WO2024252276A1 - Pyrrolopyrimidine carboxamides - Google Patents

Pyrrolopyrimidine carboxamides Download PDF

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Publication number
WO2024252276A1
WO2024252276A1 PCT/IB2024/055444 IB2024055444W WO2024252276A1 WO 2024252276 A1 WO2024252276 A1 WO 2024252276A1 IB 2024055444 W IB2024055444 W IB 2024055444W WO 2024252276 A1 WO2024252276 A1 WO 2024252276A1
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Prior art keywords
compound
cancer
formula
pharmaceutically acceptable
pkmyt1
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English (en)
Inventor
Avipsa GHOSH
Jeffrey W. JOHANNES
Martin Packer
Symon GATHIAKA
Mark O'connor
Hadi REZAEI
Iain D. G. WATSON
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AstraZeneca AB
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AstraZeneca AB
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Priority to JP2025570865A priority Critical patent/JP2026521423A/ja
Priority to CN202480037555.5A priority patent/CN121311484A/zh
Priority to EP24734130.8A priority patent/EP4720066A1/fr
Publication of WO2024252276A1 publication Critical patent/WO2024252276A1/fr
Anticipated expiration legal-status Critical
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • the disclosure further relates to pharmaceutical compositions comprising such compounds and salts; use of such compounds and salts to treat or prevent cancers, including cancers having a PKMYT1-dependency because of elevated basal levels of replication stress; kits comprising such compounds and salts; and methods for manufacturing such compounds and salts.
  • CDK1 protein kinase CDK1
  • CDC2 cell division cycle 2 protein
  • WEE1 kinase family members WEE1 and PKMYT1 inhibit CDK1 to prevent the cell from dividing until the damaged DNA is repaired (G2/M DNA damage cell cycle checkpoint arrest).
  • PKMYT1 inhibitors have the potential to induce cancer cell death, either as a monotherapy in cancers with elevated levels of endogenous or basal replication stress, or in combination with other agents that induce greater levels of replication stress in cancers.
  • PKMYT1 inhibitors particularly PKMYT1 inhibitors having pharmacologically appropriate properties (such as selectivity against WEE1, bioavailability, etc.) required for suitable administration to a subject in need of such treatment.
  • the present disclosure addresses this unmet need by providing such compounds together with corresponding pharmaceutical compositions and methods for the treatment or prevention of cancers, including cancers where there is a PKMYT1-dependency because of elevated basal levels of replication stress, either as monotherapy or in combination therapies where such a PKMYT1 dependency can be induced with DNA damaging agents or other targeted therapies such as PARP inhibitors or ATR inhibitors.
  • the present disclosure provides compounds having the structure of Formula (1): (1), or a pharmaceutically acceptable salt thereof, wherein: R 1 is selected from the group consisting of: , , , , and ; R 2 , R 3 , R 4 , R 5 , R 7 , R 8 , R 9 , R 10 , and R 12 are independently selected from the group consisting of halogen and C 1-3 -alkyl; R 6 and R 11 are independently selected from the group consisting of hydrogen and C 1-3 - alkyl; A is selected from the group consisting of: , , , , , , , and ; R 13 and R 24 are independently selected from the group consisting of hydrogen, C 1-3 -alkyl, and C 1-3 -alkoxy; R 14 , R 17 , R 18 , R 20 , R 22 , and R 28 are independently selected the group consisting of: (a) hydrogen; (b) halogen; (c) cyano;
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (2), (3), (4), (5), (6), (7), (8), and (9) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (2-A), (2-B), (2-C), (2-D), and (2- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (3-A), (3-B), (3-C), (3-D), and (3- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (4-A), (4-B), (4-C), (4-D), and (4- E): as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (5-A), (5-B), (5-C), (5-D), and (5- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (6-A), (6-B), (6-C), (6-D), and (6- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (7-A), (7-B), (7-C), (7-D), and (7- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (8-A), (8-B), (8-C), (8-D), and (8- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides compounds wherein the structure of Formula (1) is selected from the group consisting of Formulae (9-A), (9-B), (9-C), (9-D), and (9- E) as further defined in this disclosure, and pharmaceutically acceptable salts thereof.
  • the present disclosure provides pharmaceutical compositions comprising a therapeutically-effective amount of a compound having the structure of Formula (1), or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
  • the present disclosure provides pharmaceutical compositions comprising therapeutically-effective amounts of a compound having the structure of Formula (1), or a pharmaceutically acceptable salt thereof; a second pharmacological agent; and a pharmaceutically acceptable carrier.
  • the present disclosure provides methods for treating or preventing cancers by administering a therapeutically effective amount of a compound having the structure of Formula (1), or pharmaceutically acceptable salt thereof, to a subject in need thereof.
  • the cancer is a solid tumor cancer.
  • the cancer is a hematological cancer.
  • the cancer has a PKMYT1-dependency because of elevated levels of replication stress.
  • the present disclosure provides compounds having the structure of Formula (1), or pharmaceutically acceptable salts thereof, for use as a medicament for treating or preventing cancers.
  • the cancer has a PKMYT1-dependency because of elevated levels of replication stress.
  • the present disclosure provides use of compounds having the structure of Formula (1), or pharmaceutically acceptable salts thereof, to prepare a medicament for treating or preventing cancers.
  • the cancer has a PKMYT1-dependency because of elevated levels of replication stress.
  • the present disclosure provides kits comprising a compound having the structure of Formula (1), or pharmaceutically acceptable salt thereof.
  • the present disclosure provides methods for preparing compounds having the structure of Formula (1), or pharmaceutically acceptable salts thereof. BRIEF DESCRIPTION OF THE DRAWINGS [0023] Fig.1 illustrates WEE1 and PKMYT1 regulation of the mitotic cell.
  • Fig.2 illustrates the effect of treatment with a PKMYT1 inhibitor tool compound (GI 50 ) on 16 cancer cell line models having either elevated or low levels of basal replication stress.
  • Fig.3 illustrates the replication fork velocities determined for the uterine and breast cancer cell line models of Fig.2 treated with the PKMYT1 inhibitor tool compound.
  • Fig.4 illustrates the replication fork velocities determined for the same uterine and breast cancer cell lines of Fig.2 that were treated instead with Example B-1, Isomer 1 (i.e., the eutomer of Compound 1).
  • Figure 5 illustrates the dose response curves obtained for Compound 1, Isomer 1 and Compound 137, Isomer 2 in an in vitro hematopoietic stem and progenitor cell assay assessing bone marrow toxicity.
  • Fig.6 illustrates the differential expression of phosphorylated proteins in OVCAR3 cells after 24-hour treatment with Compound 137, Isomer 2 or control (adavosertib).
  • Fig.7 illustrates (i) the effect of PKMYT1 deletion from SKOV3 cells on CDK1 T14 phosphorylation (left and middle panels) after treatment with Compound 1, Isomer 1, and (ii) the effect of treatment (GI 50 ) with Compound 137, Isomer 2 on PKMYT1 knock out SKOV3 cells (i.e., cells that do not express the drug target PKMYT1) (right panel).
  • Fig.8-A illustrates the tolerability in mice of a 300 mg/kg BID dosing regimen with Compound 137, Isomer 2 over a 28-day period.
  • Fig.8-B illustrates the tolerability in mice of a 100 mg/kg BID dosing regimen with Compound 137, Isomer 2 in combination with either irinotecan hydrochloride (left panel) or gemcitabine (right panel) over a 28-day period.
  • Fig.9 illustrates PKMYT1 substrate (CDK1) phosphorylation (on Thr14) in OVCAR3 tumor bearing SCID mice following dose scheduling with Compound 137, Isomer 2 (twice-daily dosing for 21 days).
  • Fig.10 illustrates the in vivo efficacy of Compound 137, Isomer 2 in combination with gemcitabine in a human ovarian OVCAR3 xenograft model.
  • Fig.11 illustrates in vivo efficacy of Compound 137, Isomer 2 in combination with irinotecan hydrochloride in a human colorectal SW620 xenograft model.
  • Fig.12 illustrates in vitro efficacy of Compound 137, Isomer 2, alone or in combination with 1 nM exatecan, in PKMYT1 wildtype and PKMYT1 knockout SKOV3 cells.
  • FIG.13 illustrates the effect on in vivo biomarker expression (phospho-CDK1 Thr14, gH2AX, and phospho-histone H3 Ser10) in a human colorectal SW620 xenograft model after treatment with irinotecan hydrochloride, either alone or in combination with Compound 137, Isomer 2 (100 mg/kg BID).
  • Figure 14 is a metabolism scheme showing the dominant routes of biotransformations (oxidation and glucuronidation) observed in vitro of Compound 137, Isomer 2.
  • Figure 15 shows the pharmacokinetic profiles in CD-1 mice (total blood plasma concentration) of Compound 137, Isomer 2 following administration either I.V.
  • Figure 16 shows the pharmacokinetic profiles in SCID mice (total blood plasma concentration) of Compound 137, Isomer 2 following administration P.O. at three dose levels: 10 mg/kg (solid line), 30 mg/kg (dashed line), and 100 mg/kg (dotted line).
  • Figure 17 shows the pharmacokinetic profiles in Han Wistar rats (total blood plasma concentration) of Compound 137, Isomer 2 following administration I.V. (0.5 mg/ kg) to either intact (solid line) or bile-duct cannulated (dashed line) animals.
  • Figure 18 shows the pharmacokinetic profiles in Han Wistar rats (total blood plasma concentration) of Compound 137, Isomer 2 following administration P.O. at three dose levels: 10 mg/kg (solid line), 30 mg/kg (dashed line), and 100 mg/kg (dotted line).
  • Figure 20 shows the pharmacokinetic profiles in SCID mice (total blood plasma concentration) of Compound 137, Isomer 2 following administration P.O.
  • halogen means a fluorine radical (which may be depicted as F), chlorine radical (which may be depicted as Cl), bromine radical (which may be depicted as Br), or iodine radical (which may be depicted as I).
  • alkyl alone or in combination with another term(s) means a straightor branchedchain saturated hydrocarbyl substituent (i.e., a substituent containing only carbon and hydrogen).
  • Alkyl typically contains from 1 to about 20 carbon atoms, more typically from 1 to about 12 carbon atoms, even more typically from 1 to about 8 carbon atoms, and still even more typically from 1 to about 6 carbon atoms.
  • substituents include methyl, ethyl, propyl (including npropyl and isopropyl), butyl (including n-butyl, isobutyl, sec-butyl, and tert- butyl), pentyl (including n-pentyl, isoamyl, and 2,2-dimethylpropyl), and hexyl.
  • alkenyl (alone or in combination with another term(s)) means a straight or branched chain saturated hydrocarbyl substituent (i.e., a substituent containing only carbon and hydrogen) containing one or more double bonds in the alkyl chain.
  • Alkenyl typically contains from 2 to about 20 carbon atoms, more typically from 2 to about 12 carbon atoms, even more typically from 2 to about 8 carbon atoms, and still even more typically from 2 to about 6 carbon atoms.
  • substituents include ethenyl, propenyl (including 1-propenyl and 2- propenyl), butyl (including 2-butenyl and 3-butenyl), pentenyl, and hexenyl.
  • alkynyl (alone or in combination with another term(s)) means a straight or branched chain saturated hydrocarbyl substituent (i.e., a substituent containing only carbon and hydrogen) containing one or more triple bonds in the alkyl chain.
  • Alkynyl typically contains from 2 to about 20 carbon atoms, more typically from 2 to about 12 carbon atoms, even more typically from 2 to about 8 carbon atoms, and still even more typically from 2 to about 6 carbon atoms.
  • substituents include ethynyl, propynyl (including 1-propynyl and 2- propynyl), butynyl (including 1butynyl and 2butynyl), pentynyl (including penta-1,3-diene, 2- methyl-1,3-butadiene, and penta-1,2-diene), and hexynyl.
  • cycloalkyl (alone or in combination with another term(s)) means a saturated carbocyclyl substituent containing from 3 to about 14 carbon ring atoms, more typically from 3 to about 12 carbon ring atoms, and even more typically from 3 to about 8 carbon ring atoms.
  • a cycloalkyl includes a single carbon ring, which typically contains from 3 to 6 carbon ring atoms.
  • singlering cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.
  • alkoxy (alone or in combination with another term(s)) means an alkylether substituent, i.e., alkyl-O-. Examples of alkoxy include methoxy (CH 3 -O-), ethoxy, n-propoxy, iso-propoxy, n-butoxy, iso-butoxy, sec-butoxy, and tert-butoxy.
  • alkoxyalkyl (alone or in combination with another term(s)) means alkyl substituted with alkoxy such as “methoxymethyl” which may be depicted as: .
  • heterocyclyl (alone or in combination with another term(s)) means a saturated, partially saturated, or completely unsaturated (i.e., “heteroaryl”) ring structure containing a total of 3 to 14 ring atoms. At least one of the ring atoms is a heteroatom (i.e., oxygen, nitrogen, or sulfur), with the remaining ring atoms being independently selected from the group consisting of carbon, oxygen, nitrogen, and sulfur in stable combinations known to those of skill in the art.
  • the number of carbon atoms in a substituent is indicated by the prefix “C x–y “, wherein x is the minimum and y is the maximum number of carbon atoms in the substituent.
  • C 1–6 alkyl refers to an alkyl substituent containing from 1 to 6 carbon atoms.
  • C 3-6 cycloalkyl refers to a cycloalkyl substituent containing from 3 to 6 carbon ring atoms.
  • the prefix “halo” indicates that the substituent to which the prefix is attached is substituted with one or more independently selected halogen radicals.
  • haloalkyl means an alkyl substituent wherein at least one hydrogen radical is replaced with a halogen radical. Where there is more than one hydrogen replaced with halogens, the halogens may be the identical or different.
  • haloalkyls include fluoromethyl, difluoromethyl, trifluoromethyl, difluoroethyl, 1,1,1trifluoroethyl, pentafluoroethyl, difluoropropyl, heptafluoropropyl chloromethyl, dichloromethyl, trichloromethyl, difluorochloromethyl, dichlorofluoromethyl, and dichloropropyl.
  • a substituent is “substitutable” if it comprises at least one carbon or nitrogen atom that is bonded to one or more hydrogen atoms. Thus, for example, hydrogen, halogen, and cyano do not fall within this definition.
  • a substituent is described as being “substituted,” a non-hydrogen radical is in the place of a hydrogen radical on a carbon or nitrogen of the substituent.
  • a substituted alkyl substituent is an alkyl substituent wherein at least one non-hydrogen radical is in the place of a hydrogen radical on the alkyl substituent.
  • monofluoroalkyl is alkyl substituted with a fluoro radical
  • difluoroalkyl is alkyl substituted with two fluoro radicals. It should be recognized that if there is more than one substitution on a substituent, each nonhydrogen radical may be identical or different (unless otherwise stated).
  • a substituent is described as “optionally substituted”, the substituent may be either (1) not substituted, or (2) substituted. If a carbon of a substituent is described as optionally substituted with one or more of a list of substituents, one or more of the hydrogens on the carbon (to the extent there are any) may separately and/or together be replaced with an independently selected optional substituent.
  • a nitrogen of a substituent is described as optionally substituted with one or more of a list of substituents, one or more of the hydrogens on the nitrogen (to the extent there are any) may each be replaced with an independently selected optional substituent.
  • substituents are described as being “independently selected” from a group, each substituent is selected independently of the other. Each substituent therefore may be identical to or different from the other substituent(s).
  • atropisomers refers to the stereoisomers resulting from hindered rotation about one or more single bonds, where the energy barrier to rotation is high enough to allow for the isolation of the conformers.
  • the term “eutomer” refers to the more pharmacologically active conformer of an atropisomer and the term “distomer” refers to the less pharmacologically active conformer of an atropisomer.
  • pharmaceutically acceptable is used adjectivally in this disclosure to mean that the modified noun is appropriate for use as a pharmaceutical product or as a part of a pharmaceutical product.
  • pharmaceutically acceptable salts are salts that are suitable for use in mammals, particularly humans, and include salts with an inorganic base, organic base, inorganic acid, organic acid, or basic or acidic amino acid that are suitable for use in mammals, particularly humans.
  • a “therapeutically effective amount” of a pharmacological agent is an amount that is sufficient to effect beneficial or desired results, including clinical results, and, as such, will depend upon the situation in which it is being administered.
  • a therapeutically effective amount of the agent is an amount of the agent that is sufficient, either alone or in combination with additional therapies, to provide an anti-cancer effect in a subject as compared to the response obtained without administration of the agent.
  • preventing is readily understood by an ordinarily skilled physician and, with respect to treatment of a particular condition, is intended to have its normal meaning and includes primary prophylaxis to prevent the development of the condition and secondary prophylaxis whereby the condition has already developed and the patient is temporarily or permanently protected against exacerbation or worsening of the disease or the development of new symptoms associated with the condition.
  • treating is readily understood by an ordinarily skilled physician and, with respect to treatment of a particular condition, can include (1) diminishing the extent or cause of the condition being treated, and/or (2) alleviating or ameliorating one or more symptoms associated with that condition.
  • Treatment of cancer can include stabilizing (i.e., not worsening), delaying, or slowing the spread or progression of the cancer; prolonging survival as compared to expected survival if not receiving treatment; and/or otherwise ameliorating or palliating the cancer or the severity of the cancer, in whole or in part.
  • the present disclosure provides compounds having the structure of Formula (1): (1), or a pharmaceutically acceptable salt thereof, wherein: R 1 is selected from the group consisting of: , , , , and ; R 2 , R 3 , R 4 , R 5 , R 7 , R 8 , R 9 , R 10 , and R 12 are independently selected from the group consisting of halogen and C 1-3 -alkyl; R 6 and R 11 are independently selected from the group consisting of hydrogen and C 1-3 - alkyl; A is selected from the group consisting of: , , , ,
  • R 13 and R 24 are independently selected from the group consisting of hydrogen, C 1-3 -alkyl, and C 1-3 -alkoxy;
  • R 14 , R 17 , R 18 , R 20 , R 22 , and R 28 are independently selected the group consisting of: (a) hydrogen; (b) halogen; (c) cyano; (d) C 1-6 -alkyl, wherein the C 1-6 -alkyl is optionally substituted with one or more substituents independently selected from halogen; (e) -C 1-6 -alkenyl, wherein the C 1-6 -alkenyl is optionally substituted with one or more substituents independently selected from halogen; (f) C 1-6 -alkynyl, wherein the C 1-6 -alkynyl is optionally substituted with one or more substituents independently selected from halogen; (g) C 3-6 -cycloalkyl, wherein the C 3-6 -cyclo
  • the structure of Formula (1) is selected from the group consisting of Formulae (2), (3), (4), (5), (6), (7), (8), and (9): (2), (3), (4), (5), (6), and (7), (8), and (9); and wherein R 1 through R 30 are as defined for the compounds having the structure of Formula (1).
  • the structure of Formula (1) is selected from the group consisting of Formulae (2-A), (2-B), (2-C), (2-D), and (2-E): (2-A), (2-B), (2-C), (2-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (3-A), (3-B), (3-C), (3-D), and (3-E): (3-A), (3-B), (3-C), (3-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (4-A), (4-B), (4-C), (4-D), and (4-E): (4-A), (4-B), (4-C), (4-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (5-A), (5-B), (5-C), (5-D), and (5-E): (5-A), (5-B), (5-C), (5-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (6-A), (6-B), (6-C), (6-D), and (6-E): (6-A), (6-B), (6-C), (6-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (7-A), (7-B), (7-C), (7-D), and (7-E): (7-A), (7-B), (7-C), (7-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (8-A), (8-B), (8-C), (8-D), and (8-E): (8-A), (8-B), (8-C), (8-D), and
  • the structure of Formula (1) is selected from the group consisting of Formulae (9-A), (9-B), (9-C), (9-D), and (9-E): (9-A), (9-B), (9-C), (9-D), and
  • the compound having the structure of Formula (1) is an atropisomer. In one aspect, the compound is the eutomer of the atropisomer. In another aspect, the compound is the distomer of the atropisomer.
  • the present disclosure provides compounds of Formula (1), and pharmaceutically acceptable salts thereof, wherein the compound is selected from the group consisting of: In some embodiments, the compound selected is an atropisomer. In one aspect, the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound that is 6-amino-4- cyclopropyl-7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-7H-pyrrolo[2,3-d]pyrimidine-5- carboxamide having the structure: (Compound 139), and pharmaceutically acceptable salts thereof.
  • the compound is an atropisomer.
  • the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound that is 6-amino-4- (2,2-difluorocyclopropyl)-7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-7H-pyrrolo[2,3- d]pyrimidine-5-carboxamide having the structure:
  • the compound is an atropisomer. In another aspect, the compound selected is the eutomer of the atropisomer. [0083] In one embodiment, the present disclosure provides a compound that is 6-amino-7-(2- chloro-3-hydroxy-6-methylphenyl)-2-methyl-4-(trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidine-5- carboxamide having the structure: (Compound 141), and pharmaceutically acceptable salts thereof. In one aspect, the compound is an atropisomer. In another aspect, the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound that is 6-amino-7-(6- chloro-3-hydroxy-2-methylphenyl)-2-methyl-4-(trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidine-5- carboxamide having the structure: (Compound 142), and pharmaceutically acceptable salts thereof.
  • the compound is an atropisomer.
  • the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound that is 6-amino-7- (2,6-dichloro-3-hydroxyphenyl)-2-methyl-4-(trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidine-5- carboxamide having the structure: (Compound 143), and pharmaceutically acceptable salts thereof.
  • the compound is an atropisomer.
  • the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound that is 6-amino-7-(6- bromo-3-hydroxy-2-methylphenyl)-2-methyl-4-(trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidine- 5-carboxamide having the structure: (Compound 144), and pharmaceutically acceptable salts thereof.
  • the compound is an atropisomer.
  • the compound selected is the eutomer of the atropisomer.
  • the present disclosure provides a compound selected from the group consisting of: 6-amino-7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-4-(trifluoromethyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 137); 6-amino-7-(2-chloro-3-hydroxy-6-methylphenyl)-2-methyl-4-(trifluoromethyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 141); 6-amino-7-(6-chloro-3-hydroxy-2-methylphenyl)-2-methyl-4-(trifluoromethyl)-7H- pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 142); 6-amino-7-(2,6-dichloro-3-hydroxyphenyl)-2-methyl-4-(trifluoromethyl)-7H-pyrrolo[2,3- d]pyrimidine-5-carboxamide (Compound
  • the compound is an atropisomer. In another aspect, the compound selected is the eutomer of the atropisomer.
  • C. Combination of Embodiments Any embodiment of the compounds described in the present disclosure can be combined with any other suitable embodiment described herein to provide additional embodiments. For example, where one embodiment of the compounds of Formula (2) individually or collectively describes possible groups for R 13 , R 14 , and/or R 15 and a separate embodiment describes possible groups for R 1 , it is understood that these embodiments can be combined to provide an additional embodiment describing the possible groups described for R 13 , R 14 , and/or R 15 together with the possible groups described for R 1 .
  • the R 1 substituent can be as defined in any of the embodiments of R 1 described in this disclosure.
  • the compounds of the present disclosure have an IC 50 value for inhibition of PKMYT1 below about 1 ⁇ M as measured in the Target Engagement Assay described in Example C-2 below.
  • the IC 50 value is below about 500 nM.
  • the IC 50 value is below about 250 nM.
  • the IC 50 value is below about 100 nM.
  • the compounds have a pharmaceutically acceptable K d value for PKMYT1 binding affinity as measured in the SPR Direct Binding Assay described in Example C-3 below.
  • the compounds of the present disclosure have a pharmaceutically acceptable selectivity for PKMYT1 relative to WEE1 as measured in the Target Engagement Assay described in Example C-2 below.
  • the compounds are at least about 50 times more selective for PKMYT1 relative to WEE1.
  • the compounds are at least about 100 times more selective for PKMYT1 relative to WEE1.
  • the compounds are at least about 250 times more selective for PKMYT1 relative to WEE1.
  • the compounds are at least about 500 times more selective for PKMYT1 relative to WEE1.
  • the compounds of the present disclosure have an IC 50 value for inhibition of WEE1 greater than about 1 ⁇ M. In another aspect, the IC 50 value is greater than about 5 ⁇ M. In another aspect, the IC 50 value is greater than about 10 ⁇ M.
  • the compounds of the present disclosure have an IC 50 value for inhibition of CDK1 phosphorylation below about 2 ⁇ M as measured in the Functional CDK1 pThr14 AlphaLISA Assay described in Example C-4 below. In one aspect, the IC 50 value is below about 1 ⁇ M. In another aspect, the IC 50 value is below about 500 nM.
  • the IC 50 value is below about 250 nM. In another aspect, the IC 50 value is below about 100 nM.
  • the compounds of the present disclosure inhibit OVCAR3 cellular proliferation as measured in the Cellular Proliferation Assay described in Example C-5 below. In one aspect, the compounds have a GI 50 value below about 5 ⁇ M. In another aspect, the GI 50 value is below about 3 ⁇ M. In another aspect, the GI 50 value is below about 1.5 ⁇ M.
  • the compounds of the present disclosure may exist in salt form or in non-salt form (i.e., as a free base), and the present disclosure covers both salt forms and non-salt forms.
  • the compounds may form acid addition salts or base addition salts.
  • an acid addition salt can be prepared using various inorganic or organic acids. Such salts can typically be formed by, for example, mixing the compound with an acid (e.g., a stoichiometric amount of an acid) using various methods known in the art. This mixing may occur in water, an organic solvent (e.g., ether, ethyl acetate, ethanol, methanol, isopropanol, or acetonitrile), or an aqueous/organic mixture.
  • the acid addition salts are, for example, trifluoroacetate, formate, acetate or hydrochloric.
  • a base addition salt can be prepared using various inorganic or organic bases, for example an alkali or alkaline earth metal salt such as a sodium, calcium or magnesium salt, or other metal salts, such as potassium or zinc, or an ammonium salt, or a salt with an organic base such as methylamine, dimethylamine, trimethylamine, piperidine or morpholine.
  • an alkali or alkaline earth metal salt such as a sodium, calcium or magnesium salt, or other metal salts, such as potassium or zinc, or an ammonium salt
  • an organic base such as methylamine, dimethylamine, trimethylamine, piperidine or morpholine.
  • the skilled person will be aware of the general principles and techniques of preparing pharmaceutical salts, such as those described in, for example, Berge, S., et al., “Pharmaceutical Salts,” J. Pharm. Sci.66, 1 (1977).
  • the compounds and salts of the present disclosure may exist in one or more geometrical, optical, enantiomeric, and diastereomeric forms, including, but not limited to, cis- and trans-forms, E- and Z-forms, and R-, S- and meso-forms. Unless otherwise stated a reference to a particular compound includes all such isomeric forms, including racemic and other mixtures thereof.
  • a single stereoisomer is obtained by isolating it from a mixture of isomers (e.g., a racemate) using, for example, chiral chromatographic separation.
  • a single stereoisomer is obtained through direct synthesis from, for example, a chiral starting material.
  • a particular enantiomer of a compound described herein may be more active than other enantiomers of the same compound.
  • the compound, or a pharmaceutically acceptable salt thereof is a single enantiomer being in an enantiomeric excess (%ee) of ⁇ 90, ⁇ 95%, ⁇ 96%, ⁇ 97, ⁇ 98% or ⁇ 99%.
  • the single enantiomer is present in an enantiomeric excess (%ee) of ⁇ 99%.
  • the present disclosure relates to a pharmaceutical composition
  • a pharmaceutical composition comprising a compound, or a pharmaceutically acceptable salt thereof, which is a single enantiomer being in an enantiomeric excess (%ee) of ⁇ 90, ⁇ 95%, ⁇ 96%, ⁇ 97, ⁇ 98% or ⁇ 99%, or a pharmaceutically acceptable salt thereof, in association with one or more pharmaceutically acceptable excipients.
  • the single enantiomer is present in an enantiomeric excess (%ee) of ⁇ 99%.
  • G. Additional Forms [0097]
  • the compounds and salts of the present disclosure may exist in various tautomeric forms and the disclosure encompasses all such tautomeric forms.
  • “Tautomers” are structural isomers that exist in equilibrium resulting from the migration of a hydrogen atom.
  • the compounds of the present disclosure, and pharmaceutically acceptable salts thereof may exist as solvates (such as a hydrates) as well as unsolvated forms, and the present disclosure covers all such solvates.
  • the compounds of the present disclosure, and pharmaceutically acceptable salts thereof may exist in crystalline or amorphous form, and the present disclosure covers all such forms.
  • Compounds and salts of the present disclosure may be isotopically labeled (or “radio- labeled”). In that instance, one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number typically found in nature.
  • the disclosure encompasses isotopically labelled forms of compounds disclosed herein.
  • isotopes that may be incorporated include 2 H (also written as “D” for deuterium), 3 H (also written as “T” for tritium), 11 C, 13 C, 14 C, 13 N, 15 N, 15 O, 17 O, 18 O and 36 Cl.
  • the isotope that is used will depend on the specific application of that radio-labeled derivative. For example, for in vitro receptor labeling and competition assays, 3 H or 14 C are often useful. For radio-imaging applications, 11 C is often useful.
  • the radionuclide is 3 H.
  • the radionuclide is 14 C.
  • the radionuclide is 11 C.
  • the present disclosure provides additional compounds that are useful as intermediates for preparing the compounds of the present disclosure, and pharmaceutically acceptable salts thereof.
  • III. Methods of Use One consequence of cancer cell evolution is often the rewiring of the ability to control cell cycle progression which then leads to more rapid oncogenic expansion to aid oncogenesis. More than 50% of all cancers, and over 90% of cancers such as serous-like endometrial, serous ovarian, and basal-like breast cancers, carry a somatic TP53 mutation that can result in inactivation of the tumor suppressor protein p53 and partial or complete inactivation of the G1/S cell cycle checkpoint.
  • WEE1 and PKMYT1 inhibit CDK1 to prevent a cell from dividing until damaged DNA is repaired (G2/M DNA damage cell cycle checkpoint arrest).
  • PKMYT1 is selective for CDK1 relative to CDK2.
  • WEE1 also regulates CDK2 activity that normally prevents aberrant progression of cells through S-phase.
  • WEE1 inhibition can further increase replication stress (as detected by biomarkers such as phospho-RPA2 (pRPA2) and phospho-Histone H2AX ( ⁇ H2AX)) in cancers that already have elevated basal levels of replication stress, creating additional DNA damage during S-phase.
  • biomarkers such as phospho-RPA2 (pRPA2) and phospho-Histone H2AX ( ⁇ H2AX)
  • pRPA2 phospho-RPA2
  • ⁇ H2AX phospho-Histone H2AX
  • Such cancer backgrounds include those enriched for p53 mutations and/or one or more aberrations in genes and/or their related protein products including, but are not limited to, cyclin D1, cyclin E1, FBXW7, RB1, INK4a, ARF, CDK4, CDK6, SETD2, KDM4A, LKB1, TSC2, RSP6KA6, MYC, and/or KRAS.
  • cancer backgrounds are particularly enriched in tumor types such as ovarian cancer, triple negative breast cancer, uterine serous carcinoma, uterine carcinosarcoma, pancreatic cancer, small cell lung cancer, non-small cell lung cancer, esophageal cancer, colorectal cancer, soft tissue sarcoma, skin cancer, bladder cancer, head and neck cancer, glioma, and B-cell lymphoma.
  • tumor types harboring these deficiencies can be sensitive to monotherapy with WEE1 inhibitors in the clinic, the fact that WEE1 inhibition also generates S-phase replication stress and DNA damage means that certain normal tissues are also affected resulting in gastrointestinal and hematological toxicities. Consequently, monotherapy with WEE1 inhibitors can have a relatively narrow therapeutic window. Liu, J.
  • PKMYT1 regulates CDK1 and not CDK2
  • PKMYT1 inhibitors have the potential to treat cancers with elevated levels of basal replication stress, either as a single agent or in combination with DNA damaging agents or targeted agents that increase the levels of replication stress, but with an improved therapeutic window relative to WEE1 inhibitors.
  • cancer backgrounds associated with replication stress that are sensitive to PKMYT1 inhibition include those enriched for one or more aberrations in genes and/or their related protein products including, but are not limited to, cyclin E1, FBXW7, and/or KRAS.
  • cancer backgrounds associated with replication stress that are sensitive to PKMYT1 inhibition may include those enriched for TP53 mutations and/or one or more aberrations in genes and/or their related protein products including, but are not limited to, cyclin D1, cyclin E1, FBXW7, RB1, INK4a, ARF, CDK4, CDK6, SETD2, KDM4A, LKB1, TSC2, RSP6KA6, MYC, and/or KRAS.
  • PKMYT1 inhibitors that are selective for CDK1 are desirable and are within the scope of this disclosure.
  • the compounds of Formula (1), and pharmaceutically acceptable salts thereof are inhibitors of PKMYT1 activity. In one aspect, the compounds selectively inhibit PKMYT1 activity. [00107] In some embodiments, the present disclosure provides a method for treating or preventing cancers in a subject in need thereof by administering to the subject a therapeutically effective amount of a compound of Formula (1), or a pharmaceutically acceptable salt thereof. In one aspect, the cancer is a solid tumor cancer.
  • the solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), pancreatic cancer, lung cancer (including small cell lung cancer and non-small cell lung cancer), esophageal cancer, colorectal cancer, soft tissue sarcoma, skin cancer, bladder cancer, head and neck cancer, and glioma.
  • the solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), and pancreatic cancer.
  • the cancer is a hematological cancer.
  • the hematological cancer is B-cell lymphoma.
  • the present disclosure provides a method for treating or preventing cancers in a subject in need thereof by administering to the subject a therapeutically effective amount of a compound of Formula (1), or a pharmaceutically acceptable salt thereof, wherein the cancer is, in whole or in part, a PKMYT1-dependent cancer.
  • the PKMYT1-dependent cancer is a solid tumor cancer.
  • the PKMYT1-dependent solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), pancreatic cancer, lung cancer (including small cell lung cancer and non-small cell lung cancer), esophageal cancer, colorectal cancer, soft tissue sarcoma, skin cancer, bladder cancer, head and neck cancer, and glioma.
  • the PKMYT1- dependent solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), and pancreatic cancer.
  • the PKMYT1- dependent cancer is a hematological cancer.
  • the PKMYT1-dependent hematological cancer is B-cell lymphoma.
  • the present disclosure provides a method for treating or preventing cancers in a subject in need thereof by administering to the subject a therapeutically effective amount of a compound of Formula (1), or a pharmaceutically acceptable salt thereof, wherein the cancer is associated with a higher incidence of genetic or protein aberrations associated with elevated levels of basal replication stress.
  • the cancer is characterized by amplification or overexpression of the gene.
  • the cancer is characterized by one or more deletions and/or mutations in the gene.
  • the cancer is a solid tumor cancer.
  • the solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), pancreatic cancer, lung cancer (including small cell lung cancer and non-small cell lung cancer), esophageal cancer, colorectal cancer, soft tissue sarcoma, skin cancer, bladder cancer, head and neck cancer, and glioma.
  • the solid tumor cancer is selected from the group consisting of ovarian cancer, breast cancer (including triple negative breast cancer), uterine cancer (including uterine serous carcinoma and uterine carcinosarcoma), and pancreatic cancer.
  • the cancer is a hematological cancer.
  • the hematological cancer is B-cell lymphoma.
  • the present disclosure provides a method for treating or preventing cancers in a subject in need thereof by administering to the subject a therapeutically effective amount of a compound of Formula (1), or a pharmaceutically acceptable salt thereof, wherein the cancer is associated with one or more aberrations in a gene, and/or its related protein product, selected from the group consisting of cyclin E1, FBXW7, and/or KRAS.
  • the cancer is associated with one or more aberrations in the cyclin E1 gene (CCNE1) and/or its related protein product.
  • the cancer is characterized by amplification or overexpression of CCNE1.
  • the cancer is associated with one or more aberrations in the FBXW7 gene and/or its related protein product. In another aspect, the cancer is characterized by one or more deletions and/or mutations in the FBXW7 gene. In another aspect, the cancer is associated with one or more aberrations in the KRAS gene and/or its related protein product. In another aspect, the cancer is characterized by one or more deletions and/or mutations in the KRAS gene. [00111] In some embodiments, the cancer is ovarian cancer. In one aspect, the ovarian cancer is a PKMYT1-dependent ovarian cancer. In another aspect, the cancer is a platinum-sensitive or platinum-resistant ovarian cancer.
  • the cancer is breast cancer.
  • the breast cancer is a PKMYT1-dependent breast cancer.
  • the breast cancer is selected from the group consisting of hormone receptor positive (HR+) breast cancer, hormone receptor negative (HR-) breast cancer, and triple negative breast cancer.
  • the breast cancer is a chemotherapy-resistant breast cancer.
  • the breast cancer is a radiotherapy- resistant breast cancer.
  • the breast cancer is an advanced or metastatic breast cancer.
  • the cancer is uterine cancer.
  • the uterine cancer is a PKMYT1-dependent uterine cancer.
  • the uterine cancer is uterine serous carcinoma.
  • the uterine cancer is uterine carcinosarcoma.
  • the cancer is pancreatic cancer. In one aspect, the pancreatic cancer is a PKMYT1-dependent pancreatic cancer.
  • the cancer is lung cancer. In one aspect, the lung cancer is a PKMYT1-dependent lung cancer. In another aspect, the lung cancer is small cell lung cancer (SCLC). In another aspect, the lung cancer is non-small cell lung cancer (NSCLC). In another aspect, the non-small cell lung cancer (NSCLC) is squamous cell carcinoma. In another aspect, the non-small cell lung cancer (NSCLC) is adenocarcinoma.
  • the non-small cell lung cancer is large-cell carcinoma.
  • the cancer is gastric cancer.
  • the gastric cancer is a PKMYT1-dependent gastric cancer.
  • the cancer is esophageal cancer.
  • the esophageal cancer is a PKMYT1-dependent esophageal cancer.
  • the present disclosure provides a method for treating or preventing a hematological cancer in a subject in need thereof by administering to the subject a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, wherein the hematological cancer is selected from the group consisting of non-Hodgkin's lymphoma, leukemia, multiple myeloma (MM), and myelodysplastic syndrome (MDS).
  • the hematological cancer is a PKMYT1-dependent hematological cancer.
  • the hematological cancer is non-Hodgkin's lymphoma (NHL).
  • the non-Hodgkin's lymphoma is selected from diffuse large B-cell lymphoma (DLBCL), follicular lymphoma, mantle cell lymphoma (MCL), and marginal zone lymphoma.
  • the hematological cancer is leukemia.
  • the leukemia is selected from the group consisting of acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML).
  • ALL acute lymphoblastic leukemia
  • AML acute myelogenous leukemia
  • CLL chronic lymphocytic leukemia
  • CML chronic myeloid leukemia
  • the hematological cancer is multiple myeloma (MM).
  • the hematological cancer is myelodysplastic syndrome (MDS).
  • the hematological cancer is diffuse large B-cell lymphoma (DLBCL).
  • DLBCL diffuse large B-cell lymphoma
  • the compound of the present disclosure, or a pharmaceutically acceptable salt thereof is administered as first line therapy.
  • the compound of the present disclosure, or a pharmaceutically acceptable salt thereof is administered as second line (or later) therapy.
  • the subject to whom a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, is administered exhibits a partial response (PR) in response to such treatment.
  • PR partial response
  • the subject to whom a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, is administered exhibits a complete response (CR) in response to such treatment.
  • the subject to whom a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, is administered exhibits an improved progression free survival (PFS) in response to such treatment.
  • PFS progression free survival
  • the subject to whom a therapeutically effective amount of a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, is administered exhibits an improved overall survival (OS) in response to such treatment.
  • OS overall survival
  • the subject treated typically will be a human or non-human mammal, particularly a human.
  • Suitable subjects can also include domestic or wild animals; companion animals (including dogs, cats, and the like); livestock (including horses, cows and other ruminants, pigs, poultry, rabbits, and the like); primates (including monkeys such as rhesus monkeys, cynomolgus (also known as crab-eating or long-tailed) monkeys, marmosets, tamarins, chimpanzees, macaques, and the like); and rodents (including rats, mice, gerbils, guinea pigs, and the like).
  • the present disclosure provides the compounds of Formula (1), or pharmaceutically acceptable salts thereof, for use as medicaments.
  • the present disclosure provides for the use of the compounds of the Formula (1), or pharmaceutically acceptable salts thereof, for treating or preventing cancers, including PKMYT1-dependent cancers as discussed above.
  • the present disclosure provides for the use of the compounds of the Formula (1), or pharmaceutically acceptable salts thereof, for the manufacture of medicaments for treating or preventing cancer, including PKMYT1-dependent cancers as discussed above. IV.
  • the compounds and pharmaceutically acceptable salts of the present disclosure may be used in the methods described above as either as single pharmacological agents or in combination with other pharmacological agents or techniques including, for example, DNA damaging agents that increase reliance on the G2/M checkpoint and targeted therapies that induce higher levels of replication stress.
  • Such combination therapies may be achieved by way of the simultaneous, sequential, or separate dosing of the individual components of the treatment.
  • These combination therapies employ the compounds and pharmaceutically acceptable salts of the present disclosure within the dosage ranges described in this application and the other pharmacological agent(s), typically within its approved dosage range(s).
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and radiotherapy.
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and chemotherapy.
  • the chemotherapy is induction chemotherapy.
  • the chemotherapy is consolidation chemotherapy.
  • the chemotherapy comprises administration of one or more chemotherapeutics selected from the group consisting of an anthracycline, azacytidine, bleomycin, cisplatin, carboplatin, cyclophosphamide, cytarabine, dactinomycin, daunorubicin, docetaxel, doxorubicin, epirubicin, fluorouracil, gemcitabine, idarubicin, mercaptopurine, methotrexate, oxaliplatin, paclitaxel, thioguanine, and/or vincristine.
  • the chemotherapy comprises administration of a nucleoside analog.
  • the nucleoside analog is selected from the group consisting of cytarabine and gemcitabine.
  • the chemotherapy comprises administration of a taxane.
  • the taxane is selected from the group consisting of paclitaxel, docetaxel, and nab- paclitaxel.
  • the chemotherapy comprises administration of a platinum chemotherapeutic.
  • the platinum chemotherapeutic is selected from the group consisting of carboplatin and cisplatin.
  • the chemotherapy further comprises administration of a steroid.
  • the steroid is selected from the group consisting of prednisolone, dexamethasone, and hydrocortisone.
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and a topoisomerase 1 (TOP1) inhibitor, topoisomerase 2 (TOP2) inhibitor, TOP1-antibody drug conjugate (TOP1-ADC), or TOP2-antibody drug conjugate (TOP2-ADC).
  • TOP1 inhibitor is selected from the group consisting of irinotecan and topotecan.
  • the TOP1-ADC is trastuzumab deruxtecan.
  • the TOP2 inhibitor is selected from the group consisting of etoposide and teniposide.
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and a poly ADP ribose polymerase (PARP) inhibitor.
  • PARP poly ADP ribose polymerase
  • the PARP inhibitor is selected from the group consisting of olaparib, rucaparib, niraparib, talazoparib, AZD5305 (CAS No.2589531-76-8), and AZD9574 (CAS No.2756333-39-6).
  • the PARP inhibitor is olaparib.
  • the PARP inhibitor is AZD5305.
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and an ataxia telangiectasia mutated and Rad3-related kinase (ATR) inhibitor.
  • the ATR inhibitor is selected from the group consisting of camonsertib (Repare), tuvusertib (Merck), berzosertib (Merck KGaA), ceralasertib (AstraZeneca), and elimusertib (Bayer).
  • the ATR inhibitor is ceralasertib.
  • the present disclosure provides a combination suitable for use in the treatment or prevention of a PKMYT1-dependent cancer, wherein the combination comprises a compound of the present disclosure, or a pharmaceutically acceptable salt thereof, and a checkpoint kinase 1 (CHK1) inhibitor.
  • the CHK1 inhibitor is selected from the group consisting of bosutinib, crizotinib, dasatinib, erlotinib, gefitinib, lapatinib, pazopanib, ruxolitinib, sunitinib, and vemurafenib. V.
  • compositions comprising one or more pharmaceutically acceptable excipients. Therefore, in some embodiments the present disclosure provides pharmaceutical compositions comprising a compound of Formula (1), or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable excipient.
  • excipient(s) selected for inclusion in a particular composition will depend on factors such as the mode of administration and the form of the composition provided. Suitable pharmaceutically acceptable excipients are described, for example, in the Handbook of Pharmaceutical Excipients, Sixth Edition, Pharmaceutical Press, edited by Rowe, Ray C; Sheskey, Paul J; Quinn, Marian.
  • compositions may function as, for example, adjuvants, diluents, carriers, stabilisers, flavourings, colorants, fillers, binders, disintegrants, lubricants, glidants, thickening agents and coating agents.
  • certain pharmaceutically acceptable excipients may serve more than one function and may serve alternative functions depending on how much of the excipient is present in the composition and what other excipients are present in the composition.
  • compositions may be in a form suitable for oral use (for example as tablets, lozenges, hard or soft capsules, aqueous or oily suspensions, emulsions, dispersible powders or granules, syrups or elixirs), for topical use (for example as creams, ointments, gels, or aqueous or oily solutions or suspensions), for administration by inhalation (for example as a finely divided powder or a liquid aerosol), for administration by insufflation (for example as a finely divided powder) or for parenteral administration (for example as a sterile aqueous or oily solution for intravenous, subcutaneous or intramuscular dosing), or as a suppository for rectal dosing.
  • oral use for example as tablets, lozenges, hard or soft capsules, aqueous or oily suspensions, emulsions, dispersible powders or granules, syrups or elixirs
  • topical use
  • compositions intended for oral use may contain, for example, one or more coloring, sweetening, flavoring and/or preservative agents.
  • the total daily dose will necessarily be varied depending upon the subject treated, the route of administration, any therapies being co-administered, and the severity of the illness being treated, and may include single or multiple doses. Specific dosages can be adjusted, for example, depending upon the condition being treated; the age, body weight, general health condition, sex, and diet of the subject; administration routes; dose intervals; excretion rate; and other drugs being co-administered to the subject.
  • the compound of Formula (1), or a pharmaceutically acceptable salt thereof typically will be administered to a warm-blooded animal at a unit dose within the range 2.5 to 5000 mg/m 2 body area of the animal, or approximately 0.05 to 100 mg/kg, and this normally provides a therapeutically effective dose.
  • the present disclosure provides pharmaceutical compositions for use in therapy, comprising a compound of Formula (1), or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable excipient.
  • the present disclosure provides pharmaceutical compositions for use in the treatment of cancers, comprising a compound of Formula (1), or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable excipient.
  • the cancer is a PKMYT1-dependent cancer.
  • the PKMYT1-dependent cancer is a solid tumor cancer.
  • PKMYT1-dependent cancer is a hematological cancer.
  • the kit comprises a unit dosage form comprising a compound of Formula (1), or a pharmaceutically acceptable salt thereof, contained within a packaging material and a label or package insert which indicates that the pharmaceutical composition can be used for treating a cancer.
  • the cancer is a PKMYT1-dependent cancer.
  • the PKMYT1-dependent cancer is a hematological malignancy.
  • the PKMYT1-dependent cancer is a solid tumor cancer.
  • kit comprises: (a) a first unit dosage form comprising a compound of Formula (1), or a pharmaceutically acceptable salt thereof; (b) a second unit dosage form comprising a pharmacological agent selected from the group consisting of chemotherapeutics, TOP1 inhibitors, TOP2 inhibitors, TOP1-ADCs, TOP2-ADCs, PARP inhibitors, ATR inhibitors, and CHK1 inhibitors; (c) a container means for containing said first and second dosage forms; and (d) a label or package insert which indicates that the first unit dosage form and second unit dosage form can be used for treating a PKMYT1-dependent cancer. VII.
  • Scheme 1 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (B) (X, Y are halogens like fluoro, chloro, bromo, or iodo) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (D).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • a solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • a compound of formula (D) may be reacted with malononitrile of formula (E) to give a compound of formula (F).
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl2(dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable solvent such as DME, etc.
  • a compound of formula (F) may be reacted with concentrated sulfuric acid to give a compound of formula (G).
  • the protecting group in the compound of formula (G) may be removed under appropriate reaction conditions to give a compound of Formula (3).
  • Scheme 2 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (H) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (I).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • a compound of formula (I) may be reacted with malononitrile of formula (E) to give a compound of formula J.
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl2(dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable solvent such as DME, etc.
  • a compound of formula J may be reacted with concentrated sulfuric acid to give a compound of formula (K).
  • the protecting group in the compound of formula (K) may be removed under appropriate reaction conditions to give a compound of Formula (2).
  • Scheme 3 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (L) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (M).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • a compound of formula (M) may be reacted with malononitrile of formula (E) to give a compound of formula (N).
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable solvent such as DME, etc.
  • a compound of formula (N) may be reacted with concentrated sulfuric acid to give a compound of formula (O).
  • the protecting group in the compound of formula (O) may be removed under appropriate reaction conditions to give a compound of Formula (4).
  • Scheme 4 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (P) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (Q).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • a compound of formula (Q) may be reacted with malononitrile of formula (E) to give a compound of formula (R).
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • DME dimethyl methyl
  • a compound of formula (R) may be reacted with concentrated sulfuric acid to give a compound of formula (S).
  • the protecting group in the compound of formula (S) may be removed under appropriate reaction conditions to give a compound of Formula (5).
  • Scheme 5 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (T) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (U).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • a compound of formula (U) may be reacted with malononitrile of formula (E) to give a compound of formula (V).
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • a compound of formula (A1) may be reacted with boronic acid or ester of formula (B1) to give a compound of Formula (3).
  • the reaction may be catalyzed with a suitable Pd- precatalyst (such as XantPhos Pd G3, XPhos Pd G3, etc.) and ligand (such as XPhos, etc.) in the presence of a base (such as K 2 CO 3 , NaOtBu, Cs 2 CO 3 , etc.) using a suitable solvent (1,4- dioxane/water mixture, toluene/water mixture, etc.), and at temperatures typically ranging from 60°C to 120°C.
  • a compound of formula (A1) may also be reacted with alkyl or aryl amine of formula (C1) to give a compound of Formula (3).
  • the reaction may be catalyzed with a suitable Pd- precatalyst (such as Pd(dba)2, Pd-PEPPSI-iHeptCl, BrettPhos Pd G3, etc.) and ligand (such as Xphos, Cphos, BrettPhos, etc.) in the presence of a base (such as LiHMDS, etc.) using a suitable solvent (such as 2-Me THF, 1,4-dioxane, t-BuOH, etc.) and at temperatures typically ranging from 80°C to 120°C.
  • a suitable Pd- precatalyst such as Pd(dba)2, Pd-PEPPSI-iHeptCl, BrettPhos Pd G3, etc.
  • ligand such as Xphos, Cphos, BrettPhos, etc.
  • a compound of formula (A1) may also be reacted with aryl alcohol of formula (D1) to give a compound of Formula (3).
  • the reaction may be catalyzed with a suitable Pd-precatalyst (such as RockPhos Pd G3, Cphos Pd G3, etc.) and ligand (such as RockPhos, etc.) in the presence of a base (such as NaOtBu, Cs 2 CO 3 , K 3 PO 4 , etc.) using a suitable solvent (such as t- BuOH, 1,4-dioxane, etc.), and at temperatures typically ranging from 60°C to 120°C.
  • a suitable solvent such as t- BuOH, 1,4-dioxane, etc.
  • a compound of formula (E1) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (F1).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • a solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • suitable solvent such as DME, etc.
  • a compound of formula (G1) may be reacted with concentrated sulfuric acid to give a compound of formula (H1).
  • the protecting group in the compound of formula (H1) may be removed under appropriate reaction conditions to give a compound of formula (I1).
  • a compound of formula (I1) may be reacted with boronic acid or ester of formula (B1) to give a compound of Formula (2).
  • the reaction may be catalyzed with a suitable Pd- precatalyst (such as XantPhos Pd G3, XPhos Pd G3, etc.) and ligand (such as XPhos, etc.) in the presence of a base (such as K 2 CO 3 , NaOtBu, Cs 2 CO 3 , etc.) using a suitable solvent (such as 1,4- dioxane/water mixture, toluene/water mixture, etc.), and at temperatures typically ranging from 60°C to 120°C.
  • a suitable solvent such as 1,4- dioxane/water mixture, toluene/water mixture, etc.
  • the reaction may be catalyzed with a suitable Pd- precatalyst (such as Pd(dba)2, Pd-PEPPSI-iHeptCl, BrettPhos Pd G3, etc.) and ligand (such as Xphos, Cphos, BrettPhos, etc.) in the presence of a base (such as LiHMDS, etc.) using a suitable solvent (such as 2-Me THF, 1,4-dioxane, t-BuOH, etc.) and at temperatures typically ranging from 80°C to 120°C.
  • a suitable solvent such as 2-Me THF, 1,4-dioxane, t-BuOH, etc.
  • reaction may be catalyzed with a suitable Pd-precatalyst (such as RockPhos Pd G3, CPhos Pd G3, etc.) and ligand (such as RockPhos, etc.) in the presence of a base (such as NaOtBu, Cs 2 CO 3 , K 3 PO 4 , etc.) using a suitable solvent (such as t- BuOH, 1,4-dioxane, etc.), and at temperatures typically ranging from 60°C to 120°C.
  • a suitable Pd-precatalyst such as RockPhos Pd G3, CPhos Pd G3, etc.
  • ligand such as RockPhos, etc.
  • a base such as NaOtBu, Cs 2 CO 3 , K 3 PO 4 , etc.
  • suitable solvent such as t- BuOH, 1,4-dioxane, etc.
  • the reaction may be catalyzed with a suitable Pd-precatalyst (such as XantPhos Pd G3, XPhos Pd G3, etc.) and ligand (such as XPhos, etc.) in the presence of a base (such as K 2 CO 3 , NaOtBu, Cs 2 CO 3 , etc.) using a suitable solvent (1,4-dioxane/water mixture, toluene/water mixture, etc.), and at temperatures typically ranging from 60°C to 120°C. [00175] A compound of formula (K1) may be reacted with concentrated sulfuric acid to give a compound of formula (L1).
  • a suitable Pd-precatalyst such as XantPhos Pd G3, XPhos Pd G3, etc.
  • ligand such as XPhos, etc.
  • a base such as K 2 CO 3 , NaOtBu, Cs 2 CO 3 , etc.
  • a suitable solvent such as K 2
  • Scheme 8 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (M1) may be reacted with a compound of formula (N1) (X1 is typically a leaving group like iodo, bromo, chloro, or triflate) to give a compound of formula (O1).
  • the reaction may be performed in the presence of a base (typically an inorganic base such as K 2 CO 3 , etc.) or catalyzed with a suitable Pd-catalyst (such as BrettPhos Pd G3, etc.) in the presence of a base (such as Cs 2 CO 3 , etc.), using a solvent (such as t-BuOH, DMSO, acetone, etc.).
  • a base typically an inorganic base such as K 2 CO 3 , etc.
  • a suitable Pd-catalyst such as BrettPhos Pd G3, etc.
  • a base such as Cs 2 CO 3 , etc.
  • a solvent such as t-BuOH, DMSO, acetone, etc.
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a base typically an organic base such as DIPEA, etc.
  • acid typically a catalytic amount of concentrated HCl
  • a solvent such as t-BuOH, t-amyl alcohol, DMSO, etc.
  • reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • suitable solvent such as DME, etc.
  • a compound of formula (Q1) may be reacted with concentrated sulfuric acid to give a compound of formula (R1).
  • the protecting group in the compound of formula (R1) may be removed under appropriate reaction conditions to give a compound of Formula (6).
  • Scheme 9 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (S1) may be reacted with a compound of formula (N1) (X1 is typically a leaving group like iodo, bromo, chloro, or triflate) to give a compound of formula (T1).
  • the reaction may be performed in the presence of a base (typically an inorganic base such as K 2 CO 3 , etc.) or catalyzed with a suitable Pd-catalyst (such as BrettPhos Pd G3, etc.) in the presence of a base (such as Cs 2 CO 3 , etc.), using a solvent (such as t-BuOH, DMSO, acetone, etc.).
  • a compound of formula (T1) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (U1).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated. HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a compound of formula (U1) may be reacted with malononitrile of formula (E) to give a compound of formula (V1).
  • reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • suitable solvent such as DME, etc.
  • a compound of formula (V1) may be reacted with concentrated sulfuric acid to give a compound of formula (W1).
  • the protecting group in the compound of formula (W1) may be removed under appropriate reaction conditions to give a compound of Formula (7).
  • Scheme 10 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (A2) may be reacted with a compound of formula (N1) (X1 is typically a leaving group like iodo, bromo, chloro, or triflate) to give a compound of formula (B2).
  • the reaction may be performed in the presence of a base (typically an inorganic base such as K 2 CO 3 , etc.) or catalyzed with a suitable Pd-catalyst (such as BrettPhos Pd G3, etc.) in the presence of a base (such as Cs 2 CO 3 , etc.), using a solvent (such as t-BuOH, DMSO, acetone, etc.).
  • a compound of formula (B2) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (C2).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a compound of formula (C2) may be reacted with malononitrile of formula (E) to give a compound of formula (D2).
  • reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • suitable solvent such as DME, etc.
  • a compound of formula (D2) may be reacted with concentrated sulfuric acid to give a compound of formula (E2).
  • the protecting group in the compound of formula (E2) may be removed under appropriate reaction conditions to give a compound of Formula (8).
  • Scheme 11 illustrates synthetic routes to certain compounds of Formula (1).
  • a compound of formula (F2) may be reacted with a compound of formula (N1) (X1 is typically a leaving group like iodo, bromo, chloro, or triflate) to give a compound of formula (G2).
  • the reaction may be performed in the presence of a base (typically an inorganic base such as K 2 CO 3 , etc.) or catalyzed with a suitable Pd-catalyst (such as BrettPhos Pd G3, etc.) in the presence of a base (such as Cs 2 CO 3 , etc.), using a solvent (such as t-BuOH, DMSO, acetone, etc.).
  • a compound of formula (G2) may be reacted with an aryl amine of formula (C) (PG are protecting groups such as OMe, OBn, etc.) to give a compound of formula (H2).
  • the reaction may be performed in the presence of a base (typically an organic base such as DIPEA, etc.) or acid (typically a catalytic amount of concentrated HCl) using a solvent (such as t-BuOH, t-amyl alcohol, DMSO, etc.), and at temperatures typically ranging from 80°C to 120°C.
  • a compound of formula (H2) may be reacted with malononitrile of formula (E) to give a compound of formula (I2).
  • the reaction may be catalyzed with a suitable Pd-catalyst (such as PdCl 2 (dppf), etc.) in the presence of a base (such as NaOtBu, etc.) using a suitable solvent (such as DME, etc.), and at temperatures typically ranging from 80°C to 100°C.
  • a suitable Pd-catalyst such as PdCl 2 (dppf), etc.
  • a base such as NaOtBu, etc.
  • a suitable solvent such as DME, etc.
  • Flash chromatography was performed using straight phase flash chromatography on a SP1 TM Purification system from Biotage TM , CombiFlash ® Rf from ISCO or on Gilson system from Thermo Fisher using normal phase silica FLASH+ TM (40M, 25M or 12 M) or SNAP TM KP- Sil Cartridges (340, 100, 50 or 10), Flash Column silica-CS columns from Agela, with C18-flash columns or standard flash chromatography. In general, all solvents used were commercially available and of analytical grade. Anhydrous solvents were routinely used for reactions. Phase Separators used in the examples are ISOLUTE® Phase Separator columns. The starting materials were obtained from commercial sources or made via literature routes.
  • ACN Acetonitrile
  • BrettPhos 2-(Dicyclohexylphosphino)3,6-dimethoxy-2′,4′,6′-triisopropyl-1,1′-biphenyl
  • BrettPhos Pd G3 [(2-Di-cyclohexylphosphino-3,6-dimethoxy-2′,4′,6′- triisopropyl-1,1′- biphenyl)-2-(2′-amino-1,1′ -biphenyl)]palladium(II) methanesulfonate methanesulfonate
  • CPhos 2-Dicyclohexylphosphino-2′,6′-bis(N,N-dimethylamino)biphenyl
  • CPhos Pd G3 [(2-Dicyclohexylphosphino-2′,6′-bis(N,N-dimethylamino)biphenyl
  • ChemDraw Professional version 21.0.0 examples and Intermediate compounds are named using ChemDraw Professional version 21.0.0 from PerkinElmer. ChemDraw Professional version 21.0.0 generates the names of chemical structures using the Cahn-Ingold-Prelog (CIP) rules for stereochemistry and follows IUPAC rules as closely as possible when generating chemical names. Stereoisomers are differentiated from each other by stereodescriptors cited in names and assigned in accordance with the CIP rules. [00199] ChemDraw is optionally using labels in the graphical representation of stereocenters such as ‘&’ and ‘or’ to describe the configuration of the stereochemical centers present in the structure.
  • CIP Cahn-Ingold-Prelog
  • Sodium t-butoxide 0.597 g, 6.21 mmol was added slowly to a solution of malononitrile (0.431 g, 6.52 mmol) in DME (15 mL).
  • Example B-2 (S)- or (R)-6-amino-7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-4- (trifluoromethyl)-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 137)
  • Intermediate 6 5-bromo-2-methyl-6-(trifluoromethyl)pyrimidin-4(3H)-one
  • Azobisisobutyronitrile (AIBN) (0.184 g, 1.12 mmol) was added to 2-methyl-6- (trifluoromethyl)pyrimidin-4(3H)-one (Intermediate 5, 2.00 g, 11.2 mmol) and NBS (2.398 g, 13.47 mmol) in MeCN (50 mL).
  • the racemic mixture was purified by preparative chiral-HPLC on a column: CHIRALPAK IC, 2*25 cm, 5 ⁇ m; Mobile Phase A: HEX (0.5% 2M NH 3 -MeOH), Mobile Phase B: ETOH; Flow rate: 20 mL/min; Gradient: isocratic 7; Wave Length: 220/254 nm; RT1(min): 18.873; RT2(min): 22.707; Sample Solvent: EtOH; Injection Volume: 0.4 mL; Number Of Runs: 22.
  • Example B-3 (S)- or (R)-6-amino-4-(difluoromethyl)-7-(3-hydroxy-2,6-dimethylphenyl)-2- methyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 138)
  • Intermediate 13 6-(difluoromethyl)-2-methylpyrimidin-4(3H)-one
  • a mixture of sodium ethoxide (3.07 g, 45.2 mmol) in ethanol (50.0 mL) was added to a stirred mixture of ethyl 4,4-difluoro-3-oxobutanoate (Intermediate 11, 5.00 g, 30.1 mmol) and acetimidamide hydrochloride (Intermediate 12, 2.85 g, 30.1 mmol) in toluene (100 mL) at 25 °C ⁇ The resulting mixture was stirred at 80 °C for 16 hours.
  • racemic mixture was purified by preparative chiral-HPLC (Column: CHIRALPAK IE, 2*25 cm, 5 ⁇ m; Mobile Phase A: Hex (0.5% 2M NH3-MeOH), Mobile Phase B: EtOH; Flow rate: 20 mL/min; Gradient: isocratic 10; Wave Length: 220/254 nm; RT1(min): 17.882; RT2(min): 24.125; Sample Solvent: EtOH; Injection Volume: 0.4 mL; Number Of Runs: 13) to afford rotational isomer 1 and rotational isomer 2, (S) or (R)-6-amino-4-(difluoromethyl)- 7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide (Example B-3, 17.40 mg, 17.40 %) as a yellow solid.
  • Example B-4 (S)- or (R)-6-amino-4-cyclopropyl-7-(3-hydroxy-2,6-dimethylphenyl)-2-methyl- 7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide (Compound 139)
  • Intermediate 20 6-methoxy-N-(3-methoxy-2,6-dimethylphenyl)-2-methylpyrimidin-4- amine
  • Xantphos Pd G3 (4.49 g, 4.73 mmol) was added to 4-chloro-6-methoxy-2- methylpyrimidine (Intermediate 19, 15.0 g, 94.6 mmol), 3-methoxy-2,6-dimethylaniline (Intermediate 2, 15.73 g, 104.1 mmol) and Cs 2 CO 3 (92.0 g, 284 mmol) in 1,4-dioxane (300 mL) under nitrogen.
  • the racemic mixture was purified by preparative chiral-HPLC on a column: Lux 5um Cellulose-4, 2.12*25 cm, 5 ⁇ m; Mobile Phase A: Hex (0.5% 2M NH3-MeOH), Mobile Phase B: EtOH; Flow rate: 220 mL/min; Gradient: isocratic 30; Wave Length: 220/254 nm; RT1(min): 12.032; RT2(min): 17.364; Sample Solvent: ETOH; Injection Volume: 1.5 mL; Number Of Runs: 3 to afford rotational isomer 1, (S) or (R)-6-amino-4-cyclopropyl-7-(3-hydroxy-2,6- dimethylphenyl)-2-methyl-7H-pyrrolo[2,3-d]pyrimidine-5-carboxamide (Example B-4), 2.77 mg, 21%) as a white solid and rotational isomer 2.
  • GI 50 values were then correlated to basal replication stress levels for the 16 cell line models identified in Figs.2, 3, and 4. Basal replication stress levels for the uterine cancer and breast cancer cell line models used in the study were determined by DNA combing analysis described in Young, L. A., et al. (2019). "Differential Activity of ATR and WEE1 Inhibitors in a Highly Sensitive Subpopulation of DLBCL Linked to Replication Stress.” Cancer Research 79(14): 3762-3775.
  • the OVCAR3 and TOV112D FBXW7 knock out (KO) ovarian cancer cell lines express elevated levels of the CCNE1 gene product, cyclin E1 (a biomarker linked to induction of basal replication stress) compared to the SKOV3 and TOV112D ovarian cancer cell lines.
  • cyclin E1 a biomarker linked to induction of basal replication stress
  • Panc0203 pancreatic cancer cell line expresses elevated levels of phosphorylated RPA2 (a biomarker linked to elevated basal replication stress) while the pancreatic cancer KP4 cell line does not.
  • Serra, V., et al. "Identification of a Molecularly-Defined Subset of Breast and Ovarian Cancer Models that Respond to WEE1 or ATR Inhibition, Overcoming PARP Inhibitor Resistance," Clin Cancer Res 28(20): 4536-4550. (2022).
  • Figure 2 reports the GI 50 values measured in the 16 cell line models (some having elevated basal replication stress) treated with a PKMYT1 inhibitor tool compound.
  • Figure 3 reports the corresponding median replication fork velocities measured by DNA combing analysis for the uterine cancer and breast cancer cell line models treated with the tool compound.
  • the five uterine cancer and breast cancer cell lines of Figure 2 having the lowest GI 50 values also had the lowest median replication fork velocities of Figure 3.
  • Figures 2 and 3 collectively show that the cell line models with elevated levels of basal replication stress (KRAS and CCNE1-amplified uterine cancer cell lines, CCNE1-amplified breast cancer cell lines, OVCAR3 (CCNE1- amplified) and TOV112D FBXW7 KO (cyclin E1 high) ovarian cancer cell lines, and panc0203 (pRPA2 high) pancreatic cancer cell line) were the most sensitive cell lines to PKMYT1 inhibition. In contrast, the cell line models with low basal levels of replication stress were more resistant to PKMYT1 inhibition.
  • basal replication stress KRAS and CCNE1-amplified uterine cancer cell lines, CCNE1-amplified breast cancer cell lines, OVCAR3 (CCNE1- amplified) and TOV112D FBXW7 KO (cyclin E1 high) ovarian cancer cell lines, and panc0203 (pRPA2 high) pancreatic cancer cell line
  • Example C-2 Target Engagement NanoBRET (Bioluminescence Resonance Energy Transfer) Assay A. NanoBRET Assay (PKMYT1, WEE1, and EPHB3) [00234] A whole cell-based target engagement assay was used to investigate target engagement with three targets of interest using the NanoBRET technology. Specifically, compounds were tested to assess inhibition of PKMYT1, WEE1, and EPHB3 in engineered HEK293T cell lines.
  • HEK293T cell lines overexpressing PKMYT1, WEE1, and EPHB3 were thawed out from cryopreserved vials at 37°C.
  • the thawed cells were resuspended in 10mL of RPM1-1640 supplemented with 10% Fetal calf serum and 1% GlutaMax.
  • Cell lines were seeded using a Multidrop Combi with a small cassette in 384-well low volume plate (Greiner #784080) at 8000 cells/well in 10 ⁇ L (for PKMYT1 and EPHB3) and 4000 cells/well in 10 ⁇ L (for WEE1). The plates were incubated at 37°C, 5% CO 2 for 23 hours.
  • NanoBRET Assay (Kinase Panel)
  • Compound 137, Isomer 2 was further profiled (dose response) against selected kinases in a NanoBRET whole-cell target engagement assay conducted at CELLinib GmbH. IC 50 values for Compound 137, Isomer 2 are reported in Table 1-B.
  • Example C-3 PKMYT1 Surface Plasmon Resonance (SPR) Direct Binding Assay A.
  • SPR Surface Plasmon Resonance
  • Biotinylation was achieved using the BirA kit (Avidity) as per manufacturer’s instructions and biotinylation confirmed by mass spectrometry. Size exclusion chromatography was used to further purify the PKMYT1 into Storage buffer (25 mM Hepes pH 7.5, 500 mM NaCl, 0.5 mM TCEP), and fractions containing pure PKMYT1 were pooled and concentrated to 10.2 mg/ml and snap frozen on liquid nitrogen.
  • B. SPR Direct Binding Assay [00242] All SPR steps were performed on a Biacore 8K Instrument (Cytiva) at a temperature of 25 °C.
  • a Series S Sensor Chip SA (Cytiva) was functionalized with ⁇ 3000 RU biotinylated Avi-tagged PKMYT1 using a 180s injection at 5 ⁇ l min -1 .
  • the surface was conditioned with 10 start-up injections of the “running buffer” (50 mM Hepes pH 7.5, 200 mM NaCl, 5 mM MgCl 2 , 0.05% Tween-20, 1 mM TCEP, 1% DMSO).
  • a solvent correction step consisting of a 5-point titration of different DMSO containing running buffer solutions (0-2%) were injected at the beginning and the end of the screening run to correct for DMSO deviations. Three blank cycles of running buffer injections were included preceding each compound injection for blank subtraction. A threefold dilution series for each sample was injected sequentially from low to high concentration over the PKMYT1-coupled surfaces in single-cycle kinetics mode, at a flow rate of 30 ⁇ l min ⁇ 1 for 120 s, monitoring dissociation for 2500 s. [00243] All analysis was performed using the Biacore Insight Evaluation Software after applying standard double-referencing and solvent correction.
  • Example C-4 Functional CDK1 pThr14 AlphaLISA Assay
  • Compounds were tested to assess their effect on PKMYT1 activity in a cellular context. Specifically, phosphorylation of CDK1 (a PKMYT1 substrate) on Thr14 was used as a specific marker of PKMYT1 activity. Inhibition of PKMYT1 activity by inhibitors decreases levels of pCDk1(Thr14) in the cell.
  • Example C-5 Cellular Proliferation in OVCAR3 and SKOV3 Cell Lines (Hoechst Staining) [00248] An imaging-based assay was used to investigate changes in cell number. Compounds were tested to assess inhibition of cellular proliferation in a CCNE1-amplified ovarian cancer cell line and a CCNE1-nonamplified SKOV3 ovarian cancer cell line.
  • OVCAR3 and SKOV3 cells were cultured in RPMI 1640 medium with 4.5 g/ml Glucose, supplemented with 20% heat in-activated Fetal bovine serum and McCoy’s modified 5A medium supplemented with 10% heat in-activated Fetal bovine serum along with 1% GlutaMax respectively. Both the cell lines were seeded in complete growth medium using a multidrop into 384-well plates (Greiner, 781090) at the density 4000 cells/well in 40 ⁇ L/well. The plates (test compound plates + a day 0 control plate) were placed in incubator maintained at 37°C and 5% CO 2 for about 20 to 22 hours.
  • Example C-6 Kinase Profiling A.
  • Test compounds were profiled against a broad panel of kinases at ThermoFisher Scientific using the SelectScreen Kinase Profiling Services.
  • Each kinase assay used one of the following assay technology protocols: 1) Z-LYTETM Screening Protocol and Assay Conditions (Revised 23-Jan-2018 version) available at http://assets.thermofisher.com/TFS-Assets/BID/Methods-&- Protocols/20180123_SSBK_Customer_Protocol_and_Assay_Conditions.pdf; 2) AdaptaTM Screening Protocol and Assay Conditions (Revised 23-Jan-2018 version) available at http://assets.thermofisher.com/TFS-Assets/BID/Methods-&- Protocols/20180123_SSBK_Adapta_Customer_Protocol_and_Assay_Conditions.pdf ; or 3) LanthaScreenTM Eu Kinas
  • Example C-7 Metabolic Stability Assays A. Assessment Using Human Liver Cytosol (HLC) [00254] The stability of control (zaleplon and 06-benzylguanine) and test compounds was evaluated at 1 ⁇ M using 2.5 mg/mL human liver cytosol in the presence and absence of 3 ⁇ M raloxifene in 0.1 M phosphate buffer pH 7.4 at 37°C. Aliquots were removed at 0, 10, 30, 60, 90 and 120 min, quenched 1:5 with ACN containing internal standard. Samples were analyzed for loss of parent compound using LC/MS/MS. Potential hydroxylation metabolites were monitored by adding transitions of Parent+16.
  • HSC Human Liver Cytosol
  • HLM Human Liver Microsomes
  • HLM mixture containing 1.12 mg/mL HLM in phosphate buffer pH 7.4 was mixed with 25 ⁇ L 10 mM NADPH and pre-warmed at 37°C for 8 minutes prior to addition of 2.5 ⁇ L of 100 ⁇ M test compound/control prepared in DMSO.
  • Aliquots (20 ⁇ L) were removed at 0.5, 5, 10, 15, 20 and 30 minutes and quenched with 100 ⁇ L of cold stop solution (100% ACN with 100 nM of alprazolam, caffeine, and tolbutamide as internal standards). Samples were centrifuged for 20 minutes at 4,000 rpm and 4°C and a 40 ⁇ L aliquot of supernatant was mixed with 160 ⁇ L pure water for LC-MS/MS analysis.
  • cryopreserved hepatocytes are thawed in a 37° water bath, centrifuged in thawing medium and prepared in Leibovitz’s L-15 Medium (pH 7.4) to contain 1 million viable hepatocytes/ml and a final compound concentration of 1 ⁇ M.
  • Cell viability was determined using a Cellometer Vision and greater than 80% cell viability was required to proceed with the compound incubation.
  • the compound/cell solution 250 ⁇ l was incubated for 2 hours at 37 ⁇ C and shaken at 900 rpm on an Eppendorf Thermomixer Comfort plate shaker.
  • Samples (20 ⁇ l) were taken at 0.5, 5, 15, 30, 45, 60, 80, 100, and 120 minutes and quenched with 100 ⁇ l of 100% ice-cold acetonitrile. Samples were shaken at 800 rpm for 2 minutes and centrifuged at 4000 rpm for 20 minutes at 4 ⁇ C to pellet precipitated protein. The supernatant fraction was diluted 1:5 with deionized water, shaken at 1000 rpm for 2 minutes, and further diluted 1:1 with deionized water. Samples were analyzed by LC-MS/MS. Each incubation was conducted in singlicate. Peak areas were determined from extracted ion chromatograms and percent (%) parent remaining was calculated from peak area of test compound/control. D.
  • test compound was dissolved in DMSO at 10 mM, then diluted to 2 ⁇ M in dosing medium using Echo dispense. All steps were performed at room temperature, while the 2 ⁇ M test compound was preheated at 37°C before start of incubation. Prior to assay, maintenance medium was removed from the cells, followed by a washing step with 100 ⁇ L of preheated blank dosing medium without serum.
  • Example C-8 Log D [00260] The Log D values of test compounds at 10 ⁇ M were determined in octanol/PBS pH 7.4 using LC/MS/MS and nicardapine, cyclobenzaprine, and caffeine as control compounds.10 ⁇ L of test compound or control was added to a 96-deep well plate followed by addition of 500 ⁇ L of saturated octanol and 500 ⁇ L of saturated phosphate buffer. The plate was shaken at room temperature for 2h at 2,000 rpm. The samples were centrifuged at room temperature, at 4,000 rpm for 30 minutes. 100 ⁇ L of sample was removed from the octanol and buffer phase.
  • the rate of compound transport in the apical to basolateral (A-B) direction was determined from the donor well as apical (A) compartment and the receiver as basolateral (B) compartment.
  • the rate of compound transport in the basolateral to apical (B-A) direction was determined from the donor well as apical (A) compartment and the receiver as basolateral (B) compartment.
  • the rate of compound transport in the basolateral to apical (B-A) direction the donor well was the basolateral (B) compartment and the receiver was the apical (A) compartment.
  • Samples were analyzed by liquid chromatography (LC)–mass spectrometry (MS)/MS.
  • Efflux ratio P app (B to A)/P app (A to B), where: dCr/dt is the cumulative concentration of the compound in the receiver chamber as a function of time (in ⁇ M/s); Vr is the solution volume in the receiver chamber (0.1 ml on the apical side and 0.3 ml on the basolateral side); A is the surface area for the transport (i.e., 0.11 cm 2 for the area of the monolayer); and C 0 is the initial concentration in the donor chamber (in ⁇ M).
  • Substrates were, Phenacetin (30 ⁇ M), diclofenac (10 ⁇ M), S-mepheytoin (35 ⁇ M), bufuralol (5 ⁇ M) and midazolam (3 ⁇ M), for CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4/5, respectively.
  • the final concentration of HLMs was 0.2 mg/mL and the final concentrations of test compound were 0, 0.1, 0.3, 1, 3, 10 and 30 ⁇ M.
  • the incubation was intiated by addition of 20 ⁇ L of 10 mM NADPH in phosphate buffer and quenched 1:1 with ACN containing 3% formic acid and 40 nMol verapamil as internal standard, following five minutes at 37°C.
  • the samples were centrifuged at 4,000 rpm for 30 minutes, placed on ice for 20 min and recentrifuged for an additional 30 min at 4,000 rpm.
  • Supernatant 200 ⁇ L was transferred for analysis of isoform selective metabolite formation. The extent of inhibition was calculated by comparing the remaining activity in the presence of inhibitor to the DMSO only control using the following equation.
  • the concentration yielding 50% inhibition, IC 50 were calculated using non-linear regression analysis in Xlfit.
  • Example B-1 The IC 50 values for Example B-1, Isomer 1 (i.e., the eutomer of Compound 1) and Example B-2, Isomer 2 (i.e., the eutomer of Compound 137) against each of CYPs 1A2, 2C9, 2C19, 2D6, and 3A were greater than 30 ⁇ M.
  • Isomer 1 i.e., the eutomer of Compound 1
  • Example B-2 Isomer 2 (i.e., the eutomer of Compound 137) against each of CYPs 1A2, 2C9, 2C19, 2D6, and 3A were greater than 30 ⁇ M.
  • CYP Time Dependent Inhibition The potential for test compounds to inhibit Cytochrome P450 (CYP) activity in a time dependent manner was determined using 1 mg/mL of a 150-donor mixed gender human liver microsome pool (HLM), 10 ⁇ M test compound, a 30 min preincubation time, and a 10-fold dilution step to measure formation of specific substrates towards, CYPs 1A2, 2C9, 2C19, 2D6 and 3A at 3X their respective Km values.
  • HLM human liver microsome pool
  • Substrates were, Phenacetin (90 ⁇ M), diclofenac (30 ⁇ M), S-mepheytoin (105 ⁇ M), bufuralol (15 ⁇ M), and midazolam (9 ⁇ M), for CYP1A2, CYP2C9, CYP2C19, CYP2D6 and CYP3A4/5, respectively. Specific inhibitors of these pathways were included as positive controls.
  • the primary incubation consisted of 1 mg/mL HLM ⁇ 10 mM NADPH with 10 ⁇ M of test compound incubated for 30 min at 37°C.
  • the irreversible IC 50 representing the concentration where 50% inhibition would be observed, was calculated as follows: [00268] The % time dependent inhibition (TDI) values calculated for Example B-1, Isomer 1 (i.e., the eutomer of Compound 1) and Example B-2, Isomer 2 (i.e., the eutomer of Compound 137) against each of CYPs 1A2, 2C9, 2C19, 2D6, and 3A were less than 20.
  • CYP Induction [00269] The potential for test compounds to induce drug metabolizing enzymes was evaluated by assessing the Pregnane X Receptor (PXR) activation potential in DPX2 cells.
  • PXR Pregnane X Receptor
  • DPX2 cells were purchased from Puracyp and treated according to the vendor protocol.
  • Test compounds, positive control (rifampicin, 1 and 10 ⁇ M) and negative control (propranolol, 10 ⁇ M) were prepared in DMSO to a final concentration of 0.1%.
  • Test compounds were evaluated at 0.1, 0.3, 1, 3, 10 and 30 ⁇ M to enable derivation of EC 50 and E max values, corresponding to the concentration resulting in half maximal induction and the maximal induction observed, respectively.
  • Cells were treated with test compound or control for 24h. After 24h dosing media was removed and 1xCellTiter-FluorTM was added for 30 minutes at 37°C. The fluoresecence was measured using 400 nm excitation and 505 nm emission.
  • the ONE-Glo assay reagent was then added for 5 min and luminescence was determined using a luminometer.
  • the cell viability was determined using the following equation:
  • the normalized luciferase activity was determined by taking the ratios of relative luminescence units (RLU), and relative fluorescence units for three replicates at each concentration and the fold-activation of mRNA levels was determined using the following equation: [00270] No induction of PXR was observed for Example B-1, Isomer 1 (i.e., the eutomer of Compound 1).
  • Example C-11 Aqueous Solubility [00271] The thermodynamic solubility of the test compounds was measured in a shake-flask approach starting from 10 mM DMSO solutions.
  • KAT Kinase Affinity Tool
  • a Kinase Affinity Tool (“KAT”) chemical proteomics method based on the Kinobead technology 1-2 was used for in situ kinase profiling of test compound.
  • KAT is an affinity matrix made up of promiscuous ATP-competitive kinase inhibitors to enrich for a large subset of the native kinome directly from disease/safety relevant lysate.
  • Proteomic RB50 values indicate the selective affinity of the test compound for the enriched kinases.
  • Test compound was incubated with a cell lysate mix of four cell lines (K562, MV4- 11, SN-N-BE2, and COLO-205) in a dose-response format for 30 minutes at 4°C.
  • the compound-incubated lysates were then bound to KAT beads for 30 minutes at 4°C.
  • Beads were washed twice with binding buffer followed by two washes with 1X PBS. Proteins bound to beads were reduced, alkylated, and digested overnight with Trypsin/LysC enzyme at 37°C. Digested peptides were eluted with 60% ACN/0.1% formic acid in water and desalted with in- house made C18 StageTips.
  • Radioligand binding assays were used to assess the ability of Compound 137, Isomer 2 to interact with a panel of G-protein coupled receptors (GPCRs), ion channels, and transmembrane transporters. Assays measuring substrate turnover or phosphorylation by isolated proteins were used for kinase and enzyme targets, allowing direct determination of the mode of action of the compounds. Where binding activity was observed at a GPCR target, mode of action was determined using cell based functional assays with secondary messenger read outs.
  • GPCRs G-protein coupled receptors
  • the in vitro assay used the fluorescent signal generated by the reduction of non-fluorescent resazurin (7-hydroxy-3H-phenoxazin-3-one 10-oxide) to the fluorescent resorufin (Alamar blue assay) as a measure of cytotoxicity. Because cellular reduction of resazurin is dependent on a pool of reductase or diaphorase enzymes derived from the mitochondria and cytosol, resazurin can be used as an oxidation-reduction indicator in cell viability assays for mammalian cells (McMillian et al, 2002; O’Brien et al, 2000).
  • THP-1 cells were subcultured in suspension in a T175cm 2 tissue culture flask with medium (RPMI, supplemented with 1% L-glutamine and 10% quantified FBS). The sub-cultures were maintained at 37°C in a 95% humidified atmosphere with 5% CO 2 and passaged approximately every 2 to 3 days. Cell density was maintained between 50,000 cells/mL and 1 x10 6 cells/mL.
  • Test compounds 40 nL
  • Test compounds were added to a 1536-well low volume plate (Greiner #782077) at 10, half-log concentrations with a top concentration of either 100 ⁇ M or 250 ⁇ M depending on the stock concentration selected. For both, the DMSO concentration was 1%.
  • THP-1 cells were seeded into the plates at a density of 2,000 cells per well in 4 ⁇ l THP-1 media on slow speed.
  • the plates containing THP-1 cells with compounds/solvent were incubated for 48 hours under standard cell culture conditions (37°C, 5% CO 2 ).
  • a stock solution of resazurin 113 mg of resazurin in 1000 mL of PBS was prepared, warmed to 37°C, and vortexed.
  • Multidrop dispenser with the Multidrop standard cassette, 1 ⁇ L of stock resazurin solution was added to all wells at high speed to aid mixing. The plates were incubated for two hours under standard cell culture conditions. The plates were then incubated for a further one hour at room temperature, with shaking (700 rpm). [00283] The plates were then read on a BMG PheraStar reader using an excitation wavelength of 540 nm and an emission wavelength of 580nm. Controls were 100 ⁇ M Lapatinib (minimum/inhibitor control) and 1% DMSO (maximum/neutral control). The acquired raw data were loaded into Genedata analyser (Genedata AG) to determine IC 50 values.
  • Genedata AG Genedata analyser
  • Example C-15 In Vitro Hematopoietic Stem and Progenitor Cell Assay (Bone Marrow Toxicity) [00286] Test compounds were evaluated to determine their effect on hematopoietic stem and progenitor cell (“HSPC”) proliferation and differentiation.
  • HSPC hematopoietic stem and progenitor cell
  • Cryopreserved human bone marrow derived CD34 + hematopoietic stem and progenitor cells were defrosted and left to recover overnight in maintenance media (StemSpan SFEM II (Stem Cell Technologies) containing 25 ng/ml SCF, 50 ng/ml TPO, and 50 ng/ml Flt3-L human recombinant protein (all Peprotech)), in a humidified incubator at 37°C with 5% CO 2 . The next day cells were resuspended in the presence of test compound into media capable of supporting erythroid cell differentiation (Preferred Cell Systems, SEC-BFU1-40H), at a concentration of 10,000 cells/ml.
  • Example C-16 Effect on Cell Cycle in Human Ovarian OVCAR3 Cell Line
  • An immunoblotting analysis of CCNE1-amplified ovarian OVCAR3 cells was performed to determine whether short (24 hours) treatment with Compound 137, Isomer 2 results in DNA damage response (“DDR”) signaling in the replicative phase of the cell cycle.
  • DDR DNA damage response
  • Phopho- CHK1 (Ser345) and phospho-RPA2 (Ser4/Ser8) levels biomarkers considered to be specifically expressed following DNA damage formation during active replication of cells, were assessed as a measure of effect on cell cycle.
  • OVCAR3 cells were treated with Compound 137, Isomer 2 for 24 hours and then lysed in RIPA buffer (Sigma-Aldrich) supplemented with protease inhibitors (Roche), phosphatase inhibitors (Sigma-Aldrich), and benzonase (Merck). After 30 minutes (min) incubation on ice the lysates were cleared by centrifugation at 15,000 rpm 4°C for 20 minutes and the supernatants were kept for sample loading. NuPAGETM LDS Sample Buffer (ThermoFisher Scientific) and NuPAGETM Sample Reducing Agent (ThermoFisher Scientific) were added to the samples and heated to 95 °C (5 minutes).
  • H2AX Abcam, ab20669
  • gH2AX Cell Signaling Technology (CST), 2577
  • Histone H3 CST, 3638
  • phospho-Histone H3 Ser10 CST, 9701
  • RPA2 – Abcam, ab2175
  • phospho-RPA2 Ser4/Ser8 Bethyl Labs, A300-245A
  • CDK1 CST, 9116
  • phospho- CDK1 Thr14 (Abcam, ab58509) or BioLegend, 947402
  • phospho-CDK1 Tyr15 Abcam, ab275958
  • CHK1 CST, #2360
  • phospho-CHK1 Ser345 CST, 2348
  • PKMYT1 Abcam, ab307146
  • vinculin Sigma, V9131
  • Example C-17 Cellular Proliferation in PKMYT1 Knock Out SKOV3 Cells
  • the potential effect of off-target activity of Compound 137, Isomer 2 and impact on cell viability was evaluated in PKMYT1 knock out (KO) SKOV3 cells. Specifically, loss of PKMYT1 protein expression, reduction of PKMYT1 substrate phosphorylation, and growth inhibition were evaluated in PKMYT1 KO SKOV3 cells.
  • CRISPR/Cas9 technology was used to delete PKMYT1 kinase from SKOV3 cells and prepare two monoclonal PKMYT1 KO clones (B2 and H2).
  • Parental SKOV3 cells and PKMYT1 KO SKOV3 cells were either left untreated or were treated with Compound 1, Isomer 1 for 24 hours.
  • Cells were then lysed in RIPA buffer (Sigma-Aldrich) supplemented with protease inhibitors (Roche, Basel, Switzerland), phosphatase inhibitors (Sigma-Aldrich), and benzonase (Merck).
  • a CellTiter-Glo (CTG) assay was employed to evaluate the effect of Compound 137, Isomer 2 on growth inhibition of the PKMYT1 KO SKOV3 cells.
  • Parental SKOV3 and OVCAR3 cells were included as controls. Cells were seeded at 500 cells per well in a 96-well plate and exposed to Compound 137, Isomer 2 for seven days. After cell attachment on day 0, Compound 137, Isomer 2 was dispensed from compound stocks dissolved in DMSO, in seven titration dilutions with a top concentration of 30 ⁇ M. DMSO served as a vehicle control. Plates were incubated at 37oC, 5% CO 2 .
  • Example C-18 In Vivo Tolerability (Mouse Model) [00298] A tolerability study was conducted to evaluate long-term dosing of Compound 137, Isomer 2 in mice as a monotherapy, as a combination therapy with gemcitabine, and as a combination therapy with irinotecan hydrochloride.
  • A. Monotherapy Female SCID mice (18 g+ at time of first procedure) were orally dosed with 300 mg/kg BID of Compound 137, Isomer 2 for 28 days. Compound 137, Isomer 2 was formulated for dosing using a 5% DMSO, 50% of 20% Captisol, 45% water for injection (pH 3-3.2) vehicle. Bodyweights were collected and recorded daily.
  • Figure 8-B illustrates the effect of such combination dosing of Compound 137, Isomer 2 and irinotecan hydrochloride on body weight loss. No significant loss of body weight, paw or tail swelling, or other noticeable adverse events were observed at this combination dose.
  • An in vivo study in mice was conducted to evaluate PKMYT1 target engagement by Compound 137, Isomer 2.
  • Target engagement was assessed by analysis of phospho-CDK1 (T14) expression in tumor.
  • OVCAR3 tumor-bearing female SCID mice were dosed orally with 10 mg/kg BID, 30 mg/kg BID, or 100 mg/kg BID of Compound 137, Isomer 2.
  • Doses were calculated for individual animals on the day of dosing and administered with a 10 mL/kg dosing volume. After 21 days of dosing, tumors were collected, processed, and analyzed by immunoblotting: Lysis buffer (25 mM Tris/HCl, 3 mM EDTA, 3 mM EGTA, 50 mM NaF, 2 mM orthovanadate, 0.27 M sucrose, 10 mM ⁇ -glycerophosphate, 5 mM pyrophosphate, 0.5% Triton X-100), protease inhibitor (Sigma, P8340), phosphatase inhibitors (Sigma, P0044, P5726) and benzonase (Sigma, (E10141:2000) was used to lyse and homogenize (3x 30 seconds, 6.0 m/s) tumor pieces in a fast prep tube with lysing matrix.
  • Lysis buffer 25 mM Tris/HCl, 3 mM EDTA, 3
  • Lysates were sonicated and supernatants collected. Lysates were transferred and chilled in 2ml assay blocks in fast prep tubes and centrifuged again for 15 minutes for 2100 rpm at 4°C to ensure no debris transferred. Supernatants were then boiled for 5 minutes at 95°C and immunoblotted as described in Example C-16, using phospho- CDK1 Thr14 (Biolegend, 947402), and vinculin (Sigma, V9131) primary, and anti-mouse IgG, HRP-linked (CST, 7076) secondary antibodies. Following detection with G-box (Syngene), bands were quantified with Gene Tools software.
  • FIG. 9 illustrates the relative in vivo expression levels of phospho-CDK1 Thr14 for each dose.
  • Example C-20 Anti-Tumor Effect in Human OVCAR3 and SW620 Xenograft Models
  • A. Human Ovarian OVCAR3 Cell Line-Derived Xenograft Model [00309] OVCAR3 cells were implanted onto the right flank of female SCID mice.
  • Combination benefit and tumor regression (18%, P ⁇ 0.01) was achieved, however, with twice- daily dosing of 100 mg/kg of Compound 137, Isomer 2 in combination with once weekly intraperitoneal dosing of, 50 mg/kg irinotecan hydrochloride, a topoisomerase 1 inhibitor (TOP1i).
  • TOP1i topoisomerase 1 inhibitor
  • Example C-21 In Vitro Efficacy in Combination with Exatecan [00313] An in vitro assay was carried out to assess whether treatment of PKMYT1 KO isogenic SKOV3 cells (previously described in Example C-17) with Compound 137, Isomer 2 could further sensitize those cells to exatecan.
  • Treatment of the PKMYT1 KO cells with exatecan (1 nM) resulted in maximum drug activity (E max ) that did not improve further with the addition of Compound 137, Isomer 2.
  • mice were randomized to treatment groups based on tumor volume and treated with: (i) vehicle administered orally, (ii) one intraperitoneal dose of irinotecan hydrochloride (50 mg/kg), (iii) twice-daily oral doses (100 mg/kg) of Compound 137, Isomer 2 for three days, or (iv) one intraperitoneal dose of irinotecan hydrochloride (50 mg/kg) plus twice-daily oral doses (100 mg/kg) of Compound 137, Isomer 2 for three days.
  • tumors were collected and subsequently processed for immunoblotting as described in Example C-19 using phospho- CDK1 Thr14 (Biolegend, 947402), CDK1 (CST, 9116), Histone H3 (CST, 3638), phospho-Histone H3 Ser10 (CST, 9701), gH2AX (Cell Signalling Technology (CST) and vinculin (Sigma, V9131), and anti-rabbit IgG, HRP-linked (CST, 7074) and anti-mouse IgG, HRP-linked (CST, 7076) secondary antibodies. Following detection with G-box (Syngene), bands were quantified with Gene Tools software.
  • Intraperitoneal dosing of irinotecan hydrochloride resulted in DNA damage formation (gH2AX) that was not further potentiated by the addition of Compound 137, Isomer 2.
  • Irinotecan hydrochloride monotherapy resulted in a reduced mitotic population (phospho-histone H3 Ser10), a finding consistent with activation of the G2/M DNA damage cell cycle checkpoint following treatment with a TOP1i inhibitor.
  • Example C-23 In Vitro Biotransformation in Hepatocytes of Multiple Species [00319] Compound 137, Isomer 2 was incubated (5 ⁇ M) with mouse, rat, dog, minipig, and human hepatocytes (1 million cells/ml) at 37 oC for 180 minutes. Incubations were stopped by addition of organic solvent, and samples centrifuged to separate soluble and insoluble components, with the supernatant stored at -20 oC until required for analysis. A semi-quantitative analysis of the dominant biotransformations occurring to the compound in each of the incubations was performed, with liquid chromatographic separation followed by both UV and MS detection of the eluate.
  • FIG. 14 is a metabolism scheme showing the dominant routes of biotransformations observed in vitro of Compound 137, Isomer 2: oxidation and glucuronidation.
  • Example C-24 In Vivo Pharmacokinetic Studies in the CD-1 Mouse [00320] The pharmacokinetic profile of Compound 137, Isomer 2 was characterised in vivo using CD-1 mice following either a single intravenous bolus dose or a single oral bolus dose to allow the calculation of key pharmacokinetic properties. [00321] Animal experiments were conducted in accordance with the relevant welfare policies and procedures of the organisation conducting the study.
  • mice Male CD-1 mice, weighing ⁇ 20-30 g were housed in polycarbonate animal cages with absorbent rodent bedding, and allowed to acclimatize, with ad libitum access to a standard rodent chow food and sterile water. An ambient temperature of 20-25 °C and humidity of 40-70% was maintained, alongside alternating 12-hour light/dark cycles, interrupted only by study-related events (e.g., sampling).
  • the compound was formulated for administration as an aqueous solution containing 5% dimethylsulfoxide, 95% sulfobutylether- ⁇ -cyclodextrin (30% w/v) in purified water for injection (WFI), at a dose level of either 0.5 mg/kg (I.V.) or 1.0 mg/kg (P.O.) animal bodyweight, and dose volume of 2.0 mL/kg (I.V.) or 4.0 mL/kg (P.O.) animal bodyweight.
  • WFI purified water for injection
  • Blood samples ( ⁇ 0.02 mL / timepoint) were collected from the dorsal metatarsal vein into tubes containing EDTA-K2 anticoagulant, and placed on wet ice (4 °C) before centrifugation (4000 g, 5 minutes, 4oC) to obtain plasma, and then stored (-75 oC +/- 15 °C) prior to further analysis.
  • samples were collected at 2, 10, 30, 60, 120, 240, 360, 480, 720, and 1440 minute timepoints.
  • samples were collected at 5, 15, 30, 60, 120, 240, 360, 480, 720, and 1440 minute timepoints.
  • Example C-25 In Vivo Pharmacokinetic Studies in the SCID Mouse [00322] Pharmacokinetic profiles of Compound 137, Isomer 2 in female SCID mice were obtained following oral administration to establish the short-term tolerability, and confirmed the extent and duration of compound exposure. Animal husbandry, dosing, sample collection, and subsequent analytical work to determine total plasma concentrations were conducted in a similar manner as previously described In Example C-24, with minor adjustments. Compound 137, Isomer 2, formulated in 5% DMSO, 50% of 20% Captisol, and 45% purified water for injection (WFI) (pH 3-3.2), was dosed at three dose levels (10 mg/kg, 30 mg/kg, and 100 mg/kg) and a dose volume of 10 mL/kg.
  • WFI purified water for injection
  • Plasma samples were collected from the lateral tail vein at multiple timepoints (30 / 60 / 120 / 240 / 420 / 1440 minutes) following administration.
  • TABLE 18 Figure 16 shows the pharmacokinetic profiles in female SCID mice (total blood plasma concentration) of Compound 137, Isomer 2 following administration P.O. at three dose levels: 10 mg/kg (solid line), 30 mg/kg (dashed line), and 100 mg/kg (dotted line).
  • Example C-26 In Vivo Pharmacokinetic Studies in The Intact and Bile-Duct-Cannulated Han Wistar Rat [00323] The pharmacokinetic profile of Compound 137, Isomer 2 was characterized in vivo using bile-duct-cannulated Han Wistar rat following an I.V. bolus dose to determine the contribution of enterohepatic circulation to key pharmacokinetic properties. [ 00324] Animal experiments were conducted in accordance with the relevant welfare policies and procedures of the organization conducting the study.
  • Isomer 2 was formulated for administration as an aqueous solution containing 5% dimethylsulfoxide, 95% sulfobutylether- ⁇ -cyclodextrin (30% w/v) in purified water for injection (WFI), at a dose level of 0.5 mg/kg (I.V.) animal bodyweight, and dose volume of 1.0 ml/kg animal bodyweight.
  • WFI purified water for injection
  • Blood samples ( ⁇ 0.20 mL / timepoint) were collected from the jugular vein into tubes containing EDTA-K2 anticoagulant, and placed on wet ice (4°C) before centrifugation (4000 g, 5 minutes, 4 oC) to obtain plasma, and then stored (-75 oC +/- 15 oC) prior to further analysis.
  • FIG. 17 shows the pharmacokinetic profiles in male Han Wistar rats (total blood plasma concentration) of Compound 137, Isomer 2 following administration I.V. (0.5 mg/ kg) to either intact (solid line) or bile-duct cannulated (dashed line) animals.
  • Example C-27 In Vivo Pharmacokinetic Studies in the Han Wistar Rat at High Dose [00325] Pharmacokinetic profiles of Compound 137, Isomer 2 in male Han Wistar rats were obtained following oral administration to establish the short-term tolerability, and confirm the extent and duration of compound exposure at three dose levels (10 mg/kg, 30 mg/kg, and 100 mg/kg) and a dose volume of 10 mL/kg. Animal husbandry, dosing, sample collection, and subsequent analytical work to determine total plasma concentrations were conducted in a similar manner as previously described In Example C-24, with minor adjustments. Plasma samples were collected from the lateral tail vein at multiple timepoints (15 / 30 / 60 / 120 / 240 / 360 / 1440 minutes) following administration.
  • Table 18 shows the pharmacokinetic profiles in male Han Wistar rats (total blood plasma concentration) of Compound 137, Isomer 2 following administration P.O. at three dose levels: 10 mg/kg (solid line), 30 mg/kg (dashed line), and 100 mg/kg (dotted line).
  • Example C-28 In Vivo Pharmacokinetic Studies at High Dose to Assess Substance Tolerability and Effect of Repeat Dosing [00326] Pharmacokinetic profiles of Compound 137, Isomer 2 in female SCID mice were obtained following repeated daily oral administration to establish tolerability over 28 days, and confirm the extent and duration of compound exposure. Animal husbandry, dosing, sample collection, and subsequent analytical work to determine total plasma concentrations were conducted in a similar manner as previously described in Example C-24, with minor adjustments. Animals were dosed twice daily (8 hours apart) and a dose volume of 10 mL/kg. Plasma samples were collected at multiple timepoints (30 / 12 / 360 / 720 / 1440 minutes) following the first dose on day 1 and day 28.
  • Example C-29 In Vivo Pharmacokinetic Studies Comparing Substance Formulations in the SCID Mouse at High Dose [00327] Pharmacokinetic profiles of Compound 137, Isomer 2 in SCID mice were obtained following oral administration to establish the short-term tolerability at high dose, and confirm the extent and duration of compound exposure. Animal husbandry, dosing, sample collection, and subsequent analytical work to determine total plasma concentrations were conducted in a similar manner as previously described in Example C-24, with minor adjustments.
  • Compound 137, Isomer 2 formulated in either 5% DMSO, 50% of 20% Captisol, and 45% purified water for injection (WFI), pH 3-3.2), or 0.5% HPMC/0.1% Tween was dosed at 300 mg/kg and a dose volume of 10 mL/kg. Plasma samples were collected at multiple timepoints (30 / 12 / 360 / 720 / 1440 minutes) following administration.
  • Table 21 provides a summary of pharmacokinetic parameters derived following oral administration of Compound 137, Isomer 2 formulated in either 5% DMSO, 50% of 20% Captisol, and 45% WFI (pH 3-3.2), or 0.5% HPMC/0.1% Tween 80 to female SCID mice at 300 mg/kg.
  • the present disclosure encompasses not only the main group, but also the main group absent one or more of the group members.
  • the present disclosure also envisages the explicit exclusion or disclaimer of one or more of any of the group members in the claimed disclosure.
  • all ranges disclosed herein also encompass any and all possible subranges and combinations of subranges thereof as well as the individual values making up the range, particularly integer values. Any listed range can be easily recognized as sufficiently describing and enabling the same range being broken down into at least equal halves, thirds, quarters, fifths, tenths, etc.
  • each range discussed herein can be readily broken down into a lower third, middle third and upper third, etc.
  • the range C (1-6) includes the subranges C (2-6) , C (3-6) , C (3-5) , C (4-6) , etc., as well as C 1 (methyl), C 2 (ethyl), C 3 (propyl), C 4 (butyl), C 5 (pentyl) and C 6 (hexyl) individually.

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Abstract

L'invention concerne des pyrrolopyrimidine carboxamides et des sels pharmaceutiquement acceptables de ceux-ci ; des compositions pharmaceutiques comprenant de tels composés et sels ; l'utilisation de tels composés et sels pour traiter ou prévenir des cancers, y compris des cancers dépendant de PKMYT1 ; des kits comprenant de tels composés et sels ; et des procédés de fabrication de tels composés et sels.
PCT/IB2024/055444 2023-06-05 2024-06-04 Pyrrolopyrimidine carboxamides Ceased WO2024252276A1 (fr)

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EP24734130.8A EP4720066A1 (fr) 2023-06-05 2024-06-04 Pyrrolopyrimidine carboxamides

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN119751461A (zh) * 2024-12-27 2025-04-04 中国药科大学 一种pkmyt1抑制剂及其制备方法和应用
EP4543881A4 (fr) * 2022-06-23 2026-05-20 Engine Biosciences Pte Ltd Composés et procédé d'inhibition de pkmyt1

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021195781A1 (fr) * 2020-04-01 2021-10-07 Repare Therapeutics Inc. Composés, compositions pharmaceutiques et procédés de préparation et d'utilisation associés
WO2023174329A1 (fr) * 2022-03-15 2023-09-21 Insilico Medicine Ip Limited Composés hétéroaromatiques utilisés comme inhibiteurs de pkmyt1 et leur utilisation

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2021195781A1 (fr) * 2020-04-01 2021-10-07 Repare Therapeutics Inc. Composés, compositions pharmaceutiques et procédés de préparation et d'utilisation associés
WO2023174329A1 (fr) * 2022-03-15 2023-09-21 Insilico Medicine Ip Limited Composés hétéroaromatiques utilisés comme inhibiteurs de pkmyt1 et leur utilisation

Non-Patent Citations (24)

* Cited by examiner, † Cited by third party
Title
"Optimized chemical proteomics assay for kinase inhibitor profiling", J. PROTEOME RES, vol. 15, 2015, pages 1574 - 86, Retrieved from the Internet <URL:https://pubs.acs.ore/doi/10.1021/pr5012608,>
"Quantitative chemical proteomics reveals mechanisms of action of clinical ABL kinase inhibitors", NAT. BIOTECHNOL., vol. 25, 2007, pages 1035 - 44, Retrieved from the Internet <URL:htips.//ovwwnatrecom/articles/nbt>
BERGE, S. ET AL.: "Pharmaceutical Salts", J. PHARM. SCI., vol. 66, 1977, pages 1, XP002675560, DOI: 10.1002/jps.2600660104
BIOCHEM PHARMACOL, vol. 22, no. 23, pages 3099 - 108
BO MI KU ET AL.: "Mutational status of TP53 defines the efficacy of Weel inhibitor AZD1775 in KRAS-mutant non-small cell lung cancer.", ONCOTARGET, vol. 8, no. 40, 2017, pages 67526 - 67537
CHEN, X.: "Cyclin E Overexpression Sensitizes Triple-Negative Breast Cancer to Weel Kinase Inhibition", CLINICAL CANCER RESEARCH, vol. 24, no. 24, pages 6594 - 6610, XP093020937, DOI: 10.1158/1078-0432.CCR-18-1446
CHOW, J. P.R. Y. POON: "The CDK1 inhibitory kinase MYT1 in DNA damage checkpoint recovery", ONCOGENE, vol. 32, no. 40, 2013, pages 4778 - 4788, XP037750172, DOI: 10.1038/onc.2012.504
CHOW, J. PR. Y. POON: "The CDK I inhibitory kinase MYT1 in DNA damage checkpoint recovery.", ONCOGENE, vol. 32, no. 40, 2013, pages 4778 - 4788, XP037750172, DOI: 10.1038/onc.2012.504
DOSCHEDA: "a web application for interactive chemoproteomics data analysis", PEERJ COMPUTER SCIENCE, 2017, Retrieved from the Internet <URL:https://peerj.com/articles/cs-129/>
FORMENT, J. V.M. J. O'CONNOR: "Targeting the replication stress response in cancer", PHARMACOL THER, vol. 188, 2018, pages 155 - 167, XP085430789, DOI: 10.1016/j.pharmthera.2018.03.005
FORMENT, J. V.M. J. O'CONNOR: "Targeting the replication stress response in cancer.", PHARMACOL THER, vol. 188, 2018, pages 155 - 167, XP085430789, DOI: 10.1016/j.pharmthera.2018.03.005
LALLO, A. ET AL.: "The Combination of the PARP Inhibitor Olaparib and the WEE1 Inhibitor AZD1775 as a New Therapeutic Option for Small Cell Lung Cancer", CLIN CANCER RES, vol. 24, no. 20, 2018, pages 5153 - 5164, XP055875904, DOI: 10.1158/1078-0432.CCR-17-2805
LALLO, A. ET AL.: "The Combination of the PARP Inhibitor Olaparib and the WEE1 Inhibitor AZD1775 as a New Therapeutic Option for Small Cell Lung Cancer.", CLIN CANCER RES, vol. 24, no. 20, 2018, pages 5153 - 5164, XP055875904, DOI: 10.1158/1078-0432.CCR-17-2805
LIU, J. F. ET AL.: "Phase II Study of the WEE 1 Inhibitor Adavosertib in Recurrent Uterine Serous Carcinoma.", JOURNAL OF CLINICAL ONCOLOGY, vol. 39, no. 14, 2021, pages 1531 - 1539
LIU, J. F. ET AL.: "Phase II Study of the WEE1 Inhibitor Adavosertib in Recurrent Uterine Serous Carcinoma", JOURNAL OF CLINICAL ONCOLOGY, vol. 39, no. 14, 2021, pages 1531 - 1539
MCMILLIAN, MK ET AL.: "An improved resazurin-based cytotoxicity assay for hepatic cells", CELL BIOL. & TOXICOL., vol. 18, 2002, pages 157 - 173, XP037832573, DOI: 10.1023/A:1015559603643
O'BRIEN, J ET AL.: "Investigation of the Alamar Blue (resazurin) fluorescent dye for the assessment of mammalian cell toxicity", EUR J BIOCHEM., vol. 267, 2000, pages 5421 - 5426, XP002968559, DOI: 10.1046/j.1432-1327.2000.01606.x
PFISTER, SOPHIA X. ET AL.: "Inhibiting WEE1 Selectively Kills Histone H3K36me3-Deficient Cancers by dNTP Starvation.", CANCER CELL, vol. 28, no. 5, 2015, pages 557 - 568, XP093147590, DOI: 10.1016/j.ccell.2015.09.015
RICHER, A. L. ET AL.: "WEE1 Kinase Inhibitor AZD1775 Has Preclinical Efficacy in LKB1-Deficient Non-Small Cell Lung Cancer.", CANCER RESEARCH, vol. 77, no. 17, 2017, pages 4663 - 4672
SERRA, V. ET AL.: "Identification of a Molecularly-Defined Subset of Breast and Ovarian Cancer Models that Respond to WEE 1 or ATR Inhibition, Overcoming PARP Inhibitor Resistance.", CLIN CANCER RES, vol. 28, no. 20, 2022, pages 4536 - 4550
SERRA, V. ET AL.: "Identification of a Molecularly-Defined Subset of Breast and Ovarian Cancer Models that Respond to WEE1 or ATR Inhibition, Overcoming PARP Inhibitor Resistance", CLIN CANCER RES, vol. 28, no. 20, 2022, pages 4536 - 4550
T.W. GREENEP.G.M. WUTZ: "Protective Groups in Organic Synthesis", 1999, PHARMACEUTICAL PRESS
Y. P. KOK ET AL.: "Overexpression of Cyclin E1 or Cdc25A leads to replication stress, mitotic aberrancies, and increased sensitivity to replication checkpoint inhibitors.", ONCOGENESIS, vol. 9, 2020
YOUNG, L. A. ET AL.: "Differential Activity of ATR and WEE1 Inhibitors in a Highly Sensitive Subpopulation of DLBCL Linked to Replication Stress.", CANCER RESEARCH, vol. 79, no. 14, 2019, pages 3762 - 3775

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP4543881A4 (fr) * 2022-06-23 2026-05-20 Engine Biosciences Pte Ltd Composés et procédé d'inhibition de pkmyt1
CN119751461A (zh) * 2024-12-27 2025-04-04 中国药科大学 一种pkmyt1抑制剂及其制备方法和应用

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