Patent Application
20250049798
The present application belongs to the field of medicine. The invention provides compounds that degrade Wee1 protein kinase or pharmaceutically acceptable salts thereof, and uses thereof in the treatment of proliferative disorders. The invention also provides pharmaceutical compositions comprising a compound described herein or a salt thereof, and a pharmaceutically acceptable carrier.
Cell cycle refers to the entire process of cell division, which is divided into interphase composed of DNA synthesis prophase (G1 phase), DNA synthesis phase (S phase) and DNA synthesis anaphase (G2 phase), and mitosis (M phase) composed of prophase, prometaphase, metaphase, anaphase, and telophase. Cell cycle checkpoint is a key point that regulates the cell cycleand ensures that every event of the cell cycle (such as DNA replication and chromosome segregation) is completed in an orderly and quality manner. The main cell cycle checkpoints are: 1) the G1/S checkpoint, which checks and controls cells from the stationary G1 phase to the DNA synthesis phase; 2) the S-phase checkpoint, which checks whether DNA replication is completed; 3) the G2/M checkpoint, which checks and controls cells entering mitosis; 4) the Mid-anaphase checkpoint, which checks whether the spindle is properly assembled. The checkpoints will promptly sense and initiate repair when there are abnormalities such as DNA damage in certain processes of the cell cycle.
Damaged DNA is usually repaired at the G1/S checkpoint. P53 protein can repair DNA damage and prevent cells from entering S phase. Accumulation of irreversible DNA damage results in p53-dependent cell apoptosis. In normal cells, p53 protein is maintained at a low level. However, loss-of-function mutations in p53, which frequently occur in tumor cells, make G1/S checkpoint defective.
Thus, in p53 mutated tumor cells, cell cycle regulation (e.g. arresting the cell cycle or eliminating cells) relies on the G2/M checkpoint. Wee1 protein kinase is a key regulator for arresting the G2/M phase. Wee1 protein kinase is highly expressed in many cancer types. Inhibition or down-regulation of Wee1 protein kinase can lead to “mitotic catastrophe”, resulting in the apoptosis of tumor cells. The Wee1 kinase inhibitor AZD1775 has shown certain clinical activities in the treatment of tumors. However, there are still deficiencies such as low cell activity, large clinical toxicity and side effects, and poor patient tolerance.
The ubiquitin-proteasome system (UPS) is a major protein degradation system with high specificity and selectivity. The proteolysis targeting chimera (PROTAC) refers to a compound composed of a ligand of a protein of interest (POI), a ligand of an E3 ubiquitin ligase, and a linker connecting the two ligands. In other words, one end of the PROTAC molecule can bind to a POI, and the other end can bind to an E3 ubiquitin ligase, thus brining an E3 ubiquitin ligase in close proximity to a POI. This allows the E3 ubiquitin ligase targetted by the PROTAC molecule to “tag” multiple ubiquitin proteins to the POI through a process called polyubiquitination. Next, the tagged POI (polyubiquitinated POI) is recognized and degraded by the proteasome. This process does not require the POI ligand to occupy the binding site for a long time. A PROTAC molecule can rapidly induce the ubiquitination of the POI only by briefly occupying the POI site and E3 ubiquitin ligase site at the same time. Moreover, PROTAC can be recycled after the POI is degraded by the proteasome.
However, when compared to the drug-like properties of small-molecule inhibitors, many PROTAC drugs in general have several disadvantages including poor water solubility, poor membrane permeability, poor metabolic stability, low oral bioavailability, etc.
The proteolytic degradation of Wee1 can effectively blocks downstream signaling. The removal of Wee1 protein can favorably eliminate Wee1 kinase activity and any protein interaction or skeletal structure function associated with Wee1, providing wide therapeutic applications for the treatment of tumors.
The compound or a pharmaceutically acceptable salt thereof of the invention can effectively degrade Wee1 protein kinase, and showed improved water solubility, membrane permeability, metabolic stability and oral bioavailability.
The present invention provides proteolysis targeting chimera. The first embodiment of the present invention provides a compound represented by structural formula (I), or a derivative, pharmaceutically acceptable salt, isomer, solvate, hydrates adduct, complex or prodrug thereof:
W-L-D (I),
wherein:
represents an optionally substituted cyclic group;
represents a linking site;
wherein
represents an optionally substituted cyclic group;
In a second aspect of the first embodiment, W is:
wherein:
In a third aspect of the first embodiment, W is:
In a fourth aspect of the first embodiment, W is:
wherein
In a fifth aspect of the first embodiment, W is:
wherein
In a sixth aspect of the first embodiment, W is:
wherein
In a seventh aspect of the first embodiment, W is:
wherein
Preferably, W is:
In a eighth aspect of the first embodiment, W is:
In a ninth aspect of the first embodiment, W is:
R1, for each occurrence, is independently selected from absence, hydrogen, deuterium, halogen, hydroxyl, carbonyl, nitro, cyano, amino, mercapto, —COOH, C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, C1-C6 alkoxy, C2-C6 alkenyloxy, C2-C6 alkynyloxy, C2-C6 alkanoyl, C2-C6 alkylester, C1-C6 thioalkyl, C1-C6 haloalkyl, C1-C6 haloalkoxy, hydroxy C1-C6 alkyl, amino C1-C6 alkyl, (mono- and di-C1-C6 alkylamino) C0-C4 alkyl, —C0-C4 alkyl (C3-C7 cycloalkyl), —O—C0-C4 alkyl (C3-C7 cycloalkyl), C3-C12 heterocyclyl, C6-C12 aryl, and C5-C10 heteroaryl, wherein the cycloalkyl, heterocyclyl, aryl, and heteroaryl are optionally substituted with 1, 2, 3, or 4 halogens, hydroxyl, carbonyl, nitro, cyano, amino, mercapto, —COOH, or C1-C6 alkyl;
In a tenth aspect of the first embodiment, W is:
wherein
In a eleventh aspect of the first embodiment, W is:
In the twelfth aspect of the first embodiment, W is:
optionally substituted C6-C12 aryl or optionally substituted C5-C10 heteroaryl, wherein when the aryl or heteroaryl is substituted, the C6-C12 aryl and C5-C10 heteraryl are independently substituted with one or more substituents, preferably fluorine, chlorine, methyl, ethyl, methoxy, hydroxymethyl, haloalkyl.
Preferably, W is:
In a thirteenth aspect of the first embodiment, W is:
wherein
Preferably, W is:
In a fourteenth aspect of the first embodiment, W is:
wherein:
is a nitrogen-containing heteroaromatic ring selected from pyrrolyl, imidazolyl, pyrazolyl, triazolyl, or tetrazolyl.
In a fifteenth aspect of the first embodiment, W is:
wherein
Preferably, W is:
In a sixteenth aspect of the first embodiment, W is:
wherein
wherein Ra, Rb, Rc, Rd, Re, Rf, Rg, Rh, Ri, Rj, for each occurrence, are each independently selected from C1-6 alkyl, C3-5 cycloalkyl;
In a seventeenth aspect of the first embodiment, W is:
wherein
In an eighteenth aspect of the first embodiment, W is:
In a nineteenth aspect of the first embodiment, D is:
In a twentieth aspect of the first embodiment, D is.
wherein
In a 21st aspect of the first embodiment, D is:
Preferably, D is:
In a 22nd aspect of the first embodiment, D is:
wherein
In a 23rd aspect of the first embodiment, D is:
Preferably, D is:
In a 24th aspect of the first embodiment, D is:
Preferably, D is a group selected from:
In a 25th aspect of the first embodiment, D is:
is an aromatic ring or a heteroaromatic ring, wherein
Preferably, D is a group selected from:
In a 26th aspect of the first embodiment, D is:
is an aromatic ring or a heteroaromatic ring, wherein
In a 27th aspect of the first embodiment, D is:
In a 28th aspect of the first embodiment, D is:
wherein
Preferably, D is:
In a 29th aspect of the first embodiment, D is:
wherein
In a 30th aspect of the first embodiment, D is:
In a 31st aspect of the first embodiment, D is:
Preferably, D is a structure selected from:
In a 32nd aspect of the first embodiment, L is selected from:
In the second embodiment, the present invention relates to formula (II):
In the third embodiment, the present invention relates to formula (III):
In the fourth embodiment, the present invention relates to formula (IV):
In the fifth embodiment, the present invention relates to formula (V):
In the sixth embodiment, the present invention relates to formula (VI):
In the seventh embodiment, the present invention relates to formula (VIII):
Preferably, X15—R15 is CH, C or C—Cl; X14—R19 is CH, C or C—Cl; X13—R18 is CH, C or C—Cl; X12—R17 is CH, C or C—Cl; and/or X11—R16 is CH, C or Cl.
Also preferably, B1—R6 are CH2; and/or B2—R5 is NH, CH2 or N—CH3.
Also preferably, Y1 is CH or N.
Also preferably, Y5 is CH or C; Y6 is CH or C; Y7 is CH or C; and/or Y8 is CH or C.
Also preferably, Y41 is CH or N; Y42 is CH2 or ═O; and/or Y43 is NH, CH2 or N—CH3.
In the eighth embodiment, the present invention relates to formula (IX):
Preferably, X15-R15 is CH, C or C—Cl; X14—R19 is CH, C or C—Cl; X13—R8 is CH, C or C—Cl; X12—R17 is CH, C or C—Cl; and/or X11—R16 is CH, C or Cl.
Also preferably, B1—R6 are CH2; and/or B2—R5 is NH, CH2 or N—CH3.
Also preferably, Y1 is CH or N.
Also preferably, Y5 is CH or C; Y6 is CH or C; Y7 is CH or C; and/or Y8 is CH or C.
Also preferably, Y4 is CH2 or ═O.
In the nineth embodiment, the present invention relates to formula (XIV):
In the tenth embodiment, the present invention relates to formula (XV):
In the eleventh embodiment, the present invention relates to formula (XVII):
Preferably, X8-R11 is CH; R8 is Cl; and/or R9 is Cl.
Also preferably, E6 is H or absent; and/or E8 is H or absent.
Also preferably, E5 is H or —CH3.
Also preferably, Y1 is CH.
Also preferably, Y6 is CH or C; Y8 is CH or C.
In the twelfth embodiment, the present invention relates to formula (XVIII):
Preferably, X8-R11 is CH; R8 is Cl; and/or R9 is Cl.
Also preferably, E1 is H, F or absent; and/or E2 is H, F or absent.
Also preferably, E4 is H or —CH3.
Also preferably, Y1 is CH, CD or N.
Also preferably, Y6 is CH or C; and/or Y8 is CH, N or C.
In the thirteenth embodiment, the present invention relates to formula (XIX):
Preferably, X8-R11 is CH; R8 is Cl; and/or R9 is Cl.
Also preferably, J1 is H or absent; J2 is H or absent; J3 is H or absent; and/or J4 is H or absent.
Also preferably, Y1 is CH.
Also preferably, Y4 is CH2 or N—CH3.
In the fourteenth embodiment, the present invention relates to formula (XXIII):
In the fifteenth embodiment, the present invention relates to formula (XXIV):
Preferably, R4 is allyl, H or CH3.
Also preferably, R21 is CH, C—CH3 or C—Cl; and/or R22 is CH, C—CH3 or Cl.
Also preferably,
is selected from:
Also preferably, E1 is H, F or absent; and/or E2 is H, F or absent.
Also preferably, E4 is H or —CH3.
Also preferably, Y1 is CH, CD or N.
Also preferably, Y6 is CH or C; and/or Y8 is CH, N or C.
Also preferably, R2 is absent, H, F, chlorine, methyl, ethyl, methoxy, hydroxymethyl, or linked to L groups.
Also preferably, R1 is absent, H, F, chlorine, methyl, ethyl, methoxy, hydroxymethyl, or linked to L groups.
In the sixteenth embodiment, the present invention relates to formula (XXV):
In the seventeenth embodiment, the present invention relates to formula (XXVI):
Preferably, X8-R11 is CH; R8 is Cl; R9 is Cl.
Also preferably, Y21 is NH, O or absent.
Also preferably, Y10 is CH or C; Y12 is CH or C; and/or Y13 is CH or C.
In the eighteenth embodiment, the present invention relates to formula (XXVII):
Preferably, Y21 is NH, O or absent.
Also preferably, Y10 is CH or C; Y12 is CH or C; and/or Y13 is CH or C.
Also preferably, Y1 is CH or N.
Also preferably, Y10 is CH or C; Y12 is CH or C; and/or Y13 is CH or C.
Also preferably, X8-R11 is CH; R8 is Cl; R9 is Cl.
In the nineteenth embodiment, the present invention relates to formula (XXVIII):
Preferably, R4 is allyl, H or CH3.
Also preferably, Y21 is NH, O or absent.
Also preferably, Y10 is CH or C; Y12 is CH or C; and/or Y13 is CH or C.
Also preferably, R21 is CH, C—CH3 or C—Cl; and/or R22 is CH, C—CH3 or Cl.
Also preferably,
is selected from:
In the twentieth embodiment, the present invention relates to formula (XXIX):
Preferably,
is
In the 21st embodiment, the present invention relates to formula (XXX):
Preferably,
is
In the 22nd embodiment, the present invention relates to formula (XXXI):
In the 23rd embodiment, the present invention relates to formula (XXXII):
In the 24th embodiment, the present invention relates to formula (XXXIII):
Preferably,
is selected from
In the 25th embodiment, the present invention relates to formula (XXXIV):
Preferably,
is selected from
In the 26th embodiment, the present invention relates to formula (XXXV):
Preferably,
is selected from
In the 27th embodiment, the present invention relates to formula (XXXVI):
Preferably,
is selected from
In the 28th embodiment, the present invention relates to a compound represented by structural formula (XXXXI), or a derivative, pharmaceutically acceptable salt, isomer, solvate, hydrate, adduct, complex or prodrug thereof:
represents a linking site; and wherein the L group is optionally substituted with 1, 2, 3, or 4 substituents selected from halogen, hydroxyl, nitro, cyano, amino, mercapto, COOH, or C1-C6 alkyl.
In a second aspect of the 28th embodiment, the formula (XXXXI) is formula (XXXXII):
represents a linking site; and wherein the L group is optionally substituted with 1, 2, 3, or 4 substituents independently selected from halogen, hydroxyl, nitro, cyano, amino, mercapto, COOH, or C1-C6 alkyl.
In a third aspect of the 28th embodiment, the formula (XXXXII) is formula (XXXXIII):
In a fourth aspect of the 28th embodiment, the formula (XXXXI) is formula (XXXXIV):
represents a linking site; and wherein the L group is optionally substituted with 1, 2, 3, or 4 substituents independently selected from halogen, hydroxyl, nitro, cyano, amino, mercapto, COOH, or C1-C6 alkyl.
In a fifth aspect of the 28th embodiment, the formula (XXXXIV) is formula (XXXXV) or formula (XXXXVI):
In the 29th embodiment, the present application relates to a compound represented by structural formula (XXXXVII), or a derivative, pharmaceutically acceptable salt, isomer, solvate, hydrate, adduct, complex or prodrug thereof:
represents a linking site; and wherein the L group is optionally substituted with 1, 2, 3, or 4 substitutents independently selected from halogen, hydroxyl, nitro, cyano, amino, mercapto, COOH, or C1-C6 alkyl.
The structure “-L-” does not mean that L must be linked to W and/or D by a single bond. In various aspects of various embodiments of the present application, the link of L to W and/or D may be realized by one or more bonds.
The term “pharmaceutically acceptable salt” refers to pharmaceutically acceptable organic or inorganic salts of the compounds of the present invention. Exemplary salts include, but are not limited to, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acid phosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, mesylate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, pamoate (i.e., 1,1′-methylene-bis-(2-hydroxy-3-naphthoic acid) salt), alkali metal (e.g., sodium and potassium) salts, alkaline earth metal (e.g., magnesium) salts, and ammonium salt.
Pharmaceutically acceptable salts may involve the inclusion of another molecule, such as acetate ions, succinate ions, or other counterions. The counterion may be any organic or inorganic moiety that stabilizes the charge on the parent compound. In addition, pharmaceutically acceptable salts may have more than one charged atom in its structure. Examples where multiple charged atoms are part of pharmaceutically acceptable salts may have multiple counterions. Thus, pharmaceutically acceptable salts may have one or more charged atoms and/or one or more counterions.
If the compound of the present invention is a base, the desired pharmaceutically acceptable salt can be prepared by any suitable method available in the art, for example, by treating the free base with an inorganic acid (such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, methanesulfonic acid, phosphoric acid) or the organic acid (such as acetic acid, maleic acid, succinic acid, mandelic acid, fumaric acid, malonic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, pyranosidic acid (such as glucuronic acid or galacturonic acid), α-hydroxy acid (such as citric acid or tartaric acid), amino acid (such as aspartic acid or glutamic acid), aromatic acid (such as benzoic acid or cinnamic acid), sulfonic acid (such as p-toluenesulfonic acid or ethanesulfonic acid), and the like.)
If the compound of the present invention is an acid, the desired pharmaceutically acceptable salt can be prepared by any suitable method, for example, by treating the free acid with an inorganic or organic base, such as an amine (primary, secondary or tertiary amine), alkali metal hydroxide or alkaline earth metal hydroxide. Illustrative examples of suitable salts include, but are not limited to, organic salts derived from amino acids (such as glycine and arginine), ammonia, primary, secondary and tertiary amines and cyclic amines (such as piperidine, morpholine and piperazine), and inorganic salts derived from sodium, calcium, potassium, magnesium, manganese, iron, copper, zinc, aluminum and lithium.
The term “pharmaceutically acceptable” indicates that the substance or composition must be chemically and/or toxicologically compatible with other ingredients of the formulation and/or mammals treated therewith.
The compounds of the present application may exist in various stereoisomeric forms. The term “stereoisomer” refers to compounds having the same chemical composition and connectivity, but whose atoms have different orientations in space that cannot be interconverted by rotation about a single bond. “Stereoisomer” may include “diastereomer” and “enantiomer”. “Diastereomer” refers to a stereoisomer having two or more chiral centers and whose molecules are not mirror images of each other. “Enantiomer” refers to two stereoisomers of a compound, which are non-overlapping mirror images of each other. “R” and “S” represent substituent configurations surrounding one or more chiral atoms. The compounds of the present application can be prepared as individual isomers by chiral synthesis or resolution from mixtures of isomers. The disclosed compounds may have one or more stereocenters, and each stereocenter may independently exist in an R or S configuration. When the absolute stereochemistry of the stereocenter is not determined, the stereochemistry configuration can be designated (*) at the designated center. In one embodiment, the compounds described herein are present in optically active or racemic form. It should be understood that the compounds described herein include racemic, optically active, regioisomeric and stereoisomeric forms or combinations thereof having the therapeutically useful properties described herein. The chiral atoms described herein can be a chiral carbon atom, a chiral nitrogen atom, or a chiral phosphorus atom.
In particular, the compounds disclosed herein may have one or more stereocenters, and each stereocenter may independently exist in an R or S configuration. When the absolute stereochemistry at the stereocenter is not determined, the stereochemical configuration can be marked as “*” at that center, even though the compound itself has been separated as a single stereoisomer and is (non) enantiomeric pure. In other words, in some embodiments, the asterisk “*” is used to indicate that the chiral atom is in a substantially single steric configuration, and its absolute stereochemistry is not determined (even if the bond is drawn in stereochemical form). The actual absolute stereoconfiguration of atoms marked with “*” can be the same or different stereoconfiguration as the stereochemical form drawn. In other words, the atoms marked with “*” can have either an absolute R configuration or an absolute S configuration. The term “substantially” refers to variation in a range of 5%, 2%, or 1% or even 0.1% from the norm. The term “single stereoconfiguration” can be a single R configuration or a single S configuration.
In other embodiments, the stereocenter is marked as “*R” or “*S”. When the stereocenter is marked as “*R” or “*S”, it means that the absolute stereochemical properties of the stereocenter are not determined (even if the bond is drawn in stereochemical form), even though it is substantially a single stereoconfiguration. The actual absolute stereoconfiguration of the atoms marked with “*R” can be the same or different stereochemical form as the stereochemical form drawn; the actual absolute stereoconfiguration of atoms marked with “*S” can be the same or different stereochemical form as the stereochemical form drawn. In other words, “*R” can be an absolute R configuration or an absolute S configuration. Similarly, “*S” can be an absolute R configuration or an absolute S configuration. “*R” or “*S” can be randomly assigned to that molecule. The stereocenter marked with “*R” can have a single stereoconfiguration that is the same or different from another stereocenter marked with “*S”. The stereocenter marked with “*R” can have a single stereoconfiguration that is the same or different from another stereocenter marked with “*R”. The stereocenter marked with “*S” can have a single stereoconfiguration that is the same or different from another stereocenter marked with “*S”.
Similarly, The stereocenter marked with “*R” can have a single stereoconfiguration that is the same or different from another stereocenter marked with “*”. The stereocenter marked with “*S” can have a single stereoconfiguration that is the same or different from another stereocenter marked with “*”.
For example, the absolute stereoconfiguration of compound 152-A is one of the following:
In one embodiment, the compounds described herein contain one or more chiral centers. These compounds can be prepared by any means, including stereoselective synthesis, enantioselective synthesis, or separation of mixtures of enantiomers or diastereomers. Resolution of compounds and their isomers can be achieved by any means, including but not limited to chemical processes, enzymatic processes, fractional crystallization, distillation and chromatography.
The term “substituted” means that any one or more hydrogen on the designated atom or group is replaced by a moiety selected from the designated group, provided that the designated atom's normal valence is not exceeded.
“Alkyl” is a branched or straight chain saturated aliphatic hydrocarbon group. In one embodiment, the alkyl group contains 1 to about 12 carbon atoms, more typically 1 to about 6 carbon atoms or 1 to about 4 carbon atoms. In one embodiment, the alkyl group contains 1 to about 8 carbon atoms. In certain Examples, the alkyl group is C1-C2, C1-C3, or C1-C6. The specified ranges as used herein indicate an alkyl group having each member of the range described as an independent species. For example, the term C1-C6 alkyl as used herein indicates a straight or branched alkyl group having from 1, 2, 3, 4, 5, or 6 carbon atoms and is intended to mean that each of these is described as an independent species. For example, the term C1-C4alkyl as used herein indicates a straight or branched alkyl group having from 1, 2, 3, or 4 carbon atoms and is intended to mean that each of these is described as an independent species. When C0-Cn alkyl is used herein in conjunction with another group, for example, (C3-C7 cycloalkyl) C0-C4 alkyl, or —C0-C4 alkyl(C3-C7 cycloalkyl), the indicated group, in this case cycloalkyl, is either directly bound by a single covalent bond (C0 alkyl), or attached by an alkyl chain in this case 1, 2, 3, or 4 carbon atoms. Alkyls can also be attached via other groups such as heteroatoms as in —O—C0-C4 alkyl (C3-C7 cycloalkyl). Examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, t-butyl, n-pentyl, isopentyl, tert-pentyl, neopentyl, n-hexyl, 2-methylpentyl, 3-methylpentyl, 2,2-dimethylbutyl and 2,3-dimethylbutyl. In one embodiment, the alkyl group is optionally substituted as described above.
“Alkenyl” is a branched or straight chain aliphatic hydrocarbon group having one or more carbon-carbon double bonds that may occur at a stable point along the chain. Nonlimiting examples are C2-C8 alkenyl, C2-C6 alkenyl and C2-C4 alkenyl. The specified ranges as used herein indicate an alkenyl group having each member of the range described as an independent species, as described herein for the alkyl moiety. Examples of alkenyl include, but are not limited to, ethenyl, propenyl. In one embodiment, the alkenyl group is optionally substituted as described above.
“Alkynyl” is a branched or straight chain aliphatic hydrocarbon group having one or more carbon-carbon triple bonds that may occur at any stable point along the chain, for example, C2-C8 alkynyl or C2-C6 alkynyl. The specified ranges as used herein indicate an alkynyl group having each member of the range described as an independent species, as described herein for the alkyl moiety. Examples of alkynyl include, but are not limited to, ethynyl, propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1-hexynyl, 2-hexynyl, 3-hexynyl, 4-hexynyl and 5-hexynyl. In one embodiment, the alkynyl group is optionally substituted as described above.
“Alkoxy” is an alkyl group as defined above covalently bound through an oxygen bridge (—O—).
Examples of alkoxy include, but are not limited to, methoxy, ethoxy, n-propoxy, i-propoxy, n-butoxy, 2-butoxy, t-butoxy, n-pentoxy, 2-pentoxy, 3-pentoxy, isopentoxy, neopentoxy, n-hexoxy, 2-hexoxy, 3-hexoxy, and 3-methylpentoxy. Similarly an “alkylthio” or a “thioalkyl” group is an alkyl group as defined above with the indicated number of carbon atoms covalently bound through a sulfur bridge (—S—). In one embodiment, the alkoxy group is optionally substituted as described above.
“Alkenyloxy” is an alkenyl group as defined above covalently bound to the group it substitutes by an oxygen bridge (—O—).
“Alkanoyl” is an alkyl group as defined above covalently bound through a carbonyl (C═O) bridge. The carbonyl carbon is included in the number of carbons, that is C2 alkanoyl is a CH3(C═O)— group. In one embodiment, the alkanoyl group is optionally substituted as described above.
“Alkylester” is an alkyl group as defined covalently bound through an ester linkage. The ester linkage may be in either orientation, e.g., a group of the formula —O(C═O)alkyl or a group of the formula —(C═O)Oalkyl.
“Carbocyclyl”, “carbocyclic ring”, or “cycloalkyl” is a saturated or partially unsaturated (i.e., not aromatic) group containing all carbon ring atoms. A carbocyclic group typically contains 1 ring of 3 to 7 carbon atoms or 2 fused rings each containing 3 to 7 carbon atoms. Cycloalkyl substituents may be pendant from a substituted nitrogen or carbon atom, or a substituted carbon atom that may have two substituents can have a cycloalkyl group, which is attached as a spiro group. Examples of carbocyclic rings include cyclohexenyl, cyclohexyl, cyclopentenyl, cyclopentyl, cyclobutenyl, cyclobutyl and cyclopropyl rings. In one embodiment, the carbocyclic ring is optionally substituted as described above. In one embodiment, the cycloalkyl is a partially unsaturated (i.e., not aromatic) group containing all carbon ring atoms.
“Carbocycle-oxy group” is a monocyclic carbocyclic or mono- or bi-cyclic carbocyclic group as described above attached to the group it substitutes by an oxygen-O-linker.
“Haloalkyl” indicates both branched and straight-chain alkyl groups substituted with 1 or more halogen atoms, up to the maximum allowable number of halogen atoms. Examples of haloalkyl include, but are not limited to, trifluoromethyl, monofluoromethyl, difluoromethyl, 2-fluoroethyl, and penta-fluoroethyl.
“Haloalkoxy” indicates a haloalkyl group as defined herein attached through an oxygen bridge (oxygen of an alcohol radical).
“Hydroxyalkyl” is an alkyl group as previously described substituted with at least one hydroxy substituent.
“Aminoalkyl” is an alkyl group as previously described substituted with at least one amino substituent.
“Halo” indicates independently any of fluoro, chloro, bromo, and iodo.
“Aryl” indicates aromatic groups containing only carbon in the aromatic ring or rings. In one embodiment, the aryl groups contain 1 to 3 separate or fused rings and is 6 to about 18 ring atoms (such as 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18 ring atoms), without heteroatoms as ring members. When indicated, such aryl groups may be further substituted with carbon or non-carbon atoms or groups. Such substitution may include fusion to a 5 to 7-membered saturated cyclic group that optionally contains 1 or 2 heteroatoms independently chosen from N, O, and S, to form, for example, a 3,4-methylenedioxyphenyl group. Aryl groups include, for example, phenyl, naphthyl, including 1-naphthyl and 2-naphthyl. In one embodiment, aryl groups are pendant. An example of a pendant ring is a phenyl group substituted with a phenyl group. In one embodiment, the aryl group is optionally substituted as described above. Aryl groups include bicyclic groups comprising an aromatic ring fused to a saturated, partially unsaturated ring or an aromatic carbocyclic or heterocyclic ring. Typical aryl groups include, but are not limited to, those derived from benzene (phenyl), substituted benzene, naphthalene, anthracene, indenyl, indanyl, 1,2-dihydronaphthalene, 1,2,3,4-tetrahydro groups and the like. Preferably, the aryl group is a phenyl group.
The term “heterocyclyl” as used herein refers to a saturated or a partially unsaturated (i.e., having one or more double and/or triple bonds within the ring without aromaticity) carbocyclic radicals of 3 to 18 (such as 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, or 18) ring atoms wherein at least one ring atom is a heteroatom selected from nitrogen, oxygen, phosphorus and sulfur, the remaining ring atoms being C, where one or more ring atoms is optionally substituted independently with one or more substituents described above. A heterocycle may be a monocycle having 3 to 7 ring members (2 to 6 carbon atoms and 1 to 4 heteroatoms selected from N, O, P and S) or a bicycle having 6 to 10 ring members (4 to 9 carbon atoms and 1 to 6 heteroatoms selected from N, O, P and S), for example: a bicyclo[4,5], [5,5], [5,6], or [6,6] system. In one embodiment, the only heteroatom is nitrogen. In one embodiment, the only heteroatom is oxygen.
In one embodiment, the only heteroatom is sulfur. Heterocycles are described in Paquette, Leo A.; “Principles of Modern Heterocyclic Chemistry” (W. A. Benjamin, New York, 1968), particularly Chapters 1, 3, 4, 6, 7, and 9; “The Chemistry of Heterocyclic Compounds, A series of Monographs” (John Wiley & Sons, New York, 1950 to present), in particular Chapters 13, 14, 16, 19, and 28; and J. Am. Chem. Soc. (1960) 82:5566. Examples of heterocyclic rings include, but are not limited to, pyrrolidinyl, dihydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, tetrahydrothiopyranyl, piperidino, piperidonyl, morpholino, thiomorpholino, thioxanyl, piperazinyl, homopiperazinyl, azetidinyl, oxetanyl, thietanyl, homopiperidinyl, oxepanyl, thiepanyl, oxazepinyl, diazepinyl, thiazepinyl, 2-pyrrolinyl, 3-pyrrolinyl, indolinyl, 2H-pyranyl, 4H-pyranyl, dioxanyl, 1,3-dioxolanyl, pyrazolinyl, dithianyl, dithiolanyl, dihydropyranyl, dihydrothienyl, dihydrofuranyl, dihydroisoquinolinyl, tetrahydroisoquinolinyl, pyrazolidinylimidazolinyl, imidazolidinyl, 2-oxa-5-azabicyclo[2.2.2]octane, 3-oxa-8-azabicyclo[3.2.1]octane, 8-oxa-3-azabicyclo[3.2.1]octane, 6-oxa-3-azabicyclo[3.1.1]heptane, 2-oxa-5-azabicyclo[2.2.1]heptane, 3-azabicyco[3.1.0]hexanyl, 3-azabicyclo[4.1.0]heptanyl, azabicyclo[2.2.2]hexanyl, 3H-indolyl, quinolizinyl, N-pyridyl ureas, and pyrrolopyrimidine.
Spiro moieties are also included within the scope of this definition. Examples of a heterocyclic group wherein 1 or 2 ring carbon atoms are substituted with oxo (═O) moieties are pyrimidinonyl and 1,1-dioxo-thiomorpholinyl. The heterocycle groups herein are optionally substituted independently with one or more substituents described herein. “Heterocyclyl” includes “heterocycloalkyl”. “Heterocycloalkyl” is a saturated ring group. It may have, for example, 1, 2, 3 or 4 heteroatoms independently selected from N, S and O, the remaining ring atoms being carbon. In a typical embodiment, nitrogen is a heteroatom. Monocyclic heterocycloalkyl groups typically have 3 to about 8 ring atoms or 4 to 6 ring atoms. Examples of heterocycloalkyl groups include morpholino, piperazinyl, piperidino, and pyrrolinyl.
“Heterocyclyl” may contain 1, 2, 3, 4, or 5 heteroatoms selected from N, O, and S.
“Heterocyclicoxy group” is a monocyclic heterocyclic ring or a bicyclic heterocyclic group as described previously linked to the group it substitutes via an oxygen-O-linker.
“Heteroaryl” can be a stable monocyclic aromatic ring which contains from 1 to 3, or in some embodiments from 1 to 2, heteroatoms chosen from N, O, and S, with remaining ring atoms being carbon, or a stable bicyclic or tricyclic system of 5- to 10-membered (5, 6, 7, 8, 9, 10 membered) aromatic rings containing at least one 1 to 3, or in some embodiments from 1 to 2, heteroatoms chosen from N, O, and S, with remaining ring atoms being carbon. In one embodiment, the only heteroatom is nitrogen. In one embodiment, the only heteroatom is oxygen.
In one embodiment, the only heteroatom is sulfur. Monocyclic heteroaryl groups typically have from 5 to 8 ring atoms(5, 6, 7, 8 ring atoms). In some embodiments, bicyclic heteroaryl groups are 9- to 10-membered heteroaryl groups, that is, groups containing 9 or 10 ring atoms wherein one 5- to 7-membered aromatic ring is fused to a second aromatic or non-aromatic ring. When the total number of S and O atoms in the heteroaryl group exceeds 1, these heteroatoms are not adjacent to one another. In one embodiment, the total number of S and O atoms in the heteroaryl group is not more than 2. In another embodiment, the total number of S and O atoms in the aromatic heterocycle is not more than 1. Examples of heteroaryl groups include, but are not limited to, pyridinyl (including, for example, 2-hydroxypyridinyl), furyl, imidazolyl, imidazopyridinyl, pyrimidinyl (including, for example, 4-hydroxypyrimidinyl), pyrazolyl, triazolyl, pyrazinyl, tetrazolyl, furyl, thienyl, isoxazolyl, thiazolyl, oxadiazolyl, oxazolyl, isothiazolyl, pyrrolyl, quinolinyl, isoquinolinyl, tetrahydroisoquinolinyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, pyridazinyl, triazinyl, isoindolyl, pteridinyl, purinyl, oxadiazolyl, triazolyl, thiadiazolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, tetrahydrofuranyl, and furopyridinyl. Heteroaryl groups are optionally substituted independently with one or more substituents described herein. “Heteroaryloxy” is a heteroaryl group as described bound to the group it substituted via an oxygen-O-linker. “Heteroaryl” may contain 1, 2, 3, 4, or 5 heteroatoms selected from N, O, and S.
In this context,
or
and any indeterminate linking mode may represent a linking site.
The present application also covers bicyclic, tricyclic, tetracyclic, pentacyclic and other polycyclic ring systems formed by one or more monocyclic rings selected from “cycloalkyl”, “aryl” and “heteroaryl” in a valence-allowed manner.
The term cyano refers to —CN;
The term CD refers to deuterated carbon.
The compounds or pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof described herein, are useful in the treatment of proliferative disorders. The present invention also provides the use of the compound or pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof described herein in the manufacture of a medicament for the treatment of proliferative disorders. The present invention further provides the compounds or pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof for use in the treatment of proliferative disorders. The application further provides a method of treating proliferative disorders, comprising administering to a subject in need thereof a therapeutically effective amount of the compounds or pharmaceutically acceptable salts, isomers, solvates, hydrates, additions, compounds, complexes or prodrugs thereof.
The term “proliferative disorder” or “cell proliferative disorder” refers to disorders that are associated with some degree of abnormal cell proliferation, whether malignant or benign. In some embodiments, the proliferative disorder is a cancer. In some aspects, the cancer is a solid tumor. In some aspects, the cancer is a hematologic malignancy. The terms “proliferative disorder”, “cell proliferative disorder”, “cancer,” “cancerous,” and “tumor” are not mutually exclusive as referred to herein.
The term “cancer” comprises cancerous cells, and/or benign or pre-cancerous cells. Examples of cancer include, but are not limited to, breast cancer, colon cancer, brain cancer, prostate cancer, kidney cancer, pancreatic cancer, ovarian cancer, head and neck cancer, melanoma, colorectal cancer, gastric cancer, squamous cancer, small-cell lung cancer, non small-cell lung cancer, testicular cancer, Merkel cell carcinoma, glioblastoma, neuroblastoma, cancers of lymphatic organs and hematological malignancy including Leukemia (Acute lymphoblastic leukemia (ALL), Acute myelogenous leukemia (AML), Chronic lymphocytic leukemia (CLL), Chronic myelogenous leukemia (CML), Acute monocytic leukemia (AMOL), Hairy cell leukemia (HCL), T-cell prolymphocytic leukemia (T-PLL), Large granular lymphocytic leukemia, Adult T-cell leukemia), Lymphoma (small lymphocytic lymphoma (SLL), Hodgkin's lymphomas (Nodular sclerosis, Mixed cellularity, Lymphocyte-rich, Lymphocyte depleted or not depleted, and Nodular lymphocyte-predominant Hodgkin lymphoma), Non-Hodgkin's lymphomas (all subtypes), Chronic lymphocytic leukemia/Small lymphocytic lymphoma, B-cell prolymphocytic leukemia, Lymphoplasmacytic lymphoma (such as Waldenstrom macroglobulinemia), Splenic marginal zone lymphoma, Plasma cell neoplasms (Plasma cell myeloma, Plasmacytoma, Monoclonal immunoglobulin deposition diseases, Heavy chain diseases), Extranodal marginal zone B cell lymphoma (MALT lymphoma), Nodal marginal zone B cell lymphoma (NMZL), Follicular lymphoma, Mantle cell lymphoma, Diffuse large B cell lymphoma, Mediastinal (thymic) large B cell lymphoma, Intravascular large B cell lymphoma, Primary effusion lymphoma, Burkitt lymphoma/leukemia, T cell prolymphocytic leukemia, T cell large granular lymphocytic leukemia, Aggressive NK cell leukemia, Adult T cell leukemia/lymphoma, Extranodal NK/T cell lymphoma (nasal type), Enteropathy-type T cell lymphoma, Hepatosplenic T cell lymphoma, Blastic NK cell lymphoma, Mycosis fungoides/Sezary syndrome, Primary cutaneous CD30-positive T cell lymphoproliferative disorders, Primary cutaneous anaplastic large cell lymphoma, Lymphomatoid papulosis, Angioimmunoblastic T cell lymphoma, Peripheral T cell lymphoma (unspecified), Anaplastic large cell lymphoma), multiple myeloma (plasma cell myeloma or Kahler's disease). The term “cancer” also includes leukemia, carcinoma and sarcoma. Exemplary cancers include brain cancer, breast cancer, cervical cancer, colon cancer, head and neck cancer, liver cancer, kidney cancer, lung cancer, non-small cell lung cancer, melanoma, mesothelioma, ovarian cancer, sarcoma, stomach cancer, uterine cancer and Medulloblastoma. Additional examples include Hodgkin's disease, non-Hodgkin's lymphoma, multiple myeloma, neuroblastoma, ovarian cancer, rhabdomyosarcoma, essential thrombocythemia, primary macroglobulinemia, primary brain tumor, cancer, malignant pancreatic insulinoma, malignant carcinoid, bladder cancer, premalignant skin lesions, testicular cancer, lymphoma, thyroid cancer, neuroblastoma, esophageal cancer, genitourinary tract cancer, malignant hypercalcemia, endometrial cancer, adrenocortical carcinoma, tumors of the endocrine and exocrine pancreas, and prostate cancer. The term “cancer” also includes leukemia, multiple myeloma, lymphoma, liver cancer, stomach cancer, breast cancer, cholangiocarcinoma, pancreatic cancer, lung cancer, colorectal cancer, osteosarcoma, melanoma, human cervical cancer, glioma, nasopharyngeal cancer, laryngeal cancer, esophageal cancer, middle ear tumor, prostate cancer, etc.
“Link to the L group/linked to L groups/linking to L groups” described herein means that the L group is linked to the site indicated by the corresponding variable (eg, a site on a ring, such as a benzene ring atom to which R1 or R2 is attached, Y5, Y6, or Y8 itself, etc., or R1 and/or R2 themselves). Here, the corresponding variable can be present or absent.
In a preferred embodiment, the present application relates to the following diseases: acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), multiple myeloma (MM), diffuse large B cell lymphoma (DLBCL), mantle cell lymphoma (MCL), malignant melanoma, gastric cancer, gliomas, ovarian carcinoma, uterine serous carcinoma (USC), colorectal cancer, pancreatic cancer, esophageal cancer, hepatocellular carcinoma, glioblastoma, non-small-cell lung cancer (NSCLC), small-cell lung cancer (SCLC), neuroblastoma, breast cancer, triple negative breast cancer (TNBC), neuroblastoma, head and neck squamous cell carcinoma (HNSCC).
The present invention further provides a pharmaceutical composition comprising the compound or derivatives, pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof described herein, and a pharmaceutically acceptable carrier. The pharmaceutical composition described herein may be a tablet, a capsule, a granule, a syrup, a suspension, a solution, a dispersion, a slow release preparation for oral or non-oral administration, an intravenous injection preparation, a subcutaneous injection preparation, an inhalation preparation, a transdermal preparation, a rectal or vaginal suppository.
The pharmaceutically acceptable carrier described herein refers to a pharmaceutically acceptable carrier well known to those skilled in the art, and the pharmaceutically acceptable carrier of the present invention includes, but is not limited to, a filler, a wetting agent, a binder, a disintegrating agent, a lubricant, a binder, a glidant, a flavoring agent, a surfactant, a preservative, and the like. Fillers include, but are not limited to, lactose, microcrystalline cellulose, starch, powdered sugar, dextrin, mannitol, calcium sulfate, and the like. Wetting agents and binders include, but are not limited to, sodium carboxymethylcellulose, hydroxypropylcellulose, hydroxypropyl methyl cellulose, gelatin, sucrose, polyvinylpyrrolidone, and the like. Disintegrating agents include, but are not limited to, sodium carboxymethyl starch, crosslinked polyvinylpyrrolidone, croscarmellose sodium, low substituted hydroxypropylcellulose, and the like. Lubricants include, but are not limited to, magnesium stearate, aerosil, talc, hydrogenated vegetable oil, polyethylene glycol, magnesium lauryl sulfate, and the like. Binders include, but are not limited to, gum arabic, alginic acid, calcium carboxymethylcellulose, sodium carboxymethylcellulose, glucose, dextrin, dextrose, ethylcellulose, gelatin, liquid glucose, guar gum, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, magnesium aluminum silicate, maltodextrin, methyl cellulose, polymethacrylate, polyvinylpyrrolidone, pregelatinized starch, sodium alginate, sorbitol, starch, syrup and tragacanth. Glidants include, but are not limited to, colloidal silica, powdered cellulose, magnesium trisilicate, silica, and talc. Flavoring agents include, but are not limited to, aspartame, stevioside, fructose, glucose, syrup, honey, xylitol, mannitol, lactose, sorbitol, maltitol, glycyrrhizin. Surfactants include, but are not limited to, Tween-80, poloxamer. Preservatives include, but are not limited to, paraben, sodium benzoate, potassium sorbate, and the like.
Methods of preparing various pharmaceutical compositions containing various amounts of active ingredients are known, or will be apparent to those skilled in the art in light of this disclosure, as described in REMINGTON'S PHARMACEUTICAL SCIENCES, Martin, E. W., ed., Mack Publishing Company, 19th ed. (1995). Methods of preparing the pharmaceutical compositions include incorporation of suitable pharmaceutical excipients, carriers, diluents and the like. The pharmaceutical compositions described herein are made in a known manner, including conventional methods of mixing, dissolving or lyophilizing.
In the pharmaceutical compositions described herein, the amount of active ingredient may vary from about 0.01% to about 99% by weight of a given unit dosage form. In such therapeutically useful pharmaceutical composition formulations, the active ingredient is in an amount such that an effective dosage level can be obtained.
The tablet, capsule and the like described herein may comprise: a binder such as tragacanth, gum arabic, corn starch or gelatin; an excipient such as dicalcium phosphate; a disintegrating agent such as corn starch, potato starch, alginic acid, etc.; a lubricant such as magnesium stearate; and a sweetener such as sucrose, fructose, lactose or aspartame; or a flavoring agent such as mint, wintergreen or cherry flavor. When the unit dosage form is a capsule, it may contain, in addition to materials of the above types, a liquid carrier such as vegetable oil or polyethylene glycol. Various other materials may be present, as a coating, or otherwise alter the physical form of the solid unit dosage form. For example, tablets or capsules may be coated with gelatin, wax, shellac or sugar, etc. The syrup may contain an active ingredient, sucrose or fructose as a sweetener, methylparaben or propylparaben as a preservative, a dye and a flavoring agent such as cherry or orange flavor. Of course, any material used to prepare any unit dosage form should be pharmaceutically acceptable and non-toxic in the amounts employed. In addition, the active ingredient can be incorporated into a slow release formulations and a slow release device.
The active ingredient can also be administered intravenously or intraperitoneally by infusion or injection. An aqueous solution of the active ingredient or a salt thereof can be prepared, optionally with a non-toxic surfactant. Dispersions in glycerol, liquid polyethylene glycol, triacetin and mixtures thereof, and oils can also be prepared. Under ordinary conditions of storage and use, these formulations contain a preservative to prevent the growth of microorganisms.
Pharmaceutical composition dosage forms suitable for injection or infusion may comprise sterile aqueous solution or dispersion or sterile powder comprising an active ingredient suitable for the ready-to-use preparation of a sterile, injectable or infusible solution or dispersion (optionally encapsulated in a liposome). In all cases, the final dosage forms must be sterile, liquid, and stable under the conditions of manufacture and storage. The liquid carrier may be a solvent or liquid dispersion medium including, for example, water, ethanol, polyols (for example, glycerol, propylene glycol, liquid polyethylene glycol, and the like), vegetable oils, non-toxic glycerides, and suitable mixtures thereof. Proper fluidity can be maintained, for example, by liposome formation, by maintaining the desired particle size in the case of dispersion, or by the use of a surfactant. The prevention of microorganisms can be achieved by using various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal, and the like. In many cases, it will be preferred to include isotonic agents, such as sugars, buffers or sodium chloride. Prolonged absorption of the injectable composition can be brought about by the use of a composition that comprises an absorption delaying agent (for example, aluminum monostearate and gelatin).
Sterile injectable solutions are prepared by combining the active ingredient in a desired amount in a suitable solvent with the various other ingredients listed above, followed by filter sterilization. In the case of sterile powders for the preparation of sterile injectable solutions, the preferred preparation methods are vacuum drying and lyophilization techniques which result in powders of the active ingredient plus any additionally desired ingredients present in the sterile filtration solution.
Useful solid carriers include comminuted solids (e.g., talc, clay, microcrystalline cellulose, silica, alumina, etc.). Useful liquid carriers include water, ethanol or ethylene glycol or a water-ethanol/ethylene glycol mixture, and the pharmaceutical compositions of the present invention may be dissolved or dispersed in the liquid carriers in an effective amount, optionally with the aid of a non-toxic surfactant. Adjuvants (such as fragrances) and additional antimicrobial agents can be added to optimize the properties for a given use.
Thickeners (such as synthetic polymers, fatty acids, fatty acid salts and esters, fatty alcohols, modified celluloses or modified inorganic materials) can also be used with liquid carriers to form coatable pastes, gels, ointments, soaps, etc. that are used directly on the user's skin.
The therapeutically effective amount of the active ingredient will depend not only on the particular salt selected, but also on the administration route, the property of the disease to be treated, and the age and conditions of the patient, and will ultimately depend on the decision of the attending physician or clinician.
The above formulations may be presented in unit dosage form, which is a physically discrete unit containing a unit dose suitable for administration to humans and other mammalian bodies. The unit dosage form may be a capsule or a tablet. The amount of active ingredient in the unit dose may vary or be adjusted between about 0.01 to about 1000 mg or more, depending on the particular treatment involved.
The term “treatment” as used herein generally refers to the acquisition of the desired pharmacological and/or physiological effect. The effect may be prophylactic according to the prevention of the disease or its symptoms in whole or in part; and/or may be therapeutic according to the partial or complete stabilization or cure of the disease and/or the side effect due to the disease. As used herein, “treatment” encompasses any treatment for the disease of a patient, including: (a) prevention of the disease or condition in the patient that may be predisposed to the disease or condition but has not yet been diagnosed; (b) inhibition of the symptoms of the disease, i.e., preventing its development; or (c) remission of the symptoms of the disease, i.e., causing regression of the disease or symptoms in whole or in part.
The compound described herein, or pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof, may also be administered in combination with one or more additional therapeutic agents for the treatment of cancers. Such additional therapeutic agents include, but are not limited to, anthracyclines, cyclophosphamide, 5-fluorouracil, cisplatin, and the like.
Unless otherwise stated, percentages, ratios, proportions or parts used in this application are by weight or volume. The amount used in this application is a weight or volume amount. Those skilled in the art can easily determine.
In yet another aspect of the present application, the present application further includes a compound or pharmaceutically acceptable salts, isomers, solvates, hydrates, adducts, complexes or prodrugs thereof obtained by combining any group defined at any variable group of the present application.
The application also includes the following exemplary compounds.
Hereinafter, the application will illustrate its beneficial effects by examples. Those skilled in the art will recognize that these examples are illustrative and not limiting. These examples do not limit the scope of this application in any way. The experimental methods described in the following examples are conventional unless otherwise specified. Unless otherwise specified, reagents and materials are commercially available.
The compounds of this invention may be prepared or isolated as described in the following general scheme. Those compounds were synthesized by synthetic and/or semi-synthetic methods known to those skilled in the art for analogous compounds and by methods described in detail in the example, herein. All the variable groups applied in these schemes are as defined in the claims.
As described in above scheme, the applied compound may be accessed by reductive amination of aldehyde A2-1 with amine A1-1. The reductive amination conditions are generic reductive amination conditions including and not limited to conditions such as sodium cyanoborohydride, sodium triacetylborohydride, sodium borohydride, etc. The X represents a carbon or nitrogen atom, and the wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound may be accessed by reductive amination of ketone A2-2 with amine A1-1. The reductive amination conditions are generic reductive amination conditions including and not limited to conditions such as sodium cyanoborohydride, sodium triacetylborohydride, sodium borohydride, etc. The X represents a carbon or nitrogen atom, and the wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound may be accessed by reductive amination of aldehyde A1-3 with amine A2-3. The reductive amination conditions are generic reductive amination conditions including and not limited to conditions such as sodium cyanoborohydride, sodium triacetylborohydride, sodium borohydride, etc. The X represents a carbon or nitrogen atom, and the wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound may be accessed by reductive amination of ketone A1-4 with amine A2-4. The reductive amination conditions are generic reductive amination conditions including and not limited to conditions such as sodium cyanoborohydride, sodium triacetylborohydride, sodium borohydride, etc. The X represents a carbon or nitrogen atom, and the wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound can be produced by SN2 substitution of bromide B2-1 by amine B1-1 in the presence of a suitable organic base such as DIPEA, Et3N, or other inorganic base such as K2CO3, Cs2CO3 in the suitable solvent such as DMF, DMA, DMSO, THE etc. The wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound can be produced by SNAr substitution of fluoride B2-2 by secondary amine B1-2 in the presence of a suitable organic base such as DIPEA, Et3N, or other inorganic base such as K2CO3, Cs2CO3 in the suitable solvent such as DMF, DMA, DMSO, THE etc. The wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound can be produced by SN2 substitition of intermediate C1-1 with leaving group by secondary amine C2-1 in the presence of a suitable organic base such as DIPEA, Et3N, or other inorganic base such as K2CO3, Cs2CO3 in the suitable solvent such as DMF, DMA, DMSO, THE etc. The wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the applied compound can be produced by SN2 substitition of intermediate C2-2 with leaving group by secondary amine A1-1 in the presence of a suitable organic base such as DIPEA, Et3N, or other inorganic base such as K2CO3, Cs2CO3 in the suitable solvent such as DMF, DMA, DMSO, THE etc. The wavy line represents the connection of the reactive functional group with other fragments of the compound.
As described in above scheme, the appled compound can be produced by SN2 substitution of a sulfone intermediate which was formed by oxidization of thioether 2-1 with m-CPBA by aniline D1-1 in the presence of a sitable organic base such as DIPEA, Et3N, or other inorganic base such as K2CO3, Cs2CO3 in the suitable solvent such as DMF, DMA, DMSO, THE etc. The wavy line represents the connection of the reactive functional group with other fragments of the compound. Linker presents the fragment of linker.
The compounds of the present application can be synthesized by the following exemplary or the similarly general synthetic methods. Various intermediates may have various substituents described in this application as valence permits.
Synthesis of intermediate B 1 and B 2:
To a solution of intermediate B 1-1 (8.85 g, 41.49 mmol) in DMF (80 mL) was added 60% NaH in mineral oil (2.83 g, 70.79 mmol) at 0° C. The reaction mixture was stirred for 30 min at 0° C. and intermediate B 1-2 (7.8 g, 41.49 mmol) in DMF (20 mL) was added to the reaction mixture and the reaction mixture was stirred for 2 hours at 0° C. under N2 atmospheres. The reaction mixture was poured into ice-water (50 mL), extracted with MTBE (30 mL×2), the water phase was adjusted pH to 4-5, extracted with EtOAc (50 mL×2). The organic phase was washed with saturated salt solution (30 mL×2), dried over Na2SO4, filtered and concentrated to give a crude product. The crude product was added MeOH (10 mL), and the mixture was stirred for 15 min, filtered to give an intermediate B 1-3 (9.8 g, 66.50% yield) as a white solid.
1H NMR (400 MHz, DMSO-d6) δ 12.91 (s, 1H), 8.98 (s, 1H), 7.67 (d, J=8.0 Hz, 2H), 7.55-7.50 (m, 1H), 2.57 (s, 3H)
A mixture of intermediate B 1-3 (14.6 g, 41.10 mmol) in POCl3 (82.50 g, 538.05 mmol, 50 mL) and DIEA (55 mL) was stirred at 90° C. for 3 hours. The reaction mixture was concentrated by rotary distillation to remove the solvent. The residue was diluted with sat. NaHCO3 (500 mL) and extracted with EtOAc (100 mL×3). The combined organic layers were washed with sat. NaHCO3 (100 mL×3), dried over Na2SO4, filtered and concentrated by rotary distillation to give the residue intermediate B 1-4 (15.3 g, crude) as a yellow solid. The crude product was used into the next step without further purification.
A mixture of intermediate B 1-4 (15.3 g, 40.95 mmol) and intermediate B 1-5 (8.61 g, 81.90 mmol, 8.92 mL) in MeCN (150 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80° C. for 1 h under N2 atmosphere. The reaction solution was spin-dried to obtain crude product. To the crude was added PE: EtOAc: DCM (3: 1: 0.2, 50 mL). The reaction mixture was stirred for 1 hour and filtered to give a filter cake, which is intermediate B 1-6 (8.2 g, 32.60% yield) as a white solid. The mather lique was purified with chromatography on silica gel (PE: EtOAc=1: 1) to give intermediate B 1-6 (3.2 g, 17.67% yield) as a yellow solid.
MS (ESI+): m/z 441.95 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 8.92 (s, 1H), 7.80-7.75 (m, 2H), 7.71-7.64 (m, 2H), 4.58 (t, J=5.4 Hz, 1H), 3.47-3.41 (m, 2H), 3.33-3.27 (m, 6H), 2.58 (s, 3H).
intermediate B 1-6 (700 mg, 1.58 mmol) and HCl (12 M, 132 μL) were taken up into a microwave tube in MeCN (6 mL). The sealed tube was heated at 160° C. for 15 min under microwave. The reaction mixture was added EtOAc (15 mL), stirred for 15 min, filtered to give a crude product, the mather lique was washed with sat. NaHCO3 (5 mL×2), dried over Na2SO4, filtered and concentrated in vacuo to give a crude productintermediate B 1 (580 mg, crude) as a brown solid. The crude product was used into the next step without further purification.
m-CPBA (268 mg, 1.32 mmol, 85% purity) was added to a mixture consisting of intermediate B 1 (200 mg, 529 μmol) and DCM (5 mL). The reaction mixture was stirred at 25° C. for 2 hr. The reaction mixture was quenched with aq. Na2S2O3 (20 mL) and extracted with DCM (20 mL×3). The combined organic extracts were dried over anhydrous Na2SO4, filtered and concentrated to dryness under reduced pressure to give intermediate B2-1 (200 mg, 57.6% yield) as a yellow solid.
MS (ESI+): m/z 393.9 [M+H]+.
A solution of intermediate B 2-1 (200 mg, 507 μmol), intermediate B 2-3 (151 mg, 609 μmol) in DMSO (3 mL) was stirred at 120° C. for 8 hr. The reaction solution was spin-dried to obtain crude product. The reaction mixture was quenched by water (10 mL) and extracted with EA (20 mL×3). The combined organic phase was dried over anhydrous Na2SO4 and filtered and concentrated by rotary distillation to get a crude product. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 0: 1) to give intermidiate B 2-2 (100 mg, 34.1% yield) as a brown solid.
HCl/dioxane (4 M, 3 mL) was added to a mixture consisting of intermediate B 2-2 (100 mg, 173 μmol) and dioxane (1 mL). The reaction mixture was stirred at 25° C. for 1 hr. The reaction solution was spin-dried to obtain crude product. The residue was purified by prep-HPLC (Calumn: Kromasil 100-5-C18 30×150 mm, Molile Phase A: water (0.01% FA), Molile Phase B: acetonitrile, 25 mL/min, gradient condition form 3000 B to 60%) to give intermediate B 2 (45 mg, 51% yield) as a brown solid.
MS (ESI+): m/z 477.9 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ5 10.84 (s, 1H), 9.16 (s, 1H), 7.81-7.73 (m, 3H), 7.69-7.53 (m, 3H), 7.18-7.08 (m, 2H), 3.93 (s, 2H), 3.08-3.01 (m, 2H), 2.81 (s, 2H).
The following intermediates can be prepared using methods similar to the above synthetic route.
1H NMR (400 MHz,
A mixture of intermediate B 6-1 (6.76 g, 29.0 mmol), intermediate B 6-2 (5.00 g, 29.0 mmol), DIEA (9.38 g, 72.6 mmol) in THE (50 mL) was stirred at 70° C. for 16 h under N2 atmosphere.
The mixture was concentrated by rotary distillation to afford the crude product. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1: 0 to 10: 1). intermediate B 6-3 (8.50 g, 73.9% yield) was obtained as a colorless oil.
MS (ESI+): m/z 369.1 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 9.73 (s, 1H), 8.66 (s, 1H), 5.96-5.80 (m, 1H), 5.26-5.03 (m, 2H), 4.30 (q, J=7.2 Hz, 2H), 4.16-4.06 (m, 2H), 2.46 (s, 3H), 1.44-1.20 (m, 12H).
To a mixture of intermediate B 6-3 (8.50 g, 23.07 mmol) in DCM (20 mL) was added HCl/dioxane (4 M, 58 mL). The mixture was stirred at 25° C. for 1 h. The mixture was concentrated by rotary distillation. intermediate B 6-4 (7.00 g, crude, HCl salt) was obtained as a white solid.
MS (ESI+): m/z 268.7 [M+H]+.
A mixture of intermediate B 6-4 (5.00 g, 16.4 mmol, HCl salt) in aq. NaOH (6 M, 21 mL) and EtOH (30 mL) was stirred at 25° C. for 3 h under N2 atmosphere. The reaction mixture was acidified by using 6N HCl solution and EtOH was removed by rotary distillation. The mixture was extracted with DCM 100 mL for three times, washed with brine (150 mL), dried over anhydrous MgSO4, filtered and concentrated by rotary distillation. Intermediate B 6-5 (3.30 g, crude) was obtained as a yellow solid.
MS (ESI+): m/z 223.06 [M+H]+.
Intermediate B 6-5 (1.10 g, 4.95 mmol), intermediate B 6-6 (1.18 g, 5.44 mmol), CuI (943 mg, 4.95 mmol), K2CO3 (1.03 g, 7.42 mmol), and N,N-dimethylethane-1,2-diamine (873 mg, 9.90 mmol) in dioxane (10 mL) were heated at 100° C. for 1 h under microwave. The reaction was repeated for another two times in parallel. The reaction mixture was poured into water (20 mL) and extracted with EA (20 mL) in total for three times. The combined organic phase was washed with water (20 mL), dried over anhydrous Na2SO4 and concentrated by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 1: 1). intermediate B 6 (2.85 g, 53.7% yield) was obtained as a white solid.
MS (ESI+): m/z 358.1 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 9.01 (s, 1H), 8.03 (t, J=8.0 Hz, 1H), 7.79-7.73 (m, 1H), 7.69-7.60 (m, 1H), 5.75-5.59 (m, 1H), 5.39 (s, 1H), 4.98 (dd, J=1.2, 10.4 Hz, 1H), 4.84-4.72 (m, 3H), 2.55 (s, 3H), 1.45 (s, 6H).
To a solution of intermediate B 6 (2.00 g, 5.60 mmol) in toluene (10 mL) was added m-CPBA (1.36 g, 6.71 mmol, 85% purity). The mixture was stirred at 25° C. for 1 h. Tert-butyl 4-(4-aminophenyl) piperazine-1-carboxylate (1.86 g, 6.71 mmol) and DIEA (2.89 g, 22.4 mmol, 3.90 mL) was added to the mixture. The mixture was stirred at 25° C. for 12 h. The mixture was quenched with saturated Na2S2O3 and extracted with EA (10 mL×2). The combined organic phase was washed with brine (5 mL), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 3: 5) to obtained the intermediate B 7 (2.79 g, 85% yield) as a white solid.
1H NMR (400 MHz, DMSO-d6) δ5 10.28-10.11 (m, 1H), 8.83 (s, 1H), 8.13-7.97 (m, 1H), 7.81-7.71 (m, 1H), 7.64-7.55 (m, 3H), 6.95 (d, J=8.8 Hz, 2H), 5.79-5.56 (m, 1H), 5.37 (s, 1H), 4.99 (dd, J=1.2, 10.3 Hz, 1H), 4.84-4.78 (m, 1H), 4.72-4.64 (m, 2H), 3.36-3.34 (m, 4H), 3.07-3.02 (m, 4H), 1.45 (s, 6H), 1.42 (s, 9H).
To the solution of intermediate B 7 (100 mg, 170 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 2 mL), the obtained solution was stirred at 25° C. for 1 hour. Then the solution was concentrated to dry to obtained the yellow solid intermediate B 7-B (90 mg, HCl salt crude). MS (ESI+): m/z 487 [M+H]+.
The following intermediates were prepared according to the procedures described in the example above using the appropriate starting materials, and the protected intermediates can be deprotected by hydrochloride or trifluoroacetic acid to provide the corresponding amine.
1H NMR (400 MHz,
To a solution of intermediate B 11-1 (2 g, 12.3 mmol), pyridine (1.22 g, 15.4 mmol, 1.3 mL) in DCM (20 mL) was added intermediate B 11-2 (2.75 g, 12.3 mmol) under the condition of ice water, the mixture was stirred at room temperature for 48 hours. The reaction mixture was diluted with H2O (50 mL) and extracted with EA (100 mL). The combined organic layers were washed with 1 N HCl (20 mL), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was triturated with CH3CN (30 mL) to give the intermediate B 11-3 (1 g, 2.87 mmol, 23.2% yield) as a white solid.
1H NMR (400 MHz, DMSO-d6) δ 10.71 (s, 1H), 8.82 (s, 1H), 7.62 (d, J=8.0 Hz, 2H), 7.46-7.40 (m, 1H), 2.60 (s, 3H).
To a solution of intermediate B 11-3 (500 mg, 1.43 mmol), methanamine hydrochloride (290 mg, 4.30 mmol) in THE (10 mL) was added TEA (435 mg, 4.30 mmol, 600 μL) and the mixture was stirred at room temperature for 48 hours. The reaction mixture was diluted with H2O (50 mL) and extracted with EA (100 mL). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (Petroleum ether: Ethyl acetate=3: 1) to give the intermediate B 11-4 (320 mg, 49.41% yield).
MS (ESI+): m/z 343.0 [M+H]+.
To a solution of intermediate B 11-4 (100 mg, 291 μmol), dibromomethane (152 mg, 874 μmol, 61 μL), Cs2CO3 (380 mg, 1.17 mmol) in MeCN (5 mL) was stirred at 80° C. for 16 h. The reaction mixture was diluted with H2O (25 mL) and extracted with EA (50 mL). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated to give the residue. The residue was purified by column chromatography (Petroleum ether: Ethyl acetate=5: 1 to give the intermediate B 11 (60 mg, 53.59% yield).
MS (ESI+): m/z 355.2 [M+H]+.
A mixture of intermediate B 12-1 (2 g, 10.8 mmol) and tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,6-dihydro-2H-pyridine-1-carboxylate (3.32 g, 10.8 mmol) in CH3CN (20 mL) and H2O (4 mL) was added K2CO3 (4.46 g, 32.3 mmol) and Pd(dppf)Cl2 (787 mg, 1.07 mmol), then stirred at 95° C. for 6 h under N2 atmosphere. The reaction mixture was filtered and dried by rotary distillation to give the residue. The residue was purified by column chromatography on silica gel (Petroleum ether/Ethyl acetate=1/0 to 5/1) to give an intermediate B 12-2 (1.02 g, 32.8% yield) as a yellow oil. MS(ESI+):m/z 388.8 [M+H]+.
1H NMR (400 MHz, CDCl3) δ 7.56 (q, J=8.4 Hz, 2H), 4.42-4.32 (m, 2H), 2.80 (s, 3H), 1.58 (s, 12H), 1.43-1.36 (m, 3H).
A mixture of intermediate B 12-2 (500 mg, 1.73 mmol) in THE (10 mL) was added 10% Pd/C (500 mg), degassed and purged with H2 for 3 times. And then the mixture was stirred at 20° C. for 16 h under H2 atmosphere. The reaction mixture was filtered and dried by rotary distillation to give the residue, which is intermediate B 12-3 (241 mg, 83.6% yield) as a white solid.
1H NMR (400 MHz, CDCl3) δ 6.96 (d, J=7.9 Hz, 1H), 6.59-6.47 (m, 2H), 4.38-4.04 (m, 2H), 3.53 (br s, 2H), 2.84-2.69 (m, 3H), 2.27 (s, 3H), 1.76-1.67 (m, 2H), 1.54-1.51 (m, 2H), 1.49 (s, 9H).
A mixture of intermediate B 11 (100 mg, 282 μmol) in DCM (5 mL) was added 85% m-CPBA (143 mg, 704 μmol) at 0° C. and then the mixture was stirred at 20° C. for 16 h. The reaction mixture was quenched by addition Na2S2O3 (10 mL) and NaHCO3 (10 mL) at 0° C., and then diluted with H2O (20 mL) and extracted with DCM (20 mL×2). The combined organic layers were washed with saturated salt solution (10 mL×2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue as intermediate B 12-4 (246 mg, 60.2% yield) as a white solid.
A mixture of intermediate B 12-4 (60 mg, 155 μmol) and intermediate B 12-3 (45.00 mg, 155 μmol) in DMSO (1 mL) was displaced with N2 for 3 times, and then the mixture was stirred at 120° C. for 16 h under N2 atmosphere. The residue was purified by prep-HPLC to purify the residue. Intermediate B 12 (19 mg, 20.3% yield) was obtained as a white solid.
1H NMR (400 MHz, DMSO-d6) δ 11.02 (br s, 1H), 9.54-9.41 (m, 1H), 8.13 (br s, 1H), 8.04 (d, J=8.0 Hz, 1H), 7.86-7.78 (m, 1H), 7.42-7.32 (m, 8H), 7.28 (d, J=6.4 Hz, 2H), 6.51-6.37 (m, 1H), 5.18-5.11 (m, 1H), 4.57-4.48 (m, 1H), 4.44-4.36 (m, 1H), 2.96-2.87 (m, 1H), 2.60 (d, J=16.4 Hz, 1H), 2.45-2.34 (m, 1H), 2.03 (d, J=5.6 Hz, 1H).
To the solution of intermediate B 12 (19 mg, 31.8 μmol) in DCM (5 mL) was added HCl/dioxane (4M, 5 mL), the obtained solution was stirred at 20° C. under N2 atmosphere for 2 hours. Then the obtained mixture was filtered and concentrated to give a crude HCl salt of intermediate B 12-B (10 mg, 42.0% yield) as a white solid, which was used for the next step without further purification.
The following intermediates were prepared by similar methods to the above.
1H NMR (400
To a solution of intermediate B 6 (246 mg, 689 μmol) in toluene (5 mL) was added m-CPBA (327 mg, 758 μmol) and stirred at 20° C. for 0.5 hours. Then DIEA (267 mg, 2.07 mmol) and intermediate B 12-3 (200 mg, 689 μmol) was added and stirred at 20° C. for 16 hrs. The reaction mixture was quenched by adding Na2S2O3 (20 mL) and NaHCO3 (20 mL) at 0° C., and extracted with DCM 30 mL (10 mL×3). The combined organic layers were washed with water 30 mL (10 mL×3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=10:1) to give intermediate B 30-1 (287 mg, 69.5% yield) as a yellow solid. LCMS (ESI+): m/z 544.4 [M+H]+
To a solution of intermediate B 30-1 (274 mg, 457 μmol) in DCM (5 mL) was added HCl/dioxane (4 M, 5 mL). The mixture was stirred at 20° C. for 16 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. Intermediate B 30 (274 mg, crude, HCl salt) was obtained as a yellow solid, which was used for the next step directly.
The following intermediates were prepared according to the procedures described in the examples above using the appropriate starting materials.
1H NMR (400
To a mixture of intermediate B 15-1 (4 g, 26.0 mmol) in toluene (40 mL) was added POBr3 (14.9 g, 52.1 mmol). The mixture was stirred at 130° C. for 72 h. The mixture was diluted with H2O (200 mL) and poured into saturated NaHCO3 (200 mL) dropwise. Then the resulting mixture (pH>7) was stirred at room temperature for 10 min and filtered. The filtrate was extracted with EA (200 mL×3) and the combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 5: 1) to give an intermediate B 15-2 (3.0 g, 52.4% yield) as a white solid.
MS (ESI+): m/z 199.92 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 7.56 (d, J=8.0 Hz, 1H), 7.33 (d, J=8.0 Hz, 1H), 2.92-2.80 (m, 4H), 2.11-2.00 (m, 2H).
To a mixture of intermediate B 15-2 (3.44, 17.4 mmol) in DCM (40 mL) was added m-CPBA (14.9 g, 52.1 mmol, 85% purity). The mixture was stirred at 25° C. for 5 h. The reaction mixture was quenched with saturated NaHCO3 (50 mL) and saturated Na2S2O3 (50 mL), and extracted with EA (100 mL×2). The combined organic layers were dried over anhydrous Na2SO4. filtered and concentrated by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 0: 1) to give intermediate B 15-3 (3.50 g, 94.1% yield) as a white solid.
1H NMR (400 MHz, CDCl3) δ 7.46 (d, J=8.0 Hz, 1H), 6.97 (d, J=8.0 Hz, 1H), 3.26-3.18 (m, 2H), 3.04-2.97 (m, 2H), 2.26-2.17 (m, 2H)
A mixture consisting of intermediate B 15-3 (3.5 g, 16.4 mmol) and Ac20 (33 mL, 349 mmol) was stirred at 110° C. for 12 h. The mixture was dried by rotary distillation and diluted with ethyl acetate (100 mL), then washed with saturated NaHCO3 (15 mL×2) and then with brine (20 mL). The mixture was dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash chromatography on silica gel (petroleum ether: ethyl acetate=1: 0 to 4: 1) to give intermediate B 15-4 (4 g, 82.6% yield) as yellow oil.
MS (ESI+): m/z 257.89 [M+H]+.
To a solution of intermediate B 15-4 (6.20 g, 24.2 mmol) in THE (65 mL) and H2O (65 mL) was added LiOH (2.32 g, 96.8 mmol). The mixture was stirred at 25° C. for 2 hrs. The mixture was poured into water (100 mL) and extracted with ethyl acetate (20 mL×2). The combined organic extract was washed with saturated salt solution (20 mL), dried over anhydrous Na2SO4. The combined organic layers were filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash chromatography on silica gel (petroleum ether: ethyl acetate=1: 0 to 3: 1) to give intermediate B 15-5 (4.50 g, 78.2% yield) as a yellow solid.
1H NMR (400 MHz, CDCl3) δ 7.45 (d, J=8.0 Hz, 1H), 7.35 (d, J=8.0 Hz, 1H), 5.19 (t, J=6.8 Hz, 1H), 3.07-2.94 (m, 1H), 2.85-2.73 (m, 2H), 2.63-2.50 (m, 1H), 2.13-2.00 (m, 1H)
To a solution of intermediate B 15-5 (4.80 g, 22.4 mmol) in DCM (100 mL) at 0° C. was added Dess-Martin periodinane (19.0 g, 44.9 mmol). The mixture was stirred at 25° C. for 2 h. The mixture was filtered and dried by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 1: 1) to give intermediate B 15-6 (4.00 g, 75.71% yield) as a white solid.
1H NMR (400 MHz, CDCl3) δ 7.76 (d, J=8.0 Hz, 1H), 7.64 (d, J=8.0 Hz, 1H), 3.16-3.10 (m, 2H), 2.82-2.77 (m, 2H)
To a solution of intermediate B 15-6 (4 g, 18.86 mmol) in THE (80 mL) was added bromo(ethyl)magnesium (19.0 g, 44.9 mmol) at 0° C. The mixture was stirred at 25° C. for 1 h. The mixture was cooled to 0° C. and added dropwise to saturated NH4C1 (100 mL). The mixture was extracted with ethyl acetate (50 mL×2) and the combined organic extract was washed with saturated salt solution (20 mL), dried over anhydrous Na2SO4, filtered and dried by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 3: 1) to give intermediate B 15-7 (2.20 g, 43.4% yield) as a brown solid.
1H NMR (400 MHz, CDCl3) δ 7.34 (d, J=8.0 Hz, 1H), 7.25 (d, J=8.0 Hz, 1H), 2.94-2.80 (m, 1H), 2.76-2.61 (m, 1H), 2.40-2.18 (m, 2H), 2.17-2.07 (m, 1H), 1.97-1.84 (m, 1H), 1.81-1.68 (m, 1H), 0.88 (t, J=7.2 Hz, 3H)
Intermediate B 15-7 (1.1 g, 4.54 mmol), intermediate B 15-8 (1.51 g, 6.82 mmol), CuI (865 mg, 4.54 mmol), K2CO3 (1.26 g, 9.09 mmol) and N,N′-dimethylethane-1,2-diamine (801 mg, 9.09 mmol) were taken up into a microwave tube in 1,4-dioxane (10 mL). The sealed tube was heated at 100° C. for 2 hrs under microwave. The mixture was filtered and dried by rotary distillation to give a crude product, which was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 1: 1) to give intermediate B 15 (0.6 g, 32.89% yield) as a white solid.
1H NMR (400 MHz, CDCl3) δ 8.94 (s, 1H), 7.79-7.73 (m, 1H), 7.71-7.66 (m, 1H), 5.76-5.64 (m, 1H), 5.07-5.01 (m, 1H), 4.96-4.85 (m, 2H), 4.79-4.67 (m, 1H), 3.11-3.00 (m, 1H), 2.93-2.81 (m, 1H), 2.57 (s, 3H), 2.45-2.34 (m, 1H), 2.29-2.19 (m, 2H), 2.04-1.94 (m, 1H), 1.91-1.79 (m, 1H), 1.01-0.97 (m, 3H)
The intermediate B 15 (1.2 g, 3.13 mmol) was separated by prep-SFC (Column: Phenomenex-Cellulose-2 (250 mm*50 mm, 10 μm); Mobile phase: [0.1% NH3H2O/ETOH]; B %: 45%-).
Intermediate B 15-A (460 mg, 38.3% yield) as peak 1 was obtained as a yellow solid.
Intermediate B 15-B (460 mg, 38.3% yield) as peak 2 was obtained as a yellow solid.
Intermediate B 15-A optical rotation: [α]20D=−46.64, (C=0.1, CHCl3)
Chiral analysis (Column: Cellulose 2 100*4.6 mm I.D., 3 μm; Mobile Phase: A:CO2, B: EtOH (0.05% DEA), Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 220 nm, RT=1.76 min)
1H NMR (400 MHz, DMSO) δ 9.03 (s, 1H), 7.91 (d, J=8.0 Hz, 1H), 7.70 (d, J=8.0 Hz, 1H), 5.81-5.59 (m, 1H), 5.10 (s, 1H), 5.03-4.96 (m, 1H), 4.90-4.79 (m, 2H), 4.70-4.59 (m, 1H), 3.07-2.92 (m, 1H), 2.87-2.73 (m, 1H), 2.54 (s, 3H), 2.28-2.16 (m, 1H), 2.10-1.97 (m, 1H), 1.93-1.82 (m, 1H), 1.77-1.65 (m, 1H), 0.87 (t, J=7.6 Hz, 3H)
Intermediate B 15-B optical rotation: [α]20D=+48.265, (C=0.1, CHCl3)
Chiral analysis (Column: Cellulose 2 100*4.6 mm I.D., 3 μm; Mobile Phase: A:CO2, B: EtOH (0.05% DEA), Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 220 nm, RT=2.41 min)
1H NMR (400 MHz, DMSO) δ 9.03 (s, 1H), 7.91 (d, J=8.0 Hz, 1H), 7.70 (d, J=8.0 Hz, 1H), 5.81-5.59 (m, 1H), 5.10 (s, 1H), 5.03-4.96 (m, 1H), 4.90-4.79 (m, 2H), 4.70-4.59 (m, 1H), 3.07-2.92 (m, 1H), 2.87-2.73 (m, 1H), 2.54 (s, 3H), 2.28-2.16 (m, 1H), 2.10-1.97 (m, 1H), 1.93-1.82 (m, 1H), 1.77-1.65 (m, 1H), 0.87 (t, J=7.6 Hz, 3H).
Intermediate of B 26 was synthesis using the method of intermediate B 15, which through nucleophilic addition reaction by using methyl Grignard reagent instead of ethyl Grignard reagent.
Intermediate B 26 (1.10 g, 2.98 mmol) was separated by prep-SFC (Column: Phenomenex-Cellulose-2 (250 mm*50 mm, 10 μM); Mobile phase: [neutral-ETOH]; CO2%: 45%-45%, min).
Compound B 26-A (500 mg, 45%) as peak 1 was obtained as a yellow solid. Compound B 26-B (550 mg, 50%) as peak 2 was obtained as a yellow solid.
Chiral analysis (Column: Cellulose 2 100*4.6 mm I.D., 3 μm; Mobile Phase: A:CO2, B: EtOH (0.05% DEA), Isocatic: A/B=6/4 at 1.0 mL/min; Temp: 35° C.; Wavelength: 220 nm, RT=2.16 min)
Chiral analysis (Column: Cellulose 2 100*4.6 mm I.D., 3 μm; Mobile Phase: A:CO2, B: EtOH (0.05% DEA), Isocatic: A/B=6/4 at 1.0 mL/min; Temp: 35° C.; Wavelength: 220 nm, RT=2.68 min)
Intermediate B 49 was synthesis using the method of intermediate B 15 through starting material of 6,7-Dihydro-5H-quinolin-8-one.
Intermediate B 49 (212 mg, 533.34 umol, 1 eq) was purified by SFC (DAICEL CHIRALPAK IG (250 mm*30 mm, 10 um); mobile phase: [Neu-MeOH]; B %: 60%-60%, min). the first peak (Rt=4.63 min) of Compound B 49-A (110 mg, 51.9% yield) and the second peak (Rt=6.59 min) of compound B 49-B (100 mg, 47.2% yield) was obtained.
Intermediate B 50 was synthesis using the method of intermediate B 15 through starting material of 6,7-Dihydro-5H-quinolin-8-one.
Intermediate B 50 (240 mg, 627 umol) was purified by SFC (DAICEL CHIRALCEL OD-H(250 mm*30 mm, 5 um); mobile phase: [Neu-ETOH]; B %: 30%-30%, 45 min). the first peak (Rt=3.76 min) of Compound B 50-A (120 mg, 50.0% yield) and the second peak (Rt=3.97 min) of compound B 50-B (120 mg, 50.0% yield) were obtained.
The following intermediates were prepared according to the procedures described in the example of intermediate B7-B using intermediate B 15-B or intermediate B 26-B as starting materials.
1H NMR (400
The following intermediates can be prepared through methods described in literature or similar methods as expected.
B 17, B 18, B 19 and B 28 were synthesized using the method from US2019/192668A1, B 20 and B 21 were synthesized using the method from WO2018/102725A1.
1H NMR (400
A mixture of intermediate B 25-1 (25 g, 125 mmol), methylhydrazine sulfuric acid salt (36.0 g, 250 mmol) and Na2CO3 (39.7 g, 375 mmol) in EtOH (100 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 120° C. for 16 hrs under N2 atmosphere. The reaction mixture was concentrated under reduced pressure to remove EtOH. The residue was diluted with H2O (200 mL) and extracted with EtOAc (300 mL) (150 mL*2). The combined organic layers were washed with brine (200 mL) (100 mL*2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give intermediate B 25-2 (14.0 g, 45.6% yield) as a yellow solid. LCMS (ESI+): m/z 228.9 [M+H]+
A mixture of the DBU (9.16 g, 60.2 mmol, 9.07 mL) and Lac (9.26 g, 60.2 mmol) was stirred at 0° C. for 16 hrs. Then the mixture was added intermediate B 25-2 (17.0 g, 75.0 mmol) and stirred at 80° C. to obtained the solution. Then the ethyl prop-2-enoate (11.3 g, 113 mmol, 12.3 mL) was added in the stirring mixture and stirred at 80° C. for 24 hrs. Then the extra ethyl prop-2-enoate (11.3 g, 113 mmol, 12.3 mL) was added in the stirring mixture and stirred at 80° C. for 24 hrs. The reaction mixture was quenched with sodium hypochlorite (30% aq, 200 mL), diluted with H2O 200 mL and extracted with EtOAc (500 mL). The combined organic extract was washed with brine (500 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=5/1 to 3/1) to give the intermediate B 25-3 (8.00 g, 30.3% yield) as a yellow solid. LCMS (ESI+): m/z 328.7 [M+H]+.
To a solution of intermediate B 25-3 (8.00 g, 24.5 mmol) in EtOH (100 mL) was added NaOAc (4.02 g, 49.1 mmol) and BrCN (8.45 g, 79.8 mmol). The mixture was stirred at 80° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to remove EtOH. The residue was diluted with H2O (100 mL) and extracted with EtOAc (100 mL×2). The combined organic layers were washed with brine (50 mL×2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=10/1 to 3/1) to give the intermediate 25-4 (5.80 g, 67.3% yield) as a yellow solid. LCMS (ESI+): m/z 352.5 [M+H]+.
To a solution of intermediate B 25-4 (5.8 g, 16.5 mmol) in toluene (50 mL) was added (E)-acetaldehyde oxime (2.93 g, 49.5 mmol, 2.99 mL) and InCl3 (365 mg, 1.65 mmol). The mixture was stirred at 110° C. for 1 hr. The reaction mixture was concentrated under reduced pressure to remove toluene. The residue was diluted with H2O (50 mL) and extracted with EtOAc (80 mL). The combined organic layers were washed with brine (60 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give intermediate 25-5 (5.60 g, 81.7% yield) as a yellow solid. LCMS (ESI+): m/z 371.8 [M+H]+.
To a solution of intermediate B 25-5 (5.6 g, 15.2 mmol) in MeCN (150 mL) was added benzyl(trimethyl)ammonium hydroxide (9.51 g, 22.8 mmol, 10.3 mL, 40% purity). The mixture was stirred at 25° C. for 0.5 hr. The reaction mixture was concentrated under reduced pressure to remove acetonitrile and give a residue. The crude product was triturated with H2O (50 mL) at 20° C. for 5 min, filtered to give a residue which is intermediate B 25 (3.55 g, 72.4% yield) as a pale yellow solid. LCMS (ESI+): m/z 324.5 [M+H]+.
1H NMR (400 MHz, CDCl3) δ 7.95-7.76 (m, 1H), 7.66-7.52 (m, 2H), 7.05-6.94 (m, 1H), 4.41-4.33 (m, 3H), 4.11-4.04 (m, 2H), 2.94-2.85 (m, 2H).
The following intermediates B 23 and B 51 were prepared according to the procedures described in the example of intermediate B 25 using the different intermediates.
1H NMR (400
A mixture of intermediate B 23 (250 mg, 774 μmol), B 46-2 (287 mg, 928 μmol), Cs2CO3 (756 mg, 2.32 mmol) and XPhos Pd G3 (131 mg, 155 μmol) in dioxane (1.5 mL) and H2O (0.06 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 60° C. for 3 hrs under N2 atmosphere. The reaction mixture was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=1:0 to 10:1) to obtained intermediate B46-3 (285 mg, 83.12% yield). LCMS (ESI+): m/z 427.2 [M+H]+.
To a solution of intermediate B 46-3 (285 mg, 670 μmol) in THF (10 mL) was added PtO2 (250 mg, 1.10 mmol). The mixture was degassed and purged with H2 for 3 times, and then the mixture was stirred at 25° C. for 16 hr under H2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give a residue as intermediate B 46-4 (240 mg, crude).
LCMS (ESI+): m/z 372.3 [M+H−56]+.
To a solution of intermediate B 46-4 (240 mg, 561 μmol) in DCM (2 mL) was added HCl/dioxane (4 M, 2 mL). The mixture was stirred at 25° C. for 0.5 hr. The reaction mixture was concentrated under reduced pressure to give a residue as intermediate B 46 (265 mg, crude).
To a mixture of intermediate B 47-1(300 mg, 560 μmol) and intermediate B 47-2(222 mg, 719 μmol) in dioxane (5 mL) and H2O (0.1 mL) was added K2CO3 (249 mg, 1.80 mmol) and Pd(dppf)Cl2 (26.3 mg, 36.0 μmol) in one portion at 100° C. under N2. The mixture was heated at 100° C. and stirred for 16 hours. The mixture was added Na2SO4, filtered and washed with DCM (100 mL). The filter liquor was concentrated in vacuo to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/0 to 1/1) to give the intermediate B 47-3 (314 mg, 69.5% yield) as colorless oil.
1H NMR (400 MHz, CDCl3) δ 7.91 (d, J=8 Hz, 1H), 7.67 (d, J=8.8 Hz, 1H), 7.48-7.43 (m, 2H), 7.42-7.33 (m, 5H), 7.30-7.27 (m, 4H), 7.15-7.07 (m, 1H), 6.53 (d, J=8 Hz, 1H), 6.14 (br s, 1H), 5.48 (s, 2H), 5.44-5.35 (m, 2H), 4.14 (br s, 2H), 4.12-4.10 (m, 3H), 3.69 (t, J=5.2 Hz, 2H), 2.63 (br s, 2H), 1.52 (s, 9H).
To a solution of intermediate B 47-3 (314 mg, 521 μmol) in THE (10 mL) was added PtO2 (200 mg, 881 μmol) under N2. The suspension was degassed under vacuum and purged with H2 three times. The mixture was stirred under H2 (15 PSi) at 20° C. for 48 hours. The reaction mixture was filtered, washed with DCM (80 mL) and the filtrate was concentrated to the crude product. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1: 0 to 0: 1) to give the intermediate B 47-4 (65 mg, 25.2% yield) as colorless oil. LCMS (ESI+): m/z 449.3 [M+23]+.
To a solution of intermediate B 47-4 (65 mg, 152 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 1.5 mL). The mixture was stirred at 25° C. for 5 hrs. The reaction mixture was concentrated under reduced pressure to give the crude product as intermediate B 47 (60 mg, crude, HCl salt) as a brown solid which was directly used into the next step.
LCMS (ESI+): m/z 327.2 [M+H]+.
DIEA (702 mg, 5.43 mmol) was added to a mixture of intermediate 1-1 (500 mg, 1.81 mmol) and 4-piperidylmethanol (417 mg, 3.62 mmol) in NMP (8 mL). The reaction mixture was stirred at 140° C. under microwave irradiation for 2 hrs. The reaction mixture was quenched with H2O (20 mL) and extracted with EtOAc (50 mL×3), then separated. The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 10: 1) to give intermediate 1-2 (500 mg, 66.9% yield) as a yellow solid.
MS (ESI+): m/z 371.8 [M+H]+.
DMP (857 mg, 2.02 mmol) was added to a solution of intermediate 1-2 (500 mg, 1.35 mmol) in DCM (20 mL). The reaction mixture was stirred at 25° C. for 2 hrs, then quenched with NaHCO3 (20 mL) and Na2SO3 (20 mL), extracted, then separated. The combined organic layers were dried over anhydrous Na2SO4, filtered, then dried by rotary distillation to give residue. The residue was purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 10: 1) to give an intermediate 1-3 (400 mg, 79.6% yield) as a yellow solid. MS (ESI+): m/z 370.0 [M+H]+.
Intermediate 1-3 (46.3 mg, 125 μmol) was added to a mixture of intermediate B-2 (30.0 mg, 62.7 μmol) and TEA (6.35 mg, 62.7 μmol) in DCM (5 mL). The mixture was stirred at 25° C. for 10 min. Then AcOH (3.77 mg, 62.72 μmol) was added and the mixture was stirred at 25° C. for another 50 min. Finally, NaBH3CN (7.88 mg, 125.44 μmol) was added to the above mixture and stirred for 1 h. The reaction mixture was concentrated by rotary distillation to give residue. The residue was purified by prep-TLC (DCM: MeOH=20:1) to give Compound 1 (3.96 mg, 7.44% yield) as a yellow solid. MS (ESI+): m/z 831.1 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.10 (br s, 1H), 10.80 (br s, 1H), 9.15 (s, 1H), 8.35-8.29 (m, 2H), 7.85-7.73 (m, 3H), 7.68-7.62 (m, 2H), 7.61-7.55 (m, 1H), 7.36-7.30 (m, 1H), 7.28-7.20 (m, 1H), 7.11 (d, J=1.6 Hz, 1H), 5.06 (dd, J=5.2, 13.2 Hz, 1H), 4.06 (d, J=13.2 Hz, 2H), 3.62-3.57 (m, 2H), 3.00 (t, J=12.0 Hz, 2H), 2.93-2.82 (m, 3H), 2.63-2.52 (m, 3H), 2.37-2.34 (m, 1H), 2.04-1.94 (m, 2H), 1.87-1.81 (m, 1H), 1.27-1.10 (m, 3H), 1.00-0.92 (m, 2H).
Compound 2 to 25 were synthesized in a similar manner to compound 1 with various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
Intermediate B 16 (0.65 g, 2.46 mmol), 4-piperidylmethanol (340 mg, 2.95 mmol) and TEA (747 mg, 7.38 mmol) in DCM (40 mL) were added drops of T3P (1.88 g, 2.95 mmol). The mixture was stirred at 25° C. for 3 hrs. The mixture was concentrated by rotary distillation to give a crude product. The crude product was purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 9: 1) to give intermediate 26-1 (0.700 g, 78.0% yield) as a white solid.
MS (ESI+): m/z 362.0 [M+H]+.
To a solution of intermediate 26-1 (0.700 g, 1.94 mmol) in DCM (10 mL) was added DMP (1.23 g, 2.91 mmol). The mixture was stirred at 25° C. for 1.5 h. The mixture was diluted with DCM (30 mL), washed with saturated NaHCO3 (10 mL×2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a crude product, then purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 10: 1) to give intermediate 26-2 (0.350 g, crude) as a white solid, which was used into the next step without further purification.
MS (ESI+): m/z 360.0 [M+H]+.
To a solution of intermediate B4 (75.0 mg, 138 μmol) in DCM (5 mL) was added TEA (76.9 μL, 553 μmol) to adjust pH to 7˜ 8, then AcOH (31.6 μL, 553 μmol) was added to adjust pH to 5˜6. Intermediate 26-2 (75.0 mg, 138 μmol) was added to the above solution and reacted at 25° C. for 2 hours. Finally, NaBH3CN (17.4 mg, 276 μmol) was added to the mixture and reacted at 25° C. for 1 h. The mixture was diluted with DCM (30 mL), washed with saturated NaHCO3 (10 mL×2). The organic extract was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 10% B to 50% B) to give Compound 26 (29.05 mg, 23% yield) as a white solid. MS (ESI+): m/z 849.2 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.96-10.69 (m, 1H), 10.36 (s, 1H), 9.15 (s, 1H), 7.90-7.60 (m, 6H), 7.41-7.08 (m, 6H), 3.84 (s, 3H), 3.54-3.52 (m, 2H), 3.07-2.87 (m, 4H), 2.73-2.64 (m, 2H), 2.49-2.42 (m, 4H), 2.23-2.11 (m, 2H), 2.03-1.93 (m, 2H), 1.80-1.57 (m, 6H), 1.16-0.99 (m, 2H).
Compound 27 to 30 were synthesized in an analogous manner to compound 26 with various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 31-1 (2.00 g, 12.9 mmol) and 4-piperidylmethanol (1.63 g, 14.2 mmol) in NMP (5 mL) was added DIEA (5.00 g, 38.7 mmol). The mixture was reacted at 140° C. under microwave irradiation for 2 hrs. The reaction mixture was poured into water (50 mL), and extracted with EA (100 mL×3). The organic layers were collected and dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give a residue. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 1: 1) to give intermediate 31-2 (2.70 g, 74.5% yield) as a yellow solid. MS (ESI+): m/z 251.12 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 8.06-7.94 (m, 2H), 7.11 (d, J=8.4 Hz, 1H), 4.54 (t, J=5.2 Hz, 1H), 3.34-3.29 (m, 4H), 2.32 (s, 3H), 2.21-2.14 (m, 1H), 1.95-1.86 (m, 1H), 1.82-1.73 (m, 2H), 1.60-1.48 (m, 1H), 1.36-1.25 (m, 2H).
To a solution of intermediate 31-2 (1.20 g, 4.79 mmol) in EtOH (30 mL) and H2O (4 mL) was added NH4Cl (1.28 g, 24.0 mmol) and iron (1.34 g, 24.0 mmol). The mixture was stirred at 80° C. for 2 hrs. Then the mixture was filtered through Celite, and extracted with EA (30 mL×2). The organic phase was collected and washed with saturated NaHCO3 (20 mL), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to obtain Intermediate 31-3 (0.83 g, 70.7% yield) as a black liquid.
MS (ESI+): m/z 221.13 [M+H]+.
A mixture of intermediate B 12-4 (428 mg, 1.15 mmol) and intermediate 31-3 (305 mg, 1.38 mmol) in DMSO (3 mL) was heated to 120° C. and reacted for 16 hrs. The reaction mixture was diluted with DCM (100 mL) and washed with H2O (50 mL×2), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give residue. The residue was purified by flash silica gel chromatography (dichloromethane/methanol=1/0 to 10/1) to give intermediate 31-4 (260 mg, 33.4% yield) as a yellow solid.
To a solution of intermediate 31-4 (260 mg, 493 μmol) in DCM (20 mL) was added DMP (314 mg, 740 μmol) slowly. The mixture was reacted at 25° C. for 1 h. The mixture was diluted with DCM (30 mL), washed with saturated NaHCO3 (10 mL×2). The organic extract was dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give a crude product. The crude product was purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 10: 1) to give intermediate 31-5 (120 mg, 43.6% yield) as a white solid.
MS (ESI+): m/z 525.07 [M+H]+.
To a solution of intermediate B 20 (34.6 mg, 91.4 μmol, HCl) in DCM (5 mL) was added TEA (30.0 mg, 304 μmol) to adjust pH to 7˜ 8. Intermediate 31-5 (40.0 mg, 76.1 μmol) and AcOH (4.57 mg, 76.1 μmol) was added to the mixture to adjust pH to 5˜ 6. The mixture was stirred at 25° C. for 3.5 hrs. NaBH3CN (9.57 mg, 152 μmol) was added to the mixture and it was reacted for 1.5 hrs. The mixture was concentrated by rotary distillation to gice a residue, which was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% TFA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 10% B to 50% B) to give Compound 31 (30.5 mg, 46.6% yield) as a yellow solid.
MS (ESI+): m/z 851.2 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.13 (s, 1H), 10.00-9.79 (m, 1H), 8.47 (s, 1H), 7.81-7.74 (m, 1H), 7.69-7.54 (m, 4H), 7.53-7.38 (m, 3H), 7.17-6.95 (m, 1H), 5.17-5.06 (m, 1H), 4.99 (s, 2H), 3.90-3.87 (m, 2H), 3.74-3.60 (m, 2H), 3.41-3.20 (m, 6H), 3.12 (s, 5H), 2.95-2.83 (m, 1H), 2.78-2.55 (m, 3H), 2.45-2.38 (m, 1H), 2.26 (s, 3H), 2.09-1.95 (m, 2H), 1.93-1.80 (m, 2H), 1.51-1.37 (m, 2H).
Compound 32 to 33 were synthesized in a similar manner to compound 31 with various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate B 21 (70.0 mg, 192 μmol, HCl) in DCM (5 mL) was added TEA (92.4 mg, 913 μmol) to adjust pH to 7˜8. Intermediate 31-5 (120 mg, 228 μmol) and AcOH (13.7 mg, 228 μmol) was added to the above mixture to adjust pH to 5˜6. The mixture was stirred at 25° C. for 2 hrs. Then, NaBH3CN (21.5 mg, 342 μmol) was added to the mixture and it was stirred at 25° C. for 12 hrs. The mixture was concentrated by rotary distillation. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: FA, 20 mL/min, gradient condition from 10% B to 45%) to give compound 34 (8.50 mg, 4.35% yield) as a white solid.
MS (ESI+): m/z 837.2 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.96 (br s, 1H), 9.73 (s, 1H), 8.45 (s, 1H), 7.67-7.61 (m, 2H), 7.59-7.43 (m, 4H), 7.11-7.03 (m, 2H), 7.00-6.93 (m, 1H), 5.10-5.00 (m, 1H), 4.96 (s, 2H), 4.39-4.14 (m, 2H), 3.35-3.30 (m, 4H), 3.10 (s, 3H), 3.04-2.97 (m, 2H), 2.94-2.84 (m, 1H), 2.64-2.51 (m, 7H), 2.39-2.31 (m, 1H), 2.28-2.19 (m, 5H), 2.02-1.90 (m, 1H), 1.87-1.76 (m, 2H), 1.73-1.58 (m, 1H), 1.33-1.23 (m, 2H)
The compound 56 to compound 57 were synthesized according to the procedures described for Compound 34 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
DIEA (6.67 g, 51.57 mmol) was added to a mixture of 4-piperidylmethanol (1.47 g, 9.67 mmol) and intermediate 35-1 (1 g, 6.45 mmol) in DMF (10 mL). The reaction mixture was stirred at 100° C. for 12 hrs. The reaction mixture was quenched with saturated NaCl (50 mL) and then extracted with EA (50 mL×3). The combined organic layers were washed with saturated salt solution (30 mL), separared, dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1: 0 to 1: 2) to give 35-3 (1.30 g, 80.5% yield) as a white solid.
MS (ESI+): m/z 250.8 [M+H]+.
LiOH·H2O (327 mg, 7.79 mmol) was added to a solution of intermediate 35-3 (1.30 g, 5.19 mmol) in H2O (5 mL) and THE (5 mL). The reaction mixture was stirred at 25° C. for 2 hrs. The reaction mixture was neutralized by HCl (2 M) until pH to ˜3. The mixture was concentrated by rotary distillation to give intermediate 35-4 (1.20 g, 96.8% yield) as a white solid.
MS (ESI+): m/z 237.12 [M+H]+.
EDCI (609 mg, 3.17 mmol) was added to a mixture of intermediate 35-4 (500 mg, 2.12 mmol), intermediate 35-5 (418 mg, 2.54 mmol), DIEA (1.37 g, 10.58 mmol) and HOBt (572 mg, 4.23 mmol) in DMF (10 mL). The reaction mixture was stirred at 25° C. for 2 hrs. The reaction mixture was concentrated by rotary distillation to give residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 25% B to 55%) to give intermediate 35-6 (150 mg, 19.9% yield) as a white solid.
MS (ESI+): m/z 347.08 [M+H]+.
DMP (122 mg, 289 μmol) was added to a solution of intermediate 35-6 (50.0 mg, 144 μmol) in DCM (5 mL). The reaction mixture was stirred at 25° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give the residue. The residue was purified by flash silica gel chromatography (dichloromethane: methanol=1: 0 to 10: 1) to give intermediate 35-7 (50.0 mg, 74.4% yield) as a yellow oil.
MS (ESI+): m/z 345.08 [M+H]+.
AcOH (8.72 mg, 145 μmol) was added to a mixture of intermediate 35-7 (50.0 mg, 145 μmol) and intermediate B 8 (70.5 mg, 145 μmol) in DCM (5 mL). The mixture was stirred for 1 h. Then NaBH3CN (18.3 mg, 290 μmol) was added to the mixture and stirred at 25° C. for 1 h. The reaction mixture was quenched with H2O (10 mL) and extracted with DCM (10 mL×3), separated. The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30×150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 10% B to 45%) to give compound 35 (8.24 mg, 6.94% yield) as a white solid.
MS (ESI+): m/z 814.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.86 (s, 1H), 10.26 (s, 1H), 8.88 (s, 1H), 8.77-8.66 (m, 1H), 8.35-8.29 (m, 1H), 8.15-8.00 (m, 1H), 7.90-7.82 (m, 1H), 7.79-7.73 (m, 1H), 7.72-7.57 (m, 3H), 7.47-7.32 (m, 1H), 7.27-7.13 (m, 2H), 5.76-5.58 (m, 1H), 5.00 (d, J=10.0 Hz, 1H), 4.88-4.79 (m, 1H), 4.79-4.72 (m, 1H), 4.72-4.63 (m, 2H), 4.04-3.87 (m, 2H), 3.26-3.19 (m, 1H), 3.12-2.70 (m, 6H), 2.26-2.11 (m, 3H), 2.05-1.95 (m, 3H), 1.87-1.62 (m, 7H), 1.47 (s, 6H), 1.26-1.14 (m, 2H).
To a solution of intermediate 36-1 (2.00 g, 9.94 mmol) TEA (3.02 g, 29.8 mmol) in DCM (10 mL) was added 4-methylbenzenesulfonyl chloride (2.84 g, 14.9 mmol) in batches. The mixture was stirred at 25° C. for 2 hrs. The mixture was concentrated by rotary distillation. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1: 0 to 1: 1) to give intermediate36-2 (2.78 g, 73.2% yield) as a white solid. MS (ESI+): m/z 378.1 [M+Na]+.
Intermediate B 8 (140 mg, 268 μmol, HCl), intermediate 36-2 (143 mg, 402 μmol) and TEA (81.4 mg, 805 μmol) in ACN (2 mL) were added KI (134 mg, 805 μmol). The mixture was reacted at 100° C. for 2 hrs under microwave, then concentrated to give the residue. The residue was purified by flash silica gel chromatography (petroleum ether: ethyl acetate=1:0 to 0:1).to give intermediate 36-3 (150 mg, 64.4% yield) as a white solid. MS (ESI+): m/z 669.5 [M+H]+.
To a mixture of intermediate 36-3 (150 mg, 224 μmol) in DCM (2 mL) was added HCl/dioxane (4M, 4 mL) and stirred at 25° C. for 1 h. The mixture was concentrated by rotary distillation to give intermediate 36-4 (130 mg, crude, HCl salt) as a yellow solid. MS (ESI+): m/z 569.24 [M+H]+.
Intermediate 36-4 (65.0 mg, 107 μmol, HCl salt), DIEA (41.7 mg, 322 μmol), intermediate 1-1 (32.6 mg, 118 μmol) in DMSO (1 mL) was reacted at 110° C. for 1 h under microwave. The mixture was concentrated by rotary distillation to give the residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: FA, 20 mL/min, gradient condition from 15% B to 55% B). to give compound 36 (10.0 mg, 11.1% yield) as a yellow solid.
MS (ESI+): m/z 825.4 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.07 (s, 1H), 10.25 (s, 1H), 8.87 (s, 1H), 8.10-8.00 (m, 1H), 7.83-7.71 (m, 1H), 7.69-7.56 (m, 4H), 7.25-7.18 (m, 2H), 6.92-6.89 (m, 1H), 6.86-6.78 (m, 1H), 5.72-5.61 (m, 1H), 5.34 (s, 1H), 5.09-5.02 (m, 1H), 5.01-4.96 (m, 1H), 4.85-4.78 (m, 1H), 4.71-4.64 (m, 2H), 3.61-3.55 (m, 1H), 3.53-3.47 (m, 1H), 3.19-3.12 (m, 1H), 3.08-2.97 (m, 2H), 2.92-2.83 (m, 1H), 2.65-2.58 (m, 2H), 2.55-2.54 (m, 1H), 2.46-2.35 (m, 4H), 2.21-2.10 (m, 1H), 2.06-1.95 (m, 3H), 1.82-1.60 (m, 5H), 1.46 (s, 6H).
Intermediate 37-1 (2 g, 9.94 mmol), 4-methylbenzenesulfonyl chloride (2.8 g, 14.9 mmol) DMAP (364 mg, 2.98 mmol), Et3N (2 g, 19.87 mmol, 2.8 mL) in DCM (20 mL) was reacted at 20° C. for 16 hrs. The reaction mixture was concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (petroleum ether/ethyl acetate=5/1 to 3/1) to give intermediate 37-2 (4.2 g, crude) as a white solid.
MS (ESI+): m/z 355.7 [M+H]+.
Intermediate 37-2 (2 g, 5.63 mmol), intermediate 37-3 (387 mg, 1.88 mmol), Et3N (1.9 g, 18.76 mmol, 2.6 mL), KI (934 mg, 5.63 mmol) in NMP (1 mL) was reacted at 110° C. for 2 hrs under microwave. The reaction mixture was diluted with H2O (80 mL) and extracted with DCM (500 mL) in total for twice. The combined organic layers were washed with NaCl (150 mL), dried over anhydrous Na2SO4, and concentrated by rotary distillationthe to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=1/0 to 3/1) to give intermediate 37-4 (400 mg, 42.5% yield) as a yellow liquid.
MS (ESI+): m/z 390.4 [M+H]+.
A mixture of intermediate 37-4 (200 mg, 513.50 μmol) in DCM (5 mL) was added HCl/dioxane (4 M, 5 mL) and reacted at 20° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give the intermediate 37-5 (300 mg, crude) as a yellow solid, which was directly used into the next step.
MS (ESI+): m/z 290.1 [M+H]+.
A mixture of intermediate 37-5 (200 mg, 691 mol), intermediate 1-1 (210 mg, 760 μmol), DIEA (268 mg, 2.07 mmol, 361 μL) in DMSO (1 mL) was reacted at 20° C. for 2 hrs. The reaction mixture was diluted with H2O (30 mL) and extracted with EA (150 mL) in total for three times. The combined organic layers were washed with saturated salt solution (50 mL), dried over Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (Dichloromethane: Methanol=1/0 to 20/1) to give intermediate 37-6 (100 mg, 26.0% yield) as a yellow solid.
MS (ESI+): m/z 546.2 [M+H]+.
A mixture of intermediate 37-6 (100 mg, 183 μmol), Pd/C (300 mg, 10%) in THE (3 mL) was reacted under H2 atmosphere at 20° C. for 2 h. The reaction mixture was filtered and concentrated by rotary distillation to give the residue, which was purified by column chromatography (Dichloromethane: Methanol=1/0 to 20/1) to give intermediate 37-7 (30 mg, 31.7% yield) as a yellow solid.
MS (ESI+): m/z 516.2 [M+H]+.
To a solution of intermediate B 6 (20 mg, 55.96 μmol) in toluene (3 mL) was added m-CPBA (18.8 mg, 87.3 μmol, 80%). After addition, the mixture was reacted at 20° C. for 0.5 hr and then added intermediate 37-7 (30 mg, 58.2 μmol) and DIEA (22.6 mg, 175 μmol, 30 μL). The mixture was reacted at 20° C. for 16 hrs. The reaction mixture was quenched by Na2S2O3 (10 mL) and NaHCO3 (10 mL), and then extracted with EA (60 mL) in total for two times. The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by prep-HPLC (acetonitrile/water (0.1% TFA)), and the crude was further purified by prep-TLC (DCM: MeOH=10:1) to give compound 37 (4 mg, 8.15% yield) as a yellow solid.
MS (ESI+): m/z 825.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.08 (s, 1H), 10.26 (br s, 1H), 8.88 (s, 1H), 8.16-7.97 (m, 1H), 7.76 (d, J=8.0 Hz, 1H), 7.71-7.59 (m, 4H), 7.23 (d, J=12.0 Hz, 2H), 6.91 (s, 1H), 6.83 (d, J=8.0 Hz, 1H), 5.75-5.60 (m, 1H), 5.35 (s, 1H), 5.11-4.96 (m, 2H), 4.82 (d, J=16.0 Hz, 1H), 4.72-4.62 (m, 2H), 3.61-3.54 (m, 1H), 3.44-3.38 (m, 2H), 3.20-3.12 (m, 1H), 3.10-2.95 (m, 2H), 2.94-2.82 (m, 1H), 2.58 (d, J=20.0 Hz, 2H), 2.38 (d, J=8.0 Hz, 2H), 2.16 (d, J=8.0 Hz, 2H), 2.09-1.90 (m, 4H), 1.83-1.62 (m, 5H), 1.47-1.43 (s, 6H).
Compound 38 was synthesized in an analogous manner of compound 37 with different intermediate.
1H NMR (400 MHz, DMSO-d6) δ
To a solution intermediate B 8 (80.0 mg, 153 μmol, HCl) in MeOH (2 mL) was added TEA (62.0 mg, 613 μmol), and pH is about 7˜ 8. Intermediate 39-1 (114 mg, 613 μmol) and AcOH (9.20 mg, 153 μmol) was added to the above mixture and pH is about 5˜ 6. The mixture was stirred at 25° C. for 2 hrs. Then NaBH3CN (14.4 mg, 230 μmol) was added to the mixture and it was stirred at 25° C. for 12 hrs. The mixture was quenched with NH4Cl (10 mL) and extracted with DCM (20 mL×2). The combined organic layers were washed with saturated salt solution (20 mL), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1:0 to 0:1) to give intermediate 39-2 (100 mg, 97.7% yield) as a yellow solid.
MS (ESI+): m/z 655.0 [M+H]+.
To a solution of intermediate 39-2 (100 mg, 153 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 2 mL). The mixture was stirred at 25° C. for 1 h. The mixture was concentrated by rotary distillation to give intermediate 39-3 (85.0 mg, crude, HCl salt) as a yellow solid.
MS (ESI+): m/z 555.21 [M+H]+.
Intermediate 39-3 (85.0 mg, 144 μmol, HCl), intermediate 1-1 (43.7 mg, 158 μmol) and TEA (43.7 mg, 431 μmol) in NMP (2 mL) were reacted at 110° C. for 1 h under microwave. The mixture was concentrated to give the residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: ACN, 20 mL/min, gradient condition from 10% B to 50%) to give intermediate 39 (10.0 mg, 8.25% yield) as a white solid.
MS (ESI+): m/z 811.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.08 (s, 1H), 10.27 (s, 1H), 8.87 (s, 1H), 8.10-7.99 (m, 1H), 7.79-7.72 (m, 1H), 7.71-7.59 (m, 4H), 7.27-7.19 (m, 2H), 6.99-6.96 (m, 1H), 6.89-6.81 (m, 1H), 5.73-5.61 (m, 1H), 5.38-5.31 (m, 1H), 5.19-5.04 (m, 1H), 5.02-4.96 (m, 1H), 4.86-4.78 (m, 1H), 4.73-4.64 (m, 2H), 3.77-3.69 (m, 1H), 3.62-3.55 (m, 1H), 3.14-3.04 (m, 2H), 3.03-2.96 (m, 1H), 2.93-2.84 (m, 1H), 2.60-2.56 (m, 1H), 2.45-2.41 (m, 2H), 2.32-2.24 (m, 2H), 2.18-2.04 (m, 3H), 2.03-1.98 (m, 1H), 1.92-1.84 (m, 1H), 1.79-1.64 (m, 4H), 1.46 (s, 6H).
Compound 40 to 44 were synthesized in an analogous manner of compound 39, via various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 45-1 (1.00 g, 2.97 mmol), tert-butyldimethyl(prop-2-yn-1-yloxy)silane (1.01 g, 5.93 mmol), TEA (300 mg, 2.97 mmol), CuI (113 mg, 593 μmol) in DMSO (20 mL) was stirred and degassed for 3 times, and Pd(PPh3)4 (343 mg, 297 μmol) was added, then the mixture was stirred at 85° C. for 1 h under N2 atmosphere. The mixture was diluted with ethyl acetate (30 mL) and H2O (40 mL), and then extracted with EtOAc (30×2 mL). The combined organic layers were washed with saturated salt water for 2 times, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1: 0 to 3: 1) to give intermediate 45-2 (1.18 g, 92.3% yield) as a brown soild.
1H NMR (400 MHz, DMSO-d6) δ 11.14 (s, 1H), 8.17-7.65 (m, 3H), 5.16 (dd, J=5.2, 12.8 Hz, 1H), 4.75-4.38 (m, 2H), 3.02-2.77 (m, 1H), 2.64-2.55 (m, 2H), 2.14-2.02 (m, 1H), 0.90 (s, 9H), 0.15 (s, 6H).
To a solution of intermediate 45-2 (500 mg, 1.17 mmol) in THF (10 mL) was added wet Pd/C (200 mg, 606.28 μmol, 10% purity) and Pd(OH)2/C (200 mg, 142 μmol, 20% purity). The mixture was stirred at 25° C. for 12 hrs under H2 atmosphere. The mixture was filtered and concentrated by rotary distillation to give intermediate 45-3 (380 mg, 75.3% yield) as a yellow oil.
1H NMR (400 MHz, DMSO-d6) δ 11.12 (s, 1H), 7.83 (d, J=7.8 Hz, 1H), 7.76 (s, 1H), 7.70 (d, J=7.8 Hz, 1H), 5.13 (s, 1H), 3.67-3.52 (m, 2H), 2.97-2.77 (m, 3H), 2.65-2.52 (m, 2H), 2.10-2.01 (m, 1H), 1.87-1.70 (m, 2H), 0.92-0.82 (m, 9H), 0.03 (s, 6H).
To a solution of intermediate 45-3 (200 mg, 465 μmol) in DCM (8 mL) was added HCl/dioxane (12 mL, 4M). The mixture was sirred at 25° C. for 1 h. The mixture was concentrated by rotary distillation to give intermediate 45-4 (120 mg, 81.7% yield) as a yellow soild.
MS (ESI+): m/z 317.03 [M+H]+.
To a solution of intermediate 45-4 (120 mg, 379 μmol) in DCM (4 mL) was added DMP (241 mg, 569 μmol). The mixture was stirred at 25° C. for 1 h under N2 atmosphere. The mixture was added saturated Na2S2O3 (6 ml) and extracted with DCM (8 mL). The combined organic layers were washed with saturated salt solution (10 mL×2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1: 0 to 0: 1) to give intermediate 45-5 (102 mg, 85.6% yield) as a colorless oil.
1H NMR (400 MHz, DMSO-d6) δ 11.12 (s, 1H), 9.71 (s, 1H), 7.86-7.79 (m, 2H), 7.77-7.70 (m, 1H), 5.18-5.08 (m, 1H), 3.12-3.01 (m, 2H), 2.94-2.78 (m, 3H), 2.59 (s, 2H), 2.11-2.00 (m, 1H).
To a solution of intermediate B 7-B (136 mg, 260 μmol, HCl) in DCM (6 mL) was added TEA (98.5 mg, 974 μmol) to adjust pH to 7˜ 8. Then intermediate 45-5 (102 mg, 325 μmol) and AcOH (39.0 mg, 649 μmol) was added to the above mixture to adjust pH to 5˜ 6. The mixture was stirred at 25° C. for 12 hrs under N2 atmospher. Then NaBH3CN (24.5 mg, 389 μmol) was added and the mixture was stirred at 25° C. for 1 h under N2 atmospher. The mixture was diluted with DCM (16 mL) and H2O (20 mL), then extracted with DCM (30 mL×2) and the combined organic layers were washed with saturated salt solution (10 mL×2), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 40% B to 60%) to give intermediate 45 (36.0 mg, 13.4% yield) as a white soild.
MS (ESI+): m/z 785.5[M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.15 (s, 1H), 10.65-9.97 (m, 1H), 8.85 (s, 1H), 8.11-7.99 (m, 1H), 7.94-7.88 (m, 2H), 7.86-7.42 (m, 5H), 7.01 (d, J=8.8 Hz, 2H), 5.75-5.56 (m, 1H), 5.16 (dd, J=5.6, 12.8 Hz, 1H), 5.05-4.95 (m, 1H), 4.88-4.76 (m, 1H), 4.73-4.60 (m, 2H), 3.83-3.72 (m, 2H), 3.66-3.52 (m, 2H), 3.19-3.00 (m, 6H), 2.97-2.80 (m, 3H), 2.65-2.53 (m, 2H), 2.22-2.02 (m, 3H), 1.52-1.40 (m, 6H)
Compound 46 to 51 were synthesized in an analogous manner of compound 45, via various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate B 6 (200 mg, 560 μmol) in toluene (3 mL) was added m-CPBA (136 mg, 671 μmol, 85% purity). The mixture was stirred at 25° C. for 1 h. Intermediate 52-1 (139 mg, 671 μmol) and DIEA (289 mg, 2.24 mmol) was added. The mixture was stirred at room tempreture for 3 hrs. The reaction mixture was quenched by Na2S2O3 (10 mL, 1M), and then extracted with EtOAc (10 mL×3). The combined organic layers were washed with saturated salt solution (10 mL×2), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (Petroleum ether:ethyl acetate=1:0 to 1:1) to give intermediate 52-2 (230 mg, 71.8% yield) as a yellow solid.
MS (ESI+): m/z 516.19 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.15 (s, 1H), 8.82 (s, 1H), 8.15-7.97 (m, 1H), 7.82-7.70 (m, 1H), 7.63-7.47 (m, 3H), 7.09-6.83 (m, 2H), 5.74-5.53 (m, 1H), 5.36 (s, 1H), 5.08-4.93 (m, 1H), 4.88-4.77 (m, 1H), 4.74-4.59 (m, 2H), 4.57-4.48 (m, 1H), 3.73-3.56 (m, 2H), 3.31-3.29 (m, 2H), 2.63-2.58 (m, 2H), 1.80-1.70 (m, 2H), 1.50-1.42 (m, 7H), 1.29-1.21 (m, 2H).
MsCl (190 mg, 1.66 mmol) was added to a solution of intermediate 52-2 (122 mg, 237 μmol), TEA (71.8 mg, 710 μmol) in DCM (5 mL). The reaction mixture was reacted at 25° C. for 1 h. The reaction mixture was poured into H2O (20 mL) and extracted with DCM (30 mL×2). The organic layer was collected and dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the intermedaite 52-3 (130 mg, 70.7% yield) as a green solid.
1H NMR (400 MHz, DMSO-d6) δ 10.16 (s, 1H), 8.82 (s, 1H), 8.17-7.98 (m, 1H), 7.81-7.70 (m, 1H), 7.62-7.52 (m, 3H), 6.96-6.85 (m, 2H), 5.73-5.59 (m, 1H), 5.36 (s, 1H), 5.05-4.94 (m, 1H), 4.88-4.79 (m, 1H), 4.76-4.61 (m, 2H), 4.16-4.04 (m, 2H), 3.72-3.64 (m, 2H), 3.19-3.17 (m, 3H), 2.67-2.60 (m, 2H), 1.81-1.75 (m, 2H), 1.50-1.43 (m, 7H), 1.42-1.33 (m, 2H)
Intermediate 52-3 (130 mg, 153 μmol), intermediate 52-4 (52.5 mg, 153 μmol) and DIEA (59.4 mg, 460 μmol) in DMF (3 mL) was reacted at 100° C. for 1 h under MW. The reaction mixture was poured into H2O (20 mL) and extracted with DCM (30 mL×2). The organic layer was collected and dried over anhydrous Na2SO4, filtered and dried by rotaty distillation to give the residue. The residue was purified by prep-HPLC (Calumn:Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition form 20% B to 50%) to give compound 52 (10.0 mg, 15.5% yield) as a yellow solid.
MS (ESI+): m/z 840.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.09 (s, 1H), 10.05 (s, 1H), 8.86-8.68 (m, 1H), 8.08-7.95 (m, 1H), 7.90-7.19 (m, 5H), 7.16-6.85 (m, 3H), 6.54-6.47 (m, 1H), 5.72-5.59 (m, 1H), 5.33 (dd, J=5.2, 12.8 Hz, 1H), 5.05-4.94 (m, 1H), 4.88-4.76 (m, 1H), 4.74-4.60 (m, 2H), 3.32 (s, 3H), 3.05-3.00 (m, 2H), 2.93-2.82 (m, 3H), 2.67-2.61 (m, 2H), 2.46-2.25 (m, 5H), 2.18-1.94 (m, 5H), 1.81-1.58 (m, 7H), 1.45 (s, 6H).
Compound 53 to 54 were synthesized in an analogous manner of compound 52, via various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate B 17 (250 mg, 793 μmol) and intermediate 55-1 (297 mg, 961 μmol) in dioxane (5 mL) and H2O (0.2 mL) was added Cs2CO3 (723 mg, 2.22 mmol) and XPhos Pd G3 (125 mg, 148 μmol), then the mixture was displaced with N2 for 3 times, and then the mixture was stirred at 60° C. for 3 h under N2 atmosphere. The reaction mixture was dried over anhydrous Na2SO4 and filtered. The filtrate was concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (DCM: MeOH=1: 0 to 20: 1). Intermediate 55-2 (322 mg, 85.03% yield) was obtained as a white solid.
MS (ESI+): m/z 384.8 [M+H−56]+.
To a solution of intermediate 55-2 (322 mg, 629 μmol) in THE (15 mL) was added PtO2 (500 mg, 2.20 mmol). The reaction mixture was then degassed and purged with H2 for 3 times, and then the mixture was stirred at 20° C. for 32 h under H2 atmosphere. The reaction mixture was filtered and concentrated by rotary distillation to give the residue. Intermediate 55-3 (324 mg, crude) was obtained as a white solid.
MS (ESI+): m/z 387.0 [M+H−56]+.
To a solution of intermediate 55-3 (324 mg, 732 μmol) in DCM (5 mL) was added HCl/dioxane (4 M, 4 mL). The mixture was stirred at 20° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give the residue. Intermediate 55-4 (302 mg, crude) was obtained as a white solid.
A solution of intermediate 55-5 (1.0 g, 7.09 mmol), intermediate 55-6 (1.13 g, 7.09 mmol) in DMSO (5 mL) was added DIEA (4.58 g, 35.4 mmol, 6.17 mL) and stirred at 90° C. for 1 h. The reaction mixture was seprated with EA (20 mL) and water (20 mL). The organic layer was washed with saturated NaCl (30 mL), dried over anhydrous Na2SO4, concentrated by rotaty destillation to give the residue. The residue was purified by column chromatography (Petroleum ether/ethyl acetate=3/1). Intermediate 55-7 (1.50 g, 75.5% yield) was obtained as a yellow solid.
A solution of intermediate 55-7 (1.5 g, 5.35 mmol) in C4H8O (20 mL) was added Pd/C (1 g, 10% purity) and was exchanged with H2 for 3 times, then stirred at 30° C. under H2 for 1 hour. The reaction mixture was filtered and concentrated by rotaty distillation to give the intermediate 55-8 (1.35 g, crude) as a brown solid.
To a solution of intermediate B 6 (100 mg, 280 μmol) in toluene (3 mL) was added m-CPBA (62.5 mg, 308 μmol) and stirred at 30° C. for 0.5 h and then DIEA (72.3 mg, 560 μmol, 98 μL) and intermediate 55-8 (77.0 mg, 308 μmol) were added. The resulting mixture was stirred at 30° C. for 10 hours. The reaction mixture was added EA (50 mL) and saturated Na2S2O3 (30 mL), separated. The organic layer was washed with saturated NaCl (30 mL), dried over anhydrous Na2SO4 and concentrated by rotaty distillation to give the residue. The residue was purified by column chromatography (Petroleum ether/ethyl acetate=1/2) to give intermediate 55-9 (160 mg, 98.9% yield) as a yellow solid.
MS (ESI+): m/z 560.0 [M+H]+.
A solution of intermediate 55-9 (60 mg, 107 μmol,) and HCl/dioxane (4 M, 2 mL) was stirred at 30° C. for 1 hour. The reaction mixture was concentrated by rotaty distillation to give the residue. Intermediate 55-10 (65.0 mg, crude) was obtained as a yellow solid and used for the next step directly.
A solution of intermediate 55-10 (50 mg, 132 μmol), intermediate 55-4 (65 mg, 118 μmol) and DMF (1.5 mL) in DCM (4 mL) was added Et3N (120 mg, 1.18 mmol, 164 μL) and stirred at 30° C. for 0.5 h. Then AcOH (70.6 mg, 1.18 mmol, 67.6 μL) was added and the reaction mixture was stirred at 30° C. for 0.5 h. Then NaBH(OAc)3 (150 mg, 709 μmol) was added and the reaction mixture was stirred at 30° C. for 16 hours. The reaction mixture was seprated with DCM (10 mL) and water (10 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated by rotaty distillation to give the residue which was purified by prep-HPLC (acetonitrile/water (2% FA)) to give compound 55 (30.0 mg, 28.9% yield) as a yellow solid.
MS (ESI+): m/z 840.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.09 (s, 1H), 10.12 (br s, 1H), 8.82 (s, 1H), 8.22 (s, 0.476 H), 8.04 (br s, 1H), 7.82-7.69 (m, J=8.4 Hz, 1H), 7.68-7.45 (m, 3H), 7.10-6.85 (m, 5H), 5.73-5.59 (m, 1H), 5.43-5.24 (m, 2H), 5.07-4.94 (m, J=10.4 Hz, 1H), 4.82 (d, J=17.2 Hz, 1H), 4.73-4.59 (m, J=4.8 Hz, 2H), 3.64 (d, J=12.0 Hz, 2H), 3.58 (s, 3H), 2.99 (d, J=10.8 Hz, 2H), 2.93-2.81 (m, 1H), 2.76-2.57 (m, 4H), 2.45-2.38 (m, 1H), 2.28-2.17 (m, J=6.4 Hz, 2H), 2.14-1.93 (m, 3H), 1.86-1.62 (m, 7H), 1.46 (s, 6H), 1.31-1.15 (m, 2H).
The compound 95 to 96 were synthesized according to the procedures described for compound 55 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
A mixture of intermediate B 16 (500 mg, 1.89 mmol), 3-Aminopiperidine-2,6-dione (492 mg, 2.99 mmol, HCl), TEA (957 mg, 9.46 mmol, 1.3 mL), T3P (2.4 g, 3.78 mmol, 2.3 mL, 50% purity) in DCM (5 mL) was stirred at 20° C. for 6 hrs. The reaction mixture was diluted with H2O (30 mL) and extracted with Dichloromethane: Methanol=10:1 (50 mL×4). The combined organic layers were seprated and dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give intermediate 58-1 (200 mg, 14.6% yield) as a yellow solid.
MS (ESI+): m/z 362.1 [M+H]+.
A mixture of intermediate 58-1 (300 mg, 830.12 μmol) and DMP (704 mg, 1.66 mmol, 514.00 μL) in DCM (5 mL) was stirred at 20° C. for 1 h. The reaction mixture was diluted with Na2S2O3:NaHCO3=1:1 (100 mL) and extracted with DCM (30 mL). The combined organic layers were seprated, dried over anhydrous Na2SO4, and concentrated by rotary distillation to give the residue. Intermediate 58-2 (280 mg, 91.7% yield) was obtained as a yellow liquid.
MS (ESI+): m/z 360.1 [M+H]+.
To a mixture of intermediate 58-2 (200 mg, 557 μmol) and tert-butyl piperazine-1-carboxylate (103 mg, 557 μmol) in DCM (2 mL) was added AcOH (67 mg, 1.11 mmol), the mixture was stirred for 1 hour, then added NaBH(OAc)3 (354 mg, 1.67 mmol), and then the mixture was stirred at 20° C. for 16 hrs. The reaction mixture was added H2O (20 mL) and DCM (50 mL). The combined organic layers were washed dried over Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (SiO2, Dichloromethane: Methanol=1/0 to 10/1). Intermediate 58-3 (150 mg, 45.0% yield) was obtained as a yellow solid.
MS (ESI+): m/z 529.9 [M+H]+.
A mixture of intermediate 58-3 (150 mg, 283.22 μmol) and HCl/dioxane (4 M, 10 mL) in DCM (3 mL) was stirred at 20° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give intermediate 58-4 (150 mg, crude) as a gray solid, and the crude product was used into the next step.
MS (ESI+): m/z 430.1 [M+H]+.
A mixture of intermediate B 2-1 (1.6 g, 4.06 mmol and (4-aminophenyl)methanol (800 mg, 6.50 mmol) in DMSO (8 mL) was reacted at 120° C. for 3 hours. The reaction mixture was concentrated to give the intermediate 58-5 (330 mg, 12.6 yield) as a brown solid.
MS (ESI+): m/z 454.96 [M+H]+.
A mixture of intermediate 58-5 (30 mg, 66.2 μmol) and SOCl2 (40 mg, 331 μmol, 24.0 μL) in DCM (3 mL) was reacted at 20° C. for 1 h. The reaction mixture was dried by rotary distillation to give intermediate 58-6 (31 mg, 99.29% yield) as a yellow solid.
A mixture of intermediate 58-4 (30 mg, 63.60 μmol), intermediate 58-6 (30 mg, 59.71 μmol, HCl) and K2CO3 (41.26 mg, 299 μmol) in DMF (3 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 20° C. for 16 hrs under N2 atmosphere. The reaction mixture was filtered and dried by rotary distillation, then purified by prep-HPLC (acetonitrile/water (0.1% TFA)) to give compound 58 (2.43 mg, 4.66% yield) as a white solid.
MS (ESI+): m/z 864.2 [M+H]+, 1H NMR (400 MHz, DMSO-d6) δ 11.22-10.81 (m, 1H), 10.36 (s, 1H), 9.20 (s, 1H), 8.95 (br s, 1H), 7.90 (br s, 2H), 7.83-7.72 (m, 2H), 7.70-7.59 (m, 1H), 7.48 (d, J=8.0 Hz, 1H), 7.42-7.29 (m, 3H), 7.24-7.08 (m, 2H), 4.40 (br s, 1H), 4.02 (br s, 2H), 3.85 (s, 3H), 3.88-3.82 (m, 1H), 3.62-3.57 (m, 4H), 3.27 (br s, 5H), 2.74-2.60 (m, 5H), 2.33 (br s, 1H), 1.95 (br s, 1H), 1.76 (br s, 2H), 1.25-1.03 (m, 3H).
Compound 59 to 60 were synthesized in an analogous manner of compound 58 with various intermediates.
1H NMR (400 MHz, DMSO-d6) δ
A mixture of intermediate 61-1 (304 mg, 1.42 mmol), intermediate B 19 (500 mg, 1.09 mmol) and Cs2CO3 (1.07 g, 3.27 mmol) in dioxane (5 mL) and toluene (5 mL) was added Pd-PEPPSI-IPentCl (53.1 mg, 54.6 μmol), and purged with N2 for 3 times, then stirred at 100° C. for 16 hrs. The reaction mixture was diluted with CH2Cl2, filtered and concentrated by rotary distillation, then purified by column chromatography (Petroleum ether/Ethyl acetate=1/0 to 0/1) to give intermediate 61-2 (330 mg, 39.4% yield) was obtained as colorless oil.
MS (ESI+): m/z 592.2 [M+H]+.
Step 2: Synthesis of Intermediate 61-3 To a mixture of intermediate 61-2 (390 mg, 659 μmol) in toluene (0.94 mL) was added MsOH (1.27 g, 13.2 mmol, 938 μL) and degassed and purged with N2 for 3 times, and then the mixture was stirred at 120° C. for 2 hrs and stirred at 0° C. for 15 min. Then the reaction was added TEA (2.67 g, 26.4 mmol, 3.67 mL) and (Boc)20 (1.44 g, 6.59 mmol) at 0° C. and stirred at 0° C. for 15 min. The reaction mixture was diluted with DCM (30 mL) and water (50 mL). Then the organic phase was separated, and the water phase was extracted with DCM (30 mL×3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (DCM/MeOH=1/0 to 20/1) to give intermediate 61-3 (267 mg, 53.3% yield) as colorless oil.
MS (ESI+): m/z 472.3 [M+H]+.
To a solution of intermediate 61-3 (267 mg, 566 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 2.33 mL). The mixture was stirred at 30° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give the intermediate 61-4 (239 mg, HCl salt) as a yellow solid, which was used into the next step directly.
To a mixture of intermediate 55-10 (30 mg, 58.4 μmol) and intermediate 61-4 (23.8 mg, 58.4 μmol, HCl) in DCM (3 mL) was added TEA (17.7 mg, 175 μmol, 24 μL) and stirred at 25° C. for 30 min, then added AcOH (10.5 mg, 175 μmol, 10.0 μL) and stirred at 25° C. for 30 min. The reaction was added NaBH(OAc)3 (37.1 mg, 175 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was concentrated by rotary distillation and purified by prep-HPLC (acetonitrile/water (0.1% TFA condition)) to give the residue. The residue was further purified by prep-HPLC (acetonitrile/water (2% FA condition)) to give compound 61 (3 mg, 5.5% yield) as a yellow solid.
MS (ESI+): m/z 869.6 [M+H]+
1H NMR (400 MHz, DMSO-d6) δ 11.25-10.93 (m, 1H), 10.25-9.93 (m, 1H), 8.81 (s, 1H), 8.36 (s, 1.07H), 8.04 (t, J=8.0 Hz, 1H), 7.75 (d, J=6.8 Hz, 1H), 7.63-7.40 (m, 3H), 7.02-6.79 (m, 5H), 5.76-5.60 (m, 1H), 5.45-5.24 (m, 2H), 4.99 (d, J=10.0 Hz, 1H), 4.88-4.80 (m, 1H), 4.75-4.60 (m, 2H), 3.63 (s, 5H), 3.16-3.12 (m, 2H), 2.89-2.85 (m, 1H), 2.74-2.70 (m, 3H), 2.64-2.60 (m, 4H), 2.36-2.31 (m, 2H), 2.25 (s, 3H), 2.04-1.94 (m, 1H), 1.88-1.74 (m, 4H), 1.71-1.53 (m, 3H), 1.46 (s, 6H), 1.31-1.16 (m, 2H).
The compound 90, 98 and 99 were synthesized according to the procedures described for compound 61 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 62-1 (500 mg, 1.82 mmol) and 1,3-dibromopropane (1.84 g, 9.12 mmol) in DMF (2 mL) was added K2CO3 (252 mg, 1.82 mmol). The mixture was stirred at 50° C. for 16 hrs. Then water (20 mL) and EA (20 mL) was added to the mixture. The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=3/1 to 2/1) to give intermediate 62-3 (480 mg, 26.7% yield) as a colourless oil. MS (ESI+): m/z 395.20 [M+H]+.
To a solution of intermediate 62-3 (75.56 mg, 191 μmol) and intermediate B 7-B (100 mg, 191 μmol) in DMF (0.5 mL) was added K2CO3 (252 mg, 1.82 mmol). The mixture was stirred at 60° C. for 16 hrs. The reaction mixture was added water (20 mL) and EtOAc (20 mL), then the organic layer separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by prep-HPLC (TFA condition) and lyophilized to give a yellow solid. The yellow solid was diluted with DCM (5 mL) and washed with NaHCO3 (3 mL). The organic layer was dried over anhedrous Na2SO4, filtered and concentrated, then purified by chromatography on silica gel (DCM: MeOH=10: 1, 4 g silica gel column) to give compound 62 (12 mg, 14.83 μmol, 7.76% yield) as a yellow solid.
MS (ESI+): m/z 395.20 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.13 (s, 1H), 10.16 (s, 1H), 8.83 (s, 1H), 8.05 (s, 1H), 7.85 (d, J=8.4 Hz, 1H), 7.75 (d, J=8.0 Hz, 1H), 7.61 (d, J=8.0 Hz, 3H), 7.46 (d, J=2.0 Hz, 1H), 7.38 (d, J=8.4 Hz, 1H), 6.93 (d, J=8.0 Hz, 2H), 5.73-5.61 (m, 1H), 5.35 (s, 1H), 5.13 (dd, J=5.6, 13.2 Hz, H), 4.99 (d, J=10.0 Hz, 1H), 4.82 (d, J 17.2 Hz, 1H), 4.68 (d, J 4.4 Hz, 2H), 4.26 (t, J=6.0 Hz, 2H), 3.39 (s, 1H), 3.30-3.29 (m, 1H), 3.12 (s, 4H), 2.97-2.82 (m, 1H), 2.64-2.54 (in, H), 2.15-1.88 (m, 4H), 1.46 (s, 6H).
Compound 63 is similar to compound 62, and is obtained by synthesizing different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 64-1 (82.0 mg, 132.28 μmol) and intermediate 64-2 (38.0 mg, 132.28 μmol) in DCM (10 mL) was added TEA (53.5 mg, 529.12 μmol) and stirred at 25° C. for 30 min, then AcOH (23.8 mg, 396.84 μmol) was added and stirred at 25° C. for 30 min. The reaction mixture was added NaBH3CN (84.1 mg, 397 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was added water (20 mL) and EtOAc (20 mL), then the organic layer was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by prep-HPLC (FA condition) to give compound 64 (3 mg, 3.51 μmol, 2.65% yield) as a yellow solid.
MS (ESI+): m/z 877.6 [M+Na]+.
1H NMR (400 MHz, DMSO-d6) δ 11.11 (s, 1H), 10.38-9.93 (m, 1H), 8.84 (s, 1H), 8.13-7.99 (m, 1H), 7.76 (d, J=4.0 Hz, 1H), 7.61 (d, J=8.0 Hz, 3H), 7.31-6.86 (m, 5H), 5.78-5.59 (m, 1H), 5.46-5.28 (m, 2H), 5.00 (d, J=8.0 Hz, 1H), 4.83 (d, J=16.0 Hz, 1H), 4.68 (d, J=4.0 Hz, 2H), 3.72-3.62 (m, 4H), 3.31-3.29 (m, 3H), 3.09 (br s, 4H), 2.94-2.81 (m, 2H), 2.76-2.69 (m, 1H), 2.57-2.53 (m, 6H), 2.45-2.36 (m, 2H), 2.08-1.93 (m, 4H), 1.71 (br s, 2H), 1.47 (s, 6H).
A mixture of intermediate 65-2 (325 mg, 1.42 mmol), intermediate 19 (500 mg, 1.09 mmol) and Cs2CO3 (1.07 g, 3.27 mmol) in dioxane (8 mL) and toluene (8 mL) was added Pd-PEPPSI-IPentCl (53.1 mg, 54.6 μmol), then degassed and purged with N2 for 3 times, and then stirred at 100° C. for 16 hrs. The reaction was diluted with DCM, filtered and dried by rotary distillation to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=1/0 to 3/1) to give intermediate 65-3 (95 mg, 12.9% yield) as colorless oil.
1H NMR (400 MHz, DMSO-d6) δ 7.27-7.15 (m, 2H), 6.95-6.88 (m, 2H), 6.87-6.83 (m, 2H), 6.81-6.70 (m, 1H), 5.58-5.42 (m, 1H), 4.92-4.51 (m, 2H), 3.72 (s, 3H), 3.61 (s, 3H), 3.55-3.43 (m, 2H), 3.21-2.98 (m, 3H), 2.88-2.63 (m, 4H), 2.10-2.01 (m, 1H), 1.86-1.70 (m, 2H), 1.63-1.50 (m, 1H), 1.46-1.31 (m, 2H), 0.88 (s, 9H), 0.08-0.04 (m, 6H).
To a mixture of intermediate 65-3 (110 mg, 181 μmol) in toluene (2 mL) was added MsOH (1.35 g, 14.1 mmol, 1 mL) and degassed and purged with N2 for 3 times, and then stirred at 120° C. for 1.5 hrs. The reaction mixture was added additional TEA (3 mL) at 0° C. and purified by prep-HPLC (neutral condition) to give the residue. The residue was added DCM (10 mL) and adjusted to pH 8 with aq. sat. NaHCO3. Then the mixture was added H2O (5 mL), the organic phase was separated. The aqueous phase was extracted with DCM (10 mL×3). The combined organic phase dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give intermediate 65-4 (31 mg, 38.6% yield) as colorless oil.
MS (ESI+): m/z 373.2 [M+H]+.
To a solution of intermediate 65-4 (31 mg, 83.2 μmol) in DCM (2 mL) and DMSO (0.1 mL) was added DMP (70.6 mg, 166 μmol). The mixture was stirred at 25° C. for 30 min. The reaction mixture was quenched by DCM, then quenched with Na2S2O3 (5 mL) and NaHCO3 (5 mL), and extracted with DCM (15 mL×3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the intermediate 65-5 (30 mg, crude) as colorless oil, which was used into the next step directly.
To a mixture of intermediate 65-5 (30 mg, 81.0 μmol) and intermediate B 7-B (42.4 mg, 81.0 μmol, HCl) in DCM (3 mL) was added TEA (24.6 mg, 243 μmol, 33.8 μL) and stirred at 25° C. for 30 min, then added AcOH (14.6 mg, 243 μmol) and stirred at 25° C. for 30 min. The reaction was added NaBH(OAc)3 (51.5 mg, 243 μmol) and stirred at 25° C. for 16 hours. The reaction mixture was concentrated by rotary distillation to give the residue. The residue was purified by HPLC (acetonitrile/water (2% FA)). Then the residue was added MeCN (3 mL) to triturate it to give intermediate 65 (16 mg, 22.9% yield) as a yellow solid.
MS (ESI+): m/z 841.6 [M+H]+
1H NMR (400 MHz, DMSO-d6) δ 11.09 (s, 1H), 10.29-10.06 (m, 1H), 8.82 (s, 1H), 8.15-7.98 (m, 1H), 7.82-7.69 (m, 1H), 7.68-7.44 (m, 3H), 7.05-6.79 (m, 5H), 5.74-5.58 (m, 1H), 5.40-5.27 (m, 2H), 5.05-4.94 (m, 1H), 4.89-4.78 (m, 1H), 4.75-4.61 (m, 2H), 3.63 (s, 3H), 3.11 (br s, 5H), 2.95-2.83 (m, 1H), 2.76-2.62 (m, 4H), 2.59 (br s, 1H), 2.56-2.52 (m, 4H), 2.31-2.19 (m, 2H), 2.07-1.93 (m, 1H), 1.90-1.79 (m, 2H), 1.77-1.62 (m, 1H), 1.46 (s, 6H), 1.40-1.28 (m, 2H).
The compound 66 and compound 97 were synthesized according to the procedures described for compound 65 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 67-1 (3 g, 18.9 mmol) and intermediate 67-2 (5.27 g, 28.3 mmol) in NMP (10 mL) was stirred at 60° C. for 3 hrs. The reaction mixture was partitioned between EtOAc (100 mL) and water (50 mL). The combined organic layers were washed with saturated saturated salt solution, dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give intermediate 67-3 (6.01 g, crude) as a white solid. The crude product was used into the next step directly.
MS (ESI+): m/z 269.8 [M+H−56]+.
To a solution of intermediate 67-3 (6.01 g, 18.5 mmol) in THE (60 mL) was added Pd/C (2 g, 10% purity) then degassed and purged with H2 for 3 times, and then the mixture was stirred at 25° C. for 16 hr under H2 atmosphere. The reaction mixture was filtered and concentrated by rotary distillation to give intermediate 67-4 (5.66 g, crude) as a white solid. The crude product was used into the next step directly.
MS (ESI+): m/z 240.0 [M−56+H]+.
To a solution of intermediate 67-4 (1 g, 3.39 mmol) and intermediate 67-5 (1.25 g, 3.39 mmol) in 1,4-dioxane (20 mL) was added Cs2CO3 (3.31 g, 10.2 mmol) and Xphos Pd G3(141.6 mg, 169 μmol) and then the mixture was stirred at 100° C. for 16 hrs. The reaction mixture was partitioned between EtOAc (100 mL) and water (100 mL). The organic phase was washed with saturated saturated salt solution (30 mL×2), dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (PE/EA=20/1 to 3/1) to give intermediate 67-6 (908 mg, 94% yield) as a white solid.
MS (ESI+): m/z 528.9 [M+H−56]+.
To a solution of intermediate 67-6 (908 mg, 1.55 mmol) in THE (10 mL) and EtOH (10 mL) was added Pd/C (800 mg, 10% purity) and then the mixture was stirred at 25° C. for 16 hrs under H2 atmosphere. The reaction mixture was filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (PE/EA=5/1 to 1/1) to give intermediate 67-7 (324 mg, 51.3% yield) as a white solid.
MS (ESI+): m/z 350.9 [M−56]+.
Step 5: Synthesis of Intermediate 67-8 To a solution of intermediate 67-7 (324 mg, 797 μmol) in DCM (5 mL) was added HCl/dioxane (4 M, 5 mL). The mixture was stirred at 25° C. for 1 h. The reaction mixture was concentrated by rotary distillation to give intermediate 67-8 (242 mg, crude) as a white solid.
To a mixture of intermediate 67-8 (50.0 mg, 146 μmol), intermediate 67-9 (48.2 mg, 219 μmol) in DCM (2 mL) was added Et3N (44.3 mg, 438 μmol, 60.9 μL) and the reaction mixture was stirred at 20° C. for 0.5 h. Then the mixture was added AcOH (44.28 mg, 438 μmol, 25.0 μL) and stirred at 20° C. for 0.5 h. Then the mixture was added the NaBH3CN (18.3 mg, 292 μmol) and stirred at 20° C. for 16 hrs. The reaction mixture was partitioned between DCM (50 mL) and water (50 mL). The organic phase was dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (Dichloromethane/methanol=1: 0 to 10: 1) to give intermediate 67-10 (56 mg, 61.7% yield) as a yellow solid.
MS (ESI+): m/z 510.5 [M].
Step 7: Synthesis of Intermediate 67-11 To a solution of intermediate 67-10 (56 mg, 110 μmol) in THF (1 mL) and saturated NH4Cl (0.3 mL) was added iron (61 mg, 1.10 mmol), and the mixture was stirred at 80° C. for 1 h. The reaction mixture was filtered and concentrated by rotary distillation to give intermediate 67-11 (60.0 mg, crude) as a gray solid.
To a solution of intermediate 6 (60 mg, 125 μmol) in toluene (2 mL) was added m-CPBA (27.9 mg, 137 μmol). The mixture was stirred at 25° C. for 0.5 h. Then the mixture was added DIEA (32.3 mg, 250 μmol, 43.5 μL) and intermediate 67-11 (44.6 mg, 125 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was quenched by addition aqueous Na2S2O3 and NaHCO3 (1:1, 50 mL) at 20° C., and then extracted with EtOAc (50 mL). The aeparated combined organic layers were washed with saturated saturated salt solution, dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by HPLC (acetonitrile/water (2% FA)) to give compound 67 (1.00 mg, 1.01% yield) as a yellow solid.
MS (ESI+): m/z 790.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 10.23-10.11 (m, 1H), 8.83 (s, 1H), 8.38 (s, 1.897H), 8.06 (br s, 1H), 7.75 (d, J=8.4 Hz, 1H), 7.68-7.49 (m, 3H), 6.95 (d, J=8.6 Hz, 2H), 6.83 (t, J=9.6 Hz, 1H), 6.52 (d, J=16.0 Hz, 1H), 6.43 (d, J=8.4 Hz, 1H), 5.82 (d, J=7.6 Hz, 1H), 5.73-5.59 (m, 1H), 5.37 (s, 1H), 5.00 (d, J=11.2 Hz, 1H), 4.86-4.77 (m, J=16.8 Hz, 1H), 4.73-4.61 (m, J=5.2 Hz, 2H), 4.29-4.21 (m, 1H), 3.70 (d, J=11.2 Hz, 2H), 2.86 (br s, 4H), 2.64 (br s, 4H), 2.59 (d, J=4.4 Hz, 2H), 2.12-2.04 (m, 2H), 1.94-1.83 (m, 4H), 1.55 (d, J=8.4 Hz, 2H), 1.47 (s, 6H).
The compound 68 to compound 70, and compound 123, 154 and 155 were synthesized according to the procedures described for compound 67 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate67-8 (100 mg, 292 μmol) and intermediate 55-10 (160 mg, 292 μmol) in DCM (2 mL) was added Et3N (88.6 mg, 875 μmol, 122 μL), the mixture was stirred at 25° C. for 0.5 h. Then the mixture was added AcOH (52.6 mg, 875 μmol, 50 μL) and stirred at 25° C. for 0.5 h. Then the mixture was added NaBH(OAc)3 (124 mg, 583 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was partitioned between DCM (10 mL) and H2O (10 mL). The organic phase was dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by HPLC (acetonitrile/water (2% FA)) to give compound 71 (8.98 mg, 3.64% yield) as a yellow solid.
MS (ESI+): m/z 804.5 [M+H]+
1H NMR (400 MHz, DMSO-d6) δ5 10.78 (s, 1H), 10.13 (br s, 1H), 8.81 (s, 1H), 8.32 (s, 0.337 H), 8.04 (br s, 1H), 7.74 (d, J=7.2 Hz, 1H), 7.64-7.47 (m, 3H), 6.97-6.89 (b, 2H), 6.87-6.77 (m, 1H), 6.56-6.48 (m, 1H), 6.46-6.38 (m, 1H), 5.80 (d, J=7.6 Hz, 1H), 5.73-5.58 (m, 1H), 5.35 (s, 1H), 4.99 (d, J 10.0 Hz, 1H), 4.82 (d, J=18.0 Hz, 1H), 4.67 (d, J=5.2 Hz, 2H), 4.30-4.20 (m, 1H), 3.66-3.61 (m, 2H), 2.86 (br s, 4H), 2.76-2.67 (3, 1H), 2.65-2.57 (m, 2H), 2.48-2.46 (m, 5H), 2.21 (d, J=6.8 Hz, 2H), 2.12-2.00 (m, 1H), 1.89-1.76 (2, 3H), 1.66 (br s, 1H), 1.46 (s, 6H), 1.30-1.24 (i5, 2H).
The compound 72, 91, 92, 94, 140 and 147 were synthesized according to the procedures described for compound 71 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 73-2 (3 g, 18.84 mmol,) and intermediate 73-1 (4.50 g, 28.26 mmol) in DMSO (10 mL). The mixture was stirred at 90° C. for 16 hrs. The reaction mixture was partitioned between EA (20 mL) and water (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (DCM: MeOH=10:1). Intermediate 73-3 (6.93 g, crude) was obtained as a white solid, which is used for the next step directly.
MS (ESI+): m/z 298.31 [M+H]+.
To a solution of intermediate 73-3 (6.39 g, 21.4 mmol) in THE (30 mL) was added Pd/C (4 g, 10% purity). The mixture was stirred at 20° C. for 16 hrs under H2 atomosphere. The reaction mixture was filtered and concentrated under reduced pressure to give intermediate 73-4 (7.42 g, crude) as a red solid which was used to the next step directly.
MS (ESI+): m/z 268.33 [M+H]+.
A mixture of intermediate 73-4 (2.63 g, 9.80 mmol) and intermediate 73-5 (3.63 g, 9.80 mmol) in 1,4-dioxane (30 mL) was added Cs2CO3 (9.58 g, 29.41 mmol) and [2-(2-aminophenyl)phenyl]-methylsulfonyloxy-palladium-dicyclohexyl-[2-(2,6-diisopropoxyphenyl)phenyl]phosphane (410 mg, 490 μmol) and then the mixture was stirred at 100° C. for 16 hrs under N2 atmosphere. The reaction mixture was partitioned between EA (20 mL) and water (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=5/1 to 3/1). Intermediate 73-6 (3.4 g, 62.2% yield) was obtained as a red solid.
MS (ESI+): m/z 557.65 [M+H]+.
To a solution of intermediate 73-6 (3.39 g, 6.08 mmol) in THE (20 mL) was added Pd/C (2 g, 10% purity) and purged with H2 for 3 times, and then the mixture was stirred at 20° C. for 16 hrs under H2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=3/1 to 0/1) to give intermediate 73-7 (699 mg, 30.00% yield) as a pale green solid.
MS (ESI+): m/z 379.42 [M+H]+.
To a solution of intermediate 73-7 (293 mg, 772 μmol) in DCM (5 mL) was added HCl/dioxane (5 mL). The mixture was stirred at 20° C. for 3 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give intermediate 73-8 (350 mg, crude) as a yellow solid, which was used for the next step directly.
MS (ESI+): m/z 333.365 [M+H]+.
To a solution of intermediate 73-8 (80.0 mg, 216 μmol, HCl) and intermediate B 8 (113 mg, 216 μmol, HCl salt) in DCM (2 mL) was added TEA (87.6 mg, 865 μmol) and the mixture was stirred at 25° C. for 30 min, then AcOH (38.97 mg, 648.97 μmol,) was added and pH was maintain between 5 to 6. The reaction mixture was stirred at 25° C. for 30 min, then was added NaBH(OAc)3 (138 mg, 649 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give the residue. The residue was purified by prep-HPLC (FA condition) to give compound 73 (81 mg, 46.6% yield) as a white solid.
MS (ESI+): m/z 803.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H), 10.27 (br s, 1H), 8.87 (s, 1H), 8.20 (s, 1.07H), 8.07 (t, J=7.6 Hz, 1H), 7.86-7.52 (m, 4H), 7.21 (d, J=8.4 Hz, 2H), 6.83 (t, J=9.6 Hz, 1H), 6.50 (d, J=14.8 Hz, 1H), 6.41 (d, J=8.4 Hz, 1H), 5.79 (d, J=7.6 Hz, 1H), 5.73-5.64 (m, 1H), 5.35 (br s, 1H), 4.99 (d, J 10.4 Hz, 1H), 4.82 (d, J=17.2 Hz, 1H), 4.68 (d, J=5.6 Hz, 2H), 4.32-4.17 (m, 1H), 3.12 (d, J=11.2 Hz, 2H), 2.96 (d, J=10.4 Hz, 2H), 2.81-2.63 (m, 2H), 2.54 (br s, 2H), 2.46-2.41 (m, 1H), 2.21 (d, J=6.8 Hz, 2H), 2.08 (s, 1H), 1.98 (t, J=10.8 Hz, 2H), 1.90-1.81 (m, 1H), 1.80-1.69 (m, 4H), 1.62 (d, J=9.2 Hz, 2H), 1.46 (s, 6H), 1.32-1.16 ((2, 2H).
The compound 74, compound 83 to compound 88, and compound 109, 122, 125 and 158 were synthesized according to the procedures described for compound 73 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 75-1 (300 mg, 1.45 mmol) and intermediate 75-2 (434.74 mg, 2.18 mmol) in DCM (5 mL) was added Et3N (441.57 mg, 4.36 mmol) and stirred at 25° C. for 30 min. Then AcOH (349.41 mg, 5.82 mmol) was added to the mixture. The reaction mixture was stirred at 25° C. for 30 min and then NaBH3CN (182.82 mg, 2.91 mmol) was added. The mixture was stirred at 25° C. for 16 hrs. Then the reaction mixture was partitioned between EA (20 mL) and water (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (Petroleum ether/Ethyl acetate=5/1 to 3/1). Intermediate 75-3 (675 mg, crude) was obtained as a yellow oil and was used into the next step directly.
MS (ESI+): m/z 389.49 [M+H]+.
To a solution of intermediate 75-3 (675 mg, 1.73 mmol) in DCM (5 mL) was added HCl/dioxane (5 mL). The mixture was stirred at 20° C. for 2 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give intermediate 75-4 (861 mg, crude) as a white solid, which is used for the next step directly. MS (ESI+): m/z 289.37 [M+H]+.
To a solution of intermediate 75-4 (861 mg, 2.98 mmol) in ethyl 2,2,2-trifluoroacetate (5 mL) was added Et3N (903.24 mg, 8.93 mmol). The mixture was stirred at 80° C. for 16 hrs. The reaction mixture was partitioned between EA (20 mL) and water (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (DCM: MeOH=20: 1). Intermediate 75-5 (358 mg, crude) was obtained as a yellow solid and used for the next step.
MS (ESI+): m/z 385.38 [M+H]+.
To a solution of intermediate 75-5 (358.3 mg, 929.73 μmol) in THE (10 mL) was added Pd/C (300 mg, 5% purity). The mixture was stirred at 20° C. for 16 hrs under H2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give intermediate 75-6 (121 mg, crude) as black oil, which is used for the next step directly.
MS (ESI+): m/z 355.40 [M+H]+.
To a solution of intermediate 75-6 (68.66 mg, 179.06 μmol) in toluene (1 mL) was added m-CPBA (123.60 mg, 286 μmol) and the mixture was stirred at 20° C. for 30 min, then DIEA (69.43 mg, 537.17 μmol) and intermediate B 15-B (70 mg, 196.96 μmol) was added to the mixture and the obtained mixture was stirred at 20° C. for 16 hrs. The reaction mixture was quenched by addition Na2S2O3 (10 mL) and NaHCO3 (10 mL) at 0° C., partitioned between EA (30 mL) and water (30 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (DCM: MeOH=10:1). Intermediate 75-8 (149 mg, crude) was obtained as a yellow solid, which is used for next step directly.
MS (ESI+): m/z 690.76 [M+H]+.
To a solution of intermediate 75-8 (149 mg, 216 μmol) in MeOH (5 mL) was added K2CO3 (89.44 mg, 647.12 μmol). The mixture was stirred at 25° C. for 2 hrs. The reaction mixture was partitioned between EA (20 mL) and water (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (DCM: MeOH=10: 1). Compound 75-9 (120 mg, crude) was filtered and concentrated in vacuo to obtained as a yellow solid, which is used for next step directly.
MS (ESI+): m/z 594.75 [M+H]+.
To a solution of intermediate 1-1 (83.6 mg, 303 μmol) and intermediate 75-9 (120 mg, 202 μmol) in NMP (5 mL) was added DIEA (78.2 mg, 605 μmol). The mixture was stirred at 100° C. for 16 hrs. The reaction mixture was partitioned between EA (10 mL) and water (10 mL). The organic phase was separated, dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to give the residue. The residue was purified by HPLC (acetonitrile/water (0.1% TFA condition)) to give a yellow solid. The yellow solid was diluted with DCM (5 mL) and washed with NaHCO3 (3 mL). The organic layer was dried over anhydrous Na2SO4, filtered and concentrated, and then purified by combi flash (DCM: MeOH=10: 1, 4 g silica gel column) to give compound 75 (19 mg, 10.5% yield) as a yellow solid.
MS (ESI+): m/z 851.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) 11.10 (s, 1H), 10.43-10.10 (m, 1H), 8.87 (s, 1H), 7.94 (d, J=7.6 Hz, 1H), 7.75-7.60 (m, 4H), 7.39-7.31 (m, 1H), 7.27 (d, J=7.2 Hz, 1H), 7.20 (d, J=8.4 Hz, 2H), 5.81-5.56 (m, 1H), 5.11-4.95 (m, 3H), 4.85 (d, J=18.4 Hz, 1H), 4.76 (d, J=12.8 Hz, 1H), 4.63-4.50 (m, 1H), 4.12 (d, J=11.6 Hz, 2H), 3.06-2.93 (m, 5H), 2.90-2.72 (m, 3H), 2.59 (d, J=16.0 Hz, 2H), 2.32-2.15 (m, 3H), 2.08-1.97 (m, 2H), 1.96-1.83 (m, 3H), 1.83-1.44 (m, 8H), 0.87 (t, J=7.6 Hz, 3H).
To a solution of intermediate 76-1 (1 g, 2.97 mmol), and intermediate 76-2 (1.24 g, 2.97 mmol) in 1,4-dioxane (10 mL) and H2O (1 mL) was added Pd(PPh3)4 (343 mg, 297 μmol) and Na2CO3 (944 mg, 8.90 mmol), then degassed and purged with N2 for 3 times, and then the mixture was stirred at 100° C. for 16 hrs under N2 atmosphere. The reaction mixture was partitioned between EA (50 mL) and H2O (50 mL). The organic phase was washed with saturated saturated salt solution, dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (PE/EA=1/0 to 5/1) to give intermediate 76-3 (946 mg, 52.9% yield) as a yellow solid.
MS (ESI+): m/z 502.0 [M+H]+.
To a solution of intermediate 76-3 (300 mg, 600 μmol) and 4-(dimethoxymethyl)piperidine (191 mg, 1.20 mmol) in toluene (5 mL) was added Cs2CO3 (586 mg, 1.80 mmol) and RuPhos Pd G3 (50.1 mg, 60.0 μmol), then degassed and purged with N2 for 3 times, and then the mixture was stirred at 100° C. for 16 hrs under N2 atmosphere. The reaction mixture was concentrated by rotary distillation, and diluted with EA (40 mL) and extracted with water (50 mL). The organic layers were dried over anhydrous Na2SO4, concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (PE/EA=5/1 to 1/1) to give intermediate76-4 (201 mg, 55.0% yield) as a white solid.
MS (ESI+): m/z 579.3 [M+H]+.
To a solution of intermediate 76-4 (201 mg, 347 μmol) in THE (2 mL) and EtOH (2 mL) was added Pd/C (200 mg, 10% purity), then degassed and purged with H2 for 3 times, and then the mixture was stirred at 20° C. for 16 hrs under H2 atmosphere. The reaction mixture was filtered and concentrated by rotary distillation to give the residue. The residue was purified by column chromatography (PE/EA=5/1 to 1/1) to give intermediate 76-5 (82 mg, 58.9% yield) as a white solid.
MS (ESI+): m/z 401.3 [M+H]+.
To a solution of intermediate 76-5 (82 mg, 205 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 2 mL) and the mixture was stirred at 25° C. for 16 hrs. The reaction mixture was filtered and concentrated by rotary distillation to give intermediate 76-6 (113 mg, crude) as a white solid, which is used for next step directly.
To the solution of intermediate 76-6 (50 mg, 128 μmol) and intermediate B 7-B (80.3 mg, 154 μmol) in DCM (2 mL) was added Et3N (25.9 mg, 256 μmol, 36 μL), and the obtained solution was stirred at 25° C. for 0.5 h, then AcOH (15.4 mg, 256 μmol, 15 μL) was added and stirred another 0.5 h at 25° C. Then NaBH(OAc)3 (81.3 mg, 384 μmol) was added, and the mixture was stirred at 25° C. for 16 hrs. The mixture was concentrated by rotaty distillation and the residue was purified by HPLC (acetonitrile/water (2% FA)) to obtain compound 76 as a yellow solid (45 mg, 41.4%).
MS (ESI+): m/z 825.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.85 (s, 1H), 10.26-10.00 (m, 1H), 8.82 (s, 1H), 8.27 (s, 0.336 H), 8.04 (br s, 1H), 7.75 (d, J=8.4 Hz, 1H), 7.60 (d, J=7.6 Hz, 3H), 7.47 (d, J=8.8 Hz, 1H), 6.99-6.87 (m, 3H), 6.83 (s, 1H), 5.72-5.58 (m, 1H), 5.33 (s, 1H), 4.99 (d, J=10.4 Hz, 1H), 4.82 (d, J=18.4 Hz, 1H), 4.72-4.63 (m, 2H), 4.29-4.22 (m, 1H), 3.88 (s, 3H), 3.84-3.73 (m, 2H), 3.11 (br s, 4H), 2.81-2.69 (m, 2H), 2.64-2.58 (m, 1H), 2.55-2.52 (m, 4H), 2.42 (br s, 1H), 2.31-2.20 (m, 3H), 2.19-2.11 (m, 1H), 1.83 (d, J=12.4 Hz, 3H), 1.46 (s, 6H), 1.34-1.20 (m, 2H).
The compound 111, 117 and compound 144 were synthesized according to the procedures described for compound 76 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 77-1 (500 mg, 2.27 mmol) and intermediate 77-2 (443 mg, 2.27 mmol) in MeOH (10 mL) was added AcOH (130 μL, 2.27 mmol). The mixture was stirred at 25° C. for 2 hrs. Then NaBH3CN (214 mg, 3.41 mmol) was added and the mixture was stirred at 25° C. for 2 hrs. The mixture was added saturated NaHCO3 (20 mL) and extracted with ethyl acetate (40 mL×3). The organic phase was collected and dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the resiue. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1:0 to 65:35) to give intermediate77-3 (280 mg, 31.6% yield) as yellow solid.
MS (ESI+): m/z 391.14 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 8.03 (d, J=9.6 Hz, 2H), 7.01 (d, J=9.6 Hz, 2H), 4.15-3.99 (m, 2H), 3.30-3.24 (m, 4H), 3.02-2.90 (m, 2H), 2.57-2.52 (m, 1H), 2.45-2.38 (m, 4H), 1.88-1.78 (m, 2H), 1.49-1.35 (m, 11H).
To a mixture of intermediate 77-3 (500 mg, 1.28 mmol) in MeOH (10 mL) was added wet Pd/C (50 mg, 10% purity). The mixture was stirred under H2 (15 psi) at 25° C. for 1 h. The reaction mixture was filtered and concentrated by rotary distillation to give intermediate77-4 (480 mg, crude) as a brown solid.
To a solution of intermediate B 6 (262 mg, 733 μmol) in toluene (10 mL) was added m-CPBA (298 mg, 1.47 mmol, 85% purity). The mixture was stirred at 25° C. for 1 h. Intermediate 77-4 (317 mg, 879 μmol) and DIEA (379 mg, 2.93 mmol) was added to the above mixture. Then the mixture was stirred at 25° C. for 12 hrs. The reaction mixture was quenched with saturated NaHCO3 (5 mL) and saturated Na2S2O3 (5 mL), and extracted with EA (20 mL×2). The organic layers were collected and dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the crude product. The crude product was purified by flash silica gel chromatography (dichloromethane: methanol=1:0 to 10:1) to give intermediate77-5 (480 mg, 98% yield) as a yellow soild.
MS (ESI+): m/z 669.9 [M+H]+.
A solution of intermediate 77-5 (480 mg, 717 μmol) in DCM (4 mL) was added HCl/dioxane (4 M, 10 mL). The reaction mixture was stirred at 25° C. for 2 hrs. The reaction mixture was concentrated by rotary distillation to give intermediate 77-6 (408 mg, 93.9% yield, HCl salt) as a white solid.
MS (ESI+): m/z 569.6 [M+H]+.
To a solution of intermediate77-6 (200 mg, 352 μmol) in DMF (4 mL) was added intermediate 1-1 (97.0 mg, 351 μmol) and DIEA (183 μL, 1.05 mmol). The mixture was stirred at 100° C. for 2 hrs under microwave. The mixture was diluted with DCM (20 mL), washed with H2O (28 mL) and the combined organic layers were collected and dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by prep-HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 30% B to 70% B) to give compound 77 (25.36 mg, 8.66% yield) as a yellow solid.
MS (ESI+): m/z 826.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.10 (s, 1H), 10.27-10.10 (m, 1H), 8.83 (s, 1H), 8.14-8.00 (m, 1H), 7.80-7.74 (m, 1H), 7.70-7.67 (m, 1H), 7.64-7.51 (m, 3H), 7.35 (s, 1H), 7.31-7.24 (m, 1H), 7.01-6.90 (m, 2H), 5.75-5.59 (m, 1H), 5.35 (s, 1H), 5.08 (dd, J=5.2, 12.8 Hz, 1H), 5.01-4.97 (m, 1H), 4.88-4.77 (m, 1H), 4.71-4.62 (m, 2H), 3.77-3.67 (m, 2H), 3.45-3.43 (m, 4H), 2.95-2.82 (m, 1H), 2.70-2.55 (m, 8H), 2.42-2.36 (m, 1H), 2.06-1.96 (m, 1H), 1.94-1.84 (m, 2H), 1.62-1.52 (m, 2H), 1.46 (s, 6H)
The Compound 78 was synthesized according to the similar procedures described for Compound 77.
1H NMR (400 MHz, DMSO-d6) δ
Pd(dppf)Cl2 (216 mg, 296 μmol) was added to a mixture of intermediate B 18 (1 g, 2.96 mmol), intermediate 79-1 (1.19 g, 8.87 mmol) and Cs2CO3 (1.93 g, 5.91 mmol) in dioxane (5 mL) and H2O (1 mL). The mixture was degassed and purged with N2 for 5 minutes, and then the sealed tube was reacted at 80° C. for 1 h under microwave. The reaction mixture was concentrated by rotary distillation to give thr residue. The residue was purified by flash silica gel chromatography (Petroleum ether: ethyl acetate=1:1 to 0:1). Intermediate 79-2 (750 mg, 72.0% yield) was obtained as a brown solid.
MS (ESI+): m/z 286.07 [M+H]+.
NaIO4 (2.25 g, 10.5 mmol) in H2O (10 mL) was added to a solution of intermediate 79-2 (750 mg, 2.63 mmol) in THF (50 mL). The mixture was stirred at 0° C. for 10 min. K2OsO4 (48.4 mg, 131 μmol) was added to the above mixture and stirred at 25° C. for 50 min. The reaction mixture was quenched with saturated Na2SO3 (50 mL) and then extracted with DCM (50 mL×3). The combined organic layers were dried over anhydrous Na2SO4, filtered and concentrated by rotary distillation to give the residue. The residue was purified by flash silica gel chromatography (Petroleum ether:ethyl acetate=10:0 to 0:1). Intermediate 79-3 (550 mg, 30% yield) was obtained as a brown solid.
MS (ESI+): m/z 288.04 [M+H]+.
AcOH (20.9 mg, 20.0 μL) was added to a mixture of intermediate 79-3 (100 mg, 348 μmol) and 4-piperidylmethanol (60.1 mg, 522 μmol) in DMF (1 mL) and THE (2 mL). The reaction mixture was stirred at 80° C. for 0.5 h. Then NaBH3CN (43.8 mg, 696 μmol) was added to the mixture at 25° C. and stirred at 25° C. for 12 hrs. The reaction mixture was quenched with saturated NaCl (10 mL) and then extracted with DCM (20 mL×3). The aqueous phase was concentrated by rotary distillation to give the residue. The residue was purified by HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 5% B to 35%) to give intermedaite 79-5 (4.69 mg, 16% yield) as a white soild.
MS (ESI+): m/z 387.3 [M+H]+.
DMP (22.0 mg, 51.8 μmol) was added to a solution of intermediate 79-5 (10.0 mg, 25.9 μmol) in DMSO (0.1 mL) and DCM (1 mL) at 0° C. The reaction mixture was stirred at 20° C. for 2 hrs. The reaction mixture was quenched with saturated NaHCO3 (0.5 mL), filtered and concentrated by rotary distillation to give intermediate 79-6 (10 mg, crude), which was used for the next step without further purification.
MS (ESI+): m/z 385.07 [M+H]+.
TEA (5.04 mg, 49.8 μmol) was added to a mixture of intermediate B 8 (13.0 mg, 24.9 μmol, HCl) and intermediate 79-6 (9.57 mg, 24.9 μmol) in DMF (5 mL). The mixture was stirred at 25° C. for 0.5 h. Then AcOH (1.50 mg, 24.9 μmol, 1.42 μL) was added to the above mixture and stirred for another 0.5 h. Finally, NaBH3CN (3.13 mg, 49.8 μmol) was added to the mixture and reacted at 25° C. for 2 hrs. The reaction mixture was quenched with saturated NaHCO3 (0.5 mL), filtered and concentrated by rotary distillation to give the residue. The residue was purified by HPLC (Column: Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition from 10% B to 50% B) to give compound 79 (4.69 mg, 22% yield) as a white solid.
MS (ESI+): m/z 854.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.12 (br s, 1H), 10.27 (br s, 1H), 8.88 (s, 1H), 8.12-7.94 (m, 1H), 7.82-7.72 (m, 1H), 7.70-7.56 (m, 3H), 7.20 (d, J=8.4 Hz, 2H), 7.11 (s, 1H), 7.07-7.01 (m, 1H), 6.99-6.88 (m, 1H), 5.78-5.58 (m, 1H), 5.55-5.16 (m, 2H), 5.03-4.96 (m, 1H), 4.88-4.78 (m, 1H), 4.75-4.62 (m, 2H), 3.48-3.47 (m, 2H), 3.34 (s, 3H), 2.98-2.80 (m, 6H), 2.77-2.70 (m, 1H), 2.65-2.59 (m, 1H), 2.44-2.39 (m, 1H), 2.17-2.09 (m, 2H), 2.05-1.90 (m, 5H), 1.74-1.61 (m, 5H), 1.53-1.49 (m, 1H), 1.46 (s, 6H), 1.18-1.05 (m, 2H).
To a mixture of 80-1 (5 g, 20.8 mmol), 80-2 (7.73 g, 25.0 mmol), Cs2CO3 (20.36 g, 62.49 mmol) in dioxane (30 mL) and water (6 mL) was added Pd(dppf)Cl2(1.52 g, 2.08 mmol) under N2. The result mixture was stirred at 90° C. for 8 hrs. The mixture was dried over with Na2SO4 filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Petroleum ether:ethyl acetate=1:1 to 50:50) to give intermediate 80-3 (6.5 g, 91.1% yield) as a yellow solid.
LCMS (ESI+): m/z 287.8 [M−56]+.
1H NMR (400 MHz, DMSO-d6) δ 6.94 (d, J=8.4 Hz, 1H), 6.86-6.84 (m, 1H), 6.75-6.71 (m, 1H), 5.54-5.43 (m, 3H), 3.88 (br s., 2H), 3.50-3.44 (m, 2H), 2.23-2.16 (m, 2H), 1.41 (s, 9H).
To a mixture of 80-3 (2.00 g, 5.84 mmol) in EtOH (30 mL) was added Pd/C (500 mg, 10%). The suspension was degassed under vacuum and purged with H2 three times. The mixture was stirred under H2 (15 psi) at 25° C. for 20 hours. The reaction mixture was filtered and concentrated under reduced pressure to give the intermediate 80-4 (1.8 g, crude) as a brown solid, and was used into the next step directly.
1H NMR (400 MHz, DMSO-d6) δ 7.17 (d, J=8.4 Hz, 1H), 6.82 (d, J=2.4 Hz, 1H), 6.78-6.73 (m, 1H), 5.34 (s, 2H), 2.82-2.65 (m, 4H), 2.47-2.41 (m, 1H), 1.61-1.45 (m, 4H), 1.41 (s, 9H).
To a solution of 80-4 (1 g, 2.90 mmol) and 80-5 (1.12 g, 5.81 mmol) in DMF (10 mL) was added NaHCO3 (731.83 mg, 8.71 mmol). The mixture was stirred at 80° C. for 8 hrs. The reaction mixture was added water 20 mL at 25° C., and then extracted with EtOAc (30 mL×2). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography to give intermediate 80-6 (400 mg, 30.2% yield) as a green oil.
LCMS (ESI+): m/z 358.3 [M−100+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 7.27 (d, J=8.4 Hz, 1H), 6.96-6.83 (m, 2H), 6.24 (d, J=8.0 Hz, 1H), 4.45-4.36 (m, 1H), 4.15-4.00 (m, 2H), 2.97-2.90 (m, 2H), 2.84-2.77 (m, 2H), 2.59-2.55 (m, 1H), 2.13-2.03 (m, 1H), 1.96-1.87 (m, 1H), 1.61-1.48 (m, 4H), 1.41 (s, 9H).
To a solution of 80-6 (400 mg, 878.21 μmol) in DCM (2 mL) was added HCl/dioxane (4 M, 10 mL). The mixture was stirred at 25° C. for 1 hr. The reaction mixture was concentrated to dryness under reduced pressure to give intermediate 80-7 (300 mg, 87.2% yield) as a white solid. The crude product was used into the next step without further purification.
LCMS (ESI+): m/z 356.4 [M+H]+.
To a solution of 80-7 (70 mg, 179 μmol, HCl) and intermediate 55-10 (91.8 mg, 179 μmol) in DCM (4 mL) was added TEA (54.2 mg, 536 μmol, 75 μL) and AcOH (10.73 mg, 179 μmol, 10.2 μL), the mixture was stirred at 25° C. for 1 hr. Then NaBH3CN (16.8 mg, 268 μmol) was added and the obtained mixture was further stirred at 25° C. for 3 hrs. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water, (2% FA)) to give the crude, which was further purified by prep-TLC (SiO2, DCM: MeOH=10:1) to give compound 80 (12.93 mg, 8.15% yield) as a yellow solid.
LCMS (ESI+): m/z 853.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 10.14 (br s, 1H), 8.82 (s, 1H), 8.09-8.00 (m, 1H), 7.78-7.72 (m, 1H), 7.63-7.52 (m, 3H), 7.33-7.27 (m, 1H), 6.95-6.87 (m, 4H), 6.26-6.18 (m, 1H), 5.73-5.60 (m, 1H), 5.33 (s, 1H), 5.02-4.97 (m, 1H), 4.86-4.79 (m, 1H), 4.72-4.64 (m, 2H), 4.45-4.37 (m, 1H), 3.69-3.60 (m, 2H), 3.07-2.88 (m, 2H), 2.79-2.69 (m, 1H), 2.64-2.56 (m, 3H), 2.45-2.35 (m, 1H), 2.29-2.13 (m, 2H), 2.10-2.04 (m, 1H), 1.96-1.87 (m, 2H), 1.86-1.78 (m, 3H), 1.76-1.58 (m, 5H), 1.46 (s, 6H), 1.27-1.22 (m, 2H).
The compound 81, 82 and compound 145 were synthesized according to the procedures described for compound 77 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
A mixture of intermediate B 23 (100 mg, 309 μmol), intermediate 89-2 (86.5 mg, 464 μmol), RuPhos Pd G3 (51.7 mg, 61.8 μmol), Cs2CO3 (302 mg, 928 μmol) in dioxane (1 mL) was stirred at 100° C. for 2 hrs under N2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by chromatography on silica gel (DCM: MeOH=10: 1) to give intermediate 89-3 (91 mg, 54.9% yield) as a red solid.
LCMS (ESI+): m/z 429.4 [M+H]+.
To a solution of intermediate 89-3 (81 mg, 189 μmol) in DCM (2 mL) was added HCl/dioxane (4M, 2 mL). The mixture was stirred at 20° C. for 2 hrs. The reaction mixture was concentrated under reduced pressure to give a residue to give intermediate 89-4 (71 mg, crude) as a white solid which was used into next step directly.
LCMS (ESI+): m/z 329.3 [M+H]+.
Step 3: synthesis of compound 89
To a solution of intermediate 89-4(71.0 mg, 216 μmol) and intermediate 55-10 (111 mg, 216 μmol) in DCM (5 mL) was added TEA (109 mg, 1.08 mmol, 150 uL) and AcOH (64.9 mg, 1.08 mmol, 62 uL) and was stirred at 25° C. for 0.5 hour. Then the mixture was added NaBH(OAc)3 (137 mg, 648 μmol) and stirred at 20° C. for another 16 hrs. The reaction mixture was diluted with DCM (10 mL) and water (10 mL), and the organic layer was dried over Na2SO4, filtered and concentrated in vacuo to give the residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA)) to give compound 89 (20 mg, 11.1% yield) as a white solid.
LCMS (ESI+): m/z 848.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.52 (s, 1H), 10.15 (s, 1H), 8.82 (s, 1H), 8.16 (s, 0.172H), 8.05 (s, 1H), 7.75 (d, J=7.20 Hz, 1H), 7.50-7.65 (m, 3H), 7.46 (d, J=9.20 Hz, 1H), 6.93 (d, J=8.80 Hz, 3H), 6.83 (s, 1H), 5.57-5.76 (m, 1H), 5.33 (s, 1H), 4.99 (d, J=10.00 Hz, 1H), 4.82 (d, J=17.6 Hz, 1H), 4.68 (d, J=6.00 Hz, 2H), 3.86-3.92 (m, 6H), 3.65 (d, J=10.80 Hz, 2H), 3.31 (s, 2H), 3.24 (s, 5H), 2.73 (t, J=6.80 Hz, 2H), 2.59-2.65 (m, 2H), 2.25 (d, J=7.20 Hz, 2H), 1.83 (d, J=11.20 Hz, 2H), 1.71 (s, 1H), 1.46 (s, 6H), 1.23 (s, 2H).
To a solution of intermediate 93-1 (1 g, 4.16 mmol) in DCM (5 mL) was added DMP (2.65 g, 6.24 mmol) and the mixture was stirred at 25° C. for 3 hrs. The reaction mixture was quenched by addition aqueous NaHCO3 and Na2S2O3 (1:1) 10 mL at 25° C., and then diluted with H2O (20 mL) and extracted with EtOAc (40 mL×2). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give the intermediate 93-2 (956 mg, crude) as a yellow solid which was used into the next step directly.
To a solution of intermediate 93-2 (500 mg, 2.10 mmol) in THE (5 mL) was added saturated aqueous NH4Cl (1 mL) and Fe (1.17 g, 21.0 mmol). The mixture was stirred at 80° C. for 1 hr. The reaction mixture was dried over Na2SO4, filtered, washed with EtOAc and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give the intermediate 93-3 (256 mg, 28.6% yield) as a yellow solid.
To a solution of intermediate 93-3 (256 mg, 1.23 mmol) and intermediate 80-5 (708 mg, 3.69 mmol) in DMF (5 mL) was added NaHCO3 (310 mg, 3.69 mmol). The mixture was stirred at 70° C. for 3 hrs. The reaction mixture was partitioned between EtOAc (100 mL) and H2O (100 mL). The organic phase was separated, washed with brine, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give the intermediate 93-5 (151 mg, 38.5% yield) as a yellow solid. LCMS (ESI+): m/z 320.2 [M+H]+.
To a solution of intermediate 93-5 (151 mg, 424 μmol) and intermediate B 7-B (165 mg, 315 μmol) in DCM (5 mL) was added TEA (95.7 mg, 946 μmol, 132 uL), the mixture was stirred at 25° C. for 0.5 hr. Then the mixture was added AcOH (56.8 mg, 946 μmol, 54 uL) and stirred at 25° C. for 0.5 hr. Then the mixture was added NaBH(OAc)3 (134 mg, 630 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA)) to give compound 93 (10.55 mg, 4.12% yield) as a yellow solid.
LCMS (ESI+): m/z 790.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 10.18 (br s, 1H), 8.83 (s, 1H), 8.14 (s, 0.706H), 8.06 (br s, 1H), 7.82-7.71 (m, 1H), 7.66-7.46 (m, 3H), 7.00-6.89 (m, 2H), 6.88-6.78 (m, 1H), 6.55-6.46 (m, 1H), 6.45-6.36 (m, 1H), 5.85-5.76 (m, 1H), 5.72-5.59 (m, 1H), 5.35 (s, 1H), 4.99 (d, J=9.6 Hz, 1H), 4.86-4.77 (m, 1H), 4.73-4.59 (m, 2H), 4.30-4.19 (m, 1H), 3.22-3.15 (m, 2H), 3.11 (br s, 4H), 2.80-2.72 (m, 1H), 2.68 (br s, 4H), 2.63-2.56 (m, 3H), 2.33 (br s, 1H), 2.13-2.03 (m, 1H), 1.91-1.78 (m, 3H), 1.66-1.52 (m, 2H), 1.46 (s, 6H).
The compound 148 was synthesized according to the procedures described for compound 93 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
The compound 71 (12 mg, 3.13 mmol) was separated by prep-SFC (Column: DAICEL CHIRALPAK IE (250 mm*30 mm, 10 μm); Mobile phase: [IPA-ACN]; CO2%: 45%-45%).
Compound 100-A (1.18 mg, 9.83% yield) as peak 1 was obtained as a yellow solid.
Compound 100-B (0.76 mg, 6.33% yield) as peak 2 was obtained as a yellow solid.
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:isopropanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, RT=1.87 min).
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A: isopropanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, RT=2.43 min).
The compound 74 (15.20 mg, 18.91 μmol) was separated by prep-SFC (Column: DAICEL CHIRALPAK IE (250 mm*30 mm, 10 μM); Mobile phase: [IPA-ACN]; CO2%: 45%-45%.
Compound 102-A (2.10 mg, 13.82%) as peak 1 was obtained as a yellow solid. Compound 102-B (2.20 mg, 14.47%) as peak 2 was obtained as a yellow solid.
Chiral analysis (Column: chiralpak IF-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:isopropanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=3.35 min).
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:isopropanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=5.89 min).
A mixture of intermediate B 23 (500 mg, 1.55 mmol), intermediate 104-1 (320 mg, 2.01 mmol), RuPhos Pd G3 (388 mg, 464 μmol), Cs2CO3 (1.51 g, 4.64 mmol) in dioxane (5 mL) was stirred at 100° C. for 2 hrs under N2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give the intermediate 104-2 (160 mg, 17.8% yield) as a light yellow solid.
LCMS (ESI+): m/z 402.5 [M+H]+.
A mixture of intermediate 104-2 (160 mg, 399 μmol), HCl/dioxane (4 M, 3 mL) in DCM (5 mL) was stirred at 20° C. for 1 hr. The reaction mixture was concentrated under reduced pressure to give the intermediate 104-3 (160 mg, crude) as a brown solid. The crude product was used into the next step without further purification.
LCMS (ESI+): m/z 356.3 [M+H]+.
To a solution of intermediate 104-3 (80 mg, 225 μmol) and intermediate B 8 (109 mg, 225 μmol) in DCM (5 mL) was added dropwise Et3N (68.3 mg, 675 μmol) and AcOH (27 mg, 450 μmol). After addition, the mixture was stirred at this temperature for 30 min, and then NaBH(OAc)3 (143 mg, 675 μmol) was added dropwise. The resulting mixture was stirred at 20° C. for 16 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA) to give compound 104 (13 mg, 6.79% yield) as a pink solid.
LCMS (ESI+): m/z 825.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.50 (s, 1H), 10.24 (br s, 1H), 8.87 (s, 1H), 8.23 (s, 0.858FA), 8.10-8.03 (m, 1H), 7.75 (d, J=8.0 Hz, 1H), 7.69-7.59 (m, 3H), 7.43 (d, J=8.0 Hz, 1H), 7.22 (d, J=12.0 Hz, 2H), 6.91 (d, J=8.0 Hz, 1H), 6.80 (s, 1H), 5.72-5.61 (m, 1H), 5.36 (br s, 1H), 4.99 (d, J=12.0 Hz, 1H), 4.82 (d, J=16.0 Hz, 1H), 4.68 (d, J=8.0 Hz, 2H), 3.92-3.84 (m, 5H), 3.80 (d, J=12.0 Hz, 2H), 2.97 (d, J=8.0 Hz, 2H), 2.77-2.70 (m, 4H), 2.46 (br s, 1H), 2.21 (d, J=8.0 Hz, 2H), 2.04-1.93 (m, 2H), 1.83 (d, J=12.0 Hz, 2H), 1.77-1.70 (m, 3H), 1.69-1.60 (m, 2H), 1.46 (s, 6H), 1.33-1.19 (m, 2H).
The compound 105, 117, 124, 129 and compound 143 were synthesized according to the procedures described for compound 104 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
A mixture of intermediate 106-1 (500 mg, 999 μmol), tert-butyl piperazine-1-carboxylate (372 mg, 2.00 mmol), Cs2CO3 (977 mg, 3.00 mmol) and [2-(2-aminophenyl)phenyl]-methylsulfonyloxy-palladium; dicyclohexyl-[2-(2,6-diisopropoxyphenyl)phenyl]phosphane (84 mg, 99.9 μmol) in toluene (5 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100° C. for 16 hrs under N2 atmosphere. The reaction mixture was partitioned between EtOAc (50 mL) and H2O (50 mL). The organic phase was separated, washed with brine, dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=5/1 to 1/1) to give intermediate 106-2 (302 mg, 40.42% yield) as a yellow solid. LCMS (ESI+): m/z 606.5 [M+H]+.
To a solution of intermediate 106-2 (302 mg, 499 μmol), Pd/C (300 mg, 10% purity) in THF (10 mL) and EtOH (10 mL) was degassed and purged with H2 for 3 times, and then the mixture was stirred at 25° C. for 16 hrs under H2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/1 to 0/1) to give intermediate 106-3 (118 mg, 53.7% yield) as a white solid. LCMS (ESI+): m/z 428.2 [M+H]+.
To a solution of intermediate 106-3 (118 mg, 276 μmol) in DCM (2 mL) was added HCl/dioxane (4 M, 2 mL). The mixture was stirred at 20° C. for 0.5 hr. The reaction mixture was filtered and concentrated under reduced pressure to give the intermediate 106-4 (115 mg, crude) as a white solid, which was used into next step directly.
To a solution of intermediate 106-4 (115 mg, 316 μmol) and intermediate 55-10 (174 mg, 316 μmol) in DCM (5 mL) was added TEA (96 mg, 948 μmol, 132 uL) and stirred at 25° C. for 0.5 hr. Then the mixture was added AcOH (43.7 mg, 727 μmol, 41.6 uL) and stirred at 25° C. for 0.5 hr. Then the mixture was added NaBH(OAc)3 (134 mg, 632 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA)) to give compound 106 (65 mg, 23.71% yield) as a yellow solid. LCMS (ESI+): m/z 825.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.87 (s, 1H), 10.16 (br s, 1H), 8.82 (s, 1H), 8.17 (s, 0.460H), 8.05 (br s, 1H), 7.79-7.71 (m, 1H), 7.63-7.53 (m, 3H), 7.50 (d, J=8.8 Hz, 1H), 6.93 (d, J=8.8 Hz, 3H), 6.85 (s, 1H), 5.72-5.59 (m, 1H), 5.35 (br s, 1H), 4.99 (d, J=10.4 Hz, 1H), 4.86-4.78 (m, 1H), 4.72-4.61 (m, 2H), 4.30-4.23 (m, 1H), 3.89 (s, 3H), 3.69-3.60 (m, 2H), 3.23 (br s, 3H), 2.70-2.59 (m, 4H), 2.54 (br s, 5H), 2.34-2.28 (m, 1H), 2.27-2.21 (m, 2H), 2.20-2.10 (m, 1H), 1.88-1.77 (m, 2H), 1.70 (br s, 1H), 1.46 (s, 6H), 1.31-1.19 (m, 2H).
The compound 118 was synthesized according to the procedures described for compound 106 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of 107-1 (5 g, 23.91 mmol) and 107-2 (2.75 g, 23.91 mmol) in DMF (10 mL) was added K2CO3 (9.91 g, 71.74 mmol) and the mixture was stirred at 80° C. for 3 hrs under N2. The mixture was poured into water (30 mL) and extracted with ethyl acetate (40 mL×2). The combined organic extracts were washed with brine (20 mL), dried over anhydrous Na2SO4 filtered and concentrated under reduced pressure to give a residue, which was purified by flash silica gel chromatography (ISCO®; 20 g SepaFlash® Silica Flash Column, Eluent of 0˜50% Ethyl acetate/Petroleum ethergradient at 60 mL/min) to afford the title intermediate 107-3 (7.1 g, 97.6% yield) as a yellow solid
1H NMR (400 MHz, CDCl3) δ 8.51-8.48 (m, 1H), 8.32-8.26 (m, 1H), 7.26-7.22 (m, 1H), 3.62-3.54 (m, 2H), 3.48-3.39 (m, 2H), 2.93-2.90 (m, 1H), 2.88-2.84 (m, 1H), 1.92-1.81 (m, 2H), 1.72-1.66 (m, 1H), 1.53-1.41 (m, 2H).
To a solution of 107-3 (3 g, 9.86 mmol) in EtOAc (20 mL) was added Pd/C (0.325 g, 985.98 μmol, 10%) and the mixture was stirred at 25° C. for 12 hr under H2 (15 psi). The mixture was filtered and concentrated under reduced pressure to afford intermediate 107-4 (2.5 g, 92.44% yield) as a yellow solid.
LCMS (ESI+): m/z 275.9[M+H]+.
To a solution of 107-4 (2.5 g, 9.11 mmol) and intermediate 80-5 (5.25 g, 27.34 mmol) in DMF (18 mL) was added NaHCO3 (4.59 g, 54.69 mmol) and the mixture was stirred at 70° C. for 12 hrs. The mixture was poured into water (100 mL) and extracted with ethyl acetate (40 mL×2). The combined organic extracts were washed with brine (80 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to afford the crude product. The crude product was purified by flash silica gel chromatography (Ethyl acetate/Petroleum ethergradient=0/1 to 1/1) to afford the title intermediate 107-6 (1.72 g, crude) as a yellow solid.
To a solution of intermediate 107-6 (300 mg, 778.45 μmol) in DCM (10 mL) was added DMP (660.35 mg, 1.56 mmol) and the mixture was stirred at 25° C. for 10 min. The mixture was poured into aq. Na2S2O3 (Sat. 6 mL) and NaHCO3(Sat. 8 mL) and extracted with ethyl acetate (6 mL×2). The combined organic extracts were washed with brine (20 mL), dried over anhydrous Na2SO4, filtered and concentrated under reduced pressure to afford the intermediate 107-7 (170 mg, crude) as a blue solid. The residue was used for the next step reaction directly.
To a solution of intermediate 107-7 (40 mg, 104.34 μmol) and intermediate B 8 (42.86 mg, 82.10 μmol) in DCM (8 mL) was added acetic acid (66.34 mg, 313.02 μmol) and the mixture was stirred at 25° C. for 12 hrs, then sodium triacetoxyboranuide (66.34 mg, 313.02 μmol, 3 eq) was added to the mixture and the mixture was stirred at 25° C. for 1.5 hrs. The mixture was concentrated under reduced pressure to afford the crude product. The residue was purified by prep-HPLC (acetonitrile/water (0.1% TFA)) to give the desired compound 107 (18 mg, 18.87% yield) as white solids.
LCMS (ESI+): m/z 853.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.81 (s, 1H), 10.38-10.21 (m, 1H), 8.89-8.87 (m, 1H), 8.09-7.96 (m, 1H), 7.78-7.74 (m, 1H), 7.73-7.68 (m, 2H), 7.66-7.61 (m, 1H), 7.32-7.27 (m, 1H), 7.25-7.19 (m, 2H), 6.95-6.92 (m, 1H), 6.92-6.87 (m, 1H), 5.76-5.60 (m, 1H), 5.02-4.98 (m, 1H), 4.88-4.77 (m, 1H), 4.73-4.64 (m, 2H), 4.42-4.32 (m, 1H), 3.10-2.99 (m, 4H), 2.88-2.53 (m, 8H), 2.23-1.79 (m, 11H), 1.47 (s, 6H), 1.40-1.32 (m, 2H).
To a solution of intermediate 108-1 (500 mg, 2.42 mmol) in ethyl 2,2,2-trifluoroacetate (8 mL) was added TEA (491 mg, 4.85 mmol). The mixture was stirred at 80° C. for 16 hrs. The reaction mixture was partitioned between EA (30 mL) and H2O (20 mL). The organic phase was separated, washed with saturated NaCl (10 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give the intermediate 108-2 (750 mg, crude) as brown oil.
To a solution of intermediate 108-2 (750 mg, 2.48 mmol,) in THE (8 mL) was added Pd/C (500 mg, 5% purity) under N2. The suspension was degassed under vacuum and purged with H2 three times. The mixture was stirred under H2 (15 PSi) at 30° C. for 16 hours. The reaction mixture was filtered, washed with DCM (50 mL) and the filtrate was concentrated to the crude product intermediate 108-3 (670 mg, crude) as a white solid, which was directly used in next step.
To a colorless solution of intermediate B 15-B (40.0 mg, 104 μmol) in toluene (3 mL) was added m-CPBA (23.3 mg, 115 μmol, 85% purity) and stirred at 20° C. for 30 min. Then Et3N (21.5 mg, 212 μmol) and intermediate 108-3 (31.2 mg, 115 μmol) was added. The reaction mixture was stirred at 20° C. for 16 hours. The reaction mixture was concentrated to give a residue. The residue was purified by prep-TLC (DCM: MeOH=10:1) to give the intermediate 108-4 (45.0 mg, 71.0% yield) as a yellow oil.
A white suspension of intermediate 108-4 (45.0 mg, 74.1 μmol), K2CO3 (15.35 mg, 111 μmol) and MeOH (2 mL) was stirred at 30° C. for 16 hours. The reaction mixture was diluted with DCM (30 mL) and washed with water (10 mL). The organic layer was dried over Na2SO4, filtered and concentrated to give the intermediate 108-5 (24.0 mg, crude) as a yellow solid which was used into next step directly.
LCMS (ESI+): m/z 512.4 [M+H]+.
A mixture of intermediate 108-5 (24.0 mg, 46.9 μmol), intermediate 64-2 (17.4 mg, 46.9 μmol), Et3N (14.24 mg, 140.7 μmol) in DCM (2 mL) was stirred at 20° C. for 30 min and then AcOH (10.0 mg, 166.52 μmol) was added. The reaction mixture was stirred at 20° C. for 30 min and then NaBH3CN (5.90 mg, 93.8 μmol) was added. The reaction mixture was stirred at 20° C. for 16 hours. The reaction mixture was diluted with DCM (30 mL) and washed with water (30 mL). The organic layer was dried over Na2SO4, filtered and concentrated to give a residue. The residue was purified by prep-TLC (DCM: MeOH=10: 1) to give compound 108 (7 mg, 16% yield) as a white solid. LCMS (ESI+): m/z 866.6[M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.12 (s, 1H), 10.29 (br s, 1H), 9.19-8.93 (m, 1H), 8.88 (s, 1H), 7.92 (br s, 1H), 7.70 (d, J=8.0 Hz, 3H), 7.21 (d, J=8.4 Hz, 2H), 7.09-6.82 (m, 3H), 5.75-5.60 (m, 1H), 5.36 (dd, J=5.2, 12.4 Hz, 1H), 5.09 (s, 1H), 5.00 (d, J=10.4 Hz, 1H), 4.85 (d, J=17.6 Hz, 1H), 4.80-4.69 (m, 1H), 4.56 (dd, J=5.6, 15.6 Hz, 1H), 3.64 (s, 5H), 3.15 (br s, 4H), 3.01-2.84 (m, 3H), 2.83-2.68 (m, 4H), 2.44 (d, J=8.8 Hz, 1H), 2.27-2.16 (m, 2H), 2.09-1.94 (m, 5H), 1.93-1.82 (m, 4H), 1.80-1.63 (m, 2H), 1.45 (br s, 2H), 0.87 (t, J=7.6 Hz, 3H).
To a solution of intermediate 110-1 (10 g, 62.9 mmol) and intermediate 110-2 (15.0 g, 94.3 mmol) in MeCN (100 mL) was added TEA (12.7 g, 126 mmol). The mixture was stirred at 90° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/0 to 10/1) to give the intermediate 110-3 (17.8 g, 85.4% yield) as an orange solid.
1H NMR (400 MHz, chloroform-d) 7.97 (m, 1H), 7.89 (m, 1H), 6.91 (t, J=8.8 Hz, 1H), 4.10 (d, J=6.8 Hz, 1H), 3.79-3.70 (m, 2H), 3.39 (s, 6H), 2.85 (m, 2H), 1.92-1.77 (m, 3H), 1.58-1.42 (m, 2H).
Step 2: Synthesis of Intermediate 110-4 To a solution of intermediate 110-3 (17.8 g, 59.7 mmol) in THE (200 mL) was added Pd/C (10%, 4 g) under N2 atmosphere. The suspension was degassed and purged with H2 for 3 times. The mixture was stirred under H2 (15 Psi) at 15° C. for 16 hrs. The reaction mixture was filtered, wahed with DCM(200 mL) and concentrated under reduced pressure to give the intermediate 110-4 (15.8 g, crude) was obtained as an orange solid.
To a solution of intermediate 110-4 (15.8 g, 58.9 mmol) in ethyl 2,2,2-trifluoroacetate (102 g, 717 mmol) was added TEA (32.4 g, 320 mmol). The mixture was stirred at 80° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give a residue. The crude product was triturated with (PE: EA=3:1, 50 mL), filtered and dried to give the intermediate 110-5 (20.5 g, 86.0% yield) as a white solid.
1H NMR (400 MHz, CDCl3) 8.02 (br s, 1H), 7.42 (d, J=13.2 Hz, 1H), 7.18 (d, J=8.8 Hz, 1H), 6.94 (t, J=8.4 Hz, 1H), 4.15-4.05 (m, 1H), 3.46 (d, J=10.8 Hz, 2H), 3.38 (d, J=1.2 Hz, 6H), 2.63 (t, J=12 Hz, 2H), 1.85 (d, J=12.8 Hz, 2H), 1.53 (m, 2H), 1.36-1.25 (m, 1H).
A solution of intermediate 110-5 (20.5 g, 56.3 mmol) in TFA (100 mL) was stirred at 50° C. for 1.5 hrs. The reaction mixture was concentrated under reduced pressure to give the intermediate 110-6 (37 g, 3.0 eq TFA salt) as brown oil. The crude product was used into the next step without further purification. LCMS (ESI+): m/z 319.2 [M+H]+.
Step 5: Synthesis of Intermediate 110-8 To a mixture of intermediate 110-6 (37 g, 56.0 mmol, 3TFA) and intermediate 110-7 (11.6 g, 56.0 mmol) in DCM (200 mL) was added TEA (56.7 g, 560 mmol) at 25° C. and stirred at 25° C. for 30 min, then AcOH (42.0 g, 699 mmol) was added and stirred for 0.5 hr. The reaction mixture was added NaBH(OAc)3 (35.6 g, 168 mmol, 3 eq) at 25° C. and stirred at 25° C. for 16 hours. The reaction mixture was diluted with water (200 mL) and DCM (200 mL), and extracted with DCM for three times. The combined organic layers were washed with sat. aq. NaCl (100 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=1/0 to 10/1) to give the intermediate 110-8 (13.6 g, 39.5% yield) as a yellow solid.
LCMS (ESI+): m/z 509.6 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 11.25 (s, 1H), 8.14-7.96 (m, 2H), 7.49 (d, J=14.2 Hz, 1H), 7.39 (d, J=8.8 Hz, 1H), 7.15-6.91 (m, 3H), 3.45 (br s, 4H), 3.33-3.31 (m, 4H), 2.65 (t, J=11.2 Hz, 2H), 2.58-2.52 (m, 2H), 2.23 (d, J=6.8 Hz, 2H), 1.81 (d, J=12.0 Hz, 2H), 1.68 (br s, 1H), 1.31-1.20 (m, 2H).
To a solution of intermediate 110-8 (1 g, 1.96 mmol) in THE (20 mL) was added Pd/C (10%, 500 mg) under N2 atmosphere. The suspension was degassed and purged with H2 for 3 times. The mixture was stirred under H2 (15 Psi) at 20° C. for 16 hrs. The reaction mixture was filtered, washed with (DCM: MeOH=10:1, 200 mL) and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=1:0 to 10:1) to give the intermediate 110-9 (624 mg, 61.7% yield) as a gray white solid. LCMS (ESI+): m/z 480.9 [M+H]+.
A solution of intermediate B 15-B (300 mg, 782 μmol) in toluene (6 mL) was added m-CPBA (186 mg, 916 μmol, 85% purity) at 0° C. and stirred at 3 mins. Then the reaction mixture was added DIEA (202 mg, 1.56 mmol) and intermediate 110-9 (394 mg, 822 μmol) at 15° C. and the mixture was stirred at 15° C. for 16 hours. The reaction mixture was quenched by addition Na2S2O3 (15 mL) and NaHCO3 (15 mL) at 15° C., and then extracted with DCM (30 mL×3). The combined organic layers were washed with aqueous NaCl (15 mL), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM/MeOH=1:0 to 20:1) to give the intermediate 110-11 (596 mg, 84.1% yield) as a yellow solid. LCMS (ESI+): m/z 816.4 [M+H]+.
To a solution of intermediate 110-11 (596 mg, 731 μmol) in MeOH (12 mL) and H2O (4 mL) was added K2CO3 (1.52 g, 11.0 mmol). The mixture was stirred at 40° C. for 16 hrs. The reaction mixture was diluted with H2O (8 mL) and extracted with DCM 120 mL (40 mL×3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM/MeOH=1/0 to 10/1) to give the intermediate 110-12 (362 mg, 68.85% yield) as a yellow solid. LCMS (ESI+): m/z 719.2 [M+H]+.
To a solution of intermediate 110-12 (362 mg, 504 μmol) and 3-bromopiperidine-2,6-dione (1.45 g, 7.55 mmol) in DMF (8 mL) was added NaHCO3 (635 mg, 7.55 mmol). The mixture was stirred at 70° C. for 16 hrs. The reaction mixture was partitioned between H2O (30 mL) and EA (30 mL) and extracted with EA two times. The organic phase was combined, and washed with sat. aq. NaCl (15 mL×2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=1/0 to 10/1). The obtained crude product was triturated with MeCN/H2O=1:1 (8 mL) and filtered to give a cake of compound 110 (210 mg, 48.7% yield) as a yellow solid.
LCMS (ESI+): m/z 852.5 [M+Na]+.
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H), 10.29-10.05 (m, 1H), 8.82 (s, 1H), 7.93 (br s, 1H), 7.70 (d, J=8.0 Hz, 1H), 7.59 (br s, 2H), 6.92 (d, J=8.8 Hz, 2H), 6.83 (t, J=9.2 Hz, 1H), 6.49 (d, J=14.8 Hz, 1H), 6.44-6.36 (m, 1H), 5.86-5.74 (m, 1H), 5.71-5.59 (m, 1H), 5.07 (s, 1H), 4.99 (d, J=10.2 Hz, 1H), 4.92-4.67 (m, 2H), 4.64-4.49 (m, 1H), 4.31-4.18 (m, 1H), 3.10 (br s, 6H), 3.01-2.93 (m, 1H), 2.83-2.65 (m, 4H), 2.60-2.53 (m, 5H), 2.27-2.15 (m, 3H), 2.11-1.97 (m, 2H), 1.94-1.75 (m, 4H), 1.73-1.55 (m, 2H), 1.31-1.20 (m, 2H), 0.87 (t, J=7.2 Hz, 3H).
The diastereoisomers of compound 110 (50 mg, 60.2 μmol) was separated by prep-SFC (Column: DAICEL CHIRALPAK ID (250 mm*30 mm, 10 μm); Mobile phase: [MeOH-ACN]; CO2%: 50%-50% at 50 mL/min. Compound 150-A (18 mg, 36%) as peak 1 was obtained as a yellow solid.
Compound 150-B (20 mg, 40%) as peak 2 was obtained as a yellow solid.
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:methol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=3.66 min).
LCMS (ESI+): m/z 830.4 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 10.25-10.06 (m, 1H), 8.82 (s, 1H), 8.02-7.85 (m, 1H), 7.70 (d, J=8.4 Hz, 1H), 7.63-7.42 (m, 2H), 7.01-6.88 (m, 2H), 6.86-6.74 (m, 1H), 6.56-6.46 (m, 1H), 6.41 (d, J=8.8 Hz, 1H), 5.87-5.74 (m, 1H), 5.72-5.57 (m, 1H), 5.08 (s, 1H), 4.99 (d, J=10.4 Hz, 1H), 4.91-4.69 (m, 2H), 4.64-4.48 (m, 1H), 4.33-4.17 (m, 1H), 3.10 (br s, 6H), 3.00-2.93 (m, 1H), 2.81-2.66 (m, 4H), 2.62-2.54 (m, 5H), 2.26-2.16 (m, 3H), 2.10-1.98 (m, 2H), 1.94-1.76 (m, 4H), 1.73-1.58 (m, 2H), 1.30-1.18 (m, 2H), 0.87 (t, J=7.2 Hz, 3H)
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:methol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=5.92 min).
LCMS (ESI+): m/z 830.4 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.80 (s, 1H), 10.25-10.06 (m, 1H), 8.86-8.77 (m, 1H), 8.02-7.86 (m, 1H), 7.73-7.66 (m, 1H), 7.64-7.48 (m, 2H), 6.92 (d, J=8.4 Hz, 2H), 6.83 (t, J=9.2 Hz, 1H), 6.55-6.44 (m, 1H), 6.41 (d, J=9.2 Hz, 1H), 5.84-5.75 (m, 1H), 5.72-5.60 (m, 1H), 5.08 (s, 1H), 5.02-4.95 (m, 1H), 4.91-4.71 (m, 2H), 4.63-4.49 (m, 1H), 4.31-4.19 (m, 1H), 3.10 (br s, 6H), 3.00-2.94 (m, 1H), 2.84-2.69 (m, 4H), 2.56 (d, J=11.6 Hz, 5H), 2.27-2.19 (m, 3H), 2.10-1.99 (m, 2H), 1.90-1.76 (m, 4H), 1.73-1.60 (m, 2H), 1.28-1.21 (m, 2H), 0.87 (t, J=7.2 Hz, 3H)
The compound 112, compound 152-A and compound 152-B were synthesized according to the procedures described for compound 110 using the appropriated intermediates.
The compound 112 (40 mg, 49.02 μmol) was separated by prep-SFC (Column: DAICEL CHIRALPAK ID (250 mm*30 mm, 10 μM); Mobile phase: [MeOH-ACN]; CO2%: 50%-50%.
Compound 152-A (15 mg, 37.5%) as peak 1 was obtained as a yellow solid. Compound 152-B (15 mg, 37.5%) as peak 2 was obtained as a yellow solid.
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:methanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=3.43 min).
Chiral analysis (Column: chiralpak IE-3, 100*4.6 mm I.D., 3 μm; Mobile Phase: A:methanol (0.05% DEA), B: acetonitrile, Isocatic: A/B=1/1 at 1.0 mL/min; Temp: 35° C.; Wavelength: 254 nm, Rt=5.41 min).
The following compounds were synthesized according to the procedures described for compound 110 using the appropriated intermediates.
1H NMR (400 MHz, DMSO-d6) δ
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.78 (s, 1 H),
1H NMR (400 MHz, DMSO-d6) δ10.78 (s, 1H), 10.12
1H NMR (400 MHz, DMSO-d6) δ 10.77 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.78 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ10.79 (br s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.79 (s, 1H),
1H NMR (400 MHz, DMSO-d6) δ 10.77 (s, 1H),
To a solution of intermediate 114-1 (250 g, 1.01 mol) and trimethoxymethane (318.36 g, 3.0 mol, 328.88 mL) in MeOH (1 L) was added TsOH (8.70 g, 50.55 mmol). The mixture was stirred at 20° C. for 16 hrs. Then aq. NaHCO3 (150 mL) was added to the mixture, and the reaction mixture was partitioned between water (500 mL) and EA (1000 mL). The organic phase was separated, dried over Na2SO4, filtered and concentrated under reduced pressure to give a crude of intermediate 114-2 (320 g) as a yellow oil, which was used into the next step directly.
To a solution of intermediate 114-2 (320 g, 1.09 mol) in MeOH (600 mL) was added Pd(OH)2/C (7.5 g, 10% purity). The mixture was stirred at 40° C. under H2 (40 Psi) for 16 hrs. The reaction mixture was filtered and the filtrate was concentrated to give a crude of intermediate 114-3 (170 g, crude) as a yellow oil, which was used into the next step directly.
A brown suspension of intermediate B 23 (6.30 g, 19.5 mmol), intermediate 114-3 (4.04 g, 25.3 mmol), Cs2CO3 (19.1 g, 58.5 mmol), RuPhos Pd G3 (4.08 g, 4.87 mmol) in dioxane (120 mL) was degassed with N2 for 3 times and stirred at 100° C. for 6 hours under N2. The reaction mixture was diluted with DCM (100 mL) and filtered. The filtrate was concentrated to give a residue. The residue was purified by combi flash (DCM:MeOH=1:0 to 20:1, 80 g silica gel column) to give intermediate 114-4 (3.9 g, 49.8% yield) as a yellow solid.
A solution of intermediate 114-4 (3.9 g, 6.08 mmol) in TFA (50 mL) was stirred at 50° C. for 30 min. The reaction mixture was concentrated to give a crude of intermediate 114-5 (4.70 g, TFA salt) as a red oil which was used into the next step directly.
LCMS (ESI+): m/z 356.0 [M+H]+.
A yellow suspension of intermediate 114-5 (4.70 g, 10.0 mmol, TFA), intermediate 114-6 (2.70 g, 13.0 mmol), Et3N (7.09 g, 70.1 mmol) in DCM (100 mL) was stirred at 20° C. for 30 min. Then AcOH (3.01 g, 50.1 mmol) was added and the mixture was stirred at 20° C. for 1 hour. Then NaBH(OAc)3 (8.49 g, 40.1 mmol) was added and the reaction mixture was stirred at 20° C. for 1 hour. The reaction mixture was diluted with DCM (100 mL) and washed with water (100 mL). The organic layer was separated and dried over Na2SO4, filtered and concentrated to give a residue. The residue was purified by combi flash (DCM: MeOH=1: 0 to 20: 1). Intermediate 114-7 (4.90 g, 89.5% yield) was obtained as a yellow solid.
A suspension of intermediate 114-7 (3.0 g, 5.49 mmol) and Pd/C (2.0 g, 10% purity) in THE (100 mL) was stirred at 20° C. under H2 (15 Psi) for 16 hours. The black reaction mixture was filtered and the filtrate was concentrated to give a residue. The residue was purified by combi flash (DCM:MeOH=20:1 to 10:1, 20 g silica gel column) to give the intermediate 114-8 (1.70 g, 59.96% yield) as a brown solid.
To a solution of intermediate B15-B (550 mg, 1.06 mmol) in toluene (3 mL) was added m-CPBA (303 mg, 1.49 mmol, 85% purity) and the obtained mixture was stirred at 20° C. for 30 min. Then DIEA (275 mg, 2.13 mmol), THE (3 mL) and intermediate 114-8 (420 mg, 1.10 mmol) was added. The reaction mixture was stirred at 20° C. for 16 hours. The reaction mixture was quenched by Na2S2O3 (10 mL) and extrated with DCM (40 mL), dried over Na2SO4, filtered and concentrated to give a residue. The residue was purified by chromatography on silica gel (EA: MeOH=10: 1) to give the compound 114 (650 mg, 72% yield) as a pale yellow solid.
LCMS (ESI+): m/z 852.4 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.51 (s, 1H), 10.14 (br s, 1H), 8.82 (s, 1H), 7.93 (br s, 1H), 7.70 (d, J=8.0 Hz, 1H), 7.59 (br s, 2H), 7.43 (d, J=8.8 Hz, 1H), 6.98-6.84 (m, 3H), 6.81 (s, 1H), 5.76-5.56 (m, 1H), 5.06 (s, 1H), 4.99 (d, J=10.0 Hz, 1H), 4.85 (d, J=17.2 Hz, 1H), 4.75 (br s, 1H), 4.66-4.48 (m, 1H), 3.96-3.86 (m, 5H), 3.81 (d, J=12.4 Hz, 2H), 3.11 (br s, 4H), 3.02-2.90 (m, 1H), 2.82-2.71 (m, 5H), 2.62-2.54 (m, 4H), 2.28-2.18 (m, 3H), 2.09-1.97 (m, 1H), 1.93-1.81 (m, 3H), 1.79-1.61 (m, 2H), 1.31-1.21 (m, 2H), 0.87 (t, J=7.6 Hz, 3H)
The following compounds were synthesized according to the procedures described for compound 114 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 116-1 (1.36 g, 4.85 mmol) in DCM (15 mL) was added HCl/dioxane (4 M, 20 mL). The mixture was stirred at 20° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give a crude of intermediate 116-2 (1.5 g) as a yellow solid which was used into the next step directly.
LCMS (ESI+): m/z 234.9 [M+H].
A mixture of intermediate 89-4 (200 mg, 548 μmol), intermediate 116-2 (222 mg, 822 μmol) in DCM (5 mL) was added TEA (277 mg, 2.74 mmol) and HOAc (165 mg, 2.74 mmol). After addition, the mixture was stirred at this temperature for 30 min. Then NaBH(OAc)3 (349 mg, 1.64 mmol) was added and the resulting mixture was stirred at 20° C. for 16 hrs. The reaction mixture was diluted with H2O (50 mL) and extracted with EA (50 mL×2). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO2, DCM: MeOH=10: 1) to give the intermediate 116-3 (150 mg, 48.5% yield) as a yellow solid.
LCMS (ESI+): m/z 547.1 [M+H].
A mixture of intermediate 116-3 (150 mg, 274 μmol), 100% Pd/C (150 mg) in THF (10 mL) was was stirred at 20° C. for 16 hrs under H2 atmosphere (15 Psi). The reaction mixture was filtered and concentrated under reduced pressure to give a crude of intermediate 116-4 (130 mg, crude) as a white solid which was used into the next step directly.
LCMS (ESI+): m/z 517.1 [M+H]+.
To a colorless solution of intermediate B15-B (50.0 mg, 130 μmol) in toluene (10 mL) was added m-CPBA (29 mg, 143 μmol) and stirred at 20° C. for 30 min. Then Et3N (26.4 mg, 261 μmol) and intermediate 116-4 (67.3 mg, 130 μmol) was added. The reaction mixture was stirred at 20° C. for 16 hours. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-TLC (SiO2, DCM: MeOH=10:1) To give the compound 116 (23 mg, 14.5% yield) as a yellow solid.
LCMS (ESI+): m/z 852.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.52 (s, 1H), 10.13 (br s, 1H), 8.82 (s, 1H), 7.99-7.85 (m, 1H), 7.69 (d, J=8.0 Hz, 1H), 7.57 (br s, 2H), 7.46 (d, J=8.0 Hz, 1H), 6.93 (d, J=12.0 Hz, 3H), 6.83 (s, 1H), 5.74-5.58 (m, 1H), 5.06 (s, 1H), 4.99 (d, J=12.0 Hz, 1H), 4.85 (d, J=16.0 Hz, 1H), 4.75 (s, 1H), 4.56 (d, J=12.0 Hz, 1H), 3.95-3.84 (m, 5H), 3.65 (d, J=12.0 Hz, 2H), 3.28-3.19 (m, 4H), 3.03-2.91 (m, 1H), 2.84-2.76 (m, 1H), 2.75-2.70 (m, 2H), 2.65-2.58 (m, 2H), 2.54 (br s, 4H), 2.30-2.15 (m, 3H), 2.07-1.96 (m, 1H), 1.94-1.78 (m, 3H), 1.77-1.62 (m, 2H), 1.25 (d, J=16.0 Hz, 2H), 0.91-0.82 (m, 3H).
The compound 126, 127 and compound 135 were synthesized according to the procedures described for compound 116 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
To a solution of intermediate 139-1 (2.00 g, 19.8 mmol) and intermediate 139-2 (3.15 g, 19.8 mmol) in DMF (10 mL) was added K2CO3 (5.47 g, 39.6 mmol). The mixture was stirred at 70° C. for 2 hrs. The reaction mixture was poured into water (20 mL) at 25° C., and then extracted with EtOAc (20 mL×3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (Ethyl acetate/Petroleum=0/1 to 1/1) to give the intermediate 139-3 (4.5 g, 94.7% yield) as a yellow oil.
1H NMR (400 MHz, DMSO-d6) δ 7.94-7.86 (m, 2H), 6.83-6.64 (m, 1H), 4.80 (t, J=5.2 Hz, 1H), 3.68-3.44 (m, 4H), 3.39-3.32 (m, 2H), 2.46-2.31 (m, 1H), 2.09-1.99 (m, 1H), 1.79-1.66 (m, 1H).
To a mixture of intermediate 139-3 (4.5 g, 18.73 mmol) in EtOH (50 mL) was added Pd/C (450 mg, 10% purity). The suspension was degassed under vacuum and purged with H2 three times. The mixture was stirred under H2 (15 psi) at 25° C. for 1 hour. The reaction mixture was filtered and concentrated under reduced pressure to give a crude of intermediate 139-4 (4.50 g) as a white solid.
To a solution of intermediate 139-4 (1.00 g, 4.76 mmol) and intermediate 139-5 (1.83 g, 9.51 mmol) in DMF (10 mL) was added NaHCO3 (1.20 g, 14.3 mmol). The mixture was stirred at 80° C. for 8 hrs. The mixture were poured into water (60 mL), extracted with EA (40 mL×3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (ethyl acetate/petroleum=0/1 to 3/1) to give the intermediate 139-6 (800 mg, 52.3% yield) as a purple solid.
1H NMR (400 MHz, DMSO-d6) δ 10.95-10.70 (m, 1H), 6.61 (t, J=9.6 Hz, 1H), 6.51 (dd, J=2.4, 15.6 Hz, 1H), 6.40 (dd, J=2.0, 8.4 Hz, 1H), 5.57 (d, J=7.6 Hz, 1H), 4.66 (t, J=5.2 Hz, 1H), 4.30-4.14 (m, 1H), 3.44-3.37 (m, 2H), 3.20-3.02 (m, 3H), 2.97-2.90 (m, 1H), 2.73-2.65 (m, 1H), 2.61-2.54 (m, 1H), 2.38-2.26 (m, 1H), 2.15-2.05 (m, 1H), 1.98-1.74 (m, 2H), 1.64-1.49 (m, 1H).
To a solution of intermediate 139-6 (400 mg, 1.24 mmol) in DCM (5 mL) was added TEA (315 mg, 3.11 mmol), DMAP (15.2 mg, 124 μmol) and 4-methylbenzenesulfonyl chloride (119 mg, 622 μmol). The mixture was stirred at 25° C. for 4 hrs. The reaction mixture was diluted with DCM (20 mL) and washed with water (20 mL*2), The organic layer was dried over Na2SO4 and filtered, concentrated under reduce pressure to give a residue. The residue was purified by flash silica gel chromatography (ethyl acetate/petroleum=0/1 to 1/1) to give the intermediate 139-7 (300 mg, 50.68% yield) as a white solid.
MS (ESI+): m/z 475.6 [M+H]+
To a solution of intermediate 139-7 (200 mg, 420.58 μmol) and intermediate B7-B (205 mg, 391 μmol, HCl salt) in CH3CN (6 mL) was added KI (140 mg, 841 μmol) and TEA (213 mg, 2.10 mmol). The mixture was stirred at 90° C. for 2 hrs. The mixture was concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (DCM/MeOH=1/0 to 15/1) and the crude was further purified by prep-HPLC (Calumn:Kromasil 100-5-C18 30*150 mm, Mobile Phase A: water (0.01% FA), Mobile Phase B: acetonitrile, 20 mL/min, gradient condition form 15% B to 55%) to give compound 139 (30.1 mg, 8.88% yield) as a white solid.
MS (ESI+): m/z 790.5 [M+H]
1H NMR (400 MHz, DMSO-d6) δ10.78 (s, 1H), 10.31-9.98 (m, 1H), 8.83 (s, 1H), 8.13-7.99 (m, 1H), 7.82-7.71 (m, 1H), 7.66-7.49 (m, 3H), 6.98-6.86 (m, 2H), 6.67-6.57 (m, 1H), 6.55-6.47 (m, 1H), 6.42-6.35 (m, 1H), 5.72-5.60 (m, 1H), 5.59-5.54 (m, 1H), 5.40-5.28 (m, 1H), 5.03-4.95 (m, 1H), 4.86-4.77 (m, 1H), 4.73-4.63 (m, 2H), 4.26-4.16 (m, 1H), 3.23-3.21 (m, 1H), 3.15-3.08 (m, 6H), 2.95-2.89 (m, 1H), 2.76-2.67 (m, 1H), 2.61-2.52 (m, 5H), 2.44-2.32 (m, 3H), 2.13-1.98 (m, 2H), 1.88-1.77 (m, 1H), 1.65-1.53 (m, 1H), 1.46 (s, 6H).
To a solution of intermediate 141-1 (100 mg, 271 μmol) in toluene (4 mL) was added m-CPBA (60.5 mg, 298 μmol, 85% purity) and the reaction stirred at 20° C. for 0.5 hr. Then DIEA (105 mg, 812 μmol) and intermediate 141-2 (82.6 mg, 298 μmol) was added. The mixture was stirred at 20° C. for 16 hrs. The reaction mixture was quenched by addition Na2S2O3 (20 mL) and NaHCO3 (20 mL) and extracted with EA (10 mL×3). The combined organic layers were washed with water 30 mL (10 mL×3), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=10: 1) to give the intermediate 141-3 (120 mg, 74.1% yield) as a yellow solid, which was used into the next step. LCMS (ESI+): m/z 598.69 [M+H]+
Diastereoisomers of compound 141-3 (95 mg, 158.7 μmol) was separated by SFC (column: DAICEL CHIRALPAK IF (250 mm*30 mm, 10 μM); mobile phase: [IPA-ACN]; B %: 50%-50%, 45 mL/min). Peak 1 (Rt=1.13 min) as compound 141-4 (40 mg, 42.11% yield) was obtained as a pale yellow solid and Peak 2 (Rt=2.01 min) as compound 141-4A (40 mg, 42.11% yield) was obtained as a pale yellow solid. The absolute stereochemistry was arbitrarily assigned for compound 141-4 and compound 141-4A.
To a solution of compound 141-4 (40.0 mg, 66.8 μmol) in DCM (7 mL) was added HCl/dioxane (4 M, 2 mL). The mixture was stirred at 25° C. for 16 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give a crude of compound 141-5 (40 mg, crude) as a green solid which was used into the next step directly. LCMS (ESI+): m/z 498.58 [M+H]+
To a mixture of compound 141-5 (40.0 mg, 80.2 μmol) and intermediate 73-8 (26.7 mg, 80.2 μmol) in DCM (4 mL) was added TEA(56.8 mg, 562 μmol) at 20° C. and stirred at 20° C. for 30 min, then added AcOH (14.5 mg, 241 μmol) and stirred for 1 hr. The reaction was added NaBH(OAc)3 (51.0 mg, 241 μmol) stirred at 25° C. for 16 hours. The reaction mixture was partitioned between water (20 mL) and DCM (40 mL). The organic phase was separated dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA)) and lyophilized to give compound 141 (30 mg, 45.8% yield) as a white solid.
LCMS (ESI+): m/z 815.94 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.78 (s, 1H), 10.07 (br s, 1H), 8.82 (s, 1H), 8.18 (br s, 0.531H), 8.11-7.92 (m, 1H), 7.75 (d, J=8 Hz, 1H), 7.57 (t, J=7.2 Hz, 3H), 6.94-6.78 (m, 3H), 6.54-6.37 (m, 2H), 5.78 (d, J=7.6 Hz, 1H), 5.31 (br s, 3H), 4.45 (d, J=4.5 Hz, 2H), 4.29-4.19 (m, 1H), 3.13 (br s, 1H), 3.09 (br s, 4H), 2.79-2.70 (m, 1H), 2.56 (d, J=12.4 Hz, 3H), 2.50-2.47 (m, 4H), 2.22 (d, J=6.7 Hz, 3H), 2.12-1.98 (m, 2H), 1.98-1.82 (m, 2H), 1.81-1.67 (m, 4H), 1.62 (br s, 1H), 1.55 (s, 3H), 1.32-1.17 (m, 2H).
The compound 142 was synthesized according to the procedures described for compound 141 using the different intermediates.
1H NMR (400 MHz, DMSO-d6) δ
A solution of 4-bromoaniline 146-1 (10 g, 58.1 mmol) and acrylic acid (6.28 g, 87.2 mmol, 6.0 mL) in toluene (100 mL) was stirred at 100° C. for 8 hrs. The reaction mixture was concentrated under reduced pressure to give a crude of intermediate 146-2 (15 g, crude) as black oil.
A solution of intermediate 146-2 (14.5 g, 59.4 mmol) and urea (21.4 g, 356 mmol) in AcOH (100 mL) was stirred at 120° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to remove AcOH. The residue was diluted with NaHCO3 (100 mL) and extracted with EA (100 mL×3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The crude was purified by chromatography on silca gel (petroleum/EA=1/0 to 3/1) to give the intermediate 146-3 (3.69 g, 12.34 mmol, 20.8% yield) as a white solid.
1H NMR (400 MHz, DMSO-d6) δ 10.63-10.31 (m, 1H), 7.67-7.47 (m, 2H), 7.35-7.25 (m, 2 H), 5.91 (s, 1H), 3.79 (t, J=6.8 Hz, 2H), 2.71 (t, J=6.8 Hz, 2H).
A mixture of intermediate 146-3 (500 mg, 1.86 mmol), intermediate 146-4 (295 mg, 1.86 mmol), RuPhos Pd G3 (155 mg, 185 μmol), Cs2CO3 (1.82 g, 5.57 mmol) in dioxane (10 mL) was stirred at 100° C. for 2 hrs under N2 atmosphere. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM/MeOH=10/1) to give the intermediate 146-5 (34 mg, 4.21% yield) as a red solid.
LCMS (ESI+): m/z 185.7 [M+H]+.
To a solution of intermediate 146-5 (30 mg, 86.3 μmol) in DCM (1 mL) was added HCl/dioxane (5 mL). The mixture was stirred at 20° C. for 1 hr. The reaction mixture was concentrated under reduced pressure to give a crude of intermediate 146-6 (45 mg, crude) as white solid which was used into the next step directly.
LCMS (ESI+): m/z 320.4 [M+H]+.
To a solution of intermediate 146-6 (45 mg, 149 μmol) and intermediate B 8 (72.5 mg, 149 μmol) in DCM (5 mL) was added TEA (75.6 mg, 747 μmol, 104 μL) and the mixture was stirred at 25° C. for 0.5 hour. Then AcOH (44.8 mg, 747 μmol, 43 μL) was added and stirred at 25° C. for another 0.5 hour. Then the obtained mixture was added NaBH(OAc)3 (95.0 mg, 448 μmol) and stirred at 25° C. for 16 hrs. The reaction mixture was diluted with DCM (10 mL) and water (10 mL), and the organic layer was dried over Na2SO4, filtered and concentrated to give to give a residue, which was purified by prep-HPLC to give the compound 146 (20 mg, 16.0% yield) as a yellow solid.
LCMS (ESI+): m/z 793.5 [M+Na]+.
1H NMR (400 MHz, DMSO-d6) δ 10.28 (s, 1H), 8.88 (s, 1H), 8.14 (s, 0.672H), 8.06 (t, J=7.60 Hz, 1H), 7.59-7.81 (m, 4H), 7.22 (d, J=8.80 Hz, 2H), 7.13 (d, J=8.80 Hz, 2H), 6.93 (d, J=8.80 Hz, 2H), 5.60-5.74 (m, 1H), 5.36 (s, 1H), 4.99 (d, J=10.80 Hz, 1H), 4.82 (d, J=17.20 Hz, 1H), 4.68 (d, J=5.60 Hz, 2H), 3.64-3.74 (m, 4H), 3.46 (s, 4H), 3.28 (s, 1H), 3.03 (d, J=9.20 Hz, 2H), 2.66 (d, J=3.60 Hz, 2H), 2.42 (s, 1H), 2.29 (s, 2H), 2.07 (s, 2H), 1.62-1.86 (m, 7H), 1.46 (s, 6H).
A mixture of intermediate 149-1 (3 g, 10.7 mmol), tert-butyl piperazine-1-carboxylate (2.38 g, 12.8 mmol), RuPhos Pd G3 (446 mg, 534 μmol), Cs2CO3 (10.4 g, 32.0 mmol) in dioxane (40 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 100° C. for 16 hrs under N2 atmosphere. The mixture was filtered and washed with DCM (80 mL). Then the filtrate was concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/0 to 1/1) to give the intermediate 149-2 (1.14 g, 24.9% yield) as a pale yellow solid.
LCMS (ESI+): m/z 307.3 [M+H]+.
1H NMR (400 MHz, CHLOROFORM-d) δ 7.44-7.37 (m, 4H), 7.36-7.32 (m, 1H), 6.88 (t, J=9.2 Hz, 1H), 6.79-6.65 (m, 2H), 5.02 (s, 2H), 3.64-3.54 (m, 4H), 3.01-2.91 (m, 4H), 1.49 (s, 9H).
To a solution of intermediate 149-2 (1.14 g, 2.95 mmol) in THE (15 mL) and EtOH (15 mL) was added Pd/C (1 g, 10% purity) under N2. The suspension was degassed under vacuum and purged with H2 three times. The mixture was stirred under H2 (15 PSi) at 20° C. for 16 hours.
The reaction mixture was filtered, washed with DCM (100 mL) and the filterate was concentrated to the crude product. The crude product was triturated with MeCN (10 mL) and filted to give a cake of intermediate 149-3 (774 mg, 79.7% yield) as a white solid.
LCMS (ESI+): m/z 297.3 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 9.47 (br s, 1H), 6.89 (t, J=9.2 Hz, 1H), 6.61-6.46 (m, 2H), 3.43 (br s, 4H), 2.83-2.76 (m, 4H), 1.41 (s, 9H).
A mixture of intermediate 149-3 (200 mg, 675 μmol) in DMF (5 mL) was added NaH (67.5 mg, 1.69 mmol, 60% purity) at 0° C. under N2. The reaction was stirred at 0° C. for 30 mins. 3-bromopiperidine-2,6-dione (194 mg, 1.01 mmol) in DMF (0.5 mL) was added to the reaction at 0° C. under N2. The reaction was stirred at 0° C.-20° C. for 16 hrs. The reaction mixture was quenched by addition NH4Cl (10 mL) at 25° C., and then diluted with H2O (10 mL) and extracted with EA (30 mL) and extracted with EA two times. The combined organic layers were washed with sat. aq. NaCl (15 mL×2), dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=1/0 to 1/1) to give the intermediate 149-4 (160 mg, 48.9% yield) as colorless oil.
LCMS (ESI+): m/z 308.1 [M-Boc+H]+.
1H NMR (400 MHz, CHLOROFORM-d) δ 7.92-7.69 (m, 1H), 6.94-6.74 (m, 3H), 4.78 (m, 1H), 3.66-3.52 (m, 4H), 2.94 (br s, 5H), 2.75-2.63 (m, 1H), 2.38-2.27 (m, 2H), 1.49 (s, 9H).
To a solution of intermediate 149-4 (160 mg, 393 μmol) in DCM (1 mL) was added HCl/dioxane (4 M, 2 mL). The mixture was stirred at 25° C. for 16 hrs. The reaction mixture was concentrated under reduced pressure to give the crude product of intermediate 149-5 (132 mg, crude, HCl salt) as a brown solid.
LCMS (ESI+): m/z 308.1 [M+H]+.
To a mixture of intermediate 55-10 (144 mg, 262 μmol) and intermediate 149-5 (60 mg, 175 μmol, HCl salt) in DCM (3 mL) and DMF (1 mL) was added TEA (106 mg, 1.05 mmol) at 25° C. and stirred at 25° C. for 30 min, then AcOH (99.6 mg, 1.66 mmol) was added and stirred for 16 hrs. The reaction mixture was added NaBH(OAc)3 (111 mg, 524 μmol) at 25° C. and stirred at 25° C. for 16 hours. The reaction mixture was diluted with H2O (10 mL) and extracted with DCM (15 mL×3). The combined organic layers were dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by prep-HPLC (acetonitrile/water (2% FA)) to give compound 149 (53 mg, 36% yield) as a yellow solid.
LCMS (ESI+): m/z 805.5 [M+H]+.
1H NMR (400 MHz, DMSO-d6) δ 10.93 (br s, 1H), 10.26-10.00 (m, 1H), 8.82 (s, 1H), 8.04 (br s, 1H), 7.83-7.70 (m, 1H), 7.66-7.48 (m, 3H), 7.04-6.85 (m, 4H), 6.78 (d, J=8.4 Hz, 1H), 5.73-5.59 (m, 1H), 5.51-5.23 (m, 1H), 5.18-5.08 (m, 1H), 4.99 (d, J=10.4 Hz, 1H), 4.82 (d, J=17.2 Hz, 1H), 4.73-4.60 (m, 2H), 3.70-3.60 (m, 2H), 2.93 (br s, 4H), 2.70-2.58 (m, 4H), 2.42 (br s, 4H), 2.27-2.08 (m, 4H), 1.81 (d, J=12.0 Hz, 2H), 1.72-1.56 (m, 1H), 1.46 (s, 6H), 1.33-1.14 (m, 2H).
To a solution of Zn (10.0 g, 153 mmol) in THF (50 mL) was added 12 (6.35 g, 25.0 mmol) at 20° C., then tert-butyl 2-bromoacetate (6.42 g, 32.93 mmol) was added to the mixture. Then the solution of intermediate 159-1 (5 g, 25.09 mmol) in THE (20 mL) was added dropwise to the above mixture at 0° C. Then the reaction mixture was stirred at 100° C. for 16 hours. The reaction mixture was quenched by addition NaHCO3 (70 mL), and then diluted with EA (70 mL) and extracted with EA (60 mL×3), the combined organic layer was dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=2/1 to 0/1) to give the intermediate 159-2(6.81 g, crude) as a yellow solid, which was used into the next step directly.
To a solution of intermediate 159-2 (6.81 g, 21.6 mmol) in DCM (40 mL) was added TFA (12.3 g, 108 mmol). The mixture was stirred at 20° C. for 1 hr. The reaction mixture was filtered and concentrated under reduced pressure to give a crude of intermediate 159-3 (10.0 g, crude) as a red oil, which was used into the next step directly.
To a solution of intermediate 159-3 (10 g, 30.4 mmol) and 1,2-difluoro-4-nitro-benzene (4.83 g, 30.4 mmol) in DMSO (40 mL) was added TEA (6.15 g, 60.7 mmol). The mixture was stirred at 80° C. for 16 hr. The reaction mixture was partitioned between water (60 mL) and EA (20 mL×3). The organic phase was separated dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=5/1 to 3/1) to give the intermediate 159-4 (6.81 g, 63.3%) as a yellow solid. MS (ESI+): m/z 355.3
To a solution of intermediate 159-4 (6.75 g, 19.1 mmol) in THE (40 mL) was added Pd/C (10%, 5 g). The suspension was degassed and purged with H2 for 3 times. The mixture was stirred under H2 (15 Psi) at 20° C. for 16 hrs. The reaction mixture was filtered and concentrated under reduced pressure to give a crude of intermediate 159-5 (7.21 g, crude).
A mixture of intermediate 159-5 (7.21 g, 22.2 mmol), 3-bromopiperidine-2,6-dione (6.40 g, 33.3 mmol), NaHCO3 (5.60 g, 66.7 mmol) in DMF (20 mL) was degassed and purged with N2 for 3 times, and then the mixture was stirred at 80° C. for 16 hrs under N2 atmosphere. The reaction mixture was partitioned between water (10 mL) and DCM (90 mL). The organic phase was separated and dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether/Ethyl acetate=3/1 to 1/1) to give the intermediate 159-6 (7.46 g, 60.9% yield) as a green solid.
MS (ESI+): m/z 436.5 [M+H]+.
A solution of intermediate 159-6 (500 mg, 1.15 mmol) in FA (1.15 mmol, 10 mL) was stirred at 50° C. for 2 hrs. The reaction mixture was concentrated under reduced pressure to give a crude of the intermediate 159-7 (6.81 g crude) as a black solid, which was used into the next step directly.
To a mixture of intermediate 159-7 (50 mg, 118 umol) and intermediate 3 (60.25 mg, 117.53 umol) in DCM (2 mL) was added HOBt (23.82 mg, 176.30 umol) and EDCI (33.80 mg, 176.30 umol). The obtained mixture was stirred at 20° C. for 0.5 hr. Then Et3N (35.68 mg, 352.60 umol) was added to the mixture and the obtained mixture was stirred at 25° C. for 16 hours. The reaction mixture was partitioned between water (2 mL) and DCM (20 mL). The organic phase was separated and dried over Na2SO4, filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, DCM: MeOH=1/0 to 10/1) to give compound 159 (8 mg, 7.63% yield) as a white solid. MS (ESI+): m/z 874.4[M+H]+
1H NMR (400 MHz, DMSO-d6)10.79 (br s, 1H), 10.19 (br s, 1H), 8.83 (s, 1H), 7.93 (br s, 1H), 7.69 (m, 1H), 7.61 (br s, 2H), 6.96 (m, 2H), 6.85 (m, 1H), 6.49 (m, 1H), 6.41 (m, 1H), 5.79 (m, 1H), 5.72-5.60 (m, 1H), 5.08 (s, 1H), 4.99 (m, 1H), 4.90-4.71 (m, 3H), 4.57 (m, 1H), 4.30-4.19 (m, 1H), 3.68 (m, 4H), 3.59 (br s, 1H), 3.10 (m, 4H), 3.01-2.70 (m, 7H), 2.59-2.56 (m, 2H), 2.25-2.15 (m, 1H), 2.12-1.95 (m, 3H), 1.93-1.81 (m, 2H), 1.78-1.64 (m, 4H), 0.86 (in, 3H).
The following is an example of an assay that can be used to determine Wee1 degradation activity of compounds in the HT-1080 cell line. HT-1080 cell line was purchased from ATCC (Cat #CC-121), which was then engineered to stably express wild-type CRBN, GSPT (Δ1-138)/G575N mutant and HiBiT tagged Wee1. Cells were cultured in DMEM medium supplemented with 1000 heat-inactivated FBS. For Wee1 degradation assay, about 10,000 engineered HT-1080 cells in 35 μl culture medium were seeded into 384-well plates (Cat #3764, Corning), pre-spotted with compounds using the Echo 650 liquid handler (Beckman). The half-log dose-response-curve (DRC) typically has 10 points with the highest concentration at 10 μM, and the lowest concentration at 0.316 nM. Assay plates were incubated at 37° C. with 5% CO2 for 20 hours. Wee1 degradation was then assessed using the Nano-Glo HiBiT Lytic Detection Reagent according to the manufacturer's instructions. Luminescence was measured on a Pherastar microplate reader (BMG Labtech). The data was processed using Collaborative Drug Discovery Vault and DMSO control was used as the reference value to calculate degradation values for each treated sample (00 compared to DMSO control). A four Parameter Logistic Model was used to determine the compound's EC50 and DC50, using the following equation:
Dmax represented the maximum percentage of Wee1 degradation that could be achieved by a compound at its highest treatment concentration.
NCI-H1048 Cell Proliferation Assay (CTG Assay) The following is an example of an assay that can be used to determine the anti-proliferative activity of Wee1 PROTACs in a small cell lung cancer cell line, NCI-H1048 (ATCC Cat #CRL-5853). This cell line was cultured in RPMI-1640 medium supplemented with 10% FBS. About 1,200 NCI-H1048 cells in 50 μL of culture medium were seeded into a 384-well plate (Cat #3764, Corning), pre-spotted with compounds using the Echo 650 liquid handler (Beckman). The dose-response-curve (DRC) typically has 10-points with a highest concentration at 10 μM, and half-log diluted to as low as 0.316 nM, plus a DMSO control. Assay plates were incubated at 37° C. with 5% CO2 for 120 hours. After that, 20 μL of the CellTiter-Glo (CTG) Luminescent Cell Viability Assay Reagent (Cat #G7573, Promega) was added to each well and incubated at room temperature for 30 minutes. Luminescence was then measured with Pherastar microplate reader (BMG Labtech). All percentage of cell growth inhibition curves were processed using Collaborative Drug Discovery Vault.
Cell viability reads in compound-treated wells were normalized to that of DMSO control and expressed as Percent of Control (PoC, y). A four Parameter Logistic Model was used to determine the compound's EC50 and DC50, using the following equation:
Represented compounds shown in Table 2 exhibited excellent Wee1 degration activities and excellent anti-proliferative activities in H1048 cell line.
An aliquot of 30 μL stock solution (10 mM) was placed in order into their proper 96-well rack. 970 μL of PBS was added into each vial of the cap-less solubility sample plate. The assay was performed in duplicate. In each vial, a stir stick was added and sealed with a molded PTFE/Silicone plug. Then the solubility sample plate was transferred to the Eppendorf Thermomixer Comfort plate shaker, shaking at 25° C. at 1100 rpm for 2 hours. Plugs were removed after 2 hours and the stir sticks were removed using a big magnet. Then the samples were transferred from the solubility sample plate into the filter plate. All the samples were filtered using the vacuum manifold. An aliquot of 5 μL filtrate was mixed with 5 μL DMSO and 490 μL of a mixture of H2O and acetonitrile (v:v, 1:1) containing internal standard. A certain proportion of ultrapure water was used to dilute according to the peak shape. The dilution factor was changed according to the solubility values and the LC-MS/MS signals.
An aliquot of 6 μL stock solution (10 mM in DMSO) was transferred from standard (STD) stock plate into a new plate, and then 194 μL of DMSO was added to get a STD working solution of 300 μM. Then 5 μL of 300 μM working solution was mixed with 5 μL of DMSO and 490 μL of a mixture of H2O and acetonitrile (v:v, 1:1) containing internal standard and the final STD concentration was 3 μM. The STD concentration were changed according to the LC-MS/MS signals.
The sample plate was put into the autosampler and the test compound concentration was determined by LC-MS/MS.
The compounds of the present invention showed good kinetic solubility.
MDCKII (Madin-Darby Canine Kidney cell line, wild type) cell was obtained from the Netherlands Cancer Institute, which was used for P-gp substrates or inhibitors screening and predicting the permeability of compounds with high efflux barriers such as duodenum, blood-brain barrier, hepatocyte nucleus and renal unit. The MDCKII cells with 10 to 22 passages were used for the permeability investigation.
MDCKII cells were cultured using Minimum Essential Media (MEM) at 37° C., 5% CO2 and 95% relative humidity. The cells were seeded in Transwell plate with 5.45×105 cells/cm2. Then the cells were incubated in CO2 Incubator for 4-7 days for transport studies.
The complete medium was prepared by adding 10% FBS (fetal bovine serum) and 1% glutamax-1 to 500 ml MEM.
The information of reagent is shown below:
The dose concentrations of test compounds and control compounds were 1 μM. The bidirectional (A to B and B to A) permeability was evaluated. All incubations are performed in duplicate.
The regents were pre-incubated at 37±1° C. for 30 minutes. To determine the transport rate from apical to basolateral direction, 125 μL working solution (1 μM) was added to the Transwell insert (apical compartment), and 50 μL sample was immediately transferred from the apical compartment to a new 96-well plate as the initial donor sample (A-B) and mixed with 200 μL of acetonitrile or methanol containing IS. And 235 μL HBSS (10 mM HEPES, pH 7.4) was added to the receiver plate (basolateral compartment). To determine the transport rate from basolateral to apical direction, 285 μL working solution (1 μM) was added to the receiver plate (basolateral compartment), and 50 μL sample was immediately transferred from the basolateral compartment to a new 96-well plate as the initial donor sample (B-A) and mixed with 200 μL of acetonitrile or methanol containing IS. And 75 μL HBSS (10 mM HEPES, pH 7.4) was added to the Transwell insert (apical compartment). Then the plate was incubated at 37±° C. with 5% CO2 and 95% relative humidity for 120 minutes.
At the end of incubation, 50 μL samples from donor and receiver sides were transferred into a new 96-well plate. 200 μL of ice-cold acetonitrile or methanol containing appropriate internal standards (IS) was mixed with the samples. After vortex for 10 minutes, the samples are centrifuged at 3220 g for 40 minutes. An aliquot of 100 μL of the supernatant was mixed with appropriate volume of ultra-pure water and was analyzed by LC-MS/MS. All incubations are performed in duplicate.
The integrity of MDCKII cells were checked by Lucifer Yellow Rejection Assay after transport assay. The plate(s) were incubated with Lucifer Yellow solution at 37° C. for 30 minutes. The Lucifer Yellow fluorescence (to monitor monolayer integrity) was measured in a fluorescence plate reader at 480 nm excitation and 530 nm emission.
Test and reference compounds (metoprolol and digoxin) were semi-quantitatively analyzed, and the peak area ratio of the analyte and the internal standard was used to evaluate the concentration of the compounds.
The compounds of the present invention showed good permeability.
Objective: the objective of this study was to determine the pharmacokinetics of test compound following intravenous (IV) and oral (PO) administration to male CD1 mice.
Materials: Male CD1 mouse (20-30 g, 6-8 weeks, Zhejiang Vital River Laboratory Animal Technology Co., Ltd.).
The CD1 mice were administered with the test article intravenously via tail vein at 2 mg/kg or orally at 10 mg/kg. The animals had free access to food and water. After dose administration, blood was collected from the dorsal metatarsal vein at 5 min (0.0833 h, IV only), 15 min (0.25 h), 30 min (0.5 h), 60 min (1 h), 2 h, 4 h, 6 h, 8 h and 24 h. Blood samples were placed in an anticoagulant tube with EDTA-K2, mixed adequately and centrifuged at 4° C., 4000 g for 5 minutes to get the plasma. The plasma samples were stored at −75±15° C. until analyzed by LC-MS/MS. The PK parameters were calculated using non-compartmental model by Phoenix WinNonlin 6.1 (Linear/log trapezoidal).
The compounds of the present invention showed good oral exposure, low clearance rate and high bioavailability.
| Number | Date | Country | Kind |
|---|---|---|---|
| 202111322616.7 | Nov 2021 | CN | national |
| 202210360177.7 | Apr 2022 | CN | national |
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/CN2022/130700 | 11/8/2022 | WO |
