutic action. This may for example be useful for avoiding, limiting or reducing any unwanted side-effects that are associated with the use of one or more of the substances or principles when they are used in their usual amounts, while still obtaining the desired pharmacological or therapeutic effect.
Thus, in a further aspect the invention also relates to a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof—and one or more (preferably one or two, most preferably one) other pharmacologically active substance(s).
In a further aspect the invention also relates to a pharmaceutical preparation comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof—and one or more (preferably one or two, most preferably one) other pharmacologically active substance(s).
Pharmaceutical compositions to be co-administered or used in combination can also be provided in the form of a kit.
Thus, in a further aspect the invention also relates to a kit comprising
In one aspect such kit comprises a third pharmaceutical composition or dosage form comprising still another pharmacologically active substance and, optionally, one or more pharmaceutically acceptable excipient(s).
The present invention is directed to compounds inhibiting KRAS, preferably KRAS mutated at residue 12, such as KRAS G12C, KRAS G12D, KRAS G12V, KRAS G12A and KRAS G12R inhibitors, preferably inhibitors of KRAS G12V and/or KRAS G12D, or inhibitors selective for KRAS G12V, as well as compounds inhibiting KRAS wildtype, preferably amplified, KRAS mutated at residue 13, such as KRAS G13D, or KRAS mutated at residue 61, such as KRAS Q61H. In particular, compounds of formula (I) (including all embodiments thereof) are potentially useful in the treatment and/or prevention of diseases and/or conditions mediated by KRAS, preferably by KRAS mutated at residue 12, e.g. KRAS G12C, KRAS G12D, KRAS G12V, more preferably G12V, or by an amplification of KRAS wildtype, or by KRAS mutated at residue 13, e.g. KRAS G13D, or by KRAS mutated at residue 61, such as KRAS Q61H.
Thus, in a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use as a medicament.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in a method of treatment of the human or animal body.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of a disease and/or condition mediated by KRAS, preferably by KRAS mutated at residue 12, e.g. KRAS G12C, KRAS G12D, KRAS G12V, more preferably G12V, or by an amplification of KRAS wildtype, or by KRAS mutated at residue 13, e.g. KRAS G13D.
In a further aspect the invention relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof—in the manufacture of a medicament for the treatment and/or prevention of a disease and/or condition mediated by KRAS, preferably by KRAS mutated at residue 12, e.g. KRAS G12C, KRAS G12D, KRAS G12V, more preferably G12V, or by an amplification of KRAS wildtype, or by KRAS mutated at residue 13, e.g. KRAS G13D.
In a further aspect the invention relates to a method for the treatment and/or prevention of a disease and/or condition mediated by KRAS, preferably by KRAS mutated at residue 12, e.g. KRAS G12C, KRAS G12D, KRAS G12V, more preferably G12V, or by an amplification of KRAS wildtype, or by KRAS mutated at residue 13, e.g. KRAS G13D comprising administering a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof—to a human being.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in a method of treatment and/or prevention of cancer in the human or animal body.
In a further aspect the invention relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof—in the manufacture of a medicament for the treatment and/or prevention of cancer.
In a further aspect the invention relates to a method for the treatment and/or prevention of cancer comprising administering a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof—to a human being.
Preferably, the cancer as defined herein (above or below) comprises a KRAS mutation. In particular, KRAS mutations include e.g. mutations of the KRAS gene and of the KRAS protein, such as overexpressed KRAS, amplified KRAS or KRAS, KRAS mutated at residue 12, KRAS mutated at residue 13, KRAS mutated at residue 61, KRAS mutated at residue 146, in particular KRAS G12A, KRAS G12C, KRAS G12D, KRAS G12V, KRAS G12S, KRAS G13C, KRAS G13D, KRAS G13V, KRAS Q61H, KRAS Q61E, KRAS Q61P, KRAS A146P, KRAS A146T, KRAS A146V. KRAS may present one or more of these mutations/alterations.
Preferably, the cancer as defined herein (above or below) comprises a BRAF mutation in addition or in alternative to the KRAS mutation. Said BRAF mutation is in particular a class Ill BRAF mutation, e.g. as defined in Z. Yao, Nature, 2017, 548, 234-238.
Preferably, the cancer as defined herein (above or below) comprises a mutation in a receptor tyrosine kinase (RTK), including EGFR, MET and ERBB2 mutations, in addition or in alternative to the KRAS mutation.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the cancer comprises a KRAS mutation, said KRAS mutation being preferably selected from the group consisting of: KRAS G12C, KRAS G12D, KRAS G12V, KRAS G13D; or an amplification of KRAS wildtype, amplification of the KRAS gene or overexpression of KRAS.
In a further aspect the invention relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof—in the manufacture of a medicament for the treatment and/or prevention of cancer, wherein the cancer comprises a KRAS mutation, said KRAS mutation being preferably selected from the group consisting of: KRAS G12C, KRAS G12D, KRAS G12V, KRAS G13D; or an amplification of KRAS wildtype, amplification of the KRAS gene or overexpression of KRAS.
In a further aspect the invention relates to a method for the treatment and/or prevention of cancer comprising administering a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof—to a human being, wherein the cancer comprises a KRAS mutation, said KRAS mutation being preferably selected from the group consisting of: KRAS G12C, KRAS G12D, KRAS G12V, KRAS G13D; or an amplification of KRAS wildtype, amplification of the KRAS gene or overexpression of KRAS.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the cancer comprises a KRAS G12D mutation.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the cancer comprises a KRAS G12V mutation.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the cancer comprises a KRAS G13D mutation.
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the cancer comprises wildtype amplified KRAS. Another aspect is based on identifying a link between the KRAS status of a patient and potential susceptibility to treatment with a compound of formula (I). A KRAS inhibitor, such as a compound of formula (I) may then advantageously be used to treat patients with a disease dependent on KRAS who may be resistant to other therapies. This therefore provides opportunities, methods and tools for selecting patients for treatment with a compound of formula (I) particularly cancer patients.
The selection is based on whether the tumor cells to be treated possess wildtype, preferably amplified, or KRAS mutated at residue 12, preferably G12C, G12D or G12V gene, or KRAS mutated at residue 13, preferably G13D gene. The KRAS gene status could therefore be used as a biomarker to indicate that selecting treatment with a compound of formula (I) may be advantageous.
According to one aspect, there is provided a method for selecting a patient for treatment with a compound of formula (I) the method comprising
The method may include or exclude the actual patient sample isolation step.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a KRAS mutation or an amplification of KRAS wildtype.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a G12C mutant, G12D mutant, G12V mutant, G12A mutant, G13D mutant or G12R mutant KRAS gene or an amplification of KRAS wildtype.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a G12C mutant, G12D mutant, G12V mutant or G13D mutant KRAS gene or an amplification of KRAS wildtype.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a G12D mutant KRAS gene.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a G12V mutant KRAS gene.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring a G13D mutant KRAS gene.
According to another aspect, there is provided a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in treating a cancer with tumor cells harbouring wildtype amplified KRAS or overexpressed KRAS.
According to another aspect, there is provided a method of treating a cancer with tumor cells harbouring a G12C mutant, G12D mutant, G12V mutant, G12A mutant, G13D mutant or G12R mutant KRAS gene or an amplification of KRAS wildtype gene comprising administering an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof—to a human being.
According to another aspect, there is provided a method of treating a cancer with tumor cells harbouring a G12C mutant, G12D mutant, G12V mutant, G12A mutant or G12R mutant KRAS gene or an amplification of KRAS wildtype gene comprising administering an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof.
Determining whether a tumor or cancer comprises a G12C KRAS mutation can be undertaken by assessing the nucleotide sequence encoding the KRAS protein, by assessing the amino acid sequence of the KRAS, protein, or by assessing the characteristics of a putative KRAS mutant protein. The sequence of wildtype human KRAS is known in the art. Methods for detecting a mutation in a KRAS nucleotide sequence are known by those of skill in the art. These methods include, but are not limited to, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assays, polymerase chain reaction-single strand conformation polymorphism (PCR—SSCP) assays, real-time PCR assays, PCR sequencing, mutant allele-specific PCR amplification (MASA) assays, direct sequencing, primer extension reactions, electrophoresis, oligonucleotide ligation assays, hybridization assays, TaqMan assays, SNP genotyping assays, high resolution melting assays and microarray analyses. In some embodiments, samples are evaluated for G12C KRAS mutations by real-time PCR. In real-time PCR, fluorescent probes specific for the KRAS G12C mutation are used. When a mutation is present, the probe binds and fluorescence is detected. In some embodiments, the KRAS G12C mutation is identified using a direct sequencing method of specific regions (e.g. exon 2 and/or exon 3) in the KRAS gene. This technique will identify all possible mutations in the region sequenced. Methods for detecting a mutation in a KRAS protein are known by those of skill in the art. These methods include, but are not limited to, detection of a KRAS mutant using a binding agent (e.g. an antibody) specific for the mutant protein, protein electrophoresis, Western blotting and direct peptide sequencing.
Methods for determining whether a tumor or cancer comprises a G12C KRAS mutation can use a variety of samples. In some embodiments, the sample is taken from a subject having a tumor or cancer. In some embodiments, the sample is a fresh tumor/cancer sample. In some embodiments, the sample is a frozen tumor/cancer sample. In some embodiments, the sample is a formalin-fixed paraffin-embedded sample. In some embodiments, the sample is processed to a cell lysate. In some embodiments, the sample is processed to DNA or RNA. In some embodiments the sample is a liquid biopsy and the test is done on a sample of blood to look for cancer cells from a tumor that are circulating in the blood or for pieces of DNA from tumor cells that are in the blood.
Analogously it can be determined whether a tumor or cancer comprises a KRAS G12D, KRAS G12V, KRAS G12A, KRAS G13D and KRAS G12R mutation or is a KRAS wildtype, preferably amplified. Preferably, the disease/condition/cancer/tumors/cancer cells to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is selected from the group consisting of pancreatic cancer, lung cancer, colorectal cancer, cholangiocarcinoma, appendiceal cancer, multiple myeloma, melanoma, uterine cancer, endometrial cancer, thyroid cancer, acute myeloid leukaemia, bladder cancer, urothelial cancer, gastric cancer, cervical cancer, head and neck squamous cell carcinoma, diffuse large B cell lymphoma, oesophageal cancer, gastroesophageal cancer, chronic lymphocytic leukaemia, hepatocellular cancer, breast cancer, ovarian cancer, prostate cancer, glioblastoma, renal cancer and sarcomas.
Preferably, the disease/condition/cancer/tumors/cancer cells to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is selected from the group consisting of pancreatic cancer, lung cancer, ovarian cancer, colorectal cancer (CRC), gastric cancer, gastroesophageal junction cancer (GEJC) and esophageal cancer. In another aspect, the disease/condition/cancer/tumors/cancer cells to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is selected from the group consisting of pancreatic cancer (preferably pancreatic ductal adenocarcinoma (PDAC)), lung cancer (preferably non-small cell lung cancer (NSCLC)), gastric cancer, cholangiocarcinoma and colorectal cancer (preferably colorectal adenocarcinoma). Preferably, said pancreatic cancer, lung cancer, cholangiocarcinoma, colorectal cancer (CRC), pancreatic ductal adenocarcinoma (PDAC), non-small cell lung cancer (NSCLC) or colorectal adenocarcinoma comprises a KRAS mutation, in particular a KRAS G12D or KRAS G12V mutation. Preferably (in alternative or in combination with the previous preferred embodiment), said non-small cell lung cancer (NSCLC) comprises a mutation (in particular a loss-of-function mutation) in the NF1 gene.
In another aspect, the disease/condition/cancer/tumors/cancer cells to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is gastric cancer, ovarian cancer or esophageal cancer, said gastric cancer or esophageal cancer being preferably selected from the group consisting of: gastric adenocarcinoma (GAC), esophageal adenocarcinoma (EAC) and gastroesophageal junction cancer (GEJC). Preferably, said gastric cancer, ovarian cancer, esophageal cancer, gastric adenocarcinoma (GAC), esophageal adenocarcinoma (EAC) or gastroesophageal junction cancer (GEJC) comprises a KRAS mutation or wildtype amplified KRAS.
Particularly preferred, the cancer to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is selected from the group consisting of:
Preferably, “cancer” as used herein (above or below) includes drug-resistant cancer and cancer that has failed one, two or more lines of mono- or combination therapy with one or more anti-cancer agents. In particular, “cancer” (and any embodiment thereof) refers to any cancer (especially the cancer species defined hereinabove and hereinbelow) that is resistant to treatment with a KRAS G12C inhibitor.
Different resistance mechanisms have already been reported. For example, the following articles describe resistance in patients following treatment with a KRAS G12C inhibitor: (i) Awad M M, Liu S, Rybkin, II, Arbour K C, Dilly J, Zhu V W, et al. Acquired resistance to KRAS(G12C) inhibition in cancer. N Engl J Med 2021; 384:2382-93 and (ii) Tanaka N, Lin J J, Li C, Ryan M B, Zhang J, Kiedrowski L A, et al. Clinical acquired resistance to KRAS(G12C) inhibition through a novel KRAS switch-II pocket mutation and polyclonal alterations converging on RAS-MAPK reactivation. Cancer Discov 2021; 11:1913-22.
In another aspect the disease/condition/cancer/tumors/cancer cells to be treated/prevented with a compound of formula (I) or a pharmaceutically acceptable salt thereof—according to the methods and uses as herein (above and below) defined and disclosed is a RASopathy, preferably selected from the group consisting of Neurofibromatosis type 1 (NF1), Noonan Syndrome (NS), Noonan Syndrome with Multiple Lentigines (NSML) (also referred to as LEOPARD syndrome), Capillary Malformation-Arteriovenous Malformation Syndrome (CM-AVM), Costello Syndrome (CS), Cardio-Facio-Cutaneous Syndrome (CFC), Legius Syndrome (also known as NF1-like Syndrome) and Hereditary gingival fibromatosis.
Additionally, the following cancers, tumors and other proliferative diseases may be treated with compounds of formula (I) or a pharmaceutically acceptable salt thereof—without being restricted thereto. Preferably, the methods of treatment, methods, uses, compounds for use and pharmaceutical compositions for use as disclosed herein (above and below) are applied in treatments of diseases/conditions/cancers/tumors which (i.e. the respective cells) harbour a KRAS mutation at position 12 (preferably a G12C, G12D, G12V, G12A, G12R mutation) or an amplification of KRAS wildtype alternatively they have been identified to harbour a KRAS mutation at position 12 (preferably a G12C, G12D, G12V, G12A, G12R mutation) as herein described and/or referred or an amplification of KRAS wildtype:
All cancers/tumors/carcinomas mentioned above which are characterized by their specific location/origin in the body are meant to include both the primary tumors and the metastatic tumors derived therefrom.
All cancers/tumors/carcinomas mentioned above may be further differentiated by their histopathological classification:
The compounds of the invention may be used in therapeutic regimens in the context of first line, second line, or any further line treatments.
The compounds of the invention may be used for the prevention, short-term or long-term treatment of the above-mentioned diseases/conditions/cancers/tumors, optionally also in combination with radiotherapy and/or surgery.
The methods of treatment, methods, uses and compounds for use as disclosed herein (above and below) can be performed with any compound of formula (I) or a pharmaceutically acceptable salt thereof—as disclosed or defined herein and with any pharmaceutical composition or kit comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof (each including all individual embodiments or generic subsets of compounds of formula (I).
The compounds of formula (I) or the pharmaceutically acceptable salts thereof—and the pharmaceutical compositions comprising such compounds or salts may also be co-administered with other pharmacologically active substances, e.g. with other anti-neoplastic compounds (e.g. chemotherapy), or used in combination with other treatments, such as radiation or surgical intervention, either as an adjuvant prior to surgery or post-operatively. Preferably, the pharmacologically active substance(s) for co-administration is/are (an) anti-neoplastic compound(s).
Thus, in a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use as hereinbefore defined wherein said compound is administered before, after or together with one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use as hereinbefore defined, wherein said compound is administered in combination with one or more other pharmacologically active substance(s).
In a further aspect the invention relates to the use of a compound of formula (I) or a pharmaceutically acceptable salt thereof—as hereinbefore defined wherein said compound is to be administered before, after or together with one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a method (e.g. a method for the treatment and/or prevention) as hereinbefore defined wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof—is administered before, after or together with a therapeutically effective amount of one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a method (e.g. a method for the treatment and/or prevention) as hereinbefore defined wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof—is administered in combination with a therapeutically effective amount of one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a method for the treatment and/or prevention of cancer comprising administering to a patient in need thereof a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt thereof—and a therapeutically effective amount of one or more other pharmacologically active substance(s), wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof—is administered simultaneously, concurrently, sequentially, successively, alternately or separately with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a method for the treatment and/or prevention of cancer comprising administering to a patient in need thereof a therapeutically effective amount of an inhibitor of a KRAS mutated at residue 12 or 13, such as KRAS G12C, KRAS G12D, KRAS G12V, KRAS G12A, KRAS G13D and/or KRAS G12R inhibitors, preferably KRAS G12C, KRAS G12D or selective KRAS G12D inhibitors—or a pharmaceutically acceptable salt thereof—and a therapeutically effective amount of one or more other pharmacologically active substance(s), wherein the inhibitor—or a pharmaceutically acceptable salt thereof—is administered in combination with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a method for the treatment and/or prevention of cancer comprising administering to a patient in need thereof a therapeutically effective amount of an inhibitor of KRAS wildtype amplified or overexpressed—or a pharmaceutically acceptable salt thereof—and a therapeutically effective amount of one or more other pharmacologically active substance(s), wherein the inhibitor—or a pharmaceutically acceptable salt thereof—is administered in combination with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a compound of formula (I) or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof—is administered simultaneously, concurrently, sequentially, successively, alternately or separately with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to an inhibitor of a KRAS mutated at residue 12 or 13, such as KRAS G12C, KRAS G12D, KRAS G12V, KRAS G12A, KRAS G13D and/or KRAS G12R inhibitors, preferably KRAS G12C, KRAS G12D or selective KRAS G12D inhibitors—or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the inhibitor—or a pharmaceutically acceptable salt thereof—is administered in combination with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to an inhibitor of an inhibitor of KRAS wildtype amplified or overexpressed—or a pharmaceutically acceptable salt thereof—for use in the treatment and/or prevention of cancer, wherein the inhibitor—or a pharmaceutically acceptable salt thereof—is administered in combination with the one or more other pharmacologically active substance(s).
In a further aspect the invention relates to a kit comprising
In one aspect such kit for said use comprises a third pharmaceutical composition or dosage form comprising a third pharmaceutical composition or dosage form comprising still another pharmacologically active substance, and, optionally, one or more pharmaceutically acceptable excipient(s)
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered simultaneously.
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered concurrently.
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered sequentially.
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered successively.
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered alternately.
In a further embodiment of the invention the components (i.e. the combination partners) of the combinations, kits, uses, methods and compounds for use according to the invention (including all embodiments) are administered separately.
The pharmacologically active substance(s) to be used together/in combination with the compound of formula (I) or a pharmaceutically acceptable salt thereof—(including all individual embodiments or generic subsets of compounds) or in the medical uses, uses, methods of treatment and/or prevention, pharmaceutical compositions as herein (above and below) defined can be selected from any one or more of the following (preferably there is one or two additional pharmacologically active substance used in all these embodiments):
In a further embodiment of the (combined) use and method (e.g. method for the treatment and/or prevention) as hereinbefore described one other pharmacologically active substance is to be administered before, after or together with the compound of formula (I)—or a pharmaceutically acceptable salt thereof—wherein said one other pharmacologically active substance is
In a further embodiment of the (combined) use and method (e.g., method for the treatment and/or prevention) as hereinbefore described one other pharmacologically active substance is to be administered in combination with the compound of formula (I) or a pharmaceutically acceptable salt thereof—wherein said one other pharmacologically active substance is
In a further aspect of the (combined) use and method (e.g. method for the treatment and/or prevention) as hereinbefore described two other pharmacologically active substances are to be administered before, after or together with the compound of formula (I)—or a pharmaceutically acceptable salt thereof—wherein said two other pharmacologically active substances are
In a further aspect of the (combined) use and method (e.g. method for the treatment and/or prevention) as hereinbefore described two other pharmacologically active substances are to be administered in combination with the compound of formula (I) or a pharmaceutically acceptable salt thereof—wherein said two other pharmacologically active substances are
Additional pharmacologically active substance(s) which can also be used together/in combination with the compound of formula (I) or a pharmaceutically acceptable salt thereof—(including all individual embodiments or generic subsets of compounds of formula (I) or in the medical uses, uses, methods of treatment and/or prevention, pharmaceutical compositions, kits as herein (above and below) defined include, without being restricted thereto, hormones, hormone analogues and antihormones (e.g. tamoxifen, toremifene, raloxifene, fulvestrant, megestrol acetate, flutamide, nilutamide, bicalutamide, aminoglutethimide, cyproterone acetate, finasteride, buserelin acetate, fludrocortisone, fluoxymesterone, medroxyprogesterone, octreotide), aromatase inhibitors (e.g. anastrozole, letrozole, liarozole, vorozole, exemestane, atamestane), LHRH agonists and antagonists (e.g. goserelin acetate, luprolide), inhibitors of growth factors and/or of their corresponding receptors (growth factors such as for example platelet derived growth factor (PDGF), fibroblast growth factor (FGF), vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), insuline-like growth factors (IGF), human epidermal growth factor (HER, e.g. HER2, HER3, HER4) and hepatocyte growth factor (HGF) and/or their corresponding receptors), inhibitors are for example (anti-)growth factor antibodies, (anti-)growth factor receptor antibodies and tyrosine kinase inhibitors, such as for example cetuximab, gefitinib, afatinib, nintedanib, imatinib, lapatinib, bosutinib, bevacizumab and trastuzumab); antimetabolites (e.g. antifolates such as methotrexate, raltitrexed, pyrimidine analogues such as 5-fluorouracil (5-FU), ribonucleoside and deoxyribonucleoside analogues, capecitabine and gemcitabine, purine and adenosine analogues such as mercaptopurine, thioguanine, cladribine and pentostatin, cytarabine (ara C), fludarabine); antitumor antibiotics (e.g. anthracyclins such as doxorubicin, doxil (pegylated liposomal doxorubicin hydrochloride, myocet (non-pegylated liposomal doxorubicin), daunorubicin, epirubicin and idarubicin, mitomycin-C, bleomycin, dactinomycin, plicamycin, streptozocin); platinum derivatives (e.g. cisplatin, oxaliplatin, carboplatin); alkylation agents (e.g. estramustin, meclorethamine, melphalan, chlorambucil, busulphan, dacarbazin, cyclophosphamide, ifosfamide, temozolomide, nitrosoureas such as for example carmustin and lomustin, thiotepa); antimitotic agents (e.g. Vinca alkaloids such as for example vinblastine, vindesin, vinorelbin and vincristine; and taxanes such as paclitaxel, docetaxel); angiogenesis inhibitors (e.g. tasquinimod), tubuline inhibitors; DNA synthesis inhibitors, PARP inhibitors, topoisomerase inhibitors (e.g. epipodophyllotoxins such as for example etoposide and etopophos, teniposide, amsacrin, topotecan, irinotecan, mitoxantrone), serine/threonine kinase inhibitors (e.g. PDK 1 inhibitors, Raf inhibitors, A-Raf inhibitors, B-Raf inhibitors, C-Raf inhibitors, mTOR inhibitors, mTORC1/2 inhibitors, PI3K inhibitors, PI3Kα inhibitors, dual mTOR/PI3K inhibitors, STK 33 inhibitors, AKT inhibitors, PLK 1 inhibitors, inhibitors of CDKs, Aurora kinase inhibitors), tyrosine kinase inhibitors (e.g. PTK2/FAK inhibitors), protein protein interaction inhibitors (e.g. IAP inhibitors/SMAC mimetics, Mcl-1, MDM2/MDMX), MEK inhibitors, ERK inhibitors, FLT3 inhibitors, BRD4 inhibitors, IGF-1R inhibitors, TRAILR2 agonists, Bcl-xL inhibitors, Bcl-2 inhibitors (e.g. venetoclax), Bcl-2/Bcl-xL inhibitors, ErbB receptor inhibitors, BCR-ABL inhibitors, ABL inhibitors, Src inhibitors, rapamycin analogs (e.g. everolimus, temsirolimus, ridaforolimus, sirolimus), androgen synthesis inhibitors, androgen receptor inhibitors, DNMT inhibitors, HDAC inhibitors, ANG1/2 inhibitors, CYP17 inhibitors, radiopharmaceuticals, proteasome inhibitors (e.g. carfilzomib), immunotherapeutic agents such as immune checkpoint inhibitors (e.g. CTLA4, PD1, PD-L1, PD-L2, LAG3, and TIM3 binding molecules/immunoglobulins, such as e.g. ipilimumab, nivolumab, pembrolizumab), ADCC (antibody-dependent cell-mediated cytotoxicity) enhancers (e.g. anti-CD33 antibodies, anti-CD37 antibodies, anti-CD20 antibodies), t-cell engagers (e.g. bi-specific T-cell engagers (BiTEs®) like e.g. CD3×BCMA, CD3×CD33, CD3×CD19), PSMA×CD3), tumor vaccines, immunomodulator, e.g. STING agonist, and various chemotherapeutic agents such as amifostin, anagrelid, clodronat, filgrastin, interferon, interferon alpha, leucovorin, procarbazine, levamisole, mesna, mitotane, pamidronate and porfimer.
It is to be understood that the combinations, compositions, kits, methods, uses, pharmaceutical compositions or compounds for use according to this invention may envisage the simultaneous, concurrent, sequential, successive, alternate or separate administration of the active ingredients or components. It will be appreciated that the compound of formula (I) or a pharmaceutically acceptable salt thereof—and the one or more other pharmacologically active substance(s) can be administered formulated either dependently or independently, such as e.g. the compound of formula (I) or a pharmaceutically acceptable salt thereof—and the one or more other pharmacologically active substance(s) may be administered either as part of the same pharmaceutical composition/dosage form or, preferably, in separate pharmaceutical compositions/dosage forms.
In this context, “combination” or “combined” within the meaning of this invention includes, without being limited, a product that results from the mixing or combining of more than one active ingredient and includes both fixed and non-fixed (e.g. free) combinations (including kits) and uses, such as e.g. the simultaneous, concurrent, sequential, successive, alternate or separate use of the components or ingredients. The term “fixed combination” means that the active ingredients are administered to a patient simultaneously in the form of a single entity or dosage. The term “non-fixed combination” means that the active ingredients are administered to a patient as separate entities either simultaneously, concurrently or sequentially with no specific time limits, wherein such administration provides therapeutically effective levels of the compounds in the body of the patient.
The administration of the compound of formula (I) or a pharmaceutically acceptable salt thereof—and the one or more other pharmacologically active substance(s) may take place by co-administering the active components or ingredients, such as e.g. by administering them simultaneously or concurrently in one single or in two or more separate formulations or dosage forms. Alternatively, the administration of the compound of formula (I) or a pharmaceutically acceptable salt thereof—and the one or more other pharmacologically active substance(s) may take place by administering the active components or ingredients sequentially or in alternation, such as e.g. in two or more separate formulations or dosage forms.
For example, simultaneous administration includes administration at substantially the same time. This form of administration may also be referred to as “concomitant” administration. Concurrent administration includes administering the active agents within the same general time period, for example on the same day(s) but not necessarily at the same time. Alternate administration includes administration of one agent during a time period, for example over the course of a few days or a week, followed by administration of the other agent(s) during a subsequent period of time, for example over the course of a few days or a week, and then repeating the pattern for one or more cycles. Sequential or successive administration includes administration of one agent during a first time period (for example over the course of a few days or a week) using one or more doses, followed by administration of the other agent(s) during a second and/or additional time period (for example over the course of a few days or a week) using one or more doses. An overlapping schedule may also be employed, which includes administration of the active agents on different days over the treatment period, not necessarily according to a regular sequence. Variations on these general guidelines may also be employed, e.g. according to the agents used and the condition of the subject.
Terms not specifically defined herein should be given the meanings that would be given to them by one of skill in the art in light of the disclosure and the context. As used in the specification, however, unless specified to the contrary, the following terms have the meaning indicated and the following conventions are adhered to:
The use of the prefix Cx-y, wherein x and y each represent a positive integer (x<y), indicates that the chain or ring structure or combination of chain and ring structure as a whole, specified and mentioned in direct association, may consist of a maximum of y and a minimum of x carbon atoms.
The indication of the number of members in groups that contain one or more heteroatom(s) (e.g. heteroaryl, heteroarylalkyl, heterocyclyl, heterocycylalkyl) relates to the total number of atoms of all the ring members or the total of all the ring and carbon chain members.
The indication of the number of carbon atoms in groups that consist of a combination of carbon chain and carbon ring structure (e.g. cycloalkylalkyl, arylalkyl) relates to the total number of carbon atoms of all the carbon ring and carbon chain members. Obviously, a ring structure has at least three members.
In general, for groups comprising two or more subgroups (e.g. heteroarylalkyl, heterocycylalkyl, cycloalkylalkyl, arylalkyl) the last named subgroup is the radical attachment point, for example, the substituent aryl-C1-6alkyl means an aryl group which is bound to a C1-6 alkyl group, the latter of which is bound to the core or to the group to which the substituent is attached.
In groups like HO, H2N, (O)S, (O)2S, NC (cyano), HOOC, F3C or the like, the skilled artisan can see the radical attachment point(s) to the molecule from the free valences of the group itself.
The expression “compound of the invention” and grammatical variants thereof comprises compounds of formula (I), including all salts, aspects and preferred embodiments thereof as herein defined. Any reference to a compound of the invention or to a compound of formula (I) is intended to include a reference to the respective (sub)aspects and embodiments.
Alkyl denotes monovalent, saturated hydrocarbon chains, which may be present in both straight-chain (unbranched) and branched form. If an alkyl is substituted, the substitution may take place independently of one another, by mono- or polysubstitution in each case, on all the hydrogen-carrying carbon atoms.
The term “C1-5alkyl” includes for example H3C—, H3C—CH2—, H3C—CH2—CH2—, H3C—CH(CH3)—, H3C—CH2—CH2—CH2—, H3C—CH2—CH(CH3)—, H3C—CH(CH3)—CH2—, H3C—C(CH3)2—, H3C—CH2—CH2—CH2—CH2—, H3C—CH2—CH2—CH(CH3)—, H3C—CH2—CH(CH3)—CH2—, H3C—CH(CH3)—CH2—CH2—, H3C—CH2—C(CH3)2—, H3C—C(CH3)2—CH2—, H3C—CH(CH3)—CH(CH3)—and H3C—CH2—CH(CH2CH3)—.
Further examples of alkyl are methyl (Me; —CH3), ethyl (Et; —CH2CH3), 1-propyl (n-propyl; n-Pr; —CH2CH2CH3), 2-propyl (i-Pr; iso-propyl; —CH(CH3)2), 1-butyl (n-butyl; n-Bu; —CH2CH2CH2CH3), 2-methyl-1-propyl (iso-butyl; i-Bu; —CH2CH(CH3)2), 2-butyl (sec-butyl; sec-Bu; —CH(CH3)CH2CH3), 2-methyl-2-propyl (tert-butyl; t-Bu; —C(CH3)3), 1-pentyl (n-pentyl; —CH2CH2CH2CH2CH3), 2-pentyl (—CH(CH3)CH2CH2CH3), 3-pentyl (—CH(CH2CH3)2), 3-methyl-1-butyl (iso-pentyl; —CH2CH2CH(CH3)2), 2-methyl-2-butyl (—C(CH3)2CH2CH3), 3-methyl-2-butyl (—CH(CH3)CH(CH3)2), 2,2-dimethyl-1-propyl (neo-pentyl; —CH2C(CH3)3), 2-methyl-1-butyl (—CH2CH(CH3)CH2CH3), 1-hexyl (n-hexyl; —CH2CH2CH2CH2CH2CH3), 2-hexyl (—CH(CH3)CH2CH2CH2CH3), 3-hexyl (—CH(CH2CH3)(CH2CH2CH3)), 2-methyl-2-pentyl (—C(CH3)2CH2CH2CH3), 3-methyl-2-pentyl (—CH(CH3)CH(CH3)CH2CH3), 4-methyl-2-pentyl (—CH(CH3)CH2CH(CH3)2), 3-methyl-3-pentyl (—C(CH3)(CH2CH3)2), 2-methyl-3-pentyl (—CH(CH2CH3)CH(CH3)2), 2,3-dimethyl-2-butyl (—C(CH3)2CH(CH3)2), 3,3-dimethyl-2-butyl (—CH(CH3)C(CH3)3), 2,3-dimethyl-1-butyl (—CH2CH(CH3)CH(CH3)CH3), 2,2-dimethyl-1-butyl (—CH2C(CH3)2CH2CH3), 3,3-dimethyl-1-butyl (—CH2CH2C(CH3)3), 2-methyl-1-pentyl (—CH2CH(CH3)CH2CH2CH3), 3-methyl-1-pentyl (—CH2CH2CH(CH3)CH2CH3), 1-heptyl (n-heptyl), 2-methyl-1-hexyl, 3-methyl-1-hexyl, 2,2-dimethyl-1-pentyl, 2,3-dimethyl-1-pentyl, 2,4-dimethyl-1-pentyl, 3,3-dimethyl-1-pentyl, 2,2,3-trimethyl-1-butyl, 3-ethyl-1-pentyl, 1-octyl (n-octyl), 1-nonyl (n-nonyl); 1-decyl (n-decyl) etc.
By the terms propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl etc. without any further definition are meant saturated hydrocarbon groups with the corresponding number of carbon atoms, wherein all isomeric forms are included.
The above definition for alkyl also applies if alkyl is a part of another (combined) group such as for example Cx-yalkylamino or Cx-yalkyloxy.
The term alkylene can also be derived from alkyl. Alkylene is bivalent, unlike alkyl, and requires two binding partners. Formally, the second valency is produced by removing a hydrogen atom in an alkyl. Corresponding groups are for example —CH3 and —CH2—, —CH2CH3 and —CH2CH2— or >CHCH3 etc.
The term “C1-4alkylene” includes for example —(CH2)—, —(CH2—CH2)—, —(CH(CH3))—, —(CH2—CH2—CH2)—, —(C(CH3)2)—, —(CH(CH2CH3))—, —(CH(CH3)—CH2)—, —(CH2—CH(CH3))—, —(CH2—CH2—CH2—CH2)—, —(CH2—CH2—CH(CH3))—, —(CH(CH3)—CH2—CH2)—, —(CH2—CH(CH3)—CH2)—, —(CH2—C(CH3)2)—, —(C(CH3)2—CH2)—, —(CH(CH3)—CH(CH3))—, —(CH2—CH(CH2CH3))—, —(CH(CH2CH3)—CH2)—, —(CH(CH2CH2CH3))—, —(CH(CH(CH3))2)— and —C(CH3)(CH2CH3)—.
Other examples of alkylene are methylene, ethylene, propylene, 1-methylethylene, butylene, 1-methylpropylene, 1,1-dimethylethylene, 1,2-dimethylethylene, pentylene, 1,1-dimethylpropylene, 2,2-dimethylpropylene, 1,2-dimethylpropylene, 1,3-dimethylpropylene, hexylene etc.
By the generic terms propylene, butylene, pentylene, hexylene etc. without any further definition are meant all the conceivable isomeric forms with the corresponding number of carbon atoms, i.e. propylene includes 1-methylethylene and butylene includes 1-methylpropylene, 2-methylpropylene, 1,1-dimethylethylene and 1,2-dimethylethylene.
The above definition for alkylene also applies if alkylene is part of another (combined) group such as for example in HO—Cx-yalkyleneamino or H2N—Cx-yalkyleneoxy.
Unlike alkyl, alkenyl consists of at least two carbon atoms, wherein at least two adjacent carbon atoms are joined together by a C—C double bond and a carbon atom can only be part of one C—C double bond. If in an alkyl as hereinbefore defined having at least two carbon atoms, two hydrogen atoms on adjacent carbon atoms are formally removed and the free valencies are saturated to form a second bond, the corresponding alkenyl is formed.
Examples of alkenyl are vinyl (ethenyl), prop-1-enyl, allyl (prop-2-enyl), isopropenyl, but-1-enyl, but-2-enyl, but-3-enyl, 2-methyl-prop-2-enyl, 2-methyl-prop-1-enyl, 1-methyl-prop-2-enyl, 1-methyl-prop-1-enyl, 1-methylidenepropyl, pent-1-enyl, pent-2-enyl, pent-3-enyl, pent-4-enyl, 3-methyl-but-3-enyl, 3-methyl-but-2-enyl, 3-methyl-but-1-enyl, hex-1-enyl, hex-2-enyl, hex-3-enyl, hex-4-enyl, hex-5-enyl, 2,3-dimethyl-but-3-enyl, 2,3-dimethyl-but-2-enyl, 2-methylidene-3-methylbutyl, 2,3-dimethyl-but-1-enyl, hexa-1,3-dienyl, hexa-1,4-dienyl, penta-1,4-dienyl, penta-1,3-dienyl, buta-1,3-dienyl, 2,3-dimethylbuta-1,3-diene etc.
By the generic terms propenyl, butenyl, pentenyl, hexenyl, butadienyl, pentadienyl, hexadienyl, heptadienyl, octadienyl, nonadienyl, decadienyl etc. without any further definition are meant all the conceivable isomeric forms with the corresponding number of carbon atoms, i.e. propenyl includes prop-1-enyl and prop-2-enyl, butenyl includes but-1-enyl, but-2-enyl, but-3-enyl, 1-methyl-prop-1-enyl, 1-methyl-prop-2-enyl etc.
Alkenyl may optionally be present in the cis or trans or E or Z orientation with regard to the double bond(s).
The above definition for alkenyl also applies when alkenyl is part of another (combined) group such as for example in Cx-yalkenylamino or Cx-yalkenyloxy.
Unlike alkylene, alkenylene consists of at least two carbon atoms, wherein at least two adjacent carbon atoms are joined together by a C═C double bond and a carbon atom can only be part of one C═C double bond. If in an alkylene as hereinbefore defined having at least two carbon atoms, two hydrogen atoms at adjacent carbon atoms are formally removed and the free valencies are saturated to form a second bond, the corresponding alkenylene is formed.
Examples of alkenylene are ethenylene, propenylene, 1-methylethenylene, butenylene, 1-methylpropenylene, 1,1-dimethylethenylene, 1,2-dimethylethenylene, pentenylene, 1,1-dimethylpropenylene, 2,2-dimethylpropenylene, 1,2-dimethylpropenylene, 1,3-dimethylpropenylene, hexenylene etc.
By the generic terms propenylene, butenylene, pentenylene, hexenylene etc. without any further definition are meant all the conceivable isomeric forms with the corresponding number of carbon atoms, i.e. propenylene includes 1-methylethenylene and butenylene includes 1-methylpropenylene, 2-methylpropenylene, 1,1-dimethylethenylene and 1,2-dimethylethenylene.
Alkenylene may optionally be present in the cis or trans or E or Z orientation with regard to the double bond(s).
The above definition for alkenylene also applies when alkenylene is a part of another (combined) group as for example in HO—Cx-yalkenyleneamino or H2N—Cx-yalkenyleneoxy.
Unlike alkyl, alkynyl consists of at least two carbon atoms, wherein at least two adjacent carbon atoms are joined together by a C—C triple bond. If in an alkyl as hereinbefore defined having at least two carbon atoms, two hydrogen atoms in each case at adjacent carbon atoms are formally removed and the free valencies are saturated to form two further bonds, the corresponding alkynyl is formed.
Examples of alkynyl are ethynyl, prop-1-ynyl, prop-2-ynyl, but-1-ynyl, but-2-ynyl, but-3-ynyl, 1-methyl-prop-2-ynyl, pent-1-ynyl, pent-2-ynyl, pent-3-ynyl, pent-4-ynyl, 3-methyl-but-1-ynyl, hex-1-ynyl, hex-2-ynyl, hex-3-ynyl, hex-4-ynyl, hex-5-ynyl etc.
By the generic terms propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl etc. without any further definition are meant all the conceivable isomeric forms with the corresponding number of carbon atoms, i.e. propynyl includes prop-1-ynyl and prop-2-ynyl, butynyl includes but-1-ynyl, but-2-ynyl, but-3-ynyl, 1-methyl-prop-1-ynyl, 1-methyl-prop-2-ynyl, etc.
If a hydrocarbon chain carries both at least one double bond and also at least one triple bond, by definition it belongs to the alkynyl subgroup.
The above definition for alkynyl also applies if alkynyl is part of another (combined) group, as for example in Cx-yalkynylamino or Cx-yalkynyloxy.
Unlike alkylene, alkynylene consists of at least two carbon atoms, wherein at least two adjacent carbon atoms are joined together by a C—C triple bond. If in an alkylene as hereinbefore defined having at least two carbon atoms, two hydrogen atoms in each case at adjacent carbon atoms are formally removed and the free valencies are saturated to form two further bonds, the corresponding alkynylene is formed.
Examples of alkynylene are ethynylene, propynylene, 1-methylethynylene, butynylene, 1-methylpropynylene, 1,1-dimethylethynylene, 1,2-dimethylethynylene, pentynylene, 1,1-dimethylpropynylene, 2,2-dimethylpropynylene, 1,2-dimethylpropynylene, 1,3-dimethylpropynylene, hexynylene etc.
By the generic terms propynylene, butynylene, pentynylene, hexynylene etc. without any further definition are meant all the conceivable isomeric forms with the corresponding number of carbon atoms, i.e. propynylene includes 1-methylethynylene and butynylene includes 1-methylpropynylene, 2-methylpropynylene, 1,1-dimethylethynylene and 1,2-dimethylethynylene.
The above definition for alkynylene also applies if alkynylene is part of another (combined) group, as for example in HO—Cx-yalkynyleneamino or H2N—Cx-yalkynyleneoxy.
By heteroatoms are meant oxygen, nitrogen and sulphur atoms.
Haloalkyl (haloalkenyl, haloalkynyl) is derived from the previously defined alkyl (alkenyl, alkynyl) by replacing one or more hydrogen atoms of the hydrocarbon chain independently of one another by halogen atoms, which may be identical or different. If a haloalkyl (haloalkenyl, haloalkynyl) is to be further substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon atoms.
Examples of haloalkyl (haloalkenyl, haloalkynyl) are —CF3, —CHF2, —CH2F, —CF2CF3, —CHFCF3, —CH2CF3, —CF2CH3, —CHFCH3, —CF2CF2CF3, —CF2CH2CH3, —CF═CF2, —CCl═CH2, —CBr═CH2, —C≡C—CF3, —CHFCH2CH3, —CHFCH2CF3 etc.
From the previously defined haloalkyl (haloalkenyl, haloalkynyl) are also derived the terms haloalkylene (haloalkenylene, haloalkynylene). Haloalkylene (haloalkenylene, haloalkynylene), unlike haloalkyl (haloalkenyl, haloalkynyl), is bivalent and requires two binding partners. Formally, the second valency is formed by removing a hydrogen atom from a haloalkyl (haloalkenyl, haloalkynyl).
Corresponding groups are for example —CH2F and —CHF—, —CHFCH2F and —CHFCHF— or >CFCH2F etc.
The above definitions also apply if the corresponding halogen-containing groups are part of another (combined) group.
Halogen denotes fluorine, chlorine, bromine and/or iodine atoms.
Cycloalkyl is made up of the subgroups monocyclic cycloalkyl, bicyclic cycloalkyl and spiro-cycloalkyl. The ring systems are saturated and formed by linked carbon atoms. In bicyclic cycloalkyl two rings are joined together so that they have at least two carbon atoms in common. In spiro-cycloalkyl one carbon atom (spiroatom) belongs to two rings together.
If a cycloalkyl is to be substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon atoms. Cycloalkyl itself may be linked as a substituent to the molecule via every suitable position of the ring system.
Examples of cycloalkyl are cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, bicyclo[2.2.0]hexyl, bicyclo[3.2.0]heptyl, bicyclo[3.2.1]octyl, bicyclo[2.2.2]octyl, bicyclo[4.3.0]nonyl (octahydroindenyl), bicyclo[4.4.0]decyl (decahydronaphthyl), bicyclo[2.2.1]heptyl (norbornyl), bicyclo[4.1.0]heptyl (norcaranyl), bicyclo[3.1.1]heptyl (pinanyl), spiro[2.5]octyl, spiro[3.3]heptyl etc.
The above definition for cycloalkyl also applies if cycloalkyl is part of another (combined) group as for example in Cx-ycycloalkylamino, Cx-ycycloalkyloxy or Cx-ycycloalkylalkyl.
If the free valency of a cycloalkyl is saturated, then an alicycle is obtained.
The term cycloalkylene can thus be derived from the previously defined cycloalkyl. Cycloalkylene, unlike cycloalkyl, is bivalent and requires two binding partners. Formally, the second valency is obtained by removing a hydrogen atom from a cycloalkyl. Corresponding groups are for example:
The above definition for cycloalkylene also applies if cycloalkylene is part of another (combined) group as for example in HO—Cx-ycycloalkyleneamino or H2N—Cx-ycycloalkyleneoxy.
Cycloalkenyl is made up of the subgroups monocyclic cycloalkenyl, bicyclic cycloalkenyl and spiro-cycloalkenyl. However, the systems are unsaturated, i.e. there is at least one C—C double bond but no aromatic system. If in a cycloalkyl as hereinbefore defined two hydrogen atoms at adjacent cyclic carbon atoms are formally removed and the free valencies are saturated to form a second bond, the corresponding cycloalkenyl is obtained.
If a cycloalkenyl is to be substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon atoms. Cycloalkenyl itself may be linked as a substituent to the molecule via every suitable position of the ring system.
Examples of cycloalkenyl are cycloprop-1-enyl, cycloprop-2-enyl, cyclobut-1-enyl, cyclobut-2-enyl, cyclopent-1-enyl, cyclopent-2-enyl, cyclopent-3-enyl, cyclohex-1-enyl, cyclohex-2-enyl, cyclohex-3-enyl, cyclohept-1-enyl, cyclohept-2-enyl, cyclohept-3-enyl, cyclohept-4-enyl, cyclobuta-1,3-dienyl, cyclopenta-1,4-dienyl, cyclopenta-1,3-dienyl, cyclopenta-2,4-dienyl, cyclohexa-1,3-dienyl, cyclohexa-1,5-dienyl, cyclohexa-2,4-dienyl, cyclohexa-1,4-dienyl, cyclohexa-2,5-dienyl, bicyclo[2.2.1]hepta-2,5-dienyl (norborna-2,5-dienyl), bicyclo[2.2.1]hept-2-enyl (norbornenyl), spiro[4,5]dec-2-enyl etc.
The above definition for cycloalkenyl also applies when cycloalkenyl is part of another (combined) group as for example in Cx-ycycloalkenylamino, Cx-ycycloalkenyloxy or Cx-ycycloalkenylalkyl.
If the free valency of a cycloalkenyl is saturated, then an unsaturated alicycle is obtained.
The term cycloalkenylene can thus be derived from the previously defined cycloalkenyl. Cycloalkenylene, unlike cycloalkenyl, is bivalent and requires two binding partners. Formally, the second valency is obtained by removing a hydrogen atom from a cycloalkenyl. Corresponding groups are for example:
The above definition for cycloalkenylene also applies if cycloalkenylene is part of another (combined) group as for example in HO—Cx-ycycloalkenyleneamino or H2N—Cx-ycycloalkenyleneoxy.
Aryl denotes mono-, bi- or tricyclic carbocycles with at least one aromatic carbocycle. Preferably, it denotes a monocyclic group with six carbon atoms (phenyl) or a bicyclic group with nine or ten carbon atoms (two six-membered rings or one six-membered ring with a five-membered ring), wherein the second ring may also be aromatic or, however, may also be partially saturated.
If an aryl is to be substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon atoms. Aryl itself may be linked as a substituent to the molecule via every suitable position of the ring system.
Examples of aryl are phenyl, naphthyl, indanyl (2,3-dihydroindenyl), indenyl, anthracenyl, phenanthrenyl, tetrahydronaphthyl (1,2,3,4-tetrahydronaphthyl, tetralinyl), dihydronaphthyl (1,2-dihydronaphthyl), fluorenyl etc. Most preferred is phenyl.
The above definition of aryl also applies if aryl is part of another (combined) group as for example in arylamino, aryloxy or arylalkyl.
If the free valency of an aryl is saturated, then an arene is obtained.
The term arylene can also be derived from the previously defined aryl. Arylene, unlike aryl, is bivalent and requires two binding partners. Formally, the second valency is formed by removing a hydrogen atom from an aryl. Corresponding groups are for example:
The above definition for arylene also applies if arylene is part of another (combined) group as for example in HO-aryleneamino or H2N-aryleneoxy.
Heterocyclyl denotes ring systems, which are derived from the previously defined cycloalkyl, cycloalkenyl and aryl by replacing one or more of the groups —CH2— independently of one another in the hydrocarbon rings by the groups —O—, —S— or —NH— or by replacing one or more of the groups ═CH— by the group ═N—, wherein a total of not more than five heteroatoms may be present, at least one carbon atom must be present between two oxygen atoms and between two sulphur atoms or between an oxygen and a sulphur atom and the ring as a whole must have chemical stability. Heteroatoms may optionally be present in all the possible oxidation stages (sulphur→sulfoxide —SO—, sulphone —SO2—; nitrogen→N-oxide). In a heterocyclyl there is no heteroaromatic ring, i.e. no heteroatom is part of an aromatic system.
A direct result of the derivation from cycloalkyl, cycloalkenyl and aryl is that heterocyclyl is made up of the subgroups monocyclic heterocyclyl, bicyclic heterocyclyl, tricyclic heterocyclyl and spiro-heterocyclyl, which may be present in saturated or unsaturated form.
By unsaturated is meant that there is at least one double bond in the ring system in question, but no heteroaromatic system is formed. In bicyclic heterocyclyl two rings are linked together so that they have at least two (hetero)atoms in common. In spiro-heterocyclyl one carbon atom (spiroatom) belongs to two rings together.
If a heterocyclyl is substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon and/or nitrogen atoms. Heterocyclyl itself may be linked as a substituent to the molecule via every suitable position of the ring system. Substituents on heterocyclyl do not count for the number of members of a heterocyclyl.
Examples of heterocyclyl are tetrahydrofuryl, pyrrolidinyl, pyrrolinyl, imidazolidinyl, thiazolidinyl, imidazolinyl, pyrazolidinyl, pyrazolinyl, piperidinyl, piperazinyl, oxiranyl, aziridinyl, azetidinyl, 1,4-dioxanyl, azepanyl, diazepanyl, morpholinyl, thiomorpholinyl, homomorpholinyl, homopiperidinyl, homopiperazinyl, homothiomorpholinyl, thiomorpholinyl-S-oxide, thiomorpholinyl-S,S-dioxide, 1,3-dioxolanyl, tetrahydropyranyl, tetrahydrothiopyranyl, [1,4]-oxazepanyl, tetrahydrothienyl, homothiomorpholinyl-S,S-dioxide, oxazolidinonyl, dihydropyrazolyl, dihydropyrrolyl, dihydropyrazinyl, dihydropyridyl, dihydro-pyrimidinyl, dihydrofuryl, dihydropyranyl, tetrahydrothienyl-S-oxide, tetrahydrothienyl-S,S-dioxide, homothiomorpholinyl-S-oxide, 2,3-dihydroazet, 2H-pyrrolyl, 4H-pyranyl, 1,4-dihydropyridinyl, 8-aza-bicyclo[3.2.1]octyl, 8-aza-bicyclo[5.1.0]octyl, 2-oxa-5-azabicyclo[2.2.1]heptyl, 8-oxa-3-aza-bicyclo[3.2.1]octyl, 3,8-diaza-bicyclo[3.2.1]octyl, 2,5-diaza-bicyclo[2.2.1]heptyl, 1-aza-bicyclo[2.2.2]octyl, 3,8-diaza-bicyclo[3.2.1]octyl, 3,9-diaza-bicyclo[4.2.1]nonyl, 2,6-diaza-bicyclo[3.2.2]nonyl, 1,4-dioxa-spiro[4.5]decyl, 1-oxa-3,8-diaza-spiro[4.5]decyl, 2,6-diaza-spiro[3.3]heptyl, 2,7-diaza-spiro[4.4]nonyl, 2,6-diaza-spiro[3.4]octyl, 3,9-diaza-spiro[5.5]undecyl, 2,8-diaza-spiro[4,5]decyl etc.
Further examples are the structures illustrated below, which may be attached via each hydrogen-carrying atom (exchanged for hydrogen):
Preferred monocyclic heterocyclyl is 4 to 7 membered and has one or two heteroatoms independently selected from oxygen, nitrogen and sulfur.
Preferred monocyclic heterocyclyls are: piperazinyl, piperidinyl, morpholinyl, pyrrolidinyl, and azetidinyl.
Preferred bicyclic heterocyclyl is 6 to 10 membered and has one or two heteroatoms independently selected from oxygen, nitrogen and sulfur.
Preferred tricyclic heterocyclyl is 9 membered and has one or two heteroatoms independently selected from oxygen, nitrogen and sulfur.
Preferred spiro-heterocyclyl is 7 to 11 membered and has one or two heteroatoms independently selected from oxygen, nitrogen and sulfur.
The above definition of heterocyclyl also applies if heterocyclyl is part of another (combined) group as for example in heterocyclylamino, heterocyclyloxy or heterocyclylalkyl.
If the free valency of a heterocyclyl is saturated, then a heterocycle is obtained.
The term heterocyclylene is also derived from the previously defined heterocyclyl. Heterocyclylene, unlike heterocyclyl, is bivalent and requires two binding partners. Formally, the second valency is obtained by removing a hydrogen atom from a heterocyclyl. Corresponding groups are for example:
The above definition of heterocyclylene also applies if heterocyclylene is part of another (combined) group as for example in HO-heterocyclyleneamino or H2N-heterocyclyleneoxy.
Heteroaryl denotes monocyclic heteroaromatic rings or polycyclic rings with at least one heteroaromatic ring, which compared with the corresponding aryl or cycloalkyl (cycloalkenyl) contain, instead of one or more carbon atoms, one or more identical or different heteroatoms, selected independently of one another from among nitrogen, sulphur and oxygen, wherein the resulting group must be chemically stable. The prerequisite for the presence of heteroaryl is a heteroatom and a heteroaromatic system.
If a heteroaryl is to be substituted, the substitutions may take place independently of one another, in the form of mono- or polysubstitutions in each case, on all the hydrogen-carrying carbon and/or nitrogen atoms. Heteroaryl itself may be linked as a substituent to the molecule via every suitable position of the ring system, both carbon and nitrogen. Substituents on heteroaryl do not count for the number of members of a heteroaryl.
Examples of heteroaryl are furyl, thienyl, pyrrolyl, oxazolyl, thiazolyl, isoxazolyl, isothiazolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, oxadiazolyl, thiadiazolyl, pyridyl, pyrimidyl, pyridazinyl, pyrazinyl, triazinyl, pyridyl-N-oxide, pyrrolyl-N-oxide, pyrimidinyl-N-oxide, pyridazinyl-N-oxide, pyrazinyl-N-oxide, imidazolyl-N-oxide, isoxazolyl-N-oxide, oxazolyl-N-oxide, thiazolyl-N-oxide, oxadiazolyl-N-oxide, thiadiazolyl-N-oxide, triazolyl-N-oxide, tetrazolyl-N-oxide, indolyl, isoindolyl, benzofuryl, benzothienyl, benzoxazolyl, benzothiazolyl, benzisoxazolyl, benzisothiazolyl, benzimidazolyl, indazolyl, isoquinolinyl, quinolinyl, quinoxalinyl, cinnolinyl, phthalazinyl, quinazolinyl, benzotriazinyl, indolizinyl, oxazolopyridyl, imidazopyridyl, naphthyridinyl, benzoxazolyl, pyridopyridyl, pyrimidopyridyl, purinyl, pteridinyl, benzothiazolyl, imidazopyridyl, imidazothiazolyl, quinolinyl-N-oxide, indolyl-N-oxide, isoquinolyl-N-oxide, quinazolinyl-N-oxide, quinoxalinyl-N-oxide, phthalazinyl-N-oxide, indolizinyl-N-oxide, indazolyl-N-oxide, benzothiazolyl-N-oxide, benzimidazolyl-N-oxide etc.
Further examples are the structures illustrated below, which may be attached via each hydrogen-carrying atom (exchanged for hydrogen):
Preferably, heteroaryls are 5-6 membered monocyclic or 9-10 membered bicyclic, each with 1 to 4 heteroatoms independently selected from oxygen, nitrogen and sulfur.
The above definition of heteroaryl also applies if heteroaryl is part of another (combined) group as for example in heteroarylamino, heteroaryloxy or heteroarylalkyl.
If the free valency of a heteroaryl is saturated, a heteroarene is obtained.
The term heteroarylene is also derived from the previously defined heteroaryl. Heteroarylene, unlike heteroaryl, is bivalent and requires two binding partners. Formally, the second valency is obtained by removing a hydrogen atom from a heteroaryl. Corresponding groups are for example:
The above definition of heteroarylene also applies if heteroarylene is part of another (combined) group as for example in HO-heteroaryleneamino or H2N-heteroaryleneoxy.
By substituted is meant that a hydrogen atom which is bound directly to the atom under consideration, is replaced by another atom or another group of atoms (substituent). Depending on the starting conditions (number of hydrogen atoms) mono- or polysubstitution may take place on one atom. Substitution with a particular substituent is only possible if the permitted valencies of the substituent and of the atom that is to be substituted correspond to one another and the substitution leads to a stable compound (i.e. to a compound which is not converted spontaneously, e.g. by rearrangement, cyclisation or elimination).
Bivalent substituents such as ═S, ═NR, ═NOR, ═NNRR, =NN(R)C(O)NRR, =N2 or the like, may only be substituents on carbon atoms, whereas the bivalent substituents ═O and =NR may also be a substituent on sulphur. Generally, substitution may be carried out by a bivalent substituent only at ring systems and requires replacement of two geminal hydrogen atoms, i.e. hydrogen atoms that are bound to the same carbon atom that is saturated prior to the substitution. Substitution by a bivalent substituent is therefore only possible at the group —CH2— or sulphur atoms (═O group or =NR group only, one or two ═O groups possible or, e.g., one ═O group and one ═NR group, each group replacing a free electron pair) of a ring system.
Isotopes: It is to be understood that all disclosures of an atom or compound of the invention include all suitable isotopic variations. In particular, a reference to hydrogen also includes deuterium.
Stereochemistry/solvates/hydrates: Unless specifically indicated, throughout the specification and appended claims, a given chemical formula or name shall encompass tautomers and all stereo, optical and geometrical isomers (e.g. enantiomers, diastereomers, E/Z isomers, etc.) and racemates thereof as well as mixtures in different proportions of the separate enantiomers, mixtures of diastereomers, or mixtures of any of the foregoing forms where such isomers and enantiomers exist, as well as salts, including pharmaceutically acceptable salts thereof and solvates thereof such as for instance hydrates including solvates and hydrates of the free compound or solvates and hydrates of a salt of the compound.
In general, substantially pure stereoisomers can be obtained according to synthetic principles known to a person skilled in the field, e.g. by separation of corresponding mixtures, by using stereochemically pure starting materials and/or by stereoselective synthesis. It is known in the art how to prepare optically active forms, such as by resolution of racemic forms or by synthesis, e.g. starting from optically active starting materials and/or by using chiral reagents.
Enantiomerically pure compounds of this invention or intermediates may be prepared via asymmetric synthesis, for example by preparation and subsequent separation of appropriate diastereomeric compounds or intermediates which can be separated by known methods (e.g. by chromatographic separation or crystallization) and/or by using chiral reagents, such as chiral starting materials, chiral catalysts or chiral auxiliaries.
Further, it is known to the person skilled in the art how to prepare enantiomerically pure compounds from the corresponding racemic mixtures, such as by chromatographic separation of the corresponding racemic mixtures on chiral stationary phases, or by resolution of a racemic mixture using an appropriate resolving agent, e.g. by means of diastereomeric salt formation of the racemic compound with optically active acids or bases, subsequent resolution of the salts and release of the desired compound from the salt, or by derivatization of the corresponding racemic compounds with optically active chiral auxiliary reagents, subsequent diastereomer separation and removal of the chiral auxiliary group, or by kinetic resolution of a racemate (e.g. by enzymatic resolution); by enantioselective crystallization from a conglomerate of enantiomorphous crystals under suitable conditions, or by (fractional) crystallization from a suitable solvent in the presence of an optically active chiral auxiliary.
Salts: The phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgement, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication and commensurate with a reasonable benefit/risk ratio.
As used herein “pharmaceutically acceptable salts” refers to derivatives of the disclosed compounds wherein the parent compound is modified by making acid or base salts thereof. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like.
For example, such salts include salts from benzenesulfonic acid, benzoic acid, citric acid, ethanesulfonic acid, fumaric acid, gentisic acid, hydrobromic acid, hydrochloric acid, maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonic acid, 4-methyl-benzenesulfonic acid, phosphoric acid, salicylic acid, succinic acid, sulfuric acid and tartaric acid.
Further pharmaceutically acceptable salts can be formed with cations from ammonia, L-arginine, calcium, 2,2′-iminobisethanol, L-lysine, magnesium, N-methyl-D-glucamine, potassium, sodium and tris(hydroxymethyl)-aminomethane.
The pharmaceutically acceptable salts of the present invention can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base form of these compounds with a sufficient amount of the appropriate base or acid in water or in an organic diluent like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile, or a mixture thereof.
Salts of other acids than those mentioned above which for example are useful for purifying or isolating the compounds of the present invention (e.g. trifluoro acetate salts), also comprise a part of the invention.
In a representation such as for example
the letter A has the function of a ring designation in order to make it easier, for example, to indicate the attachment of the ring in question to other rings.
For bivalent groups in which it is crucial to determine which adjacent groups they bind and with which valency, the corresponding binding partners are indicated in brackets where necessary for clarification purposes, as in the following representations:
or (R2)—C(═O)NH— or (R2)—NHC(═O)—.
If such a clarification is missing then the bivalent group can bind in both directions, i.e., e.g., —C(═O)NH—also includes —NHC(═O)— (and vice versa).
Groups or substituents are frequently selected from among a number of alternative groups/substituents with a corresponding group designation (e.g. Ra, Rb etc). If such a group is used repeatedly to define a compound according to the invention in different parts of the molecule, it is pointed out that the various uses are to be regarded as totally independent of one another.
By a therapeutically effective amount for the purposes of this invention is meant a quantity of substance that is capable of obviating symptoms of illness or of preventing or alleviating these symptoms, or which prolong the survival of a treated patient.
Features and advantages of the present invention will become apparent from the following detailed examples which illustrate the principles of the invention by way of example without restricting its scope:
Unless stated otherwise, all the reactions are carried out in commercially obtainable apparatus using methods that are commonly used in chemical laboratories. Starting materials that are sensitive to air and/or moisture are stored under protective gas and corresponding reactions and manipulations therewith are carried out under protective gas (nitrogen or argon).
If a compound is to be represented both by a structural formula and by its nomenclature, in the event of a conflict the structural formula is decisive.
Microwave reactions are carried out in an initiator/reactor made by Biotage or in an Explorer made by CEM or in Synthos 3000 or Monowave 3000 made by Anton Paar in sealed containers (preferably 2, 5 or 20 mL), preferably with stirring.
The thin layer chromatography is carried out on ready-made silica gel 60 TLC plates on glass (with fluorescence indicator F-254) made by Merck.
The preparative high pressure chromatography (RP HPLC) of the example compounds according to the invention is carried out on Agilent or Gilson systems with columns made by Waters (names: SunFire™ Prep C18, OBD™ 10 μm, 50×150 mm or SunFire™ Prep 018 OBD™ 5 μm, 30×50 mm or XBridge™ Prep C18, OBD™ 10 μm, 50×150 mm or XBridge™ Prep C18, OBD™ 5 μm, 30×150 mm or XBridge™ Prep C18, OBD™ 5 μm, 30×50 mm) and YMC (names: Actus-Triart Prep C18, 5 μm, 30×50 mm).
Different gradients of H2O/ACN are used to elute the compounds, while for Agilent systems 5% acidic modifier (20 mL HCOOH to 1 L H2O/ACN (1/1)) is added to the water (acidic conditions). For Gilson systems the water is added 0.1% HCOOH.
For the chromatography under basic conditions for Agilent systems H2O/ACN gradients are used as well, while the water is made alkaline by addition of 5% basic modifier (50 g NH4HCO3+50 mL NH3 (25% in H2O) to 1 L with H2O). For Gilson systems the water is made alkaline as follows: 5 mL NH4HCO3 solution (158 g in 1 L H2O) and 2 mL NH3 (28% in H2O) are replenished to 1 L with H2O.
The supercritical fluid chromatography (SFC) of the intermediates and example compounds according to the invention is carried out on a JASCO SFC-system with the following columns: Chiralcel OJ (250×20 mm, 5 μm), Chiralpak AD (250×20 mm, 5 μm), Chiralpak AS (250×20 mm, 5 μm), Chiralpak IC (250×20 mm, 5 μm), Chiralpak IA (250×20 mm, 5 μm), Chiralcel OJ (250×20 mm, 5 μm), Chiralcel OD (250×20 mm, 5 μm), Phenomenex Lux C2 (250×20 mm, 5 μm).
The analytical HPLC (reaction control) of intermediate and final compounds is carried out using columns made by Waters (names: XBridge™ C18, 2.5 μm, 2.1×20 mm or XBridge™ C18, 2.5 μm, 2.1×30 mm or Aquity UPLC BEH C18, 1.7 μm, 2.1×50 mm) and YMC (names: Triart C18, 3.0 μm, 2.0×30 mm) and Phenomenex (names: Luna C18, 5.0 μm, 2.0×30 mm). The analytical equipment is also equipped with a mass detector in each case.
The retention times/MS-ESI+ for characterizing the example compounds according to the invention are produced using an HPLC-MS apparatus (high performance liquid chromatography with mass detector). Compounds that elute at the injection peak are given the retention time tRet.=0.00.
The compounds according to the invention and intermediates are prepared by the methods of synthesis described hereinafter in which the substituents of the general formulae have the meanings given hereinbefore. These methods are intended as an illustration of the invention without restricting its subject matter and the scope of the compounds claimed to these examples. Where the preparation of starting compounds is not described, they are commercially obtainable or their synthesis is described in the prior art or they may be prepared analogously to known prior art compounds or methods described herein, i.e. it is within the skills of an organic chemist to synthesize these compounds. Substances described in the literature can be prepared according to the published methods of synthesis. If a chemical structure in the following is depicted without exact configuration of a stereo center, e.g. of an asymmetrically substituted carbon atom, then both configurations shall be deemed to be included and disclosed in such a representation. The representation of a stereo center in racemic form shall always deem to include and disclose both enantiomers (if no other defined stereo center exists) or all other potential diastereomers and enantiomers (if additional, defined or undefined, stereo centers exist).
To a suspension of 5-chloropentanenitrile (22.9 g, 195 mmol, 1.00 equiv.) in EtOH (136 mL) is added acetyl chloride (111 mL, 1.56 mol, 8.00 equiv.) dropwise at 0° C. The reaction mixture is allowed to warm to rt and stirred for 12 h. The mixture is concentrated under reduced pressure and washed with Et2O and the crude product A-2a is used as the HCl salt directly in the next step without further purification (HPLC method: A; tret=1.03 min; [M+H]+=164).
Crude A-2a (HCl salt) (28.0 g, 140 mmol, 1.00 equiv.) and ethylene glycol (7.38 g, 119 mmol, 0.90 equiv.) are dissolved in DCM (300 mL) and stirred at rt for 6 d. The resulting suspension is concentrated under reduced pressure, diluted with Et2O (200 mL) and filtered. The filtrate is concentrated under reduced pressure, taken up in DCM (200 mL) and treated with a KOH solution (2 M in water, 150 mL). The mixture is stirred at rt overnight keeping the phases intact. The phases are separated, the water phase is extracted with DCM (2×) and the combined organic phases are dried over magnesium sulfate, filtered and concentrated under reduced pressure. The crude orthoester A-3a is used for the next step without further purification (HPLC method: A; tret=1.37 min; [M+H]+=163).
Crude A-3a (22.3 g, 107 mmol, 1.00 equiv.), 1-cyclohexenyloxytrimethylsilane (16.4 mL, 82.3 mmol, 0.80 equiv.) and zinc chloride (10.2 g, 74.8 mmol, 0.70 equiv.) are dissolved in DCM (120 mL) and stirred at rt for 5 h. The reaction mixture is treated by addition of sat. sodium bicarbonate solution. The organic phase is separated, dried over magnesium sulfate, filtered and concentrated under reduced pressure. The crude product is purified by NP-chromatography to give the desired compound A-4a (HPLC method: A; tret=1.25 min; [M+H]+=283).
A-4a (14.9 g, 57.1 mmol, 1.00 equiv.) and sodium iodide (25.9 g, 171 mmol, 3.00 equiv.) are dissolved in acetone (120 mL) and stirred under reflux for 16 h. The reaction mixture is concentrated under reduced pressure, diluted with DCM and washed with a sat. sodium thiosulfate solution. The organic phase is separated, dried over magnesium sulfate, filtered and concentrated under reduced pressure. The crude product A-5a is used for the next step without further purification.
A-5a (30.0 g, 85.0 mmol, 1.00 equiv.) is dissolved in THF. The mixture is treated with potassium tert-butoxide (28.7 g, 256 mmol, 3.0 equiv.) at 0° C. and stirred at rt overnight. The reaction mixture is quenched by addition of water (2 mL), diluted by addition of Et2O and sat. sodium hydrogencarbonate solution. The organic phase is separated, dried over magnesium sulfate, filtered and concentrated under reduced pressure. The crude product is purified by NP-chromatography to give (racemic) compound A-6a (HPLC method: A; tret=1.17 min; [M+H]+=225).
Reaction sequence A-1a→A-6a is based on Marko et al., THL 2003, 44, 3333-3336 and Maulide et al., Eur. J. Org. Chem. 2004, 19:3962-3967.
Enantiomer A-6b can then be obtained after chiral separation via SFC (using a Lux Cellulose-4 column (250×30 mm, 5 μm), 30° C. column temperature, 90% CO2, 10% ACN as cosolvent) with enantiomer A-6b (HPLC method: A; tret=1.17 min; [M+H]+=225/SFC method: SFC-1; tret=2.99 min) eluting as the 2nd peak after the other enantiomer.
A dry and clean reactor is charged with toluene (234 L) under nitrogen (note: 2.5 V toluene in total for this reaction). Water (1.56 kg, 85.5 mol, keep H2O:Pd=160:1) is added followed by rinsing the charging line with 1,1,3,3-tetramethylguanidine (175.5 kg, 1527.6 mol, 2.0 equiv.) under nitrogen and then toluene (13 L). A-7a (130.0 kg, 763.8 mol) is added under nitrogen followed by rinsing with toluene (13 L). Allyl acetate (98.8 kg, 992.9 mol, 1.3 equiv.) is added under nitrogen and rinsed with toluene (13 L). Under agitation, the mixture is cooled to 10° C. in 0.5 h. The batch is degassed by sparging the solution with nitrogen for ˜30 min. (S,S)-DACH-Ph Trost ligand (0.429 kg, 0.619 mol, 0.081 mol %) in degassed toluene (13 L) (note: keep Pd:ligand=1:1.15) is added followed by rinsing with degassed toluene (13 L). Allylpalladium(II) chloride dimer (97.5 g, 267 mol, 0.035 mol %) in degassed toluene (13 L) is added followed by rinsing with degassed toluene (13 L). The batch is kept at 10-15° C. at least 8 h. After the reaction is complete by HPLC, a solution of N-acetyl-L-cysteine (3.9 kg, 22.9 mol, 0.03 equiv.) in water (260 L) below 25° C. is added. The resulting solution is warmed to 20-25° C. and kept at 20-25° C. at least for 1 h. After phase cut to discard the bottom aq. layer, 10 wt % NH4Cl aq. solution (260 L) is added. After the mixture is agitated for 10 min, the bottom aq. layer is drained. The organic phase is further washed with water (130 L). The organic layer is filtered through a very short pad of Celite® and the reactor and Celite® bed is rinsed with toluene (65 L). The filtrate is charged into a clean reactor and then toluene is distilled off under vacuum at 40-50° C. The crude product is directly used for the next step or the product is drained into a container with the help of minimum amount of toluene (65 L) and stored at 20-23° C. 150 kg of A-8a is usually obtained as a light yellow oil in 96% yield with 90:10 enantomeric ratio (chiral method W).
1H NMR (500 MHz, CDCl3): δ 5.75 (ddt, J=14.8, 9.4, 7.5 Hz, 1H), 5.06-5.00 (m, 2H), 4.19 (q, J=7.1 Hz, 2H), 2.61 (dd, J=13.9, 7.1 Hz, 1H), 2.51-2.43 (m, 3H), 2.33 (dd, J=13.9, 7.9 Hz, 1H), 2.03-1.98 (m, 1H), 1.78-1.60 (m, 3H), 1.50-1.42 (m, 1H), 1.25 (t, J=7.1 Hz, 3H). 13C NMR (125 MHz, CDCl3): δ 207.7, 171.6, 133.5, 118.4, 61.3, 61.0, 41.3, 39.4, 35.9, 27.7, 22.6, 14.3. ESI-MS: m/z 211 [M+H]+.
To the reactor containing A-8a (150 kg, 713.4 mol) from step 1 (less than 1 V toluene if used) is added ethylene glycol (600 L) to give a yellow biphasic mixture. After the mixture is cooled to 10-15° C., TMSCI (193.5 kg, 1783.5 mol, 2.5 equiv.) is added over not less than 15 min, at a rate to maintain the internal temperature between 20-30° C. (orange biphasic mixture obtained). Sufficient agitation is needed to achieve mixing. After the batch is kept at 20-25° C. for 2 h, agitation is stopped and kept for at least 15 min at 20-25° C. The batch is cooled to 0-5° C. NaOH (96 kg, 1854.8 mol, 2.6 equiv.) in water (600 L) is added at a rate to maintain the internal temperature below 20° C. (light yellow cloudy biphasic mixture obtained). Toluene (300 L) is added and then the batch is agitated for 10 min. After phase cut to drain the bottom aq. layer (note: some precipitate may form at interphase), the organic layer is washed with water (300 L) two times. The organic phase is filtered through a short pad of Celite® to remove insoluble solids/interphase. The organic solution is charged into a clean and dry reactor and then the solvent is distilled off at 40-50° C. to a minimum stirrable volume. The crude product A-9a (189 kg, 95.2 wt %, 100% yield) is drained to a container with the help of minimum amount of toluene.
1H NMR (500 MHz, CDCl3): δ 5.65 (ddt, J=14.7, 8.1, 6.6 Hz, 1H), 5.07-4.98 (m, 2H), 4.20-4.10 (m, 2H), 3.97-3.88 (m, 4H), 2.81 (dd, J=13.9, 6.6 Hz, 1H), 2.35 (dd, J=13.9, 8.1 Hz, 1H), 2.04-1.98 (m, 1H), 1.75-1.35 (m, 7H), 1.26 (t, J=7.1 Hz, 3H). 13C NMR (125 MHz, CDCl3): δ 173.7, 134.3, 117.6, 110.9, 65.0, 64.7, 60.5, 54.6, 36.2, 32.3, 30.3, 23.3, 20.9, 14.4. ESI-MS: m/z 255 [M+H]+.
To a dry and clean reactor is added 9-BBN (688.5 kg, 401 mol, 1.2 equiv.) under nitrogen. The solution is cooled to 0-5° C. to obtain a slurry. A-9a (85.0 kg, 334.2 mol) from step 2 is added at 0-5° C. and rinsed with THF (40 L). The mixture is warmed to 20-23° C. in 1 h and kept at 20-23° C. for not less than 1 h. After that the mixture is cooled to −45 to −40° C., methyl chloroacetate (69.6 kg, 1.3 equiv) is added in one portion followed by dropwise addition of LiHMDS (909.5 kg, 1102.9 mol) while keeping temperature below −35° C. The batch is then warmed to 20-23° C. in 1 h and then kept at 20-23° C. at least for 18 h. ˜12-13 V solvent is removed by distillation under vacuum with heating (35° C.). EtOH (255 kg) is added followed by a solution of NaOH (13.4 kg) in H2O (212.5 L). The mixture is heated at reflux (at 66-70° C.) for at least 14 h.
After that ˜5-6 V solvent is removed by distillation at reflux, the batch is cooled to 20-25° C. and then filtered through a short pad of Celite® to remove insoluble material and rinsed with heptane (160 L). ˜5-6 V solvent (or most of the residual THF and ethanol) is distilled under vacuum at 40-50° C. The batch is cooled to 20-25° C. After that, water (255 L) is added, the crude product is extracted twice with heptane (2364.8 kg). The combined heptane layers are washed once with water (85 L). After solvent removal by distillation under vacuum at 40-50° C., the crude product (52.7 kg, 87.5 wt %) is obtained as a yellow oil in 52.6% assay yield with 90:10 enantiomeric ratio (method: SFC-3). The crude product A-6b is used for next step directly.
1H NMR (500 MHz, CDCl3): δ 4.01-3.82 (m, 4H), 2.50-2.44 (m, 1H), 2.382.34 (m, 1H), 2.28-2.22 (m, 1H), 2.11-2.05 (m, 1H), 2.01-1.95 (m, 1H), 1.92-1.86 (m, 1H), 1.81-1.58 (m, 6H), 1.54-1.43 (m, 3H), 1.27-1.18 (m, 1H). ESI-MS: m/z 225 [M+H]+.
A dry and clean reactor is charged with LiHMDS (1 M in THF) (406.4 kg, 456.1 mol, 1.1 equiv.). The solution is cooled to 0-5° C., crude A-6b (93.0 kg, 414.6 mol) is charged below 5° C. and rinsed with THF (46.5 kg). After 30 min, at 0-5° C., diethyl oxalate (72.5 kg, 497.5 mol, 1.2 equiv.) is added at below 5° C. After the reaction mixture is warmed to 20-25° C. in 1 h, the mixture is kept at 20-25° C. for not less than 3 h. After the reaction mixture is cooled to 10-15° C., cooled HCl solution [prepared by adding acetyl chloride (73.6 kg, 932.9 mol, 2.25 equiv.) in EtOH (293.9 kg) at 0-5° C.] is added below 25° C. to reach final pH ˜6-7 of the yellow slurry. Solid NH2OH·HCl (28.8 kg, 414.4 mol, 1.05 equiv.) is added in one portion and the resulting mixture is heated to reflux for 6-10 h. The reaction mixture is concentrated under reflux at 66-70° C. and residuals of THF by coevaporation with EtOH (73.5 kg). The residue is taken up in water (372.0 kg) and EtOH (293.9 kg). After 3-6 h at 70-75° C., the mixture is cooled to 30-35° C. and 0.5-1% A-10a crystals are seeded. After 2-4 h at 30-35° C., heptane (63.2 kg) is added in not less than 1 h. After 60 min at 20-25° C., water (279.0 kg) is added over 4-6 h. After 1 h at 20-25° C., the solid is collected and washed with 1:2 EtOH/water (51.2 kg EtOH and 130.2 kg water) and then heptane (63.2 kg) two times. The solid is dried under vacuum under nitrogen stream to give the product (93.0 kg) with 65% yield.
1H NMR (500 MHz, CDCl3): δ 4.42 (q, J=7.1 Hz, 2H), 2.73 (dt, J=16.8, 5.1 Hz, 1H), 2.64 (dt, J=14.3, 6.0 Hz, 1H), 2.60-2.51 (m, 2H), 2.43-2.30 (m, 2H), 2.09-1.96 (m, 3H), 1.91-1.81 (m, 3H), 1.76-1.67 (m, 1H), 1.65-1.58 (m, 1H), 1.40 (t, J=7.1 Hz, 3H). ESI-MS: m/z 278 [M+H]+.
A dry and clean reactor is charged with A-10a (72.0 kg, 259.6 mol), EtOH (56.9 kg) and NH4OH (aq) (280.8 kg). The reaction mixture is kept at 20-25° C. for not less than 16 h. After water (144.0 kg) is added over 30 min, the slurry is kept at 20-25° C. for 30 min. The solid is collected by filtration, washed with 1:3 EtOH/water (28.5 kg EtOH and 108 kg water) and heptane (97.9 kg). After dried under vacuum for 1 h at 23° C., the solid is dried under vacuum at 50-55° C. overnight to give the product (61.4 kg, 87.2% yield) with ≥95:5 er (254 nm) and water content ≤0.5% based on Karl Fischer titration.
A dry and clean reactor is charged crude A-11a (60.0 kg, 1.0 equiv.), 1,4-dioxane (240.0 kg) and activated carbon (3.0 kg, 5 wt %). The reaction mixture is stirred at 55-65° C. for 2-4 h. After filtration at high temperature (55-65° C.), the filter cake is washed with 1,4-dioxane (33.0 kg). The filtrate is transferred into a clean reactor. The temperature is adjusted to 45-55° C. and stirred at 45-55° C. for 1-2 h. Water (240.0 kg) is added over 2 h. The temperature is adjusted to 45-55° C. and stirred at 45-55° C. for 1-2 h. The reaction mixture is cooled to 35-45° C. and stirred at 35-45° C. for 2-4 h. Water (87.0 kg) is charged over 4 h. The mixture is cooled to 15-25° C. and stirred at 15-25° C. for 12-14 h. The solid is collected by centrifuge, washed with water (120.0 kg) and dried under vacuum at 50-55° C. overnight to give the product (44.8 kg, 71% yield) as a light yellow to off-white solid.
1H NMR (500 MHz, DMSO-d6): δ 7.99 (s, 1H), 7.71 (s, 1H), 2.80-2.69 (m, 1H), 2.60-2.53 (m, 1H), 2.50-2.42 (m, 1H), 2.40-2.28 (m, 2H), 2.26-2.18 (m, 1H), 2.05-1.70 (m, 7H), 1.48-1.39 (m, 1H). ESI-MS: m/z 249 [M+H]+.
A dry and clean reactor is charged with A-11a (40.0 kg, 161.1 mol), MeCN (96.0 kg) and pyridine (30.8 kg, 386.6 mol, 2.4 equiv.). After the mixture is cooled to 0-5° C., TFAA (40.8 kg, 193.3 mol, 1.2 equiv.) is added slowly at below 5° C. After 5 min at 0-5° C., water (120.0 kg) is added over 30 min at 0-5° C. and seeded with 0.5% A-12a crystals. After 15 min at 0-5° C., water (120.0 kg) is added over 30 min. After 30 min at 0-5° C. for 30 min, the solid is collected by filtration, washed with 1:3 MeCN/water (15.6 acetonitrile and 60.0 kg water) and then water (80.0 kg). The solid is dried under vacuum to give crude product A-12a (33.0 kg, 93.6% yield) as a tan solid. The crude product (32.5 kg, 1.0 equiv.) is dissolved in MTBE (48.1 kg), the slurry is agitated at 20-25° C. for 30 min. Heptane (132.6 kg) is added over 1 h. After 30 min at 20-25° C., the solid is collected, dried under vacuum to give the desired A-12a (26.6 kg, 82% yield) as a white solid with >99:1 er (254 nm, method: SFC-4) and >98 A % (220 nm) purity.
1H NMR (500 MHz, DMSO-d6): δ 2.83-2.73 (m, 1H), 2.60-2.40 (m, 3H), 2.34-2.20 (m, 2H), 2.06-1.75 (m, 7H), 1.53-1.43 (m, 1H). ESI-MS: m/z 231 [M+H]+.
A dry and clean reactor is charged with 9-BBN (387 mL, 193.5 mmol, 1.2 equiv., 0.5 M in THF) under nitrogen. The solution is cooled to 0-5° C. to obtain a slurry. A-9a (41.0 g, 161.2 mmol) is added at 0-5° C. and rinsed with THF (20.5 mL). The mixture is warmed to 20-23° C. in 1 h and kept at 20-23° C. for not less than 1 h. After the mixture is cooled to −45 to −40° C., methyl chloroacetate is added in one portion followed by addition of dropwise LiHMDS (355 mL, 532.0 mmol, 3.3 equiv.) while keeping temperature below −35° C. The reaction mixture is then warmed to 20-23° C. in 1 h and then kept at 20-23° C. at least for 18 h. The reaction mixture is cooled to 5-10° C., AcOH (30.4 mL, 3.3 equiv.) is added below 20° C. followed by water (41 mL) below 20° C. AcOH (30.4 mL, 3.3 equiv.) is charged below 20° C. to reach pH ˜6-7. ˜15-16 V of THF is removed under vacuum below 35° C. MTBE (246 mL) and water (205 mL) is added. After phase separation (discard of the bottom aq. layer) the organic layer is cooled to 0-5° C., a solution of sodium percarbonate (37.2 g, 322.4 mmol, 2.0 equiv.) in water (320 mL) is added below 20° C. After 1 h at 20-23° C., 20 wt % sodium sulfite solution (31 mL) is added. After 15 min at 20-23° C., the bottom aq. layer is separated and discarded. The organic layer is washed with 5 wt % ammonium chloride solution (123 mL) and water (328 mL). The organic layer is treated with 5% activated carbon for 30 min prior to filtration. After ˜4-5 V of solvent is removed under vacuum below 35° C. to give the crude product A-13a (80% yield) as orange-brown oil.
A reactor is charged with A-13a (45.5 g, 161.2 mmol), Ethanol (91.0 mL), NaOAc (39.7 g, 483.6 mmol, 3.0 equiv.), water (45.5 mL) and NH2OH·HCl (33.6 g, 483.6 mmol, 3.0 equiv.). The mixture is heated at 73-78° C. for not less than 16 h. After the batch is cooled to 20-23° C., water (227.6 mL) is added over 0.5 h. MTBE (136.5 mL) is added over 0.5 h followed by heptane (113.8 mL) over 1 h. After 0.5 h at 20-23° C., the solid is collected by filtration. The solid is washed successively with MTBE (45 mL) and water (91.0 mL). The solid is dried under vacuum to give the product A-14a as an off-white solid (18.27 g, 93.2 wt %) in 40% yield. 1H NMR (500 MHz, DMSO-d6): δ 11.48 (br s, 1H), 4.04 (q, J=6.0 Hz, 1H), 3.89-3.80 (m, 2H), 3.60 (q, J=6.8 Hz, 1H), 2.10-1.95 (m, 2H), 1.92-1.76 (m, 4H), 1.69-1.39 (m, 8H). ESI-MS: m/z 266 [M+H]+.
A clean reactor is charged with A-14a (100.0 g, 376.9 mmol, 1.0 equiv.), K3PO4 (240.0 g; 1130.8 mmol; 3.0 equiv.) in (499.0 g; 500.0 mL) and toluene (432.5 g; 500.0 mL). The bi-phase mixture is agitated. After the mixture is cooled to 0-5° C., Tf2O (186.0 g; 110.9 mL; 659.6 mmol; 1.750 equiv.) is added with syringe pump over 2 h below 5° C. After phase separation, the organic layer is filtered through a Celite® bed with Na2SO4. After rinse with toluene (50 mL), the crude product A-15a (149.8 g, 100% yield) is used for next step without further purification.
1H NMR (500 MHz, CDCl3): δ 3.95-3.91 (m, 3H), 3.77-3.74 (m, 1H), 2.51-2.44 (m, 2H), 2.16-1.80 (m, 4H), 1.77-1.48 (m, 8H). ESI-MS: m/z 398 [M+H]+.
Lithium chloride (4.95 g, 116.7 mmol, 3.5 equiv.) is predried in a flask at 50° C. under vacuum. A-15a (13.25 g, 33.34 mmol, 1.0 equiv.) is dissolved in dry dioxane (70 mL) and hexamethylditin (8.47 mL, 40.01 mmol, 1.2 equiv.), tetrakis(triphenylphosphine)palladium(0) (1.95 g, 1.67 mmol, 0.05 equiv.), and LiCl are added. The mixture is stirred for 3 h at 100° C. After complete conversion the mixture is diluted with DCM and the crude product is purified via NP followed by RP chromatography yielding A-16a (HPLC method: A; tret=1.67 min; [M+H]+=414).
Pd(OAc)2 (10.17 g, 45.3 mmol, 0.02 equiv.), BINAP (28.21 g, 45.3 mol, 0.02 equiv.), EtOH (1040 g, 22.65 mol, 10 equiv.) and DIPEA (439.08 g, 3.40 mol, 592 mL, 1.5 equiv.) are added to a mixture of A-15-a (900 g, 2.26 mol, 1 equiv.) in 2-MeTHF (3600 mL) at 20° C. After that, the reactor is purged with nitrogen (100 psi) 2 times followed by two additional purges with CO (100 psi). The reactor is then pressurized to 200 psi under CO and the reaction is stirred for 36 h at 60° C. Next, the reaction mixture is filtered through a Celite pad and solvent is removed. The obtained residue is purified by column chromatography to deliver A-17a (SiO2, Petroleum ether/EtOAc=3:1, Rf=0.50). The desired product is confirmed via LCMS (Method A.1,tret: =0.456 min, m/z 322 [M+H]+).
Ethyl 3-oxothiane-2-carboxylate (324 g, 1721 mmol, 1 equiv.) and tBuOK (193 g, 1721 mmol, 1 equiv.) are added to THF (3240 mL) and then allyl bromide (208 g, 1721 mmol, 1 equiv.) is added drop-wised. The reaction is then brought to 70° C. and stirred for 4 h. After complete conversion of the starting materials the pH is adjusted to 5 with HCl (2M). Next, the reaction mixture is extracted with EtOAc (1000 mL) and the combined organic layers is washed with brine (500 mL×2), dried over Na2SO4, filtered and concentrated under reduced pressure. The obtained residue is purified by column chromatography (SiO2, Petroleum ether/EtOAc=10/1·Rf=0.60) to yield A-19a as a yellow oil.
1H NMR: (400 MHz, CDCl3): δ 5.75 (tdd, J=7.2, 10.0, 17.2 Hz, 1H), 5.16-5.05 (m, 2H), 4.34-4.19 (m, 2H), 3.04 (ddd, J=2.8, 11.6, 14.0 Hz, 1H), 2.82 (dd, J=7.2, 14.4 Hz, 1H), 2.62-2.33 (m, 6H), 1.29 (t, J=7.2 Hz, 3H)
A-19a (335 g, 1419 mmol, 1 equiv.) and 1,2-ethanediol (1296 mL) are dissolved in toluene (335 mL) followed by the addition of TMSCI (385 g, 3548 mmol, 2.5 equiv.). The reaction is then stirred at 50° C. for 16 h. The mixture is then cooled to 0-5° C. and a solution of NaOH (150 g, 2.5 equiv.) in H2O (1300 mL) is added. Next, toluene (500 mL) is also added and the mixture is agitated for 10 min. After that, the organic and the aq. phase are separated and the organic layer is washed with water. After removal of solvent under reduced pressure, the obtained residue is purified by column chromatography (SiO2, Petroleum ether/EtOAc=10/1, Rf=0.40) to yield A-20a as a yellow oil.
1H NMR: (400 MHz, CDCl3): δ 5.81 (tdd, J=7.2, 10.0, 17.2 Hz, 1H), 5.15-5.00 (m, 2H), 4.28-4.15 (m, 2H), 4.05-3.94 (m, 4H), 3.02-2.82 (m, 2H), 2.58-2.36 (m, 2H), 2.07-1.99 (m, 3H), 1.76-1.61 (m, 1H), 1.29 (t, J=7.2 Hz, 3H)
A-20a (30.0 g, 110 mmol, 1 equiv.) is added to a 0° C. solution of 9-BBN (0.5 M, 264 mL, 1.2 equiv.) in THF (15 mL). Next, the resulting mixture is allowed to warm up to 25° C. and it is stirred for 16 h. The solution is cooled down to −45° C., followed by drop-wise addition of ethyl chloroacetate (17.6 g, 143 mmol, 1.3 equiv.) and drop-wise addition of LiHMDS (1 M, 364 mL, 3.3 equiv.). The obtained mixture is again allowed to warm up to 25° C. and stirred for 16 h. After complete conversion the mixture is poured into sat. NH4Cl solution, extracted with EtOAc. The organic layers are then combined, washed with brine, dried over Na2SO4, filtered and the solvent is removed under reduced pressure. The obtained residue is then purified by column chromatography (SiO2, Petroleum ether/EtOAc=3/1·Rf=0.40) to yield A-21a as a yellow oil.
1H NMR: (400 MHz, CDCl3): δ 13.24 (s, 1H), 4.29-4.16 (m, 2H), 4.15-3.91 (m, 5H), 2.49-2.42 (m, 2H), 2.18-1.64 (m, 10H), 1.33-1.26 (m, 3H)
A-21a (180 g, 572 mmol, 1 equiv) is added to a mixture of H2O (562 mL) and EtOH (540 g, 11.7 mol, 20.47 eq) and the reaction mixture is cooled to 0° C. Next, NaOH (28.4 g, 709 mmol, 1.24 eq) is added to the and the temperature is raised to 75° C. The reaction is stirred for 2 h. After complete conversion of the starting material the reaction is cooled to room temperature and solvent is removed under reduced. The obtained crude is poured into water and extracted with EtOAc. The combined organic layers are washed with brine, dried over Na2SO4, filtered and concentrated under reduced pressure. The obtained residue is the purified by column chromatography (SiO2, Petroleum ether/EtOAc=2/1, Rf=0.50) to deliver A-22a as a yellow oil.
1H NMR: (400 MHz, CDCl3): δ 4.14-3.90 (m, 4H), 3.33-3.20 (m, 1H), 2.57-2.33 (m, 3H), 2.27-1.68 (m, 10H).
A dry and clean reactor is charged with LiHMDS (1 M in THF) (57 g, 439.9 nmol, 1.1 equiv.).
The solution is cooled to 0-5° C., A-22a (75.0 g, 309 mmol, 1.0 equiv) is charged below 5° C. and rinsed with THF (32 mL). After 30 min, at 0-5° C., diethyl oxalate (72.5 kg, 497.5 mol, 1.2 equiv.) is added at below 5° C. After the reaction mixture is stirred at 50° C. for 4 h. After the reaction mixture is cooled to 10-15° C., cooled HCl solution [prepared by adding acetyl chloride (54.7 g, 696 mmol, 2.25 eq) in EtOH (225 mL) at 0-5° C.] is added below 25° C. to reach final pH ˜6-7 of the yellow slurry. Solid NH2OH·HCl (22.6 g, 325 mmol, 1.05 eq) is added in one portion and the resulting mixture is heated to 75° C. for 10 h. Next, the reaction mixture is allowed to cool to room temperature and solvent is removed under reduced pressure. The left residue is then poured into water and extracted with EtOAc. The combined organic layers are washed with brine, dried over Na2SO4, filtered and concentrated under reduced pressure. The residue is then purified by column chromatography (SiO2, Petroleum ether/EtOAc=3/1·Rt=0.58) to deliver compound A-23a.
1H NMR: (400 MHz, CDCl3): δ 4.43 (q, J=7.2 Hz, 2H), 4.10-3.84 (m, 4H), 3.03-2.77 (m, 2H), 2.66-2.52 (m, 2H), 2.32-2.10 (m, 4H), 2.03-1.89 (m, 4H), 1.41 (t, J=7.2 Hz, 3H). ESI-MS: m/z 340 [M+H]+.
A mixture of A-17a (500 g, 1.56 mol, 1.00 equiv.) and NH3·H2O (2000 mL) in EtOH (1500 mL) is degassed and purged with N2 for 3 times at 15° C. The reaction is then stirred at 15° C. for 16 h and then solved is removed to give A-24a as a white solid.
Other intermediates A-24 (Table 1) are available in an analogous manner. The crude product is purified by chromatography if necessary.
HCl (2M, 18.4 mL, 36.7 mmol, 0.76 equiv) is added to a solution of A-24b (15.0 g, 48.3 mmol, 1 equiv.) in THF (55 mL) the reaction mixture is stirred for 18 h at 70° C. After complete conversion of the starting material the reaction solution is diluted with EtOAc, washed with NaHCO3 and brine and dried, filtered, and concentrated under reduced pressure to give the crude product, which is purified by column chromatography (SiO2, Petroleum ether/EtOAc=0/1, Rf=0.70) to obtain A-25a as a white solid.
1H NMR: (400 MHz, CDCl3): 6.71 (s, 1H), 5.87 (s, 1H), 3.65-3.50 (m, 1H), 3.01 (dt, J=4.4, 12.4 Hz, 1H), 2.85-2.43 (m, 7H), 1.99-1.79 (m, 2H), 1.70-1.60 (m, 1H)
A dry and clean reactor is charges with A-24a (720 g, 2.46 mol, 1.00 equiv.) in MeCN (2160 mL). After that, the mixture is cooled to 0-5° C. and pyridine (468 g, 5.91 mol, 2.4 equiv.) and TFAA (621 g, 2.96 mol, 1.2 equiv.) is added below 5° C. After stirring the reaction for 1 hour, 4 L of water is added bellow 15° C. The mixture is extracted with MTBE (4 L) and the organic phase is washed with 5% sodium bicarbonate solution (2 L). The combined organic layers are washed with brine 3000 mL, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. A-26a (1380 g, crude) is obtained as a colourless oil.
A-26b (Table 2) is available in an analogous manner by using A-25a as the starting material followed by SFC separation of the resulting racemic mixture (column: DAICEL CHIRALPAK IG (250 mm*30 mm, 10 um); mobile phase: [CO2+40%-MeOH(0.1% NH3H2O)]; isocratic elution mode, A-26b eluting first, other enantiomer eluting afterwards).
1H NMR:
A-26a (345 g, 1.26 mol, 1.0 equiv.) and NaOMe (3.45 g, 63.86 mmol, 0.05 equiv.) are dissolved in MeOH (2760 mL). The mixture is degassed and purged with N2 at 15° C. and stirred at 15° C. for 1 h. NH4Cl (134.6 g, 2.52 mol, 2.0 equiv.) is then added and the mixture is further stirred for 1 h at 60-70° C. After complete conversion, solvent is removed under reduced pressure and the obtained residue is diluted with 3000 mL DCM and filtered. The filtrate is then concentrated and A-27a (1.37 kg, crude) is obtained as a gray solid.
Other intermediates A-27 (Table 3) are available in an analogous manner. The crude product is purified by chromatography if necessary.
B-1a (4.92 g, 19.1 mmol, 1.0 equiv.), N,N′-carbonyldiimidazole (5.14 g, 28.6 mmol, 1.5 equiv.) and molecular sieves (3A, 500 mg) are dissolved in DCM (30 mL) and stirred for 40 min at rt. After complete activation, N,O-Dimethylhydroxylamine hydrochloride (2.79 g, 28.6 mmol, 1.5 equiv.) is added and the reaction is stirred again for 2 h at rt. After complete conversion, water (100 mL) and DCM (150 mL) are added and the phases are separated, the water phase is extracted with DCM (2×). The combined organic phase is washed with brine and concentrated under reduced pressure. The residue is purified by NP chromatography to give the product B-2a.
Other intermediates B-2 (Table 4) are available in an analogous manner. The crude product is purified by chromatography if necessary.
B-2a (4.88 g, 16.9 mmol, 1.0 equiv.) is dissolved in THF (15 mL) under an argon atmosphere and cooled to −10° C. Bromo(methyl)magnesium (3.4 M in MeTHF, 6.46 mL, 22.0 mmol, 1.3 equiv.) is added and stirred for 1 h at −10° C. After complete conversion, the reaction mixture is cooled to −20° C. and quenched by addition of brine. The resulting mixture is extracted with DCM (3×). The combined organic phase is concentrated under reduced pressure to obtain B-3a.
Other intermediates B-3 (Table 5) are available in an analogous manner. The crude product is purified by chromatography if necessary.
To a stirred solution of tetrahydro-pyrrolizine-7a-carbonitrile (1.00 g, 0.007 mol, 1.0 equiv.) in dry THF (5 mL), MeLi (13.77 mL, 0.022 mol. 3.0 equiv.) is added at −78° C. and the reaction mixture is then stirred at same temperature for 2 h. After complete conversion the reaction mixture is quenched with sat. NH4Cl solution and extracted with EtOAc. Organic layers are combined, dried, filtered and concentrated and the crude product is purified via NP chromatography yielding B-3h (HPLC method: H; tret=0.28 min; [M+H]+=154).
(R)-Methyl oxazaborolidine (0.99 g, 3.3 mmol, 0.20 equiv.) is dissolved in THF (2 mL) under an argon atmosphere and cooled to −5° C. Borane-dimethyl sulfide complex (1.0 M, 22 mL 22 mmol, 1.3 equiv.) is added. The mixture is stirred for 30 min at rt. The mixture is cooled to −5° C. and B-3a (4.1 g, 17 mmol, 1 equiv.) is added slowly, dropwise. The reaction is stirred at rt for 1 h. After complete conversion of starting material, the reaction is cooled to −10° C. and quenched by addition of MeOH. The mixture is concentrated under reduced pressure. The residue is dissolved in water (150 mL) and formic acid (0.5 mL) and extracted with DCM (3×). The combined organic phase is concentrated under reduced pressure and purified by NP chromatography to give the product B-4a.
Other intermediates B-4 (Table 6) are available in an analogous manner. The crude product is purified by chromatography if necessary.
B-4a (306 mg, 12.5 mmol, 1.0 equiv.) is dissolved in THF, (31 mL) under an argon atmosphere. Lithium aluminium hydride (1 M in THF, 24.9 mL, 25.0 mmol, 2.0 equiv.) is added slowly. The reaction is stirred at 60° C. for 1 h. After complete conversion, the reaction is cooled to rt, Rochelle salt solution and KOH is added and stirred for 1 h. The existing suspension is extracted with DCM (3×), the combined organic phase is concentrated under reduced pressure to yield B-5a.
Other intermediates B-5 (Table 7) are available in an analogous manner. Deuterated intermediates B-5 are obtained analogously but lithium aluminium hydride is exchanged by lithium aluminium deuteride. The crude product is purified by chromatography if necessary.
(S)-(+)-3-hydroxytetrahydrofuran (3.0 g, 33 mmol, 1.0 equiv.) is dissolved in MTBE (7 mL) at 0° C. and DABCO (4.63 g, 112 mmol, 1.2 equiv.) and p-toluenesulfonylchloride (7.07 g, 37 mmol, 1.1 equiv.) are added. The reaction mixture is warmed to rt and stirred for 4 h. Morpholine is added to quench residual p-toluenesulfonylchloride. The reaction mixture is diluted with aq. NaHCO3 solution and EtOAc and stirred for 15 minutes. The organic layer is separated and washed with NaHCO3, NH4Cl, and NaCl solutions. The organic layer is dried with Na2SO4, filtered, and concentrated under reduced pressure. Purification by RP chromatography provides the product B-7a (HPLC method: C; tret=0.52 min; [M+H]+=260).
To a stirred solution of (2S,3S)-1-tert-butoxycarbonyl-3-hydroxy-pyrrolidine-2-carboxylic acid (25.0 g, 108 mmol, 1.0 equiv.) in THF (300 mL) is added N,O-dimethyl hydroxylamine HCl (21.09 g, 216 mmol, 2.0 equiv.) and HATU (23.01 g, 216 mmol, 2.0 equiv.) at 0° C., then DIPEA (75.5 mL, 432 mmol, 4.0 equiv.) is added dropwise at same temperature and the mixture is stirred at rt for 12 h. After complete conversion the mixture is concentrated, dissolved in EtOAc, washed with water and sat. NaCl solution, and the organic layer is dried, filtered, and concentrated. The crude material is purified via NP chromatography to obtain the desired product B-9a (HPLC method: H; tret=1.34 min; [M+H]+=275).
To a stirred solution of B-9a (50.00 g, 182 mmol, 1.0 equiv.) in THF (500 mL) at 0° C. methyl magnesium bromide (1 M in THF, 911 mL, 911 mmol, 5.0 equiv.) is added and the mixture is stirred for 3 h. After complete conversion the reaction mixture is quenched with sat. ammonium chloride solution and extracted with DCM. The organic layer is concentrated and the crude material is purified by NP chromatography to obtain the desired product B-10a (HPLC method: H; tret=1.40 min; [M+H-tBu]+=174).
To a stirred solution of (3a-R)-1-methyl-3,3-diphenyl-hexahydropyrrolo[1,2-c][1,3,2]oxazaborole (1.92 mL, 17.4 mmol, 0.2 equiv.) in THF (200 mL) at −20° C. is added BH3*THF (1 M in THF, 113 mL, 113 mmol, 1.3 equiv.). After 30 min the mixture is cooled to −60° C., a THF solution of B-10a (20.00 g, 87.2 mmol, 1.0 equiv.) is added dropwise and the mixture is stirred for 3 h while allowed to reach rt. After complete conversion the mixture is quenched with methanol, diluted with water, extracted with DCM, filtered, and concentrated. The crude material is purified by NP chromatography to obtain the desired product B-11a (HPLC method: H; tret=1.35 min; [M+H]+=232).
To a stirred solution of B-11a (15.0 g, 64.9 mmol, 1.0 equiv.) in DMF (80 mL) at 0° C. is added sodium hydride (3.11 g, 130 mmol, 2.0 equiv.) and the resulting mixture is stirred at rt for 16 h. After complete conversion the reaction mixture is quenched with ice water, extracted with DCM and the organic layer is concentrated. The crude material is purified by NP chromatography to obtain the desired product B-12a (HPLC method: H; tret=0.87 min; [M+H]+=158).
To a stirred solution of B-12a (10.0 g, 63.6 mmol, 1.0 equiv.) in THF (400 mL) at 0° C. is added diethylazodicarboxylate (12.2 g, 70.0 mmol) followed by benzoic acid (11.7 g, 95.4 mmol, 1.5 equiv.) and triphenylphosphine (30.0 g, 114.5 mmol, 1.8 equiv.) and the mixture is stirred at rt for 2 h. After complete conversion the mixture is diluted with water, extracted with DCM, washed with sat. NaCl solution, filtered and concentrated. The crude material is purified by NP chromatography to obtain the desired product B-13a (HPLC method: H; tret=1.67 min; [M+H]+=262).
A freshly prepared solution of KOH (1M in MeOH, 28.7 mL, 28.7 mmol, 1.5 equiv.) is added to a stirred solution of B-13a (5.00 g, 19.1 mmol, 1.0 equiv.) in methanol (50 mL) and the mixture is stirred for 2 h at rt. After complete conversion the reaction mixture is neutralized with 1 N HCl, MeOH is evaporated, and the mixture is extracted with EtOAc. The organic layers are washed with sat. NaCl solution, filtered and concentrated. The crude material is purified by NP chromatography to obtain the desired product B-14a (HPLC method: H; tret=0.95 min; [M+H]+=158).
(Diethylamino)sulfurtrifluoride (30.2 mL, 229 mmol, 2.0 equiv.) is added slowly to a stirred solution of B-14a (18.0 g, 115 mmol, 1.0 equiv.) in DCM (360 mL) at −78° C. and the mixture is stirred for 3 h. After complete conversion the mixture is quenched with sat. NaHCO3 solution, diluted with water and extracted with DCM. The organic layer is washed with sat. NaCl solution, filtered and concentrated and the crude material is purified by NP chromatography to obtain the desired product B-15a.
The following intermediates B-15 (Table 8) are available in an analogous manner. The crude product is purified by chromatography if necessary.
To a stirred solution of B-15a (4.00 g, 25.1 mmol. 1.0 equiv.) in THF (1 mL) at 0° C. is added lithium aluminium hydride (1 M in THF, 37.7 mL, 37.7 mmol, 1.5 equiv.) and the reaction mixture is stirred for 2 h. After complete conversion the reaction mixture is quenched with 4 M NaOH solution, diluted with ice water (2 mL), filtered, and concentrated under reduced pressure to get the crude material which is purified by NP chromatography to obtain the desired product B-16a.
The following intermediates B-16 (Table 9) are available in an analogous manner. The crude product is purified by chromatography if necessary.
To an in a glove box stirred solution of 1-acetonyloxypropan-2-one (10.0 g, 76.84 mmol, 1.0 equiv) in 1,4-dioxane (100 mL), p-phenetidine (14.8 mL, 115.26 mmol, 1.5 equiv), diethyl 1,4-dihydro-2,6-dimethyl-3,5-pyridinedicarboxylate (42.8 g, 169.05 mmol, 2.2 equiv) are added followed by (11bS)-4-hydroxy-2,6-bis(2,4,6-triisopropylphenyl)dinaphtho[2,1-d:″,″-f][1,3,2]dioxaphosphepine 4-oxide (5.8 g, 7.68 mmol, 0.1 equiv). The reaction is stirred at 60° C. for 72 h. After complete conversion the reaction is diluted with water and EtOAc. The combined organic layers are washed with brine, dried over Na2SO4 and concentrated under reduced pressure to afford B-18a. (HPLC method: AD; tret=0.888 min; [M+H]+=236).
To a stirred solution of B-18a (15.0 g, 63.74 mmol, 1.0 equiv) in ACN (150 mL) and water (150 mL), periodic acid (21.8 g, 95.61 mmol, 1.5 equiv) is added at 0° C. followed by H2SO4 (1 M, 32 mL). The reaction is stirred at rt for 16 h. After complete conversion the reaction is washed with DCM. The organic phase is discarded and 5 M aq. KOH solution Is added dropwise until pH 11. The aqueous solution is extracted with EtOAc. The combined organic layers are brought to pH 1 by adding i-PrOH/HCl and then dried over MgSO4. The solvent is concentrated under reduced pressure to afford the HCl salt of B-19a. TLC: 100% EtOAc, Rf: 0.2, UPLC method: AE, tret=4.292 min)
Trichloropyrimidine-5-amine (2.0 g, 10.1 mmol, 1.0 equiv.) is dissolved in THF (16 mL) under argon and (R)-tetrahydrofuran-3-amine hydrochloride (2.57 g, 20.2 mmol, 2.0 equiv.) and DIPEA (8.6 mL, 50.4 mmol, 5.0 equiv.) are added. The reaction mixture is stirred at 100° C. overnight. Next, 1,1′-carbonyldiimidazole (4.21 g, 25.2 mmol, 2.50 equiv.) and DIPEA (6 mL, 35.3 mmol, 3.5 equiv.) are added, and the reaction mixture is stirred at 50° C. for a further 6.5 h. The reaction mixture is concentrated under reduced pressure and purified by RP chromatography to give product C-2a.
The following intermediates C-2 (Table 10) are available in an analogous manner. The crude product is purified by chromatography if necessary.
(R)-tetrahydrofuran-3-amine hydrochloride (300 mg, 2.43 mmol, 1.0 equiv.), ethyl bromoacetate (302 μL, 2.67 mmol, 1.1 equiv.) and DIPEA (1.04 mL, 6.07 mmol, 2.5 equiv.) are dissolved in THF (1 mL) and stirred at 40° C. overnight. After complete conversion the mixture is filtered, the residue is washed with THF and the solution is concentrated to give crude C-3a, which is used for the next step without purification. Trichloropyrimidin-5-amine (300 mg, 1.51 mmol, 0.62 equiv.), crude C-3a (420 mg, 1.94 mmol, 0.80 equiv.), and DIPEA (0.52 mL, 3.02 mmol, 1.24 equiv.) in THF (2 mL) is heated to 100° C. overnight. The reaction mixture is concentrated under reduced pressure and purified by RP chromatography to give product C-4a (HPLC method: C; tret=0.34 min; [M+H]+=289).
C-2a (200 mg, 0.58 mmol, 1.0 equiv.) is dissolved in DMF (5 mL), 2-bromoethyl methyl ether (81 μL, 0.87 mmol, 1.5 equiv.) and Cs2CO3 (227 mg, 0.70 mmol, 1.2 equiv) are added. The reaction mixture is stirred at room temperature for 2.5 h and then heated to 55° C. overnight. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted three times with DCM. The combined organic phases are dried over magnesium sulfate, filtered, and concentrated under reduced pressure to give the crude product C-5a which is used without further purification.
The following N-alkyl intermediates C-5 (Table 11) are available in an analogous manner from the corresponding NH-analogues C-2 and C-4. The crude product is purified by chromatography if necessary.
(R)-Tetrahydrofuran-3-amine hydrochloride (3.9 g, 30 mmol, 2.0 equiv.) and DIPEA (12 mL, 73 mmol, 5.0 equiv.) is added to trichloropyrimidin-5-amine (2.9 g, 14 mmol, 1.0 equiv.) in THF (40 mL). The reaction mixture is heated to 70° C. overnight. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted three times with DCM. The combined organic phases are dried over magnesium sulfate, filtered, and concentrated under reduced pressure. Purification by RP chromatography gives the final product C-6a (HPLC method: C; tret=0.34 min; [M+H]+=249).
C-6a (200 mg, 0.80 mmol, 1.0 equiv.), triethyl orthoformate (3 mL, 17 mmol, 22.0 equiv.), and p-toluenesulfonic acid monohydrate (15 mg, 0.08 mmol, 0.1 equiv.) is stirred at 100° C. overnight. The reaction mixture is diluted with DMF and purified by RP chromatography to give the final product C-7a.
The following intermediates C-7 (Table 12) are available in an analogous manner. The crude product is purified by chromatography if necessary.
2,4-Dichloro-5-methyl-7H-pyrrolo[2,3-D]pyrimidine (688 mg, 3.30 mmol, 1.10 equiv.), (S)-(+)-3-hydroxytetrahydrofuran (270 mg, 3.00 mmol, 1.0 equiv.) and triphenylphosphine (1.18 g, 4.5 mmol, 1.5 equiv.) in THF (30 mL) is stirred at room temperature for 10 minutes. The solution is cooled to 0° C., DIAD (909 mg, 4.5 mmol, 1.5 equiv.) is added and the reaction mixture stirred at room temperature for 2 h. The reaction mixture is diluted with water and extracted three times with DCM. The organic layers are concentrated under reduced pressure, dissolved in ACN and purified by RP chromatography to obtain the final product C-9a (HPLC method: A; tret=1.27 min; [M+H]+=272).
(1S)-1-[(2S)-1-methylpyrrolidin-2-yl]ethan-1-ol (83 mg, 0.61 mmol, 1.1 equiv.) and sodium hydride (60% in mineral oil, 46 mg, 1.16 mmol, 2.0 equiv.) are stirred in THF (1.5 mL) at room temperature followed by addition of C-5a (193 mg, 0.58 mmol, 1.0 equiv.) in THF (1.5 mL). The reaction mixture is stirred at 50° C. for 2 h. The reaction mixture is diluted with DCM and poured onto sat. NaHCO3. It is extracted with DCM (3×) and dried over magnesium sulfate, filtered and concentrated to give the crude product D-1a, which is used without further purification.
The following ethers 0-1 (Table 13) are available in an analogous manner from the corresponding dichloropyrimidine intermediates C and the suitable alcohol. The crude product is purified by chromatography if necessary.
D-1s (515 mg, 1.01 mmol, 1.0 equiv.) is dissolved in THF (5.0 mL) and tetrabutylammonium fluoride (1 M in THF, 1514 μL, 1.51 mmol, 1.5 equiv.) is added. The light brown reaction mixture is stirred at rt for 45 min. After complete conversion the reaction mixture is extracted with DCM and aq. sat. NaHCO3 solution. The organic phase is dried over Na2SO4 and the solvent is removed under vacuum. The crude product D-2a can be obtained as a light brown solid and is used without further purification for the next step (HPLC method: C; tret=0.65 min; [M+H]+=396) which is used in the next step without further purification.
2,4-dichloro-6-methyl-7H-pyrrolo[2,3-d]pyrimidine (1.50 g, 7.28 mmol, 1.0 equiv.), (1S)-1-[(2S)-1-methylpyrrolidin-2-yl]ethan-1-ol (1.57 g, 10.91 mmol, 1.5 equiv.) and DIPEA (2.48 mL, 14.5 mmol, 2.0 equiv) are combined in ACN (12 mL) and stirred at 80° C. for 72 h. After complete conversion the reaction mixture is diluted with DCM and poured onto water. It is extracted 3 times with DCM and dried over magnesium sulfate, filtered and concentrated. The crude product is purified by NP followed by RP chromatography to give product D-3a (HPLC method: A; tret=1.26 min; [M+H]+=295).
(1S)-1-[(2S)-1-methylpyrrolidin-1-yl]ethanol (422 mg, 3.1 mmol, 1.3 equiv.) in THF (3 mL) is cooled to 0° C., KOtBu (1 M, 4.8 mL, 4.78 mmol, 2.0 equiv.). The reaction mixture is warmed to room temperature and stirred for 15 minutes. Next, C-11a (500 mg, 2.4 mmol, 1.0 equiv.) is added and the reaction mixture stirred overnight at room temperature. The reaction mixture is neutralised with NaCl (sat., aq.) and extracted three times with DCM. The combined organic layers are dried over Na2SO4, filtered, and concentrated under reduced pressure. Purification by RP chromatography gives the product D-4a.
The following intermediates D-4 (Table 14) are available in an analogous manner. The crude product is purified by chromatography if necessary.
D-1a (214 mg, 0.50 mmol, 1.0 equiv.) and A-16a (261 mg, 0.60 mmol, 1.2 equiv.) and XPhos-Pd-G3 (42.5 mg, 0.60 mmol, 0.1 equiv.) in 1,4-dioxane (3 mL) are degassed with argon and heated to 100° C. for 2 h. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted 3 times with DCM. The combined organic phases are concentrated and purified by RP chromatography to give the product E-1a.
The following intermediates E-1 (Table 15) are available in an analogous manner from the corresponding chloropyrimidine intermediates D. The crude product is purified by chromatography if necessary.
E-1ao (120 mg, 0.232 mmol, 1.0 equiv.) is dissolved in dry ACN (3 mL), Cs2CO3 (302 mg, 0.927 mmol, 4.0 equiv.) is added and the mixture is stirred for 30 min at rt. Then B-7a (112 mg; 0.463 mmol; 2.0 equiv.) is added and the mixture is stirred for 1.5 h at 85° C. After complete conversion the reaction mixture is filtered and purified via RP chromatography yielding E-2a (HPLC method: C; tret=0.96 min; [M+H]+=578).
E-1ap (134 mg, 0.26 mmol, 1.00 equiv.) and Cs2CO3 (343 mg, 1.05 mmol, 4.00 equiv.) in DMSO (1 mL) is stirred for 10 minutes at room temperature before B-7a (127 mg, 0.53 mmol, 2.00 equiv.) is added. The reaction mixture is heated to 85° C. for 1 h. The reaction mixture is diluted with NaHCO3 and extracted three times with DCM. The combined organic layers are dried over Na2SO4, filtered, and concentrated under reduced pressure. Purification by RP chromatography gives the product E-3a (HPLC method: C; tret=1.79 min; [M+H]+=579).
E-1a (139 mg, 0.218 mmol, 1.0 equiv.) and HCl (4 M in 1,4-dioxane, 272 μL, 1.09 mmol, 5.0 equiv.) in methanol (3 mL) is stirred at 50° C. for 1 h. After complete conversion the reaction mixture is quenched with NaHCO3 and extracted 3 times with DCM. The organic phases are combined and concentrated under reduced pressure to yield the desired product F-1a, which is used for the next step without further purification.
The following intermediates F-1 (Table 16) are available in an analogous manner from the corresponding ketals E. The crude product is purified by chromatography if necessary.
For Compound F-1aj and F-1ak instead of MeOH, 1,4-dioxane is used as solvent for the reaction.
E-1y (105 mg, 0.169 mmol, 1.0 equiv.) is dissolved in ACN (1 mL), TFA (440 μL, 1.76 mmol, 10.0 equiv.) is added and the mixture is stirred at rt for 0.5 h. The reaction mixture is quenched with sat. aq. NaHCO3 solution and extracted 3 times with DCM. The organic phases are combined and concentrated under reduced pressure to yield F-2a. The crude material is used directly in the next step.
The following intermediates F-2 (Table 17) are available in an analogous manner from the corresponding ketals E. The crude product is purified by chromatography if necessary.
E-1aq (762 mg, 1.32 mmol, 1.0 equiv.) and HCl (4 M in 1,4-dioxane, 1.98 mL, 7.90 mmol, 6.0 equiv.) in methanol (2 mL) is stirred at 60° C. for 2 h. The reaction mixture is quenched with NaHCO3 and extracted 3 times with DCM. The organic phases are combined and concentrated under reduced pressure and the crude product is purified via RP chromatography to yield F-3a (HPLC method: A; tret=1.34 min; [M+H]+=451).
F-3a (100 mg, 0.22 mmol, 1.0 equiv.) and Cs2CO3 (289 mg, 0.888 mmol, 4.0 equiv.) in DMSO (1 mL) is stirred for 10 minutes at room temperature before B-7a (107 mg, 0.44 mmol, 2.0 equiv.) is added. The reaction mixture is stirred at 85° C. for 1 h. After complete conversion the reaction mixture is diluted with sat. aq. NaHCO3 and extracted three times with DCM. The combined organic layers are dried over Na2SO4, filtered, and concentrated under reduced pressure to yield F-4a (HPLC method: C; tret=0.84 min; [M+H]+=521) which is used for the next step without further purification.
E-1x (288 mg, 0.50 mmol, 1.0 equiv.) is dissolved in methanol (3 mL) and HCl (4 M in 1,4-dioxane, 620 μL, 2.48 mmol, 5.0 equiv.) is added. The reaction mixture is heated to 50° C. for 3 h. The reaction mixture is quenched with NaHCO3 and extracted 3 times with DCM. The organic phases are combined and concentrated under reduced pressure to yield crude F-5a (HPLC method: C; tret=0.78 min; [M+H]+=573) which is used for the next step without purification.
A solution of F-5a (284 mg, 0.50 mmol, 1.0 equiv.) and NaH (60% in mineral oil, 48 mg, 1.19 mmol, 2.4 equiv.) in DMF (5 mL) is stirred at 50° C. for 3.5 h. The reaction mixture is diluted with water and extracted three times with DCM. The organic layers are combined, dried over Na2SO4, filtered, and concentrated under reduced pressure. The crude product is purified via RP chromatography to obtain the desired product F-6a (HPLC method: C; tret=0.71 min; [M+H]+=537).
To a solution of A-27a (70.0 g, 240 mmol, 1.0 equiv) in DMF (350 mL), dimethyl malonate (63.5 g, 481 mmol, 2.0 equiv) and DBU (109.6 g, 721 mmol, 3.0 quiv) are added. The reaction is stirred at 90° C. for 16 h. After complete conversion the reaction is cooled to rt, ice cold water (500 mL) is added and the pH is adjusted to 3-4 with 6 N HCl. MTBE (300 mL) is added and the mixture is stirred at rt for 1 h. The precipitate is collected by filtration and stirred in diethyl ether (500 mL) at rt for 20 min, filtered and dried under reduced pressure. Then the crude product is washed with n-pentane (500 mL) and co-distilled with toluene (250 mL) under reduced pressure to give F-7a as desired product, which is used for the next step without any further purification. (HPLC method: H; tret=1.64 min; [M+H]+=360).
Fuming HNO3 (25 mL, 587.2 mmol, 4.2 equiv.) is added dropwise at 0° C. to F-7a (50.0 g, 139.1 mmol, 1.0 equiv.) in acetic acid (25 mL) and the mixture is stirred at rt for 1 h. After complete conversion the reaction is quenched with ice water (300 mL) and stirred for 30 min. The precipitated compound is collected by filtration and dried under reduced pressure. The crude product is slurried in diethyl ether (500 mL), stirred for 25 min, filtered, dried under reduced pressure and co-distilled with toluene (2×200 mL) under reduced pressure to yield F-8a (HPLC method: AD, tret=0.68 min; [M+H]+=361). The crude product is used for the next step without further purification.
To a stirred solution of F-8a (4.0 g, 11.1 mmol, 1.0 equiv) in toluene (50 mL), DIPEA (19 mL, 111 mmol, 10.0 equiv) followed by POCl3 (2.3 ml, 55.5 mmol, 5.0 equiv) are added at −10° C. The reaction is stirred at rt for 3 h. After complete conversion the reaction is quenched with ice water (50 mL), stirred for 10 min and extracted with MTBE (3×50 mL). The organic layer is washed with NaHCO3 (sat.), brine (50 mL) and dried over Na2SO4. After filtration the solution is concentrated under reduced pressure, washed with n-pentane (30 mL) and dried under reduced pressure to afford F-9a (HPLC method: A; tret=1.53 min; [M+H]+=397). The crude product is used for the next step without any further purification.
To a stirred solution of F-9a (2.0 g, 5.04 mmol, 1.0 equiv.) in ethanol (5 mL) and water (2 mL), iron nanopowder (2.8 g, 50.35 mmol, 10.0 equiv.) followed by ammonium chloride (2.69 g, 50.4 mmol, 10.0 equiv) are added at rt. The reaction is stirred at rt for 6 h. After complete conversion the reaction is filtered through Celite @ pad and washed with EtOAc (20 mL). The organic layer is washed with brine (5 mL) and dried over Na2SO4. The mixture is filtered and concentrated under reduced pressure to afford F-10a (HPLC method: C; tret=0.676 min; [M+H]+=367). The crude product is used for the next step without any further purification.
F-10a (482 mg, 1.31 mmol, 1.0 equiv.) is dissolved in dry DMSO (4.0 mL) under argon and 5-methyloxolan-3-amine (112 mg, 1.05 mmol, 0.8 equiv.) and DIPEA (0.67 mL, 3.96 mmol, 3.0 equiv.) are added. The reaction mixture is stirred at 100° C. for 3 h. 1,1′-Carbonyldiimidazole (1.0 g, 5.98 mmol, 4.5 equiv.) and DIPEA (0.67 mL, 3.96 mmol, 3.0 equiv.) are added and the mixture is stirred at 50° C. overnight. After complete conversion, the reaction is diluted with NaHCO3 (sat.) and extracted with DCM. The organic phase is dried over Na2SO4 and concentrated under reduced pressure to afford F-11a. The crude product is used for the next step without any further purification. (HPLC method: C; tret=0.503 min; [M+H]+=458)
To a stirred solution of F-9a (15.0 g, 37.76 mmol, 1.0 equiv) in DCM (150 mL) under N2 gas, B-19a (4.0 g, 26.43 mmol, 0.7 equiv) and DIPEA (29.2 g, 226.58 mmol, 6.0 equiv) are added at 0° C. The reaction is stirred at rt for 2 h. After complete conversion the reaction is concentrated under reduced pressure, quenched with ice water and extracted with DCM. The organic layer is dried with Na2SO4 and concentrated under reduced pressure to give F-12a. (HPLC method: AD; tret=1.276 min; [M+H]+=476). The crude product is used without further purification.
To a stirred solution of F-12a (20.0 g, 42.03 mmol, 1.0 equiv) in EtOH (180 mL) and water (20 mL) under N2 gas, iron powder (14.1 g, 252.14 mmol, 6.0 equiv) and ammonium chloride (13.5 g, 252.1 mmol, 6.0 equiv) are added at rt. The reaction is stirred at 70° C. for 2 h. After complete conversion the reaction is filtered through Celite® bed and washed with EtOAc. The filtrate is concentrated under reduced pressure. The residue is dissolved in water and extracted with EtOAc. The combined organic layers are washed with brine and dried over Na2SO4. The solvent is removed under reduced pressure to afford F-13a. (HPLC method: AD; tret=1.146 min; [M+H]+=446). The crude product is used without further purification.
To a stirred solution of F-13a (20.5 g, 45.97 mmol, 1.0 equiv) in THF (200 mL), DIPEA (17.8 g, 137.9 mmol, 3.0 equiv) and triphosgene (20.5 g, 68.954 mmol, 1.5 equiv) are added at 0° C., then stirred under N2 gas at rt of 2 h. After complete conversion the reaction is quenched with ice water and extracted with EtOAc. The combined organic layers are washed with brine and dried over Na2SO4. The solvent is concentrated under reduced pressure to afford F-14a. (HPLC method: AD; tret=1.121 min; [M+H]+=472). The crude product is used without further purification.
F-11a (601 mg, 1.31 mmol, 1.0 equiv.) is dissolved in DMF (4 mL), iodomethane (123 μL, 1.97 mmol, 1.5 equiv.) and Cs2CO3 (641.0 mg, 1.97 mmol, 1.5 equiv) are added, and the reaction mixture is stirred at rt overnight. After complete conversion the reaction mixture is diluted with water and extracted three times with DCM. The combined organic phases are dried over magnesium sulfate, filtered, concentrated under reduced pressure, and purified via RP chromatography to yield F-15a.
The following products F-15 (Table 18) are available in an analogous manner from the corresponding ketones F-11a and F-14a. The crude product is purified by chromatography if necessary.
Compound F-15a is obtained initially in a mixture of four diastereomers which are then separated via Chiral HPLC. (Column: Daicel Chiralpak IC, 5 μm, flow: 40 mL/min, solvent A: Heptane, solvent B: EtOAc/Ethanol 3:1+0.1% DEA, Gradient: 17 min 80% A/20% B, then 8 min 60% A/40% B, total runtime 25 min, 310 nm). Peak 4 is assigned to F-15a and taken forward in the next steps of the synthesis.
To a mixture of B-5d (22.5 mg, 0.127 mmol, 3.0 equiv.) and F-15a (20 mg, 0.042 mmol, 1 equiv.) in THF (1 mL) sodium tert-butoxide (2 M in THF, 127.1 mg, 0.254 mmol, 6.0 equiv.) is added and the reaction is stirred at room temperature for 10 min. After complete conversion the reaction mixture is diluted with water and some KHSO4, extracted with DCM and dried over sodium sulfate. The mixture is concentrated under reduced pressure to give F-16a. The crude product is used for the next step without further purification.
The following products F-16 (Table 19) are available in an analogous manner from the corresponding ketones F-15. The crude product is purified by chromatography if necessary.
F-1a (129 mg, 0.22 mmol, 1.0 equiv.), sulfur (12 mg, 0.39 mmol, 2.0 equiv.) and ammonium acetate (47 mg, 0.61 mmol, 2.8 equiv.) in ethanol (1.5 mL) are heated to 70° C. Malonodinitrile (195 μL, 0.39 mmol, 1.8 equiv.) in ethanol (0.5 mL) is added and the reaction mixture is stirred for 1 h at 70° C. then overnight at 60° C. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted three times with DCM. The combined organic phases are concentrated and purified via RP chromatography to give the desired product 1-1.
The following products I (Table 20) are available in an analogous manner from the corresponding ketones F. The crude product is purified by chromatography if necessary.
Tetramethylammonium cyanide (615 mg, 3.90 mmol, 2.0 equiv.) and DABCO (552 mg, 4.92 mmol, 2.5 equiv.) is added to a solution of D-1w (854 mg, 1.97 mmol, 1.0 equiv.) in ACN (6 mL) and DMSO (0.5 mL). The reaction mixture is heated to 40° C. overnight. The reaction mixture is diluted with sat. aq. NaHCO3 solution and extracted three times with DCM. The organic phases are combined and concentrated under reduced pressure. The residue is dissolved in in ACN and water and purified by basic RP chromatography to give the desired product G-1a.
The following products G-1 (Table 21) are available in an analogous manner from the corresponding chloropyrimidines D. The crude product is purified by chromatography if necessary.
G-1a (575 mg, 1.43 mmol, 1.0 equiv.) is dissolved in methanol (4 mL) and sodium hydroxide (4 M aq., 642 μL, 2.57 mmol, 1.8 equiv.) and stirred at room temperature for 2 h. Next, HCl (4 M aq., 1.07 mL, 4.3 mmol, 3.0 equiv.) is added and stirred for 1 h. The mixture is neutralized with NH3 in MeOH, concentrated, and the crude product is purified via RP chromatography to give the desired product G-2a.
The following products G-2 (Table 22) are available in an analogous manner from the corresponding chloropyrimidines G-1. The crude product is purified by chromatography if necessary.
A solution of A-6b (180 mg, 0.76 mmol, 1.8 equiv.) and molecular sieves (3 Å) in dry THF (1 mL) is heated to 50° C. for 20 minutes followed by addition of LiHMDS (1 M in THF, 2.08 mL, 2.08 mmol, 3.3 equiv.) and stirred for a further 10 minutes at the same temperature. This solution is added to a pre-stirred (50° C. for 20 mins) solution of G-2a (275 mg, 0.63 mmol, 1.00 equiv.), magnesium bromide diethyl etherate (247 mg, 0.95 mmol, 1.5 equiv.), and molecular sieves (3 Å) in dry THF (5 mL) and the resulting reaction mixture is stirred at 50° C. for 1 h. After complete conversion the reaction mixture is quenched with water, then adjusted to pH 7 with HCl (2 M) and extracted three times with DCM. The organic phases are combined and concentrated under reduced pressure. The crude product is purified by RP chromatography to give the desired product H-1a as a mixture of 3 tautomers.
The following products H-1 (Table 23) are available in an analogous manner from the corresponding chloropyrimidines G-2. The crude product is purified by chromatography if necessary.
A solution of H-1a (354 mg, 0.56 mmol, 1.0 equiv.), hydroxylamine (50% in water, 44 μL, 0.73 mmol, 1.3 equiv.), and formic acid (26 μL, 0.70 mmol, 1.2 equiv.) in 1,4-dioxane (5 mL) is stirred at 50° C. overnight. After complete conversion the reaction mixture is diluted with NaHCO3 and extracted three times with DCM. The organic phases are combined and concentrated under reduced pressure. The crude product H-2a is used in the subsequent step without further purification.
The following products H-2 (Table 24) are available in an analogous manner from the corresponding chloropyrimidines H-1. The crude product is purified by chromatography if necessary.
H-1c (2.75 g, 3.90 mmol, 1.0 equiv.) is dissolved in pyridine (20 mL), hydroxylamine hydrochloride (767 mg, 11.03 mmol, 2.8 equiv.) is added and the mixture is stirred at rt overnight. After complete conversion the reaction mixture is diluted with NaHCO3 and extracted three times with DCM. The organic phases are combined and concentrated under reduced pressure to yield crude H-3a (HPLC method: C; tret=0.91 min; [M+H]+=597) which is used for the next step without purification.
H-2a (362 mg, 0.56 mmol, 1.0 equiv.) is dissolved in HCl (4 M in 1,4-dioxane, 4 mL, 16 mmol, 28 equiv.) and stirred overnight at room temperature. Next, water (1 mL) is added and the reaction mixture is heated to 50° C. for 1 h. The reaction mixture is diluted with sat. aq. NaHCO3 solution and extracted three times with DCM. The organic layers are combined and concentrated, and the crude product is purified by RP chromatography to give the desired product J-1a.
The following products J-1 (Table 25) are available in an analogous manner from the corresponding oximes H-2. The crude product is purified by chromatography if necessary.
H-3a (717 mg, 1.20 mmol, 1.0 equiv.) is dissolved in MeOH (8.5 mL), HCl (37% aq., 15.6 mmol, 13.0 equiv.) is added and the mixture is stirred for 1 h at 65° C. After complete conversion the mixture is diluted with water, extracted 3 times with DCM, and the combined organic layers are washed with sat. aq. NaHCO3 solution, dried, filtered, and concentrated. in 1,4-dioxane (5 mL) at 0° C. The crude product is purified by RP chromatography to yield J-2a (HPLC method: C; tret=0.91 min; [M+H]+=597).
(S)-(+)-3-hydroxytetrahydrofuran (25.7 mg, 0.286 mmol, 1.5 equiv.) is dissolved in dry THF (2.0 mL). J-2a (100.0 mg, 0.190 mmol, 1.0 equiv.) and triphenylphosphine (75.6 mg, 0.286 mmol, 1.5 equiv.) is added and the mixture is cooled to 0° C. Diisopropylazodicarboxylate (57.38 μL, 0.286 mmol, 1.5 equiv.) is added and the mixture is stirred for 2 h while allowed to reach rt. After complete conversion the reaction mixture is diluted with water and extracted three times with DCM. The organic phases are combined and concentrated under reduced pressure to yield crude product which is purified via RP chromatography to yield J-3a (HPLC method: A; tret=1.49 min; [M+H]+=521).
A-12a (9.31 g, 39.9 mmol, 1.0 equiv) is dissolved in MeOH (90 mL), NaOMe (25% in MeOH, 432 mg, 2.00 mmol, 0.05 equiv.) is added and the mixture is stirred at 50° C. for 1 h. NH4Cl (2.99 g, 55.9 mmol, 1.4 equiv.) is added and the mixture is stirred for additional 1 h at 50° C. Next, dimethyl malonate (6.97 mL, 60 mmol, 1.5 equiv.) is added followed by NaOMe (25% in MeOH, 21.58 g, 99.9 mmol, 2.5 equiv.) and the solution heated to reflux overnight. After complete conversion, the reaction mixture is diluted with water, and acidified to pH=2 using HCl (4 M aq.). The acidic solution is cooled in the fridge until a yellow precipitate occurs. The precipitate is filtered, washed with water and MTBE and dried to give the product. J-4a (HPLC method: C; tret=0.20 min; [M+H]+=316) which is used for the next step without further purification.
Fuming HNO3 (1.50 mL, 35.9 mmol, 1.3 equiv.) is added to J-4a (8.60 g, 27.3 mmol, 1.0 equiv.) in TFA (50 mL) and the mixture is stirred at rt for 15 min. After complete conversion the mixture is concentrated, and the residue is dried under reduced pressure. Then DIPEA (15 mL, 86.1 mmol, 2.2 equiv.) is added followed by portionwise addition of POCl3 (15 mL, 164.4 mmol, 6.0 equiv.) and the mixture is stirred for 30 min at 40° C. After complete conversion the mixture is slowly added to water at rt and stirred for 20 min. The resulting aq. Mixture is extracted 3 times with DCM. The combined organic layers are dried over Na2SO4 and concentrated under reduced pressure. The crude product is purified by NP chromatography to obtain J-5a (HPLC method: C; tret=0.92 min; [M+H]+=397).
J-5a (7.60 g, 19.1 mmol, 1.0 equiv.) and iron powder (7.48 g, 133.9 mmol, 7.0 equiv.) in acetic acid (50 mL) and HCl (4 M aq., 100 μL) is stirred at room temperature for 15 minutes while the temperature of the reaction rises to 60° C. After complete conversion the solution is diluted with water and adjusted to pH=5 with aq. NaHCO3. The aq. Mixture is extracted 3 times with DCM, the combined organic layers are washed with aq. NaHCO3 solution, dried over Na2SO4, filtered, and concentrated under reduced pressure. The crude product is purified by NP chromatography to yield J-6a (HPLC method: C; tret=0.68 min; [M+H]+=367).
J-6a (600 mg, 1.63 mmol, 1.0 equiv.) is dissolved in dry DMSO (4.0 mL) under argon and (R)-tetrahydro-2H-pyran-3-amine hydrochloride (459 mg, 3.27 mmol, 2.0 equiv.) and DIPEA (1.14 mL, 6.54 mmol, 4.0 equiv.) is added. The reaction mixture is stirred at 100° C. for 3 h. 1,1′-Carbonyldiimidazole (1.37 g, 8.17 mmol, 5.0 equiv.) and DIPEA (0.854 mL, 4.90 mmol, 3.0 equiv.) are added and the mixture is stirred at 50° C. overnight. After complete conversion, the mixture is diluted with MeOH, filtered, and purified via RP chromatography to yield J-7a.
The following products J-7 (Table 26) are available in an analogous manner using a suitable amine. The crude product is purified by chromatography if necessary.
J-7a (269 mg, 0.59 mmol, 1.0 equiv.) is dissolved in DMF (4 mL), iodomethane (55 μL, 0.88 mmol, 1.5 equiv.) and Cs2CO3 (230 mg, 0.71 mmol, 1.2 equiv) are added, and the reaction mixture is stirred at rt for 2.5 h. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted three times with DCM. The combined organic phases are dried over magnesium sulfate, filtered, concentrated under reduced pressure, and purified via RP chromatography to yield J-8a.
The following products J-8 (Table 27) are available in an analogous manner from the corresponding intermediates J-7. The crude product is purified by chromatography if necessary.
Compound J-8b is obtained initially in a mixture of four diastereomers which are then separated via Chiral HPLC. For that, the product is separated in multiple runs. The first run result on the separation of peak 1 and peak 2 (Column: IA, flow: 15 mL/min, 60% Heptane/10% Ethanol/30% Ethylacetate+0.1% DEA, 296 nm) and peak 3 and 4 are collected together and separated in a subsequent run (Column: IH, flow: 20 mL/min, 75% Heptane/25% THF+0.1% DEA, 296 nm.). Peak 1 from the first separation is assigned to J-8b and taken forward in the next steps of the synthesis.
B-5d (32.4 mg, 0.19 mmol, 1.5 equiv.) and sodium hydride (60% in mineral oil, 10.2 mg, 0.25 mmol, 2.0 equiv.) are stirred in THF (1.5 mL) at room temperature followed by addition of J-8a (60 mg, 0.13 mmol, 1.0 equiv.) in THF (0.5 mL). The reaction mixture is stirred at 50° C. for 2 h. After complete conversion the reaction mixture is diluted with DCM and poured onto sat. NaHCO3. The mixture is extracted 3 times with DCM and dried over magnesium sulfate, filtered, concentrated, and purified via RP chromatography to yield J-9a (HPLC method: C; tret=0.78 min; [M+H]+=595).
The following products J-9 (Table 28) are available in an analogous manner from the corresponding intermediates J-7. The crude product is purified by chromatography if necessary.
To a mixture of (1S)-1-[(2S)-1-methylpyrrolidin-2-yl]ethan-1-ol (61.4.4 mg, 0.447 mmol, 1.5 equiv.) and J-8b (145 mg, 0.298 mmol, 1 equiv.) in THF (1 mL) sodium tert-butoxide (2 M in THF, 10.2 mg, 0.25 mmol, 2.0 equiv.) is added and the reaction is stirred at room temperature for 10 min. After complete conversion the reaction mixture is diluted with DCM and poured onto sat. NaHCO3. Next. The aq. phase is extracted 3 times with DCM and dried over magnesium sulfate, filtered, concentrated, and purified via RP chromatography to yield J-10a (HPLC method: C; tret=0.76 min; [M+H]+=565).
J-1a (171 mg, 0.29 mmol, 1.0 equiv.), ammonium acetate (63 mg, 0.83 mmol, 2.8 equiv.), and sulfur (17 mg, 0.532 mmol, 1.8 equiv.), in EtOH (4 mL) are treated with malononitrile (266 μL, 0.532 mmol, 1.8 equiv.) dissolved in ethanol (1 mL) and the mixture is stirred at 70° C. for 2 h. After complete conversion the reaction mixture is diluted with sat. NaHCO3 solution and extracted three times with DCM. The combined organic phases are concentrated and purified by RP chromatography to give the product II-1
The following products II (Table 29) are available in an analogous manner from the corresponding ketones J. The crude product is purified by chromatography if necessary.
NaOH (1.67 g, 41.77 mmol, 2.2 equiv.) is dissolved in water (35 mL), cooled to 0° C. and methyl-3-aminooxolane-3-carboxylate hydrochloride K-1a (3.63 g, 18.99 mmol, 1.0 equiv.) is added. After stirring for 5 min acrylonitrile (1.89 mL, 28.48 mmol, 1.5 equiv.) is added, the mixture is allowed to reach rt and stirred overnight. After complete conversion the mixture is concentrated and the crude product K-2a (method C, tret=0.09 min; [M+H]+=185) is used for the next step without further purification.
K-2a (3.50 g, 19.0 mmol, 1.0 equiv.) is dissolved in a mixture of methanol (26 mL) and sulfuric acid (conc., 5 mL) and stirred at 70° C. for 2 h. After complete conversion aq. sat. NaHCO3 solution is slowly added and the mixture is extracted with DCM. The organic phases are combined, dried, filtered and concentrated under reduced pressure and the crude product K-3a (method C, tret=0.20 min; [M+H]+=232) is used for the next step without further purification.
K-3a (6.76 g, 29.23 mmol, 1.0 equiv.) and DIPEA (11.96 mL, 87.70 mmol, 3.00 equiv.) in THF (65 mL) under nitrogen atmosphere are cooled to 0° C., benzyl chloroformate (3.20 mL, 21.79 mmol, 1.5 equiv.) is added and the mixture is stirred for 3 h while allowed to reach rt. After complete conversion, the reaction mixture is diluted water and extracted with DCM. The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product K-4a (HPLC method: A, tret=1.17 min; [M−H]−=366).
Add K-5a (5.80 g, 39.6 mmol, 1.0 equiv.) and methyl 5-bromopentanoate (15.5 g, 79.4 mmol, 1.0 equiv.) in DMF (50 mL) to a bottle. Add NaH (2.38 g, 59.5 mmol, 1.5 equiv.) to the reaction mixture. After full conversion, the reaction mixture is quenched at 0° C. with sat. aq. ammonium chloride (300 mL) and the reaction mixture is allowed to warm to rt. The layers are separated and the aq. layer is extracted with EtOAc (2×). The combined organic layers are dried over sodium sulfate and concentrated under reduced pressure. The reaction mixture is purified by column chromatography to give K-6a.
1H NMR: (CDCl3 400 MHz): δ 4.01-3.94 (m, 4H), 3.78 (s, 3H), 3.67 (s, 3H), 3.44-3.38 (m, 2H), 2.42-2.33 (m, 3H), 2.22-2.16 (m, 1H), 1.74-1.70 (m, 2H), 1.67-1.60 (m, 2H); TLC: Petroleum ether/EtOAc=1/1, Rf=0.35.
Step 1: Tetrahydrofuran-3-carboxylic acid (350 g, 3.01 mol, 1.00 equiv.), SOCl2 (366 g, 3.08 mol, 1.02 equiv.) and pyridine (7.15 g, 90.43 mmol, 0.03 equiv.) are dissolved in DCM (1.40 L) at 20° C., under nitrogen atmosphere for 0.5 h. After that, the reaction mixture is concentrated under vacuum and compound K-10a is used in the next step without further purification.
Step 2: To a solution of 1,3,5-tribenzyl-1,3,5-triazinane (K-7a, 368 g, 1.03 mol, 1.00 equiv.) in DCM (1.50 L) BF3·Et2O (438 g, 3.09 mol, 380 mL, 3.00 equiv.) is added at 20° C. within 5 min. The mixture is then stirred at 20° C. for 0.5 h. The mixture is used directly for the next step without workup.
Step 3: K-10a (413 g, 3.07 mol, 1.00 equiv.) is dissolved in DCM (4.10 L) and the mixture is cooled to −58° C. Next, triethylamine (933 g, 9.22 mol, 3.00 equiv.) is added dropwise into the mixture while keeping the temperature below at −58° C. Then the solution of N-benzylmethanimine (K-8a, 366 g, 3.07 mol, 1.00 equiv.) prepared in the procedure above is slowly added to this mixture. The temperature is monitored throughout the process to insure it does not exceed −37° C. After that, the reaction is allowed to warm up to room temperature and after complete conversion of the starting material the reaction mixture is quenched by addition NaHCO3. The mixture is then extracted with DCM 2.0 L (1.0 L*2) and the combined organic layers is washed with brine 1.0 L, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue is the purified by column chromatography (SiO2, Petroleum ether/EtOAc=5/1) to yield K-11a.
1H NMR: (CDCl3 400 MHz) δ ppm 7.43-7.18 (m, 5H), 4.49-4.31 (m, 2H), 4.00 (q, J=9.6 Hz, 2H), 3.94-3.83 (m, 2H), 3.29-3.23 (m, 1H), 3.23-3.17 (m, 1H), 2.44 (td, J=7.6, 12.8 Hz, 1H), 2.12 (td, J=6.8, 13.2 Hz, 1H);
LCMS: Method AA; tret=0.30 min, m/z=218.
K-11a (243 g, 1.12 mol, 1.00 equiv.) and NaOMe (90.6 g, 1.68 mol, 1.50 equiv.) are dissolved in MeOH (2.40 L) and degassed and purged with N2 for 3 times at 20° C. Next, the mixture is stirred at 45° C. for 12 h. After complete conversion, solvend is removed under reduced pressure and then quenched into NH4Cl 1.0 L. The resulting mixture is then extracted two times with DCM 2.0 L (1.00 L*2). The combined organic layers are then washed with 1 L of brine, dried over Na2SO4, filtered and concentrated under reduced pressure to give K-12a as a residue.
LCMS: Method AB: tret=0.25 min, m/z=250.
1H NMR: (CDCl3 400 MHz) δ ppm 7.31-7.18 (m, 5H), 4.02 (d, J=8.8 Hz, 1H), 3.82 (t, J=7.6 Hz, 2H), 3.78-3.78 (m, 1H), 3.76 (s, 2H), 3.71 (d, J=9.2 Hz, 1H), 3.68 (s, 2H), 2.86-2.71 (m, 2H), 2.33 (td, J=6.4, 12.8 Hz, 1H), 1.86 (td, J=7.2, 12.8 Hz, 1H)
K-12a (161 g, 646 mmol, 1.0 equiv.), DBU (98.3 g, 646 mmol, 1.0 equiv.) and methyl acrylate (83.2 g, 975. mmol, 1.5 equiv.) are dissolved in can (1.6 L) and the mixture is degassed and purged with N2 for 3 times at 20° C. Next, the mixture is stirred at 70° C. for 24 h. After complete conversion, solvent is removed, and the reaction is purified via normal phase chromatography to obtain K-13a (HPLC method: AB; tret=0.34 min, m/z=336).
1H NMR: (CDCl3 400 MHz): δ ppm 7.33-7.23 (m, 5H), 4.09 (d, J=8.8 Hz, 1H), 3.88-3.71 (m, 3H), 3.70 (s, 3H), 3.63-3.61 (m, 3H), 3.57 (s, 2H), 2.93-2.81 (m, 2H), 2.76 (t, J=7. Hz, 2H), 2.50-2.39 (m, 3H), 1.90 (td, J=7.6, 12.4 Hz, 1H).
K-4a (211 mg, 0.577 mmol, 1.0 equiv.) is dissolved in dry THF, cooled to 0° C., LiHMDS (1 M in THF, 1155 μL, 1.16 mmol, 2.0 equiv.) is added, and the mixture is stirred for 90 min while the mixture is allowed to warm to rt. After complete conversion, the reaction mixture is diluted water, brought to pH=5 using citric acid (5% in water) and extracted with DCM. The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product K-14a.
The following intermediates K-14 (Table 30) are available in an analogous manner from the corresponding diesters K—6a and K-13a. The crude product is purified by chromatography if necessary.
K-14c (123 g, 407.5 mmol, 1.0 equiv.) is dissolved in MeOH (600 mL) followed by the addition of Pd/C (13.0 g, 12.22 mmol, 10% purity, 10%) and Boc2O (178 g, 815.6 mmol, 2.0 equiv.). The mixture is then stirred at 20° C. for 5 h under 15 psi H2. After that, solvent is removed under reduced pressure to give a reside that is further purified by column chromatography (SiO2, Petroleum ether/EtOAc=12/1-8/1) to deliver pure K-15a.
1H NMR: (CDCl3 400 MHz): δ ppm 4.34-4.15 (m, 1H), 4.04-3.80 (m, 5H), 3.78 (s, 3H), 3.74-3.69 (m, 1H), 3.63 (d, J=13.6 Hz, 1H), 3.31 (d, J=10.8 Hz, 1H), 2.41-2.29 (m, 1H), 1.78-1.69 (m, 1H), 1.48 (s, 9H); TLC: Petroleum ether:EtOAc, 3/1, Rf=0.50)
2,6-dioxaspiro[4.5]decan-10-one (300 mg, 1.82 mmol, 1.0 equiv.) under argon atmosphere is dissolved in dry THF (1.5 mL). LiHMDS (1.0 M in THF, 2.74 mL, 2.74 mmol, 1.5 equiv.) is added and the mixture is stirred for 10 min at rt. Methyl chloroformate (214 μL, 2.74 mmol, 1.5 equiv.) is added and the mixture is stirred at 65° C. overnight. After complete conversion, the reaction mixture is diluted aq. sat. NaHCO3 solution and water and extracted with DCM. The organic phase is concentrated under reduced pressure and purified by column chromatography to give the desired product K-17a.
The following intermediates K-17 (Table 31) are available in an analogous manner from the corresponding ketones. The crude product is purified by chromatography if necessary.
To a solution of NaHMDS (1 M, 692 mL, 1.0 equiv.) in THF (320 mL) at −78° C. under N2 atmosphere is added a solution of ((chloromethoxy)methyl)benzene (119 g, 760 mmol, 105 mL, 1.1 equiv.) and methyl oxolane-3-carboxylate (90.0 g, 760 mmol, 1.0 equiv.) in THF (160 mL) stirred for 3 h at −70-−78° C. After full conversion, the reaction is quenched with Aq. ammonium chloride solution, diluted with EtOAc and the phases are separated. The organic phase is washed with water, brine, dried over anhydrous sodium sulfate and concentrated under reduced pressure.
The residue is dissolved methanol (300 mL) and Pd/C (8.20 g, 10% wt, 0.1 equiv.) is added and hydrogenated. After full conversion, the reaction mixture is filtered under celatom and concentrated under reduced pressure to yield K-19a.
1H NMR: (CDCl3-d400 MHz) δ ppm 3.91-3.72 (m, 5H) 3.70-3.67 (m, 3H) 3.66 (s, 3H) 2.21 (ddd, J=12.8, 7.54, 5.82 Hz, 1H) 1.83 (dt, J=8.40, 7.41 Hz, 1H), TLC: Petroleum ether/EtOAc=1/1, Rf=0.43.
To a solution K-19a (46.0 g, 287 mmol, 1.0 equiv.) and Cs2CO3 (187 g, 574 mmol, 2.0 equiv.ACN (1000 mL) at 20° C. is added methyl acrylate (270 g, 3.14 mol, 282 mL, 10.9 equiv.) The reaction is quenched with water diluted with EtOAc (50.0 mL). The layers are separated, and the organic phase is washed with water, brine, dried over anhydrous sodium sulfate, and concentrated. The residue is purified by NP chromatography to yield K-12a.
1H NMR: (CDCl3-d-d 400 MHz) δ ppm 4.00 (d, J=9.20 Hz, 1H) 3.86-3.77 (m, 2H) 3.73-3.64 (m, 8H) 3.60 (s, 2H) 2.53 (t, J=12.8 Hz, 2H) 2.35-2.22 (m, 1H) 1.88 (dt, J=12.8, 7.24 Hz, 1H), TLC: Petroleum ether/EtOAc=1/1, Rf=0.7.
Sodium hydride (14.6 g, 610 mmol, 60% purity, 3.0 equiv.) is dissolved THF (250 mL) and stirred for 30 min at rt. A solution of K-20a (50.0 g, 203 mmol, 1.0 equiv.) in THF (250 mL) is slowly added dropwise, and the addition is completed stirred at 20-30° C. for 1 h. After complete conversion the reaction mixture solution is adjusted −o pH 6-7 with 2% dilute hydrochloric acid, extracted with dichloromethane (3×50 mL), and the organic layer are combined and washed with sat. sodium chloride solution (3×50 mL), dried over sodium sulfate and the filtrate is concentrated. The residue is purified by NP chromatography to yield K-21a.
1H NMR: (CDCl3-d, 400 MHz) δ ppm 4.42-3.38 (m, 12H), 2.42-2.23 (m, 1H), 1.96-1.85 (m, 1H), 1.62 (td, J=7.6, 12.6 Hz, 1H); TLC: Petroleum ether/EtOAc=5/1, Rf=0.6.
A-12a (60.0 mg, 0.259 mmol, 1.0 equiv.) is dissolved in dry MeOH (0.5 mL), sodium methoxide (25% in MeOH, 8.9 μL, 0.039 mmol, 0.15 equiv.) is added and the mixture is stirred for 30 min at rt. ammonium chloride (18.0 mg, 0.337 mmol, 1.3 equiv.) is added and the mixture is stirred for 40 min at 60° C. K-14a (95.1 mg, 0.285 mmol, 1.1 equiv.), dissolved in MeOH (0.5 mL) and triethylamine (0.181 mL, 1.30 mmol, 5.0 equiv.). is added and the mixture is stirred for 5 h at 90° C. in a microwave. After complete conversion, the reaction mixture is diluted with ACN and water, filtered and the crude product is purified by RP chromatography to give the desired product L-1a.
The following intermediates L-1 (Table 32 are available in an analogous manner from the corresponding keto esters. The crude product is purified by chromatography if necessary.
A-12a (373 mg, 1.61 mmol, 0.67 equiv.) is dissolved in dry MeOH (0.5 mL), sodium methoxide (25% in MeOH, 17 μL, 0.07 mmol, 0.03 equiv.) is added and the mixture is stirred for 15 min at rt. Ammonium chloride (129 mg, 2.41 mmol, 1.0 equiv.) is added and the mixture is stirred for 15 min at 60° C. K-17a (515 mg, 2.40 mmol, 1.0 equiv.), dissolved in MeOH (0.5 mL) and DBU (0.818 mL, 5.77 mmol, 2.4 equiv.). is added and the mixture is stirred overnight at 90° C. After complete conversion, the reaction mixture is diluted with water and extracted with DCM. The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product L-2a.
The following intermediates L-2 (Table 33) are available in an analogous manner from the corresponding keto esters. The crude product is purified by chromatography if necessary.
L-2b and L-2c are obtained as a diastereomeric mixture which can be separated by SFC column: DAICEL CHIRALPAKAD (250 mm*30 mm, 10 um); mobile phase: [CO2-iPrOH (0.1% NH3H2O)]; B %:35%, isocratic elution, to obtain L-2b (eluting 1st as peak1) and L-2c (eluting afterwards as peak2).
L-2d and L-2e are obtained as a diastereomeric mixture which can be separated by SFC column: Chiralcel OJ-3(50×4.6 mm I.D, 3 um); mobile phase:[CO2-MeOH(0.05% DEA)]; B %:5%-40%,
K-15a (30.0 g, 95.7 mmol, 1.0 equiv.) is dissolved in MeOH (300 mL) and DBU (58.3 g, 383.0. mmol, 4.0 equiv.′ and ′A-12a (30.7 g, 124.14 mmol, 1.30 equiv.) are added. The mixture is then stirred at 55° C. for 12 h and after complete conversion solvent is removed under reduced pressure. Lastly, the residue is triturated with MeOH at 20° C. for 30 min to delivery L-3a and L-3b as a diastereomeric mixture which can be separated by SFC (column: (s,s) WHELK-O1 (250 mm*30 mm, 10 um); mobile phase: [CO2+40% ACN/EtOH(0.1% NH3H2O)]; isocratic elution mode). Peak 1 (L-3a, HPLC-MS method S, tret=0.47 min; [M−H]−=511) is taken further in the next synthetic steps.
A-27b (3 g, 11.31 mmol, 1.0 equiv.) and K-21a (3.45 g, 16.11 mmol, 1.4 equiv.) are diluted in 1,4-dioxane (30 mL) and DBU (6.02 g, 39.57 mmol, 5.96 mL, 3.5 equiv.) is then added. Next, the reaction is stirred at 70° C. for 16 h. After complete conversion the pH of the reaction mixture is adjusted to 4-5 with HCl (1M) and extracted with EtOAc (50 mL). The combined organic layers are then washed with brine (30 mL×2), dried over Na2SO4, filtered and concentrated under reduced pressure to obtain the diastereomeric mixture of L-4a and its diastereomer as a residue.
The diastereomeric mixture is then separated via chiral SFC (column: DAICEL CHIRALCEL OD(250 mm*30 mm, 10 um); mobile phase: [CO2+50% ACN/MeOH(0.1% NH3H2O)], isocratic elution mode), and then further separated by SFC (column: (s,s) WHELK-O1 (250 mm*30 mm, 10 um); mobile phase: [CO2+42% ACN/i-PrOH(0.1% NH3H2O)], isocratic elution mode) to obtain L-4a (HPLC-MS method: AA, tret=0.38 min; [M−H]−=430) which is used for the following steps as the first eluting peak before the other diastereomer.
L-2b (200 mg, 0.486 mmol, 1.0 equiv.) and N,N-bis(trifluoromethylsulfonyl)aniline (213 mg, 0.583 mmol, 1.2 equiv.) are dissolved in DMA (2.0 mL), DIPEA (248.0 μL, 1.458 mmol, 3.0 equiv.) is added and the mixture is stirred for 30 min at rt. After complete conversion, the crude product is purified by RP chromatography to give the desired product M-1a.
The following intermediates M-1 (Table 34) are available in an analogous manner. The crude product is purified by chromatography if necessary.
M-1a (226 mg, 0.416 mmol, 1.0 equiv.) is dissolved in ACN (4.5 mL), DIPEA (217.3 μL, 1.25 mmol, 3.0 equiv.) and (S)-1-((S)-1-methyl pyrrolidin-2-yl) ethan-1-ol (525.6 μL, 3.74 mmol, 9.0 equiv.) are added and the mixture is stirred for 4 h at 80° C. After complete conversion, the reaction mixture is diluted with DMF and the crude product is purified by RP chromatography to give the desired product M-2a.
The following intermediates M-2 (Table 35) are available in an analogous manner. The crude product is purified by chromatography if necessary.
M-1a (18.68 g; 31.8 mmol, 1.0 equiv.) is dissolved in DCM (180 mL), then HCl, 37% in water (18.4 mL; 223 mmol, 7.0 equiv.) is slowly added. The reaction is then stirring vigorously for 5 min at room temperature. After full conversion to the desired product, the reaction mixture is carefully neutralized with a sat. NaHCO3 solution and the aq. phase is extracted 3 times with DCM. The organic phases are combined, dried over MgSO4 and the solvent is completely evaporated. The residue is further diluted in ACN and purified via RP chromatography to give the desired product M-3a (HPLC-MS method: N, tret=0.81 min; [M−H]−=430)
M-1d (575 mg, 0.89 mmol, 1.0 equiv) is dissolved in DCM (5 mL) and HCl (0.5 mL, 37% in water, 5.81 mmol, 6.5 equiv) is added. The reaction is stirred at room temperature overnight and after that neutralized with NaHCO3 (sat.) and extracted with DCM. The crude product M-4a (HPLC-MS method: C, tret=0.60 min; [M−H]−=429) is then utilized in the next reaction without further purification.
M-1e (127 mg; 0.23 mmol; 1.0 equiv.) is dissolved in HCl 4 M in Dioxan (0.28 μl; 1.14 mmol; 5.0 equiv.) and stirred at room temperature over night. After full conversion, the mixture is quenched with sat. NaHCO3 solution, extracted three times with DCM and evaporated to give the crude product which is then purified by RP chromatography to deliver M-5a (HPLC-MS method: C, tret=0.73 min; [M−H]−=448) as a pure product.
To a solution of M-4a (100 mg, 0.25 mmol, 1.0 equiv) in DCM (1 mL), 3 oxetanone (26 mg, 0.35 mmol, 1.5 equiv.) and sodium triacetoxyborohydride (78 mg, 0.35 mmol, 1.5 equiv.) are added and the mixture is stirred for 30 min at room temperature. The reaction is then quenched with water, DCM is removed under reduced pressure and the residue is further diluted in ACN and purified via RP chromatography to give the desired product M-6a (HPLC-MS method: C, tret=0.69 min; [M−H]−=485).
M-3a (4.10 g, 9.54 mmol, 1.0 equiv.) is dissolved in THF (130 mL) and B-5a (1.98 g, 12.4 mmol, 1.3 equiv.) is added. The mixture is cooled to 0° C. and NaOtBu (2.0M in THF, 6.44 mL, 12.9 mmol, 1.45 equiv.) is added slowly. After 15 min the reaction is allowed to warm to room temperature and is stirred for 1 h. After complete conversion, the reaction is diluted with 50 mL of a sat. solution of NaHCO3 and 100 mL of water and extracted two time with DCM. The organic phases are combined, solvent is removed and the obtained crude product M-7a is used in the next step without further purification.
The following intermediates M-7 (Table 36) are available in an analogous manner. The crude product is purified by chromatography if necessary.
L-1a (458 mg, 0.863 mmol, 1.0 equiv.) is dissolved in dry DMA (4.5 mL), N,N-bis(trifluoromethylsulfonyl)aniline (373 mg, 1.036 mmol, 1.2 equiv.) and DIPEA (451 μL, 2.59 mmol, 3.0 equiv.) are added and the mixture is stirred for 30 min at rt. (S)-1-((S)-1-methylpyrrolidin-2-yl)ethan-1-ol (359 μL, 2.59 mmol, 3.0 equiv.) dissolved in dry DMA (4.5 mL) is added and the mixture is stirred for 1 h at 60° C. After complete conversion, the reaction mixture is filtered and the crude product is purified by RP chromatography to give the desired product M-8a (HPLC-MS method: A, tret=1.66 min; [M+H]+=642).
M-8a (277 mg, 0.422 mmol, 1.0 equiv.) is dissolved in THF (30 mL) and MeOH (0.3 mL), palladium (10% on carbon, 42.4 mg, 0.1 equiv.) is added and the reaction is stirred for 7 h at rt under 3 bar hydrogen atmosphere. After complete conversion, the reaction mixture is filtered over Celite® and concentrated under reduced pressure. The residue is dissolved in DMSO and purified by RP chromatography to give product M-9a (HPLC-MS method: A, tret=1.27 min; [M+H]+=508).
M-9a (100 mg, 0.197 mmol, 1.0 equiv.) and triethylamine (82.4 μL, 0.591 mmol, 3.0 equiv.) are dissolved in in dry 2-propanol (1.0 mL), 2-fluoropyrimidine (59.76 mg, 0.591 mmol, 3.0 equiv.) are added and the mixture is stirred overnight at 100° C. in a microwave. After complete conversion, the reaction mixture is extracted with DCM/water. The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product M-10a (HPLC-MS method C: tret=0.87 min; [M+H]+=586).
L-1b (79.5 mg, 0.20 mmol, 1.0 equiv.), sulfur (13.0 mg, 0.40 mmol, 2.0 equiv.) and malononitrile (30.0 mg, 0.45 mmol, 2.2 equiv.) are dissolved in EtOH (2 mL), ammonium acetate (31 mg, 0.40 mmol, 2.0 equiv.) is added and the mixture is stirred at 70° C. overnight under argon atmosphere. After complete conversion, the reaction mixture is extracted with DCM/water. The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product N-1a.
The following examples N-1 (Table 37) are available in an analogous manner from the corresponding ketones. The crude product is purified by chromatography if necessary.
N-1b is obtained as a diastereomeric mixture which can be separated by RP HPLC (XBridge Prep C18 10 μm OBD™ 50×150 mm column, eluent: A=>2 mL of 25% aq. NH4OH+5 mL of NH4HCO3 stock solution (=158 g NH4HCO3+1 L H2O) are replenished to 1 L water, B=>ACN) with the desired analogue eluting first.
M-10a (14.0 mg, 0.024 mmol, 1.0 equiv.), malonodinitrile (5.64 mg, 0.084 mmol, 3.5 equiv.), ammonium acetate (18.43 mg, 0.239 mmol, 10.0 equiv.), and sulfur (3.83 mg, 0.120 mmol, 5.0 equiv.) are combined in 2-propanol (0.2 mL) and stirred for 2 h at 50° C. After complete conversion the mixture is diluted with DMF and purified by basic RP chromatography to give the desired products II-7a and II-7b.
Instead of 2-propanol, dry ethanol is used as solvents in the synthesis of compounds II-9-II-14. For this same compounds, the reaction is stirred at 80° C. instead of 50 00.
The following examples II (Table 38) are available in an analogous manner from the corresponding ketones. The crude product is purified by chromatography if necessary.
The diastereomeric mixture of II-7a and II-7b can be separated via chiral HPLC (Chiralpack ID, 250×20 mm, 5 μm; solvent: heptane/ethanol 80:20) to obtain II-7a (eluting 1st as peak 1) and II-7b (eluting afterwards as peak 2).
N-1a (30.8 mg, 0.06 mmol, 1.0 equiv.), N,N-bis(trifluoromethylsulfonyl) aniline (35 mg, 0.10 mmol, 1.5 equiv.) and DIPEA (90 μL, 0.52 mmol, 8.0 equiv.) are dissolved in ACN and stirred for 1 h at rt. (S)-1-((S)-1-methylpyrrolidin-2-yl) ethan-1-ol (51.0 mg, 0.39 mmol, 6.0 equiv.) is added and the mixture is stirred overnight at 60° C. After complete conversion the mixture is diluted with DMF and purified by basic RP chromatography to give the desired product II-18.
The following examples II (Table 39) are available in an analogous manner. The crude product is purified by chromatography if necessary.
To a solution of II-19 (51 g, 0.071 mmol, 1.0 equiv.) in dry methanol (0.5 mL) is added HCl solution (4 M in 1,4-dioxane, 0.106 mL, 0.425 mmol, 6.0 equiv.) and the mixture is stirred at 65° C. for 1 h. After complete conversion the mixture is diluted with ACN and water, basified with NaHCO3 solution and purified via RP chromatography to give the desired product 11-20 (HPLC method: A, tret=1.37 min; [M+H]+=620).
A-27a (65 g, 223.1 mmol, 1.00 equiv.) and DBU (118.9 g, 780.9 mmol, 3.50 equiv.) are dissolved in 1,4-dioxane (650 mL) and the mixture is degassed and purged with N2 at 15.0° C. Next, K-21a (62.1 g, 290.08 mmol, 1.30 equiv.) is added and the mixture is stirred at 70° C. for 15 hrs. After complete conversion, the mixture is allowed to warm up to room temperature and 1 L of water is used to quench the reaction. Next an extraction with EA (500 mL*3) is performed and the combined organic layers is washed with brine (500 mL), dried over sodium sulfate, filtered and concentrated under reduced pressure. Lastly, the obtained residue is purified by column chromatography (SiO2, EtOAc=1, Rf=0.43) to deliver a racemic mixture of O-1a and O-1b as a yellow solid. (UPLC-MS Method: AA (tret: 0.428 min, [M+H]+: 456)
The racemic mixture is further separated by SFC (column: (s,s) WHELK-O1 (250 mm*30 mm, 10 um); mobile phase: [CO+35% ACN/MeOH (0.1% NH3H2O)]; isocratic elution mode). Peak 1 (O-1a) is further used into the next steps.
A-27a (30.0 g, 95.7 mmol, 1.00 equiv.) is dissolved in MeOH (300 mL) and DBU (58.3 g, 383.0 mmol, 4.00 equiv.) and K-15a (36.0 g, 123.6 mmol, 1.30 equiv.) are added. The mixture is then stirred at 55° C. for 12 h and after complete conversion solvent is removed under reduced pressure. Lastly, the residue is triturated with MeOH at 20° C. for 30 min to give the desired product O-2a and O-2b as a diastereomeric mixture which can be separated by SFC (column: DAICEL CHIRALPAK AD (250 mm*50 mm, 10 um); mobile phase: [CO2+45% ACN/i-PrOH(0.1% NH3H2O)], isocratic elution mode. Peak 1 (O-2a) is taken further in the next synthetic steps.
O-1a (200 mg, 0.486 mmol, 1.0 equiv.) and N,N-bis(trifluoromethylsulfonyl)aniline (213 mg, 0.583 mmol, 1.2 equiv.) are dissolved in DMA (2.0 mL), DIPEA (248.0 μL, 1.458 mmol, 3.0 equiv.) is added and the mixture is stirred for 1 h at rt. After complete conversion, the crude product is purified by RP chromatography to give the desired product P-1a.
The following intermediates P-1 (Table 40) are available in an analogous manner. The crude product is purified by chromatography if necessary.
P-1a (18.68 g; 31.79 mmol, 1.0 equiv.) is dissolved in DCM (180 mL), then HCl, 37% in water (18.43 mL; 222.56 mmol, 7.0 equiv.) is slowly added. The reaction is then stirring vigorously for 5 h at room temperature. After full conversion to the desired product, the reaction mixture is carefully neutralized with a sat. NaHCO3 solution and the aq. phase is extracted 3 times with DCM. The organic phases are combined, dried over MgSO4 and the solvent is completely evaporated. The residue is further diluted in ACN and purified via RP chromatography to give the desired product P-2a. (HPLC-MS method: A; tret=1.40 min; [M+H]+=430)
P-1b (1.2 g, 1.75 mmol, 1.0 equiv) is dissolved in DCM (10 mL) and HCl (1 mL, 37% in water, 11.53 mmol, 6.6 equiv) is added. The reaction is stirred at 50° C. overnight and after that neutralized with NaHCO3 (sat.) and extracted twice with DCM. The crude product is purified via RP chromatography to give the desired product P-3a (HPLC-MS method: A; tret=1.24 min; [M+H]+=429)
To a solution of P-3a (365 mg, 0.851 mmol, 1.0 equiv) in DCM (4 mL), 3 oxetanone (95 mg, 1.277 mmol, 1.5 equiv.) and sodium triacetoxyborohydride (285 mg, 1.277 mol, 1.5 equiv.) are added followed by 2 drops of TFA. The mixture is stirred for 1 h at room temperature. The reaction is then that quenched with NaHCO3 (sat.) and extracted twice with DCM to give P-4a (HPLC method: C; tret=0.708 min; [M+H]+=485) as a product. The crude product is the utilized in the next reaction without further purification.
P-2a (3.4 g, 7.862 mmol, 1.0 equiv.) is dissolved in THF (110 mL) and (S)-1-((S)-1-methylpyrrolidin-2-yl)ethan-1-ol (1435 μL, 8.255 mmol, 1.3 equiv.) is added and the mixture. The mixture is cooled to 0° C. and NaOtBu is added slowly. After 15 min the reaction is allowed to warm to room temperature and is stirred for 1 h. After complete conversion, the reaction is diluted with 50 mL of a sat. solution of NaHCO3 and 100 mL of water and extracted two times with DCM. The organic phases are combined, solvent is removed to give P-5a as a product. The crude product is used in the next step without further purification.
The following examples P-5 (Table 41) are available in an analogous manner from the corresponding chlorides. The crude product is purified by chromatography if necessary.
P-5a (104.5 mg, 0.20 mmol, 1.0 equiv.), sulfur (13.0 mg, 0.40 mmol, 5.0 equiv.) and malononitrile (30.0 mg, 0.45 mmol, 3.5 equiv.) are dissolved in 2-propanol (1 mL), ammonium acetate (31 mg, 0.40 mmol, 10.0 equiv.) is added and the mixture is stirred at 50° C. for 2 h under argon atmosphere. After complete conversion, the reaction mixture is extracted with DCM/NaHCO3 (sat.). The organic phase is concentrated under reduced pressure and purified by RP chromatography to give the desired product 1-54.
The following examples I (Table 42) are available in an analogous manner from the corresponding ketones. The crude product is purified by chromatography if necessary.
This assay can be used to examine the potency with which compounds according to the invention binding to (mutated) KRAS inhibit the protein-protein interaction between SOS1 and (mutated) KRAS e.g. KRAS G12C, KRAS G12D. This inhibits the GEF functionality of SOS1 and locks the corresponding (mutated) KRAS protein in its inactive, GDP-bound state. Low IC50 values in this assay setting are indicative of strong inhibition of protein-protein interaction between SOS1 and KRAS:
These assays measure the inhibitory effect of compounds on KRAS mutant protein-protein interactions using the Alpha Screen technology by Perkin Elmer.
The following (mutant) enzyme forms of KRAS and interacting proteins are used in these assays at the given concentrations:
KRAS (G12D) 1-169, N-terminal 6His-tag, C-terminal avi-tag (Xtal BioStructures, Inc.); final assay concentration 10 nM and SOS1 564-1049, N-terminal 229 GST-tag, TEV cleavage site (Viva Biotech Ltd); final assay concentration 5 nM;
KRAS (G12C) 1-169, N-terminal 6His-tag for purification, cleaved off, C-terminal avi-tag, biotinylated, mutations: C51S, C80L, C118S (in house); final assay concentration 7.5 nM and SOS1 564-1049, N-terminal 229 GST-tag, TEV cleavage site (Viva Biotech Ltd); final assay concentration 5 nM;
Test compounds dissolved in DMSO are dispensed onto assay plates (Proxiplate 384 PLUS, white, PerkinElmer; 6008289) using an Access Labcyte Workstation with the Labcyte Echo 55x. For the chosen highest assay concentration of 100 μM, 150 nL of compound solution are transferred from a 10 mM DMSO compound stock solution. A series of eleven fivefold dilutions per compound are transferred to the assay plate, compound dilutions are tested in duplicates. DMSO are added as backfill to a total volume of 150 nL.
The assays run on a fully automated robotic system in a darkened room below 100 Lux. To 150 nl of compound dilution 10 μl of a mix including KRAS mutant protein, SOS1 (final assay concentrations see above) and GDP nucleotide (Sigma G7127; final assay concentration 10 μM) in assay buffer (1×PBS, 0.1% BSA, 0.05% Tween 20) are added into columns 1-24.
After 30 minutes incubation time 5 μl of Alpha Screen bead mix in assay buffer are added into columns 1-23. Bead mix consists of AlphaLISA Glutathione Acceptor Beads (PerkinElmer, Cat No AL109) and AlphaScreen Streptavidin Donor Beads (PerkinElmer Cat No 6760002) in assay buffer at a final assay concentration of 10 μg/mL each.
Plates are kept at room temperature in a darkened incubator. After an additional 60 minutes incubation time the signal is measured in a PerkinElmer Envision HTS Multilabel Reader using the AlphaScreen specs from PerkinElmer.
Each plate contains up to 16 wells of a negative control depending on the dilution procedure (platewise or serial) (DMSO instead of test compound; with KRAS mutant::SOS1 GOP mix and bead mix; column 23) and 16 wells of a positive control (DMSO instead of test compound; with KRAS mutant::SOS1 GOP mix w/o bead mix; column 24).
As internal control known inhibitors of KRAS mutant::SOS1 interaction can be measured on each compound plate.
IC50 values are calculated and analyzed with Boehringer Ingelheim's MEGALAB C50 application using a 4 parametric logistic model.
Tables of example compounds disclosed herein contain IC50 values determined using the above assays (see Table 43).
This assay measures the inhibitory effect of compounds on KRAS::CRAF protein::protein interactions in the presence of GTP using Time-resolved fluorescence energy transfer (TR-FRET).
The following wildtype or mutant alleles of KRAS and CRAF protein have been used at the given concentrations:
KRAS (G12D) 1-169, N-terminal 6His-tag, C-terminal avi-tag (Xtal BioStructures, Inc.); final assay concentration 15 nM,
KRAS (G12V) 1-169, C118S, N-terminal 6His-tag, C-terminal avi-tag (Boehringer Ingelheim in house protein prep); final assay concentration 15 nM.
KRAS (WT) 1-169, N-terminal 6His-tag, C-terminal avi-tag (Boehringer Ingelheim in house protein prep); final assay concentration 15 nM.
CRAF Ras binding domain, GST-tag, TEV cleavage site (Boehringer Ingelheim in house protein prep); final assay concentration 15 nM.
Test compounds dissolved in DMSO were dispensed onto assay plates (Proxiplate 384 PLUS, white, PerkinElmer; 6008289) using an Access Labcyte Workstation with the Labcyte Echo 55x. For the chosen highest assay concentration of 100 μM, 150 nl of compound solution was transferred from a 10 mM DMSO compound stock solution. A series of eleven fivefold dilutions per compound was transferred to the assay plate, compound dilutions were tested in duplicates. DMSO was added as backfill to a total volume of 150 nl.
The assay runs on a fully automated robotic system. To 150 nl of compound dilution 15 μl of a mix including the respective KRAS allele, CRAF (e.g. KRAS (G12D)::CRAF; final assay concentrations see above), GTP nucleotide (Sigma G8877; final assay concentration 10 μM), Lance Eu—W1024 labeled Streptavidin (PerkinElmer, Cat No AD0063; final assay concentration 1.5 nM) and Anti-GST surelight APC (PerkinElmer, Cat No AD0059G; final assay concentration 30 nM) in assay buffer (1×PBS, 0.1% BSA, 0.05% Tween 20) are added into columns 1-23 and 15 μL of the solution without the respective KRAS allele were added to row 24.
Plates are kept at room temperature in a darkened incubator. After 60 minutes incubation time the signal is measured in a PerkinElmer Envision HTS Multilabel Reader using the TR-FRET LANCE Ultra specs from PerkinElmer.
Each plate contains 16 wells of a negative control (diluted DMSO instead of test compound; w respective KRAS allele; column 23) and 16 wells of a positive control (diluted DMSO instead of test compound; w/o respective KRAS allele; column 24).
IC50 values were calculated and analyzed with Boehringer Ingelheim's MEGALAB 50 application using a 4 parametric logistic model.
Tables of example compounds disclosed herein contain IC50 values determined using the above assays (see Table 44).
Ba/F3 cells are ordered from DSMZ (ACC300, Lot17) and grown in RPMI-1640 (ATCC 30-2001)+10% FCS+10 ng/mL IL-3 at 37° C. in 5% 002 atmosphere. Plasmids containing KRASG12 mutants (i.e. G121D, G12C, G12V) are obtained from GeneScript. To generate KRASG12-dependent Ba/F3 models, Ba/F3 cells are transduced with retroviruses containing vectors that harbor KRASG12 isoforms. Platinum-E cells (Cell Biolabs) are used for retrovirus packaging. Retrovirus is added to Ba/F3 cells. To ensure infection, 4 μg/mL polybrene is added and cells are spinfected. Infection efficiency is confirmed by measuring GFP-positive cells using a cell analyzer. Cells with an infection efficiency of 10% to 20% are further cultivated and puromycin selection with 1 μg/mL is initiated. As a control, parental Ba/F3 cells are used to show selection status. Selection is considered successful when parental Ba/F3 cells cultures died. To evaluate the transforming potential of KRASG12 mutations, the growth medium is no longer supplemented with IL-3. Ba/F3 cells harboring the empty vector are used as a control. Approximately ten days before conducting the experiments, puromycin is left out.
For proliferation assays, Ba/F3 cells are seeded into 384-well plates at 1.5×1 (M cells/60 μL in growth media (RPMI-1640+10% FCS). Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Treated cells are incubated for 72 h at 37° C. with 5% CO2 and cell viability is measured in the PerkinElmer Envision Multimode Reader using AlamarBlue™ (ThermoFisher) viability stain. Viability (stated as percent of control) is defined as relative fluorescence units RFU of each well divided by the RFU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
IC50 values of representative compounds according to the invention measured with this assay are presented in Table 45.
Additional Proliferation Assays with Mutant Cancer Cell Lines
NCI-H358 cells (ATCC No. CRL-5807) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 200 cells per well in 60 μl RPMI-1640 ATCC-Formulation (Gibco #A10491)+10% FCS (fetal calf serum). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
AsPC-1 cells (ATCC CRL-1682) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 500 cells per well in 60 μl RPMI ATCC-Formulation (PAN P04-18047)+10% FCS (fetal calf serum, HyClone, SH30084.03). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
MKN1 cells (JCRB0252) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 200 cells per well in 60 μl RPMI-1640 ATCC-Formulation (Gibco #A10491)+10% FCS (fetal calf serum, HyClone, SH30084.03). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
SK—CO-1 cells (ATCC HTB-39) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 500 cells per well in 60 μl EMEM (Sigma M5650)+10% FCS (fetal calf serum, HyClone, SH30084.03). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
NCI-H727 cells (ATCC No. CRL-5815) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 500 cells per well in 60 μl RPMI ATCC-Formulation (PAN P04-18047)+10% FCS (fetal calf serum, HyClone, SH30084.03). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
LOVO cells (ATCC No. CCL-229) are dispensed into black 384-well plates, flat and clear bottom (Greiner, PNr. 781091) at a density of 1000 cells per well in 60 μl DMEM (Sigma D6429)+10% FCS (fetal calf serum, HyClone, SH30084.03). Cells are incubated overnight at 37° C. in a humidified tissue culture incubator at 5% CO2. Compounds (10 mM stock in DMSO) are added at logarithmic dose series using the ECHO acoustic liquid handler system (Beckman Coulter), normalizing for added DMSO and including DMSO controls. All compound treatments are performed in technical duplicates. For the TO time point measurement, untreated cells are analyzed at the time of compound addition. Plates are incubated for 120 h, and cell viability is measured using CellTiter-Glo luminescent cell viability reagent (Promega product code G7570). Viability (stated as percent of control) is defined as relative luminescence units RLU of each well divided by the RLU of cells in DMSO controls. IC50 values are determined from viability measurements by non-linear regression using a four-parameter model.
IC50 values of representative compounds according to the invention measured with these assays in the indicated cell lines are presented in Table 46.
The metabolic degradation of the test compound is assayed at 37° C. with pooled liver microsomes (mouse (MLM), rat (RLM) or human (HLM)). The final incubation volume of 48 μL per time point contains TRIS buffer (pH 7.5; 0.1 M), magnesium chloride (6.5 mM), microsomal protein (0.5 mg/mL for mouse/rat, 1 mg/mL for human specimens) and the test compound at a final concentration of 1 μM. Following a short preincubation period at 37° C., the reactions are initiated by addition of 12 μL beta-nicotinamide adenine dinucleotide phosphate, reduced form (NADPH, 10 mM) and terminated by transferring an aliquot into solvent after different time points (0, 5, 15, 30, 60 min). Additionally, the NADPH-independent degradation is monitored in incubations without NADPH, terminated at the last time point by addition of acetonitrile. The quenched incubations are pelleted by centrifugation (4,000 rpm, 15 min). An aliquot of the supernatant is assayed by LC-MS/MS to quantify the concentration of parent compound in the individual samples.
In vitro intrinsic clearance (CLint,in vitro) is calculated from the time course of the disappearance of the test drug during the microsomal incubation. Each plot is fitted to the first-order elimination rate constant as C(t)=C0*exp(−ke*t), where C(t) and C0 are the concentration of unchanged test drug at incubation time t and that at preincubation and ke is the disappearance rate constant of the unchanged drug. Subsequently, CLint, in vitro (μL min−1·amount protein) values are converted to predicted CLint,in vivo (mL min−1·kg−1) from incubation parameters according to the equation CLint, in vivo=CLint, in vitro×(incubation volume (mL)/amount protein (mg))×(amount protein (mg)/g liver tissue)×(liver weight/body wt.).
For better across species comparison the predicted clearance is expressed as percent of the liver blood flow [% QH] (mL min−1·kg−1) in the individual species. In general, high stability (corresponding to low % QH) of the compounds across species is desired.
Table 47 shows metabolic stability data obtained with the disclosed assay in HLM for a selection of compounds (I) according to the invention.
Binding of test compounds to plasma was determined using equilibrium dialysis (ED) and quantitative mass spectrometry interfaced with liquid chromatography (LC-MS). In brief, ED was performed with dialysis devices consisting of two chambers separated by a semipermeable membrane with a molecular weight cut-off of 5-10 kg/mol. One chamber was filled with 10% FCS in PBS containing 1-10 μmol/L test compound and the other chamber was filled with phosphate-buffer saline (PBS) with or without dextran. The dialysis chamber was incubated for 3-5 hours at 37° C. After incubation, protein was precipitated from aliquots of each chamber and the concentration of test compound in the supernatant of the plasma-containing compartment (cserum) and of the buffer-containing compartment (cbuffer) was determined by LC-MS. The fraction of unbound test compound (not bound to plasma) (f) was calculated according to the following equation:
Table 48 shows metabolic stability data obtained with the disclosed assay for a selection of compounds (I) according to the invention.
The time dependent inhibition towards CYP3A4 is assayed in human liver microsomes (0.02 mg/mL) with midazolam (15 μM) as a substrate. The test compounds and water control (wells w/o test compound) are preincubated in presence of NADPH (1 mM) with human liver microsomes (0.2 mg/mL) at a concentration of 25 uM for 0 min and 30 min. After preincubation, the incubate is diluted 1:10 and the substrate midazolam is added for the main incubation (15 min). The main incubation is quenched with acetonitrile and the formation of hydroxy-midazolam is quantified via LC/MS-MS. The formation of hydroxy-midazolam from the 30 min preincubation relative to the formation from the 0 min preincubation is used as a readout. Values of less than 100% mean that the substrate midazolam is metabolized to a lower extent upon 30 min preincubation compared to 0 min preincubation. In general low effects upon 30 min preincubation are desired (corresponding to values close to 100%/not different to the values determined with water control).
Table 49 shows data obtained with the disclosed assay for a selection of compounds (1) according to the invention.
A 10 mM DMSO stock solution of a test compound is used to determine its aq. solubility. The DMSO solution is diluted with an aq. medium (McIlvaine buffer with pH=4.5 or 6.8) to a final concentration of 250 μM. After 24 h of shaking at ambient temperature a potentially formed precipitate is removed by filtration. The concentration of the test compound in the filtrate is determined by LC-UV methods by calibrating the signal to the signal of a reference solution with complete dissolution of the test compound in acetonitrile/water (1:1) with known concentration.
Table 50 shows data obtained with the disclosed assay for a selection of compounds (1) according to the invention.
The assay provides information on the potential of a compound to pass the cell membrane, on the extent of oral absorption as well as on whether the compound is actively transported by uptake and/or efflux transporters. Permeability measurements across polarized, confluent Caco-2 cell monolayers grown on permeable filter supports (Corning, catalog #3391) are used. 10 μM test compound solution in assay buffer (128.13 mM NaCl, 5.36 mM KCl, 1 mM MgSO4, 1.8 mM CaCl2), 4.17 mM NaHCO3, 1.19 mM Na2HPO4, 0.41 mM NaH2PO4, 15 mM 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid (HEPES), 20 mM glucose, pH 7.4) was added to the donor compartment of the cell chamber containing a monolayer of Caco-2 cells in between the donor and the receiver compartment. The receiver and donor compartments contain 0.25% bovine serum albumine (BSA) in assay buffer. Passive diffusion and/or active transport of compounds across the monolayer is measured in both apical to basolateral (a-b) and basolateral to apical (b-a) direction. a-b permeability (PappAB) represents drug absorption from the intestine into the blood and b-a permeability (PappBA) drug secretion from the blood back into the intestine via both passive permeability as well as active transport mechanisms mediated by efflux and uptake transporters that are expressed on the Caco-2 cells. After a pre-incubation of 25-30 min at 37° C., at predefined time points (0, 30, 60 and 90 min), samples were taken from the receiver and donor compartment, respectively. Concentrations of test compounds in samples were measured by HPLC/MS/MS, samples from the donor compartment were diluted 1:50 (v:v) with assay buffer, samples from receiver compartment were measured without dilution.
Apparent permeabilities in a-b (PappAB) and b-a (PappBA) directions are calculated according to the formula:
Caco-2 efflux ratios (ER) are calculated as the ratio of PappBA/PappAB.
Table 51 shows data obtained with the disclosed assay for a selection of compounds (1) according to the invention.
The formulation examples which follow illustrate the present invention without restricting its scope:
The finely ground active substance, lactose and some of the corn starch are mixed together. The mixture is screened, then moistened with a solution of polyvinylpyrrolidone in water, kneaded, wet-granulated and dried. The granules, the remaining corn starch and the magnesium stearate are screened and mixed together. The mixture is compressed to produce tablets of suitable shape and size.
The finely ground active substance, some of the corn starch, lactose, microcrystalline cellulose and polyvinylpyrrolidone are mixed together, the mixture is screened and worked with the remaining corn starch and water to form a granulate which is dried and screened. The sodium carboxymethyl starch and the magnesium stearate are added and mixed in and the mixture is compressed to form tablets of a suitable size.
The active substance, lactose and cellulose are mixed together. The mixture is screened, then either moistened with water, kneaded, wet-granulated and dried or dry-granulated or directly final blend with the magnesium stearate and compressed to tablets of suitable shape and size. When wet-granulated, additional lactose or cellulose and magnesium stearate is added and the mixture is compressed to produce tablets of suitable shape and size.
The active substance is dissolved in water at its own pH or optionally at pH 5.5 to 6.5 and sodium chloride is added to make it isotonic. The solution obtained is filtered free from pyrogens and the filtrate is transferred under aseptic conditions into ampoules which are then sterilised and sealed by fusion. The ampoules contain 5 mg, 25 mg and 50 mg of active substance.
| Number | Date | Country | Kind |
|---|---|---|---|
| 23175994.5 | May 2023 | EP | regional |
| EP23175999.4 | May 2023 | EP | regional |
