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WO2025199434A1 - Methods of treating breast cancer using a pyrazole compound - Google Patents

Methods of treating breast cancer using a pyrazole compound

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Publication number
WO2025199434A1
WO2025199434A1 PCT/US2025/020907 US2025020907W WO2025199434A1 WO 2025199434 A1 WO2025199434 A1 WO 2025199434A1 US 2025020907 W US2025020907 W US 2025020907W WO 2025199434 A1 WO2025199434 A1 WO 2025199434A1
Authority
WO
WIPO (PCT)
Prior art keywords
compound
mdm2
pharmaceutical composition
patient
copy number
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Pending
Application number
PCT/US2025/020907
Other languages
French (fr)
Inventor
David N. Cook
Dung Thai
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Edgewood Oncology
Original Assignee
Edgewood Oncology
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Filing date
Publication date
Application filed by Edgewood Oncology filed Critical Edgewood Oncology
Publication of WO2025199434A1 publication Critical patent/WO2025199434A1/en
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/506—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim not condensed and containing further heterocyclic rings
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • This disclosure relates to methods of treating, preventing, or ameliorating one or more symptoms of breast cancer using a pyrazole compound.
  • the tumor suppressor, p53 is a transcription factor that induces cell cycle arrest and activates apoptotic cell death through target gene activation in response to DNA damage and cellular stress.
  • Fields S Jang SK, Presence of apotent transcription activating sequence in the p53 protein, Science. Aug 31 1990; 249(4972): 1046-9. doi: 10.1126/science.2144363; Kastenhuber ER, Lowe SW.
  • p53 is mutated in about half of solid tumors, cancers that have a wild type p53 must adopt other mechanisms to inhibit p53 activity.
  • Casein kinases are serine/threonine kinases that phosphorylate proteins to mediate normal biological functions and malignant transformation.
  • Casein kinase 1 alpha (CKla) functions as a tumor inducer in several cancers through negative regulation of Wnt/p-catenin signaling and p53. Ebert & Krbnke, N. Engl. J. Med. 2018, 379, 1873-1874.
  • CKla phosphorylates [3-catenin at serine 45, leading to ubiquitination and degradation of P-catenin.
  • CKla also phosphorylates murine double minute X (MDMX) at serine 289, resulting in enhanced binding of MDMX to p53. Wu et al., Mol. Cell. Biol. 2012, 32, 4821- 4832. Additionally, a complex of CKla and mouse double minute 2 homolog (MDM2) inhibits p53. Elyada et al., Nature 2011, 470, 409-413.
  • MDMX murine double minute X
  • MDM2 mouse double minute 2 homolog
  • SEs Super-enhancers
  • Tumor cells acquire SEs at key oncogenes like c-Myc and at genes that produce the hallmark features of cancer. Hnisz et al. 2013. Given the dependency on high levels of gene expression, tumor cells are particularly sensitive to disturbances in transcription.
  • Transcriptional kinases are essential components of the transcription apparatus.
  • CDK7 cyclin dependent kinase 7
  • CTD carboxy terminal domain
  • RNA Pol II ribonucleic acid polymerase II
  • MDM2 is a negative regulator of p53 that promotes p53 ubiquitination and subsequent degradation.
  • Haupt Y, Maya R, Kazaz A, Oren M. Mdm2 promotes the rapid degradation of p53. Nature. 1997/05/01 1997;387(6630):296-299. doi:10.1038/387296a0
  • One mechanism for inhibiting wild type p53 is to amplify the gene expression of MDM2 and thus increase the turnover of p53. Mutations that result in increased expression of MDM2 may be in transcription factors or DNA sequences, e.g., selecting for changes in promoter or enhancer function that increase MDM2 transcription.
  • Tumors that utilize Super Enhancer- mediated transcription have been characterized as “transcriptionally addicted” due to their need for high rates of expression of proteins with short half-lives, such as MDM2.
  • MDM2 gene amplification is observed in almost all patients and activates p53-independent growth and stress response genes.
  • Bevill SM Casani-Galdon S, El Farran CA, et al. Impact of supraphysiologic MDM2 expression on chromatin networks and therapeutic responses in sarcoma. Cell Genom. Jul 12 2023;3(7): 100321. doi: 10.1016/j.xgen.2023.10032).
  • liposarcoma is the canonical example of MDM2 gene amplification, the phenomenon occurs in tumors originating from many different tissues.
  • MDM2 genes are amplified by 4-fold or more include, but are not limited to, breast, non-small-cell lung cancer, bladder (urothelial), stomach, esophageal, melanoma, prostate, cholangiocarcinoma, pancreatic, colorectal, salivary gland, and cervical. Furthermore, a higher copy number of MDM2, e.g., >8, is associated with decreased survival rates.
  • MDM2 inhibitors For example, Rain Oncology tested milademetan in MDM2-amplified solid tumors (clinicaltrials.gov NCT05012397). This multicenter, single-arm, open-label basket study was designed to evaluate the safety and efficacy of milademetan in patients with advanced or metastatic solid tumors refractory or intolerant to standard-of-care therapy that exhibit wildtype (WT) TP53 and MDM2 copy number (CN) > 8 using prespecified biomarker criteria. Interim results for this study showed intriguing anti-tumor activity.
  • WT wildtype
  • CN MDM2 copy number
  • Rates of grade 3/4 hematologic toxicity were significant and reported as follows: 39.5% of patients with thrombocytopenia; 25.5% of patients with neutropenia; and 18.6% of patients with anemia.
  • brigimadlin is dosed infrequently with a long treatment holiday in an attempt to overcome toxicity. As monotherapy, 45 mg is administered only on the first day of a 21-day cycle.
  • brigimadlin (BI 907828), an MDM2- -p53 antagonist, in patients (pts) with advanced biliary tract cancer: Data from two phase la/Ib dose-escalation/expansion trials, Journal of Clinical Oncology Volume 42, Number 3_supp).
  • Early results for this agent reported the following rates of Grade 3/4 hematologic toxicity: 19% thrombocytopenia; 20% neutropenia; 10% anemia. While these data appear to be improved compared to milademetan, there remains a need to provide better tolerated agents that can achieve continuous pharmacologic coverage of the drug target.
  • Sirimadlin is a direct MDM2 inhibitor that was evaluated in 115 solid tumor patients and 93 patients with hematologic malignancies.
  • Grade 3/4 Adverse Events suspected to be treatment related were observed in 52 (45%) patients with solid tumors and 66 (71%) patients with hematologic tumors.
  • Hematologic toxicities were the most common grade 3/4 AE for all indications and were more frequent in patients with hematologic tumors compared with those with solid tumors.
  • thrombocytopenia is an on-target effect of MDM2 inhibition.
  • GATA3 is a transcription factor that is important in maintaining the normal differentiation status of luminal cells in mammary tissue. Kouros-Mehr et al., GATA-3 maintains the differentiation of the luminal cell fate in the mammary gland, Cell 2006 127(5): 1041-55.
  • GATA3 Women with mutations in GATA3 have worse progression-free and overall survival (Velimirovic et al., Landscape of GATA3 mutations identified from circulating tumor DNA clinical testing and their impact on disease outcomes in estrogen receptor-positive (ER+) metastatic breast cancers treatedwith endocrine therapies, J. Clin. Oncol. 2021 39(suppl 15): abstract 1065) and, importantly, GAT A3 mutations downstream of the splice site at amino acid 307 are mutually exclusive with wild-type P53 and demonstrate higher levels of MDM2 expression than other mutational subgroups (Li et al. Genomic Alteration in Metastatic Breast Cancer and Its Treatment, Am. Soc. Clin. Oncol. Educ. Book 2020 40: 1-14).
  • MDM2 plays an essential role in tumor cell survival.
  • Pharmacologic inhibition of MDM2 in an ER+ GATA3mt background leads to tumor cell growth inhibition in both cell lines and patient-derived xenografts and induces apoptosis in vitro.
  • Bianco et al., GATA3 andMDM2 are synthetic lethal in estrogen receptor-positive breast cancers, Commun. Biol. 2022 5(1):373.
  • This disclosure provides methods of treating a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
  • the breast cancer is refractory. In some embodiments, the breast cancer is metastatic. In some embodiments, the breast cancer is drug-resistant. In some embodiments, the patient has failed a prior therapy. In some embodiments, the patient is a human.
  • the compound is formulated as a -toluenesulfonate salt or a di- -toluenesulfonate salt.
  • the pharmaceutical composition is administered orally.
  • the pharmaceutical composition is administered as a tablet or capsule.
  • the therapeutically effective amount is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 25 mg, about 30 mg, or about 35 mg.
  • the pharmaceutical composition is administered in a cycle. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle. In some embodiments, the pharmaceutical composition is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week. In some embodiments, the pharmaceutical composition is administered for 3 days or 5 days per week. In some embodiments, the pharmaceutical composition is administered on three non-consecutive days in a week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 3 days per week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 3 non-consecutive days per week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 5 days per week.
  • the pharmaceutical composition is administered in a 28-day cycle on Days 1, 2, 3, 4, and 5 per week.
  • the therapeutically effective amount is about 21 mg. In some embodiments, the therapeutically effective amount is about 30 mg. In some embodiments, the therapeutically effective amount is about 35 mg.
  • this disclosure provides methods of inhibiting MDM2 in a patient having ER+, HER- metastatic breast cancer and having GATA3mt, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
  • the compound is formulated as a p-toluenesulfonate salt. In some embodiments, the compound is formulated as a di- -toluenesulfonate salt.
  • the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg.
  • the pharmaceutical composition is administered three times per week.
  • the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
  • this disclosure provides methods of inhibiting CDK7 in a patient having ER+, HER- metastatic breast cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
  • the compound is formulated as a -toluenesulfonate salt. In some embodiments, the compound is formulated as a di- -toluenesulfonate salt. [0030] In some embodiments, the breast cancer has been resistant to a CDK4/6 inhibitor.
  • the patient has TP53wt. In some embodiments, the patient has GATA3wt. In some embodiments, the patient has GATA3mt.
  • the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
  • Compound A22 is administered three times per week. In some embodiments, the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg per day.
  • this disclosure provides methods of treating cancer in a patient having amplified expression of MDM2 or having a >4 copy number of the MDM2 gene, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
  • the compound is formulated as a -toluenesulfonate salt or a di- -toluenesulfonate salt.
  • this disclosure provides methods of inhibiting MDM2 expression in a patient having cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
  • the patient has breast cancer with GATA3mt.
  • the compound is formulated as -toluenesulfonate salt or a di-/?-toluenesulfonate salt.
  • the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
  • administering comprises administering about 21 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 30 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 35 mg of Compound A22 three times per week.
  • compositions comprising Compound A22, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, for use in the treatment of a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer.
  • ER+ Estrogen Receptor positive
  • HER2- Human Epidermal Growth Factor Receptor 2 negative
  • the patient has GAT3mt.
  • the treatment comprises administration of the pharmaceutical composition to the patient on three non-consecutive days in a week. In some embodiments of the use, the administration occurs for one or more cycles. In some embodiments, the cycle is 28 days.
  • the pharmaceutical composition is orally administered to the patient.
  • the pharmaceutical composition is formulated as an oral unit dosage form. In some embodiments, the oral unit dosage form comprises about 21 mg of Compound A22. In some embodiments, the oral unit dosage form comprises about 30 mg of Compound A22. In some embodiments, the oral unit dosage form comprises about 35 mg of Compound A22.
  • this disclosure provides a method of inducing apoptosis in an ER+ HER2- breast cancer cell, comprising contacting the cell with an effective amount of a compound of Compound A22, or a pharmaceutically acceptable salt thereof.
  • FIGS. 1A and IB shows the results of inhibition or suppression of certain biomarkers from application of Compound A22.
  • FIGS. 2A and IB shows the results of inhibition or suppression of certain biomarkers from application of Compound A22.
  • FIGS. 3 A and 3B show the results of CDK2 and phospho-CDK2 signaling in the presence of Compound A22.
  • subject refers to an animal, including, but not limited to, a primate (e.g., human), cow, pig, sheep, goat, horse, dog, cat, rabbit, rat, or mouse.
  • primate e.g., human
  • cow, pig, sheep, goat horse
  • dog cat
  • rabbit rat
  • patient are used interchangeably herein in reference, for example, to a mammalian subject, such as a human subject.
  • the subject is a human.
  • treat includes alleviating, improving, inhibiting the progression of, or abrogating a disorder, disease, or condition, or one or more of the symptoms associated with the disorder, disease, or condition; or alleviating, improving, inhibiting the progression of, or eradicating the cause(s) of the disorder, disease, or condition itself. Treating includes curing, improving, or at least partially ameliorating the disorder, disease, or condition.
  • prevent are meant to include a method of delaying and/or precluding the onset of a disorder, disease, or condition, and/or its attendant symptoms; barring a subject from acquiring a disorder, disease, or condition; or reducing a subject’s risk of acquiring a disorder, disease, or condition.
  • the terms “alleviate” and “alleviating” refer to easing or reducing one or more symptoms (e.g., pain) of a disorder, disease, or condition.
  • the terms can also refer to reducing adverse effects associated with an active ingredient.
  • the beneficial effects that a subject derives from a prophylactic or therapeutic agent do not result in a cure of the disorder, disease, or condition.
  • contacting or “contact” is meant to refer to bringing together of a therapeutic agent and cell or tissue such that a physiological and/or chemical effect takes place as a result of such contact. Contacting can take place in vitro, ex vivo, or in vivo.
  • a therapeutic agent is contacted with a cell in cell culture (in vitro to determine the effect of the therapeutic agent on the cell.
  • the contacting of a therapeutic agent with a cell or tissue includes the administration of a therapeutic agent to a subject having the cell or tissue to be contacted.
  • terapéuticaally effective amount or “effective amount” is meant to include the amount of a compound that, when administered, is sufficient to treat, prevent development of, or alleviate a disorder, disease, or condition, or one or more of the symptoms of the disorder, disease, or condition being treated.
  • therapeutically effective amount or “effective amount” also refers to the amount of a compound that is sufficient to elicit a biological or medical response of a biological molecule (e.g., a protein, enzyme, RNA, or DNA), cell, tissue, system, animal, or human, which is being sought by a researcher, veterinarian, medical doctor, or clinician.
  • a biological molecule e.g., a protein, enzyme, RNA, or DNA
  • pharmaceutically acceptable carrier refers to a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, solvent, or encapsulating material.
  • each component is “pharmaceutically acceptable” in the sense of being compatible with the other ingredients of a pharmaceutical formulation, and suitable for use in contact with the tissue or organ of a subject (e.g., a human or an animal) without excessive toxicity, irritation, allergic response, immunogenicity, or other problems or complications, commensurate with a reasonable benefit/risk ratio.
  • the term “about” or “approximately” means an acceptable error for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined. In certain embodiments, the term “about” or “approximately” means within 1, 2, 3, or 4 standard deviations. In certain embodiments, the term “about” or “approximately” means within 50%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.05% of a given value or range.
  • solvate refers to a complex or aggregate formed by one or more molecules of a solute, e.g., a compound provided herein, and one or more molecules of a solvent, which are present in stoichiometric or non-stoichiometric amount.
  • Suitable solvents include, but are not limited to, water, methanol, ethanol, ⁇ -propanol, isopropanol, and acetic acid.
  • the solvent is pharmaceutically acceptable.
  • the complex or aggregate is in a crystalline form.
  • the complex or aggregate is in a noncrystalline form.
  • the solvent is water
  • the solvate is a hydrate. Examples of hydrates include, but are not limited to, a hemihydrate, monohydrate, dihydrate, trihydrate, tetrahydrate, and pentahydrate.
  • Compound A22 of this disclosure (lr,4r)-A 1 -(5-chloro-4-(5-(cyclopropylmethyl)- 1 -methyl- 177-pyrazol-4-yl)pyrimidin-2-yl)cy cl ohexane-l,4-diamine, has the following structure:
  • Compound A22 is a CKla, CDK7, and CDK9 inhibitor. Minzel et aL, Cell 2018, 775, 1-15. The Compound can be prepared according to the procedures described in Minzel et al, Cell ' 2018, 775, 1-15; or U.S. Pat. Appl. Publ. No. 2018/0214447 Al. The entire contents of those disclosures are incorporated herein by reference. [0060] Compound A22 may also be referred to as compound A51 or BTX-A51.
  • Compound A22 is formulated as a non-hygroscopic crystalline salt of (lr,4r)-A 1 -(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-U/-pyrazol-4- yl)pyrimidin-2-yl)cyclohexane-l,4-diamine with an acid, or a pharmaceutically acceptable solvate thereof.
  • Compound A22 is formulated as a non-hygroscopic crystalline tosylate salt.
  • Compound A22 is formulated as a non- hygroscopic crystalline ditosylate salt.
  • the disclosure provides Compound A22, chloro-4-(5-(cyclopropylmethyl)- 1 -methyl- U/-pyrazol-4-yl)pyrimidin-2-yl)cyclohexane- 1 ,4- diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
  • Compound A22 is provided as a -toluensulfonate of (lr,4r)-A 1 -(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-lJ7-pyrazol-4-yl)pyrimidin-2- yl)cyclohexane-l,4-diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically solvate or hydrate.
  • Compound A22 is provided as a di- -toluensulfonate of (lr,4r)-A 1 -(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-lJ7-pyrazol-4-yl)pyrimidin-2- yl)cyclohexane-l,4-diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically solvate or hydrate.
  • the -toluenesulfonate and di- -toluensulfonate salts are described in U.S. Pub. No.
  • Compound A22 is provided as a toyslate salt. In some embodiments, Compound A22 is provided as a ditosylate salt.
  • Compound A22 is isolated or purified.
  • a compound described herein has a purity of at least about 50%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 98%, at least about 99%, or at least about 99.5% by weight.
  • the compounds described herein are intended to encompass all possible stereoisomers unless a particular stereochemistry is specified.
  • the compound may exist as one or mixture of geometric cisltrans (or ZZE) isomers.
  • structural isomers are interconvertible, the compound may exist as a single tautomer or a mixture of tautomers. This can take the form of proton tautomerism in the compound that contains, for example, an imino, keto, or oxime group; or so-called valence tautomerism in the compound that contain an aromatic moiety. It follows that a single compound may exhibit more than one type of isomerism.
  • Compound A22 can be enantiomerically pure, such as a single enantiomer or a single diastereomer, or be stereoisomeric mixtures, such as a mixture of enantiomers, e.g., a racemic mixture of two enantiomers; or a mixture of two or more diastereomers.
  • a compound in its (A) form is equivalent, for compounds that undergo epimerization in vivo, to administration of the compound in its (5) form.
  • Conventional techniques for the preparation/isolation of individual enantiomers include synthesis from a suitable optically pure precursor, asymmetric synthesis from achiral starting materials, or resolution of an enantiomeric mixture, for example, chiral chromatography, recrystallization, resolution, diastereomeric salt formation, or derivatization into diastereomeric adducts followed by separation.
  • Compound A22 can also be provided as a pharmaceutically acceptable salt. Salts of Compound A22 are described in U.S. Pub. No. 2022/0332702, the entire contents of which are incorporated herein by reference.
  • Compound A22 may also be provided as a prodrug, which is a functional derivative of a compound, and is readily convertible into the parent compound in vivo.
  • Prodrugs are often useful because, in some situations, they may be easier to administer than the parent compound. They may, for instance, be bioavailable by oral administration whereas the parent compound is not.
  • the prodrug may also have enhanced solubility in pharmaceutical compositions over the parent compound.
  • a prodrug may be converted into the parent drug by various mechanisms, including enzymatic processes and metabolic hydrolysis.
  • compositions can be prepared, isolated, or obtained by any method known to one of ordinary skill in the art, for example, by following the procedures described in U.S. Pat. No. 10,376,511.
  • compositions comprising Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and a pharmaceutically acceptable excipient.
  • a pharmaceutical composition comprises the -toluenesulfonate or di- -toluensulfonate salt of Compound A22.
  • a pharmaceutical composition provided herein can be formulated in various dosage forms, including, but not limited to, dosage forms for oral, parenteral, and topical administration.
  • the pharmaceutical composition can also be formulated as modified release dosage forms, including delayed-, extended-, prolonged-, sustained-, pulsatile-, controlled-, accelerated-, fast-, targeted-, programmed-release, and gastric retention dosage forms.
  • These dosage forms can be prepared according to conventional methods and techniques known to those skilled in the art. See, e.g., Remington: The Science and Practice of Pharmacy, supra, Modified-Release Drug Delivery Technology, 2nd ed.; Rathbone et al., Eds.; Drugs and the Pharmaceutical Sciences 184; CRC Press: Boca Raton, FL, 2008.
  • a pharmaceutical composition provided herein is formulated in a dosage form for oral administration.
  • the pharmaceutical composition is a tablet or a capsule.
  • a pharmaceutical composition provided herein is formulated in a dosage form for parenteral administration.
  • a pharmaceutical composition provided herein is formulated in a dosage form for intravenous administration.
  • a pharmaceutical composition provided herein is formulated in a dosage form for intramuscular administration.
  • a pharmaceutical composition provided herein is formulated in a dosage form for subcutaneous administration.
  • a pharmaceutical composition provided herein is formulated in a dosage form for topical administration.
  • a pharmaceutical composition provided herein can be provided in a unit-dosage form or multiple-dosage form.
  • a unit-dosage form refers to physically discrete a unit suitable for administration to a subject, and packaged individually as is known in the art. Each unit-dose contains a predetermined quantity of an active ingredient(s) (e.g., a compound provided herein) sufficient to produce the desired therapeutic effect, in association with the required pharmaceutical excipient(s).
  • an active ingredient(s) e.g., a compound provided herein
  • Examples of a unit-dosage form include, but are not limited to, an ampoule, syringe, and individually packaged tablet and capsule.
  • a unitdosage form may be administered in fractions or multiples thereof.
  • a multiple-dosage form is a plurality of identical unit-dosage forms packaged in a single container to be administered in a segregated unit-dosage form.
  • Examples of a multiple-dosage form include, are not limited to, a vial, bottle of tablets or capsules, or bottle of pints or gallons.
  • a pharmaceutical composition provided herein can be administered at once or multiple times at intervals of time. It is understood that the precise dosage and duration of treatment may vary with the age, weight, and condition of the subject being treated, and may be determined empirically using known testing protocols or by extrapolation from in vivo or in vitro test or diagnostic data. It is further understood that for any particular individual, specific dosage regimens should be adjusted over time according to the subject’s need and the professional judgment of the person administering or supervising the administration of the pharmaceutical composition.
  • a pharmaceutical composition provided herein comprises Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and sugar beads, talc, and povidone.
  • a pharmaceutical composition provided herein comprises Compound A22, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and sugar beads, talc, and povidone.
  • a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt; and sugar beads, talc, and povidone.
  • a pharmaceutical composition comprises a tosylate salt of Compound A22.
  • a pharmaceutical composition comprises a ditosylate salt of Compound A22.
  • the pharmaceutical composition is formulated as a capsule. In some embodiments, the pharmaceutical composition is formulated as a tablet.
  • a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.1 to about 50, from about 0.2 to about 40, from about 0.5 to about 35, or from about 0.5 to about 30 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.1 to about 50 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.2 to about 35 mg per capsule or tablet.
  • a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.5 to about 30 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.5 to about 5 mg per capsule.
  • a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount of about 0.5, about 1, or about 2 mg per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 21 mg of Compound A22 per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 30 mg of Compound A22 per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 35 mg of Compound A22 per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 or a di- -toluenesulfonate salt of Compound A22; and sugar beads, talc, and povidone.
  • the pharmaceutical composition is formulated as a capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50, from about 0.2 to about 20, from about 0.5 to about 10, or from about 0.5 to about 5 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.2 to about 40 mg per capsule.
  • a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 30 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 25 mg per capsule.
  • a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises a p- toluenesulfonate salt of Compound A22 in an amount of about 21, 30, or 35 mg per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22; and sugar beads, talc, and povidone.
  • the pharmaceutical composition is formulated as a capsule.
  • a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50, from about 0.2 to about 20, from about 0.5 to about 10, or from about 0.5 to about 5 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.2 to about 40 mg per capsule.
  • a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 30 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- - toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 25 mg per capsule.
  • a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount of about 21, 30, or 35 mg per capsule or tablet or unit oral dosage form.
  • a pharmaceutical composition provided herein is formulated as an immediate-release capsule with a size of, e.g., size 1.
  • This disclosure provides methods of treating Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of Compound A22.
  • This disclosure is based on the surprising discovery of multiple mechanisms of action of Compound A22.
  • the methods of this disclosure are based on the discovery that Compound A22 inhibits CKla, inducing p53 activation and apoptosis, and also inhibits CDK7 and CDK9, which inhibits MDM2 expression and MCL1 expression, thereby also activating p53 and inducing apoptosis.
  • the methods of this disclosure exploit the surprising synergistic ability of Compound A22 to effect multiple mechanisms of action.
  • the methods of this disclosure are further based on the discovery that in patients with GATA3mt, the P53 gene is almost always wild type. Treating GATA3mt tumors in preclinical studies with MDM2 inhibitors and MDM2 siRNAs has shown that these tumors, unlike GATA3wt tumors, are sensitive to MDM2 inhibition or knockdown. Thus, GATA3 mutations are synthetic lethal with MDM2 inhibition. Despite this observation and the fact that patients with GATA3 mutations have poor survival outcomes, there are no approved agents approved for the treatment of GAT A3 mt ER+ Her2- breast cancer.
  • the breast cancer is refractory. In some embodiments, the breast cancer is metastatic. In some embodiments, the breast cancer is drug-resistant. In some embodiments, prior therapy has failed to stop progression of the breast cancer.
  • the subject has GATA3/7?/. In some embodiments, the subject has p53vi7.
  • the breast cancer is drug-resistant.
  • the breast cancer is resistant to CDK4/6 inhibitors, anti-estrogen therapies including aromatase inhibitors, PI3 kinase inhibitors, mTOR inhibitors, taxane chemotherapy, fluoropyrimidine chemotherapy, anti-HER2 antibodies, and/or antibody drug conjugates targeting HER2 or TROP2.
  • the subject is newly diagnosed with GAT A3 mt breast cancer.
  • the subject has failed a prior therapy.
  • the subject has failed more than one prior therapy.
  • a prior therapy has failed to stop progression of the breast cancer.
  • the subject is a mammal. In certain embodiments, the subject is a human.
  • Methods provided herein encompass treating a subject regardless of patient’s age, although some diseases are more common in certain age groups.
  • the therapeutically effective amount of Compound A22 is ranging from about 0.001 to about 10 mg/kg per day, from about 0.002 to about 5 mg/kg per day, from about 0.005 to about 2 mg/kg per day, from about 0.01 to about 1 mg/kg per day, or from about 0.01 to about 0.5 mg/kg per day. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.001 to about 10 mg/kg per day. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.002 to about 5 mg/kg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.005 to about 2 mg/kg per day.
  • the therapeutically effective amount of Compound A22 ranges from about 0.01 to about 1 mg/kg per day. In yet another embodiment, the therapeutically effective amount of a Compound A22 ranges from about 0.01 to about 0.5 mg/kg per day. In still another embodiment, the therapeutically effective amount of Compound A22, is about 0.01, about 0.02, about 0.03, about 0.05, about 0.08, about 0.1, about 0.12, about 0.15, about 0.17, about 0.2, or about 0.25 mg/kg per day.
  • the therapeutically effective amount of Compound A22 ranges from about 0.1 to about 200 mg per day, from about 0.2 to about 100 mg per day, from about 0.5 to about 50 mg per day, or from about 1 mg every other day to about 20 mg per day. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.1 to about 200 mg per day. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.2 to about 100 mg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.5 to about 50 mg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 1 to about 20 mg per day.
  • the therapeutically effective amount of Compound A22 is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, or about 40 mg per day.
  • the therapeutically effective amount of Compound A22 ranges from about 1 to 500, from about 2 to 250, from about 5 to about 105, from about 10 to about 90, or from about 20 to about 65 mg per week. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 1 to 500 mg per week. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 2 to 250 mg per week. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 5 to about 105 per week. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 10 to about 90 mg per week.
  • the therapeutically effective amount of Compound A22 ranges from about 10 to about 63 or 65 mg per week. In still another embodiment, the therapeutically effective amount of Compound A22, is about 5, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 50, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, about 100, about 105 mg per week.
  • the compound is administered at a dose of about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 28 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 40 mg, about 42 mg, about 45 mg, about 48 mg, about 51 mg, about 52 mg, about 55 mg, about 58 mg, about 60 mg, about 62 mg, about
  • the compound is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week.
  • the administered dose of Compound A22 can also be expressed in units other than mg/kg every other day.
  • doses for parenteral administration can be expressed as mg/m 2 per day.
  • doses for parenteral administration can be expressed as mg/m 2 per day.
  • One of ordinary skill in the art would readily know how to convert doses from mg/kg per day to mg/m 2 per day to given either the height or weight of a subject or both. For example, a dose of 1 mg/m 2 per day for a 65 kg human is approximately equal to 58 mg/kg per day.
  • Compound A22 may be administered by oral, parenteral (e.g., intramuscular, intraperitoneal, intravenous, CIV, intracistemal injection or infusion, subcutaneous injection, or implant), inhalation, nasal, vaginal, rectal, sublingual, or topical (e.g., transdermal or local) routes of administration.
  • parenteral e.g., intramuscular, intraperitoneal, intravenous, CIV, intracistemal injection or infusion, subcutaneous injection, or implant
  • inhalation nasal, vaginal, rectal, sublingual, or topical (e.g., transdermal or local) routes of administration.
  • Compound A22 is administered orally. In some embodiments, Compound A22 is administered as tablet or capsule. In another embodiment, Compound A22, is administered parenterally. In yet another embodiment, Compound A22, is administered intravenously. In yet another embodiment, Compound A22, is administered intramuscularly. In yet another embodiment, Compound A22, is administered subcutaneously. In still another embodiment, Compound A22, is administered topically.
  • a compound described herein, e.g., Compound A22 can be delivered as a single dose such as, e.g., a single bolus injection, or oral tablets or pills; or over time such as, e.g., continuous infusion over time or divided bolus doses over time.
  • a compound described herein, e.g., Compound A22 can be administered repetitively if necessary, for example, until the subject experiences stable disease or regression, or until the subject experiences disease progression or unacceptable toxicity. Stable disease or lack thereof is determined by a method known in the art such as evaluation of subject’s symptoms, physical examination, visualization of the cancer that has been imaged using X-ray, CAT, PET, or MRI scan and other commonly accepted evaluation modalities.
  • a compound described herein e.g., Compound A22
  • the administration can be continuous, i.e., every day, or intermittently.
  • the term “intermittent” or “intermittently” as used herein is intended to mean stopping and starting at either regular or irregular intervals.
  • intermittent administration of Compound A22 is administration for one to six days per week, administration in cycles (e.g., daily administration for two to eight consecutive weeks, then a rest period with no administration for up to one week), or administration on alternate days.
  • Compound A22 is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week. In certain embodiments, Compound A22 is administered for 3 days or 5 days per week. In some embodiments, Compound A22 is administered for 3 non- consecutive days per week. In certain embodiments, Compound A22 is administered on days 1, 2, and 3 in a week. In some embodiments, Compound A22 is administered on days 1, 2, 3, 4, and 5 in a week. In some embodiments, Compound A22 is administered on days 1, 3, and 5 in a week.
  • Compound A22 is cyclically administered to a subject to be treated. Cycling therapy involves the administration of the compound for a period of time, followed by a rest for a period of time, and repeating this sequential administration. Cycling therapy can reduce the development of resistance to one or more of the therapies, avoid or reduce the side effects of one of the therapies, and/or improves the efficacy of the treatment.
  • Compound A22 is administered for a cycle of about one week, about two weeks, about three weeks, about four weeks, about five weeks, about six weeks, about eight weeks, or about ten weeks, with a rest period of 0 days to about four weeks. In one embodiment, Compound A22, is administered for a cycle of three weeks, four weeks, five weeks, or six weeks with a rest period of 0, 1, 3, 5, 7, 9, 12, or 14 days. In some embodiments, Compound A22 is administered for a cycle of 21-42 days. In some embodiments, Compound A22 is administered for a cycle of 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 days. In some embodiments, a cycle has no rest period.
  • Compound A22 is administered for consecutive cycles with no rest period between cycles. In some embodiments, Compound A22 is administered for multiple cycles with a rest period. In certain embodiments, the rest period is 7 days. In certain embodiments, the rest period is 14 days. In certain embodiments, the rest period is a period that is sufficient for bone marrow recovery. In certain embodiments, no rest period is required between cycles. The frequency, number, and length of dosing cycles can be increased or decreased.
  • Compound A22 is administered for three weeks in a 28-day cycle with a 7-day rest period. In one embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered every day for five days of a week. In another embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered on Days 1, 2, 3, 4, 5, 8, 9, 10, 11, 12, 15, 16, 17, 18, and 19. In one embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered every other day for three days of a week. In another embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered on Days 1, 3, 5, 8, 10, 12, 15, 17, and 19.
  • Compound A22 is administered 3 days per week for four weeks in a 28-day cycle. In some embodiments, Compound A22 is administered for multiple cycles. In some embodiments, when Compound A22 is administered for multiple cycles, there is a rest period between cycles. In some embodiments when Compound A22 is administered for multiple cycles, there is no rest period between cycles. [00109] In certain embodiments, the subject is treated with Compound A22, from about 1 to about 50, from about 2 to about 20, from about 2 to 10, or from about 4 to about 8 cycles. In certain embodiments, the subject is treated with Compound A22 from about 1 to about 50 cycles. In certain embodiments, the subject is treated with Compound A22 from about 2 to about 20 cycles.
  • the subject is treated with Compound A22 from about 2 to 10 cycles. In certain embodiments, the subject is treated with Compound A22, from about 4 to about 8 cycles. In some embodiments, Compound A22 is administered with a rest period between cycles. In some embodiments, Compound A22 is administered with no rest period between cycles.
  • a method of inhibiting the growth of a cell comprising contacting the cell with an effective amount of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
  • a method of modulating the activity of CKla in a cell comprising contacting the cell with a compound of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
  • a method of inducing apoptosis in a cell comprising contacting the cell with a compound of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
  • the cell is a breast cancer cell. In certain embodiments, the cell is a relapsed or refractory breast cancer cell. In certain embodiments, the cell is a relapsed breast cancer cell. In certain embodiments, the cell is a refractory breast cancer cell. In certain embodiments, the cell is a metastic breast cancer cell. In certain embodiments, the cell is a drug-resistant breast cancer cell.
  • the disclosures relates to one or more of the following enumerated embodiments:
  • a method of treating a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
  • A22 or a pharmaceutically acceptable salt thereof is provided.
  • the method of any one of embodiments 1 to 15, wherein the therapeutically effective amount is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 25 mg, about 30 mg, or about 35 mg.
  • A22 or a pharmaceutically acceptable salt thereof.
  • the method of any one of embodiments 37 to 47, wherein the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg per day.
  • a method of treating cancer in a subject having amplified expression of MDM2 or having a >4 copy number of the MDM2 gene comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
  • the method of embodiment 49 wherein the compound is formulated as a di- - toluenesulfonate salt.
  • the method of any one of embodiments 49 to 50 wherein the subject over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
  • the method of any one of embodiments 49 to 51 wherein the subject over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
  • the method of any one of embodiments 49-53, wherein administering comprises administering about 21 mg of Compound A22 three times per week.
  • the method of any one of embodiments 46-50, wherein administering comprises administering about 30 mg of Compound A22 three times per week.
  • the method of any one of embodiments 46-50, wherein administering comprises administering about 35 mg of Compound A22 three times per week.
  • a method of inhibiting MDM2 expression in a subject having cancer comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
  • the method of embodiment 57 wherein the compound is formulated as a di- - toluenesulfonate salt.
  • the method of any one of embodiments 57 to 60 wherein the subject over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
  • the method of any one of embodiments 54 to 59, wherein administering comprises administering about 21 mg of Compound A22 three times per week.
  • the method of any one of embodiments 54 to 59, wherein administering comprises administering about 30 mg of Compound A22 three times per week.
  • administering comprises administering about 35 mg of Compound A22 three times per week.
  • a pharmaceutical composition comprising Compound A22, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, for use in the treatment of a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer.
  • ER+ Estrogen Receptor positive
  • HER2- Human Epidermal Growth Factor Receptor 2 negative
  • compositions 66-68 The pharmaceutical composition for use according to any one of embodiments 66-68, wherein the administration occurs for one or more cycles.
  • composition 72 for use according to any one of embodiments 66-71, wherein the pharmaceutical composition is formulated as an oral unit dosage form.
  • composition for use according to any one of embodiments 72, wherein the oral unit dosage form comprises about 21 mg of Compound A22.
  • composition for use according to embodiment 72, wherein the oral unit dosage form comprises about 35 mg of Compound A22.
  • Cells were seeded into 96-well plates (Corning 3904) with 100 pL media at 250, 500, 1000, 2000, 5000 and 10,000 cells/well on Day -1. Cell growth was measured by Cell Titer Glow® 2.0 Assay (Promega, G9243) on Days 0, 1, 2 and 3 (total 96 hours) measured with an Envision plate reader. The experiment was performed with 5 replicates.
  • a serial titration of Compound A22 and Milademetan at ten concentrations was performed to determine the IC50 of each compound in the two cell lines.
  • Milademetan is a direct MDM2 inhibitor. Duplicate wells were evaluated for each compound concentration and two biological replicates were performed. Cell growth was measured by Cell Titer Glow® 2.0 Assay.
  • MCF7 cells were seeded into 10 cm cell culture dishes and cultured for 24 hours. Compound A22 or vehicle (DMSO) were added and incubated with cells for 16 hours. After incubation, the media was aspirated and cells were harvested with a trypsin-EDTA solution. Cells were transferred to tubes and washed with cold PBS.
  • Compound A22 or vehicle (DMSO) were added and incubated with cells for 16 hours. After incubation, the media was aspirated and cells were harvested with a trypsin-EDTA solution. Cells were transferred to tubes and washed with cold PBS.
  • the washed cells were lysed by the addition of 500 pL pre-chilled RIPA buffer (Boston BioProducts BP115D) with protease and phosphatase inhibitors (Roche 05892791001; Roche 0490068337001). Cell were incubated for 30 minutes on ice.
  • Lysates were clarified by spinning at 12000 rpm for 10 minutes at 4 degrees and the supernatant was transferred to a fresh tube. Protein concentrations were determined by BCA (Thermo Scientific) and samples were adjusted to the same final protein concentration with RIPA.
  • Samples were combined with a loading buffer and heated at 100°C for 10 minutes before cooling to room temperature. Samples were loaded onto a polyacrylamide gel and run for 20 minutes at 80V, followed by an increase to 120 V for 120 minutes.
  • Results are shown in FIGS. 1 and 2 and were as follows: pRbpl S2/5 was reduced by 73% compared to DMSO control; MCL-1 was reduced by 70% compared to DMSO control; Cleaved PARP was increased by 12 fold and p53 was increased by 5 fold compared to DMSO treatment.
  • Gamma-142 AX a marker of DNA damage response, was increased by approximately 2-fold (data not shown).
  • Rb phosphyorylation was also decreased by Compound A22 treatment. Phosphorylation at S780 was decreased approximately 80% and phosphorylation at Ser807/810 was decreased by approximately 90%. Rb itself was decreased by approximately 50%. However, Actin levels were unchanged, suggesting that these changes in Rb are specific to the mechanism of action of Compound A22 rather than being non-specific effects.
  • Example 3
  • CDK7 is the catalytic component of the CAK (Cyclin Activating Kinase) protein complex. One of its roles is to phosphorylate CDK2 to enable progression from G1 into S phase in the cell cycle.
  • CAK Cyclin Activating Kinase
  • Detection of CDK2 and phospho-CDK2 in cell lysates required the use of an immunoprecipitation step prior to a Western blot analysis.
  • Cells were seeded into culture dishes and cultivated for 24 hours as in Example 3. After treatment with 100 nM Compound A22 for 16 hours at 37°C, cells were processed using trypsin-EDTA solution, washed with cold PBS, and lysed in 0.5 mL ice cold IP -MS Cell Lysis Buffer (ThermoFisher #87787) with protease inhibitor cocktail and phosphatase inhibitor cocktail for cell lysis for 30 min on ice. Samples were clarified by centrifugation as above and the supernatant evaluated for protein concentration by BCA assay. 50 pg of the lysate was retained as input to the immunoprecipitation.
  • Pierce Protein A/G magnetic beads (100 uL per IP) were prepared by washing 3 times in TBST followed by incubation with the anti-Cdk2 antibody (Cell Signaling Technology #2546) for 1 hour at room temperature. The immunoprecipitation was carried out by adding the antibody conjugated magnetic beads to 1 mg of cell lysate. Samples were incubated overnight at 4°C with rotation on a ThermoMixer at 300 rpm. The beads were recovered magnetically and washed 3 times with TBST.
  • the study has two phases: Phases la and lb.
  • safety is evaluated by a Dose Escalation Committee (DEC) that includes the Principal Investigator(s), the Sponsor’ s physician (in consultation with the Sponsor’s pharmacologist/ pharmacokineticist as needed), and/or independent experts.
  • the DEC reviews all cumulative available data and authorizes plans for the dosing of each subsequent cohort. Treatment decisions for individual subjects is the responsibility of the Investigators.
  • Phase la is designed to determine the dose limiting toxi cities (DLTs), maximum tolerated dose (MTD), and recommended Phase 2 dose (RP2D) of orally administered Compound A22.
  • the Compound A22 starting dose for Cohort 1 is ⁇ 21 mg and does not exceed the highest dose deemed safe as determined in the BTX-A51-001 study of Compound A22 in subjects with acute myeloid leukemia and myelodysplastic syndrome.
  • Compound A22 is administered once daily on a weekly schedule of 5 days on/2 days off. Dose escalation proceeds according to a modified 3+3 design. Each cycle consists of 28 days (4 weeks), and the DLT observation period is the first cycle (i.e., 28 days after initiation of dosing).
  • a DLT may be observed in no more than 0 out of 3 or 1 out of 6 subjects who have completed the DLT observation period before the next cohort initiates accrual.
  • sequential dose escalation of Compound A22 proceeds with up to a total of 6 dose levels to a maximum of 200 mg; on the basis of these an MTD is identified.
  • the MTD is defined as the highest dose level with a subject incidence of DLTs of 0 or 1 out of 6 during the first 28 days of study drug dosing. A minimum of 6 subjects are treated at a dose level before this dose level can be deemed as the MTD.
  • Subjects who receive less than 75% of planned doses due to reasons other than a DLT are replaced for the determination of the MTD but remain part of the safety population if they received at least one dose of study drug.
  • dose levels equal to or below the MTD up to 6 additional subjects per dose level cohort may be enrolled to obtain additional safety, PK, and pharmacodynamic (PD) data.
  • Toxicity severity is graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events (NCI CTCAE) Version 5.0. For purposes of dose escalation, the totality of accrued safety information across all cycles completed at the time of DEC data review is taken into consideration.
  • a DLT is defined as a severe or clinically significant AE or abnormal laboratory value (Grade 3 or greater, unless otherwise specified), unless it is clearly related to disease progression, intercurrent illness, preexisting condition, or concomitant medications.
  • Phase la the RP2D has been determined to be 21 mg administered 3x/week based on a comprehensive analysis of available safety, tolerability, PK/PD, and preliminary efficacy as well as the MTD from the dose escalation phase.
  • Phase lb up to 40 additional subjects are enrolled to evaluate safety and preliminary efficacy of Compound A22 in subjects with estrogen receptor positive (ER+), human epidermal growth factor receptor 2 negative (HER2-), GATA3 mutant (mt) and wildtype (wt) metastatic breast cancer (mBC). Based on the Phase la results, the RP2D for Compound A22 was determined to be 21 mg administered orally 3-times-weekly (3x/week) on a Monday /Wednesday /Friday (MWF) schedule for 4 weeks in a 28 day cycle; this dose and schedule is evaluated in Phase lb.
  • ER+ estrogen receptor positive
  • HER2- human epidermal growth factor receptor 2 negative
  • mt wildtype metastatic breast cancer
  • Phase lb is conducted in a staged manner with 22 subjects enrolled to Stage 1. If an objective response (i.e., PR or CR) is observed in >2 subjects of the first 22 enrolled, then an additional 18 subjects are enrolled. Of the first 22 subjects enrolled, a minimum of 13 subjects should have a pathogenic GAT A3 mutation based on next generation sequencing (NGS) of tumor tissue. Of the 40 subjects enrolled to Stages 1 and 2, a minimum of 24 subjects should have a GAT A3 mutation.
  • NGS next generation sequencing
  • Dosing in this phase of the study consists of the first cycle of therapy (i.e., 28 days).
  • the DEC reviews cumulative safety and available PK data in subjects treated in Phase lb for DLTs, with DEC reviews scheduled after the 6th, 12th, 22nd (interim analysis), 30th, and 40th subjects complete a cycle of Compound A22.
  • Subjects who complete one cycle of Compound A22 in either Phase la or Phase lb are offered continued access to study drug until disease progression or unacceptable toxicity.
  • the DEC continues to review accruing safety/PK data, inclusive of all cycles, for subjects who continue with treatment.
  • the duration of the treatment phases of this study is approximately 38 months.
  • the follow up Phase is 5 years after the last subject is enrolled.
  • the Phase lb includes a cohort of up to approximately 40 subjects with advanced ER+, HER2-, GATA3 mt and wt mBC to further characterize the safety, tolerability, and preliminary efficacy when administered at the recommended dose (21 mg 3x/week) identified in the Dose-Escalation Phase la.
  • Eligible subjects for the study demonstrate an understanding by voluntarily signing an informed consent form and are 18 years old or older with histologically or cytologically documented, incurable or metastatic solid tumor that is refractory to or intolerant of all standard therapy or for which no standard therapy is available.
  • subjects To be eligible for Phase lb, subjects have histologically confirmed diagnosis of ER+, HER2- mBC not amenable to resection or radiation therapy with curative intent. Further, the subject has (i) adequate tumor tissue sample from locally recurrent or metastatic site; (ii) documented evidence of ER+ mBC confirmed with the most recently available tumor biopsy from a locally recurrent or metastatic site.
  • ER should be more than 10% ER positive or Allred >5 by local laboratory testing; (iii) documented evidence of HER2- status defined as a negative in situ hybridization test or an IHC status of 0, 1+ or 2+. If IHC is 2+, a negative in situ hybridization (FISH, CISH, or SISH) test is required by local laboratory testing and based on the most recently analyzed tissue sample; and (iv) GATA3 and p53 mutational status based on NGS data obtained prior to or during screening.
  • Participants in the Phase lb expansion phase have progressed on or been intolerant of standard therapy (at least 2 prior lines of endocrine therapy and up to 3 lines of chemotherapy).
  • Female participants of childbearing potential may participate provided they have a negative serum pregnancy test at screening and a negative serum pregnancy test within 72 hours prior to starting on treatment; they also must agree to either abstain from sexual intercourse or use a highly effective method of contraception (for up to 3 months after the last dose of study drug) such as: established use of oral or long acting injected or implanted hormonal methods of contraception for at least 90 days prior to Cycle 1 Dose 1 (participants in Part lb should not be on any form of hormonal contraception); placement of an intrauterine device or intrauterine system; barrier methods: condom, diaphragm, or cervical/vault cap with spermicidal foam/gel/film/cream/suppository; male partner sterilization (with the appropriate post vasectomy documentation of the absence of spermatozoa in the ejaculate [the vasectomized partner should be the sole partner for that subject]); or true abstinence (when this is in line with the preferred and usual lifestyle of the subject).
  • the study also excludes subjects undergoing treatment with any local or systemic antineoplastic therapy (including chemotherapy, hormonal therapy, or radiation) within 3 weeks prior to first dose of Compound A22, with the following exceptions: hormonal therapy with gonadotropinreleasing hormone (GnRH) agonists or antagonists for prostate cancer; hormone-replacement therapy or oral contraceptives; herbal therapy intended as anti -cancer >1 week prior to first dose of Compound A22; palliative radiotherapy for bone metastases >2 weeks prior to first dose of Compound A22.
  • GnRH gonadotropinreleasing hormone
  • the study also excludes subjects who experienced adverse events from prior anti-cancer therapy that have not resolved to Grade ⁇ 1 except for alopecia or Grade ⁇ 2 immunotherapy related thyroid toxicity.
  • CNS central nervous system
  • Subject with known CNS metastases that are treated and stable (without evidence of CNS toxicity) and are not requiring systemic steroids are allowed to be enrolled.
  • Subjects with left or right bundle branch block, where QT cannot be accurately assessed, will be allowed into the study with the approval of the Medical Monitor; LVEF ⁇ 40%.
  • the study drug in phases la and lb is orally administered, immediate-release capsules of 1.0 mg, 2.0 mg, and 7.0 mg.
  • the capsules contain Compound A22 as the active moiety as well as the following inactive compendial excipients: Suglets (Sugar beads), Talc (USP/EP), and Povidone (USP).
  • Suglets Sugar beads
  • Talc USP/EP
  • Povidone USP
  • the study drug is dispensed in separate bottles for each capsule strength, including a sufficient amount to complete the treatment cycle (with the exception that the 0.5-mg and 1.0-mg strengths are not be dispensed together).
  • the 1.0 mg and 2.0 mg capsules are visually distinguishable from one another with the 1.0 mg capsules being white, and the 2.0 mg capsules being orange.
  • the 7 mg capsules are also white but are much bigger.
  • the 7 mg is a size 000 capsule, whereas the 1 mg is a size 1.
  • the capsules strengths on the study drug bottle label are highlighted in different colors: 1 mg - blue, 2 mg - red, and 7 mg - green.
  • TBD to be determined.
  • a De-escalation in dose if required in response to DLT, will be to a mid-dose between the current dose and the previous lower dose with DLT subject incidence of ⁇ 33%.
  • b Dose level increases will be 2-fold or less.
  • Dosing in Phase lb is at the RP2D, which is 21 mg administered 3x/week. To the greatest extent practicable, Day 1 (and thus the first dose) of each cycle is scheduled to occur preferably on a Monday. For a given subject, one or more doses may be held as needed to manage toxicity. In some cases, the dose and/or schedule is lowered in subjects in response to toxicity.
  • Subjects are instructed to take their whole daily dose at approximately the same time each day and to swallow capsules whole. Subjects are instructed to open the Compound A22 packaging as close as possible to when they are going to take Compound A22. Subjects may take study drugs with or without food. Subjects are advised that if study drug is taken with food, they should avoid consuming a high fat meal.
  • Dosing may be interrupted in response to toxi cities or tolerability issues not amounting to a DLT. In such subjects, dosing will resume at the original dose; no dose reductions are permitted for non-DLT AE. In the event of a dose interruption due to a DLT, treatment is recommenced at a lower dose or schedule following appropriate resolution of toxicity. Subjects who experience any Grade > 3 skin drug-related AE or suspected Stevens Johnson syndrome (SJS) or toxic epidermal necrolysis (TEN) are withdrawn.
  • SJS Stevens Johnson syndrome
  • TEN toxic epidermal necrolysis
  • Treatment is discontinued in the event of disease progression, unacceptable toxicity or prolonged time required for recovery from toxicity, confirmed pregnancy, withdrawal of consent, loss to follow-up or noncompliance with protocol, or if the Investigator determines removal from the study is in the subject’s best interest.
  • HIV human immunodeficiency virus
  • LVEF left ventricular ejection fraction
  • MRI magnetic resonance imaging
  • PBMC peripheral blood mononuclear cells
  • PE physical examination
  • PK pharmacokinetics
  • SAE serious adverse event.
  • Medical history includes significant past medical events (e.g., prior hospitalizations or surgeries), a review of the disease under study, prior anti-cancer therapies, and any concurrent medical illnesses.
  • Screening and end of treatment physical examinations include a complete physical examination. Beginning on CID 1, a modified physical examination is performed to monitor for any changes (e.g., lymph nodes, lung, cardiac, abdomen, skin, neurologic, and any systems, as clinically indicated). Weight (without shoes) is measured at each physical examination. Height (without shoes) is measured at screening only.
  • any changes e.g., lymph nodes, lung, cardiac, abdomen, skin, neurologic, and any systems, as clinically indicated.
  • Weight (without shoes) is measured at each physical examination. Height (without shoes) is measured at screening only.
  • Serum pregnancy is conducted at screening for female subjects of childbearing potential. Urine pregnancy is conducted predose on Day 1 of each cycle and at EOT.
  • HIV, HBsAg, and HCV antibody viral testing in subjects with current or prior history of HIV, HBV, and HCV infection, respectively, is conducted at screening.
  • Plasma samples for Compound A22 PK are collected prior to the dose administered in the clinic on that day in all patients that are not having intensive PK collection. Blood samples are collected from each subject for measurement of plasma concentrations of Compound A22 (and its demethylated metabolite, if applicable) on the days indicated in the Schedule of Assessments.
  • Intensive PK is collected in all subjects in Phase la dose escalation cohorts and in the first 10 subjects in the Phase lb dose expansion.
  • Plasma samples for intensive PK are collected on C1D1 and C1D5 at pre-dose (within -2 hrs) and 1 ( ⁇ 10 min), 2 ( ⁇ 15 min), 3 ( ⁇ 20 min), 6 ( ⁇ 30 min), and 24 ( ⁇ 1 hr) hours post dose.
  • a 48 ( ⁇ 3hr) hour post dose sample is collected prior to the dose scheduled for Day 3.
  • the Day 8 sample ( ⁇ 1 day) is collected prior to the dose scheduled for Day 8. Actual sample collection time is recorded.
  • PK blood samples are collected from the affected subject in order to measure levels of Compound A22.
  • Exploratory biomarker blood samples are collected for future analyses, in efforts to identify novel biomarkers of response in breast cancer and other solid tumors and to better understand the biological effect of Compound A22.
  • Exploratory biomarker samples are collected: pre-dose on Day 1, on Day 2 (24-hours post-dose), 4 hours post-dose on Day 5 of Cycle 1, and 4 hours postdose on Day 1 at the beginning of Cycles 2 and 3.
  • Collection timepoints may be adjusted in response to emerging data.
  • Biological specimens are collected for the evaluation of exploratory systemic and/or tissue specific biomarkers, to explore association with study drug response, including efficacy and/or AEs, and to increase knowledge and understanding of the biology of Compound A22.
  • Samples for pharmacodynamic/biomarker assessments are collected predose on Day 1, on Day 2 (24 hours post-dose), 4 hours post-dose on Day 5 of Cycle 1, and 4 hours post-dose on Day 1 at the beginning of Cycles 2 and 3. Collection timepoints may be adjusted in response to emerging data.
  • the specific analyses include, but are not limited to, the biomarkers and assays described in the following assessments: serum biomarkers (including but not limited to MIC-1, Macrophage Inhibitory Cytokine- 1) as a surrogate of p53 since it is a p53 -regulated secreted cytokine; plasma biomarkers; whole blood genomic DNA; whole blood RNA; circulating tumor DNA; exploratory peripheral blood mononuclear cells (PBMC) to evaluate MCL-1, MDM2, and p53 protein expression; cytogenetics and mutation panel may including: gene expression levels of target SE genes (i.e., Mell, MYC, MYB, and MDM2) and gene mutation analysis by NGS.
  • serum biomarkers including but not limited to MIC-1, Macrophage Inhibitory Cytokine- 1
  • plasma biomarkers including but not limited to MIC-1, Macrophage Inhibitory Cytokine- 1
  • plasma biomarkers including but not limited to MIC-1, Macrophage In
  • Genomic DNA is collected predose on CID 1 or may be collected at any time during the course of the study.
  • Archival tumor tissue for biomarker analysis is collected.
  • the tissue is obtained either at the time of or after the diagnosis of advanced disease has been made and from a site not previously irradiated.
  • Tissue is collected between screening and Cycle 2.
  • an ECHO to be performed for the evaluation of LVEF and GLS, is performed within the 14 days prior to starting study treatment. ECHO testing is also conducted during Cycles 2, 4, and 6 (no earlier than the Day 15 Visit) and at the end of Cycle 8 (no earlier than the Day 22 Visit), or more frequently if clinically indicated. It is acceptable for ECHO results to be obtained within 7 days prior to the next cycle’s Day 1 Visit; however, results and interpretation must be received prior to the start of the next cycle (Cycles 3, 5, and 7).
  • ECHO is performed within the 14 days prior to the start of treatment, i.e., Day 1 of Cycle 1. If troponin levels are elevated, post-treatment ECHO is conducted at the time of the elevation and every 2 months thereafter until troponin levels have returned to baseline.
  • responses are evaluated using RECIST vl.l.
  • tumor evaluation by CT/MRI is performed during screening (within 28 days of Day 1) and every 8 weeks on Day 1 of each odd numbered cycle. A baseline scan within 6 weeks of Day 1 is acceptable.
  • Responses are evaluated using RECIST vl.l, or the appropriate response evaluation for the corresponding malignancy, which may include a bone marrow biopsy.
  • the same radiographic procedure used to define measurable lesions is used throughout the study for each subject.
  • Response to disease (CR or PR) is confirmed by the same imaging technique no less than 4 weeks after the criteria for response are first met.
  • a baseline CT/MRI of the head is performed during screening.
  • EOT Visit The End of Treatment (EOT) Visit is conducted within 14 to 28 days after the last dose of study drug is administered in a given living subject, regardless of the reason for discontinuation.
  • Adverse events > Grade 2 ongoing at the EOT Visit are followed until the event resolves to ⁇ Grade 1, stabilizes, subjects start alternate therapy, returns to a status that is clinically acceptable in the judgment of Investigator, is lost to follow-up, or terminates with the subject death.
  • discontinuation of Compound A22 all surviving subjects are contacted by telephone once every 3 months thereafter for up to 2 years or until death, withdrawal of consent, loss to follow-up, or end of study. Subjects with continued response (i.e., beyond the 2-year survival follow-up period) are to be followed until this study is closed.
  • the patient was treated with two cycles of Ibrance/letrozole and 1 cycle of ribociclib but showed disease progression.
  • the patient also received Enhertu for 3 cycles but developed shortness of breath associated with disease progression.
  • Biopsy of a metastatic lesion in the right lung showed breast cancer origin and GATA3 mutation (P409fs) with an allele frequency of 43%.
  • the patient was treated with 21 mg Compound A22 on a 3x/week schedule (Monday-Wednesday-Friday).
  • the target lesion linear diameter measured 6.4 cm.
  • a scan conducted after 2 cycles of Compound A22 treatment showed a reduction of target lesion linear diameter to 5.5 cm, a -14% reduction in the lesion.

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Abstract

Provided herein is a method of treating, preventing, or ameliorating one or more symptoms of breast cancer using a pyrazole having the formula: (I)

Description

METHODS OF TREATING BREAST CANCER USING A PYRAZOLE COMPOUND
PRIORITY
[0001] The application claims priority to U.S. Provisional Application No. 63/568,074, filed March 21, 2024, and U.S. Provisional Application No. 63/654,569, filed May 31, 2024, each of which is incorporated by reference in its entirety.
FIELD
[0002] This disclosure relates to methods of treating, preventing, or ameliorating one or more symptoms of breast cancer using a pyrazole compound.
BACKGROUND
[0003] The tumor suppressor, p53, is a transcription factor that induces cell cycle arrest and activates apoptotic cell death through target gene activation in response to DNA damage and cellular stress. Fields S, Jang SK, Presence of apotent transcription activating sequence in the p53 protein, Science. Aug 31 1990; 249(4972): 1046-9. doi: 10.1126/science.2144363; Kastenhuber ER, Lowe SW. Putting p53 in Context. Cell. Sep 7 2017; 170(6): 1062-1078. doi: 10.1016/j. cell.2017.08.028) While p53 is mutated in about half of solid tumors, cancers that have a wild type p53 must adopt other mechanisms to inhibit p53 activity.
[0004] Casein kinases are serine/threonine kinases that phosphorylate proteins to mediate normal biological functions and malignant transformation. Schittek et al., Mol. Cancer 2014, 13, 231-245. Casein kinase 1 alpha (CKla) functions as a tumor inducer in several cancers through negative regulation of Wnt/p-catenin signaling and p53. Ebert & Krbnke, N. Engl. J. Med. 2018, 379, 1873-1874. CKla phosphorylates [3-catenin at serine 45, leading to ubiquitination and degradation of P-catenin. Schittek et al., Mol. Cancer 2014, 13, 231-245; Ebert & Krbnke, N. Engl. J. Med. 2018, 379, 1873-1874; Elyada et al., Nature 2011, 470, 409-413. CKla also phosphorylates murine double minute X (MDMX) at serine 289, resulting in enhanced binding of MDMX to p53. Wu et al., Mol. Cell. Biol. 2012, 32, 4821- 4832. Additionally, a complex of CKla and mouse double minute 2 homolog (MDM2) inhibits p53. Elyada et al., Nature 2011, 470, 409-413. Thus, enhanced inhibition of CKla with subsequent p53 activation has the potential to be effective in treating a wide array of cancers. [0005] Super-enhancers (SEs) are large clusters of transcriptional enhancers that normally drive expression of genes that define cell identity. Hnisz et al., Cell 2013 155(4): 934-947. Tumor cells acquire SEs at key oncogenes like c-Myc and at genes that produce the hallmark features of cancer. Hnisz et al. 2013. Given the dependency on high levels of gene expression, tumor cells are particularly sensitive to disturbances in transcription.
Transcriptional kinases are essential components of the transcription apparatus. In particular, cyclin dependent kinase 7 (CDK7) phosphorylates the carboxy terminal domain (CTD) of ribonucleic acid polymerase II (RNA Pol II) at Ser5 and Ser7, enabling transcriptional initiation and CDK9 phosphorylates RNA Pol II CTD at Ser2 to facilitate transcription elongation. Ferguson and Gray, Nat. Rev. Drug. Discov., 2018 17(5): 353-377.
[0006] The dual mechanism of action of preventing the transcription of key oncogenic genes via the inhibition of CDK7 and CDK9 and activating p53 via inhibition of CKla may result in tumor responses in patients with a wide variety of tumor types.
[0007] MDM2 is a negative regulator of p53 that promotes p53 ubiquitination and subsequent degradation. (Haupt Y, Maya R, Kazaz A, Oren M. Mdm2 promotes the rapid degradation of p53. Nature. 1997/05/01 1997;387(6630):296-299. doi:10.1038/387296a0) One mechanism for inhibiting wild type p53 is to amplify the gene expression of MDM2 and thus increase the turnover of p53. Mutations that result in increased expression of MDM2 may be in transcription factors or DNA sequences, e.g., selecting for changes in promoter or enhancer function that increase MDM2 transcription. Tumors that utilize Super Enhancer- mediated transcription have been characterized as “transcriptionally addicted” due to their need for high rates of expression of proteins with short half-lives, such as MDM2. SE- mediated transcription regulates MDM2 expression in some tumors.
[0008] Another mechanism that increases MDM2 is amplification of its gene copy number. In liposarcoma, MDM2 gene amplification is observed in almost all patients and activates p53-independent growth and stress response genes. (Bevill SM, Casani-Galdon S, El Farran CA, et al. Impact of supraphysiologic MDM2 expression on chromatin networks and therapeutic responses in sarcoma. Cell Genom. Jul 12 2023;3(7): 100321. doi: 10.1016/j.xgen.2023.10032). While liposarcoma is the canonical example of MDM2 gene amplification, the phenomenon occurs in tumors originating from many different tissues. A wide range of cancer types in which MDM2 genes are amplified by 4-fold or more include, but are not limited to, breast, non-small-cell lung cancer, bladder (urothelial), stomach, esophageal, melanoma, prostate, cholangiocarcinoma, pancreatic, colorectal, salivary gland, and cervical. Furthermore, a higher copy number of MDM2, e.g., >8, is associated with decreased survival rates.
[0009] Multiple biopharmaceutical companies have developed MDM2 inhibitors. For example, Rain Oncology tested milademetan in MDM2-amplified solid tumors (clinicaltrials.gov NCT05012397). This multicenter, single-arm, open-label basket study was designed to evaluate the safety and efficacy of milademetan in patients with advanced or metastatic solid tumors refractory or intolerant to standard-of-care therapy that exhibit wildtype (WT) TP53 and MDM2 copy number (CN) > 8 using prespecified biomarker criteria. Interim results for this study showed intriguing anti-tumor activity. Boehringer-Ingelheim presented data at the American Society of Oncology meeting in 2023 for its MDM2 inhibitor, BI-907828 also known as brigimadlin, in patients with biliary cancer. [Yamamoto et al, J Clin Oncol 2023 41 :4 suppl, 543], About 5-8% of biliary cancers have amplified MDM2. In two parallel studies, brigimadlin was either given as monotherapy, 45 mg every 3 weeks, or at lower doses in combination with an anti-PDl antibody. In addition, brigimadlin has been evaluated in an MDM2-amplified basket study of solid tumors. It demonstrated clinical activity in both well-differentiated liposarcoma and dedifferentiated liposarcoma (LoRusso et al, The MDM2-p53 Antagonist Brigimadlin (BI 907828) in Patients with Advanced or Metastatic Solid Tumors: Results of a Phase la, First-in-Human, Dose-Escalation Study. Cancer Disc. 2023; 13: 1802-13 doi: 10.1158/2159-8290.CD-23-0153).
[0010] Despite intriguing clinical activity, direct MDM2 inhibitors have proven to be challenging to develop due to hematologic toxicity. For example, milademetan was evaluated at various doses and regimens to obviate undue toxicity. A phase 3 study in liposarcoma was conducted using the optimized regimen (260 mg for 3 consecutive days followed by 11 days off treatment; cycles repeated every 14 days). [NCT04979442] This randomized, multicenter, open-label study was designed to evaluate the safety and efficacy of milademetan compared to trabectedin in patients with unresectable or metastatic dedifferentiated liposarcoma that had progressed on 1 or more prior systemic therapies. The study was terminated early. A significant contributing factor to the lack of efficacy was the high rate of hematologic toxicity which led to treatment interruptions and patient discontinuations. Rates of grade 3/4 hematologic toxicity were significant and reported as follows: 39.5% of patients with thrombocytopenia; 25.5% of patients with neutropenia; and 18.6% of patients with anemia.
[0011] Similarly, brigimadlin is dosed infrequently with a long treatment holiday in an attempt to overcome toxicity. As monotherapy, 45 mg is administered only on the first day of a 21-day cycle. (Macarulla et al., Efficacy and safety of brigimadlin (BI 907828), an MDM2- -p53 antagonist, in patients (pts) with advanced biliary tract cancer: Data from two phase la/Ib dose-escalation/expansion trials, Journal of Clinical Oncology Volume 42, Number 3_supp). Early results for this agent reported the following rates of Grade 3/4 hematologic toxicity: 19% thrombocytopenia; 20% neutropenia; 10% anemia. While these data appear to be improved compared to milademetan, there remains a need to provide better tolerated agents that can achieve continuous pharmacologic coverage of the drug target.
[0012] Sirimadlin (HDM 201) is a direct MDM2 inhibitor that was evaluated in 115 solid tumor patients and 93 patients with hematologic malignancies. [Stein et al Clin Cancer Res 2022;28:870-81 doi: 10.1158/1078-0432.CCR-21-1295] Grade 3/4 Adverse Events (AEs) suspected to be treatment related were observed in 52 (45%) patients with solid tumors and 66 (71%) patients with hematologic tumors. Hematologic toxicities were the most common grade 3/4 AE for all indications and were more frequent in patients with hematologic tumors compared with those with solid tumors. A total of 50 patients (43.5%) with solid tumors and 38 patients (40.9%) with hematologic tumors had at least one AE leading to dose adjustment or interruption. Importantly, researchers concluded that thrombocytopenia is an on-target effect of MDM2 inhibition.
[0013] Therefore, despite promising anti-tumor activity of targeted MDM2 inhibitors, there remains a pressing need for agents that inhibit MDM2 by a mechanism that does not cause unacceptable Grade 3/4 toxicity, particularly hematologic toxicities including thrombocytopenia. In addition, there remains a need for agents that can be dosed continuously rather than with long holiday periods, such as 1, 2 or 3 weeks between doses. A need exists for a better tolerated agent with more frequent dosing that provides more uniform exposure and improved pharmacology.
[0014] Some patients with metastatic breast cancer have mutations in GATA3, in 13.6% to 18.7% of metastatic HR+/HER2- tumors. Li et al. Genomic Alteration in Metastatic Breast Cancer and Its Treatment, Am. Soc. Clin. Oncol. Educ. Book 2020 40: 1-14. GATA3 is a transcription factor that is important in maintaining the normal differentiation status of luminal cells in mammary tissue. Kouros-Mehr et al., GATA-3 maintains the differentiation of the luminal cell fate in the mammary gland, Cell 2006 127(5): 1041-55. Women with mutations in GATA3 have worse progression-free and overall survival (Velimirovic et al., Landscape of GATA3 mutations identified from circulating tumor DNA clinical testing and their impact on disease outcomes in estrogen receptor-positive (ER+) metastatic breast cancers treatedwith endocrine therapies, J. Clin. Oncol. 2021 39(suppl 15): abstract 1065) and, importantly, GAT A3 mutations downstream of the splice site at amino acid 307 are mutually exclusive with wild-type P53 and demonstrate higher levels of MDM2 expression than other mutational subgroups (Li et al. Genomic Alteration in Metastatic Breast Cancer and Its Treatment, Am. Soc. Clin. Oncol. Educ. Book 2020 40: 1-14). Recent evidence suggests that in a GATA3mt P53wt setting, MDM2 plays an essential role in tumor cell survival. Pharmacologic inhibition of MDM2 in an ER+ GATA3mt background leads to tumor cell growth inhibition in both cell lines and patient-derived xenografts and induces apoptosis in vitro. Bianco et al., GATA3 andMDM2 are synthetic lethal in estrogen receptor-positive breast cancers, Commun. Biol. 2022 5(1):373.
[0015] Despite advances, a need exists for a breast cancer treatment that suppresses oncogenes and causes tumor regression.
SUMMARY
[0016] This disclosure provides methods of treating a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22, or a pharmaceutically acceptable salt thereof.
[0017] In some embodiments, the patient has GAT A3 mt (mt = mutant). In some embodiments, the patient has TP53wt (wt = wild type). [0018] In some embodiments, the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
[0019] In some embodiments, the breast cancer is refractory. In some embodiments, the breast cancer is metastatic. In some embodiments, the breast cancer is drug-resistant. In some embodiments, the patient has failed a prior therapy. In some embodiments, the patient is a human.
[0020] In some embodiments, the compound is formulated as a -toluenesulfonate salt or a di- -toluenesulfonate salt.
[0021] In some embodiments, the pharmaceutical composition is administered orally. In some embodiments, the pharmaceutical composition is administered as a tablet or capsule. In some embodiments, the therapeutically effective amount is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 25 mg, about 30 mg, or about 35 mg.
[0022] In some embodiments, the pharmaceutical composition is administered in a cycle. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle. In some embodiments, the pharmaceutical composition is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week. In some embodiments, the pharmaceutical composition is administered for 3 days or 5 days per week. In some embodiments, the pharmaceutical composition is administered on three non-consecutive days in a week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 3 days per week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 3 non-consecutive days per week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle for 5 days per week. In some embodiments, the pharmaceutical composition is administered in a 28-day cycle on Days 1, 2, 3, 4, and 5 per week. [0023] In some embodiments, the therapeutically effective amount is about 21 mg. In some embodiments, the therapeutically effective amount is about 30 mg. In some embodiments, the therapeutically effective amount is about 35 mg.
[0024] In another aspect, this disclosure provides methods of inhibiting MDM2 in a patient having ER+, HER- metastatic breast cancer and having GATA3mt, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22, or a pharmaceutically acceptable salt thereof.
[0025] In some embodiments, the compound is formulated as a p-toluenesulfonate salt. In some embodiments, the compound is formulated as a di- -toluenesulfonate salt.
[0026] In some embodiments, the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg. In some embodiments, the pharmaceutical composition is administered three times per week.
[0027] In some embodiments, the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
[0028] In another aspect, this disclosure provides methods of inhibiting CDK7 in a patient having ER+, HER- metastatic breast cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
[0029] In some embodiments, the compound is formulated as a -toluenesulfonate salt. In some embodiments, the compound is formulated as a di- -toluenesulfonate salt. [0030] In some embodiments, the breast cancer has been resistant to a CDK4/6 inhibitor.
[0031] In some embodiments, the patient has TP53wt. In some embodiments, the patient has GATA3wt. In some embodiments, the patient has GATA3mt.
[0032] In some embodiments, the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
[0033] In some embodiments, Compound A22 is administered three times per week. In some embodiments, the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg per day.
[0034] In another aspect, this disclosure provides methods of treating cancer in a patient having amplified expression of MDM2 or having a >4 copy number of the MDM2 gene, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
[0035] In some embodiments, the compound is formulated as a -toluenesulfonate salt or a di- -toluenesulfonate salt.
[0036] In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene. In some embodiments, administering comprises administering about 21 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 30 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 35 mg of Compound A22 three times per week.
[0037] In yet another aspect, this disclosure provides methods of inhibiting MDM2 expression in a patient having cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof. In some embodiments, the patient has breast cancer with GATA3mt. [0038] In some embodiments, the compound is formulated as -toluenesulfonate salt or a di-/?-toluenesulfonate salt.
[0039] In some embodiments, the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. In some embodiments, the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
[0040] In some embodiments, administering comprises administering about 21 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 30 mg of Compound A22 three times per week. In some embodiments, administering comprises administering about 35 mg of Compound A22 three times per week.
[0041] In a further aspect, this disclosure provides pharmaceutical compositions comprising Compound A22, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, for use in the treatment of a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer.
[0042] In some embodiments of the use, the patient has GAT3mt. In some embodiments of the use, the treatment comprises administration of the pharmaceutical composition to the patient on three non-consecutive days in a week. In some embodiments of the use, the administration occurs for one or more cycles. In some embodiments, the cycle is 28 days. In some embodiments, the pharmaceutical composition is orally administered to the patient. In some embodiments, the pharmaceutical composition is formulated as an oral unit dosage form. In some embodiments, the oral unit dosage form comprises about 21 mg of Compound A22. In some embodiments, the oral unit dosage form comprises about 30 mg of Compound A22. In some embodiments, the oral unit dosage form comprises about 35 mg of Compound A22.
[0043] In yet another aspect, this disclosure provides a method of inducing apoptosis in an ER+ HER2- breast cancer cell, comprising contacting the cell with an effective amount of a compound of Compound A22, or a pharmaceutically acceptable salt thereof. BRIEF DESCRIPTION OF THE DRAWINGS
[0044] FIGS. 1A and IB shows the results of inhibition or suppression of certain biomarkers from application of Compound A22.
[0045] FIGS. 2A and IB shows the results of inhibition or suppression of certain biomarkers from application of Compound A22.
[0046] FIGS. 3 A and 3B show the results of CDK2 and phospho-CDK2 signaling in the presence of Compound A22.
DETAILED DESCRIPTION
[0047] To facilitate understanding of the disclosure set forth herein, a number of terms are defined below.
[0048] Generally, the nomenclature used herein and the laboratory procedures in organic chemistry, medicinal chemistry, biochemistry, biology, and pharmacology described herein are those well-known and commonly employed in the art. Unless defined otherwise, or context dictates otherwise, all technical and scientific terms used herein generally have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs.
[0049] The term “subject” refers to an animal, including, but not limited to, a primate (e.g., human), cow, pig, sheep, goat, horse, dog, cat, rabbit, rat, or mouse. The terms “subject” and “patient” are used interchangeably herein in reference, for example, to a mammalian subject, such as a human subject. In one embodiment, the subject is a human.
[0050] The terms “treat,” “treating,” and “treatment” are include alleviating, improving, inhibiting the progression of, or abrogating a disorder, disease, or condition, or one or more of the symptoms associated with the disorder, disease, or condition; or alleviating, improving, inhibiting the progression of, or eradicating the cause(s) of the disorder, disease, or condition itself. Treating includes curing, improving, or at least partially ameliorating the disorder, disease, or condition.
[0051] The terms “prevent,” “preventing,” and “prevention” are meant to include a method of delaying and/or precluding the onset of a disorder, disease, or condition, and/or its attendant symptoms; barring a subject from acquiring a disorder, disease, or condition; or reducing a subject’s risk of acquiring a disorder, disease, or condition.
[0052] The terms “alleviate” and “alleviating” refer to easing or reducing one or more symptoms (e.g., pain) of a disorder, disease, or condition. The terms can also refer to reducing adverse effects associated with an active ingredient. Sometimes, the beneficial effects that a subject derives from a prophylactic or therapeutic agent do not result in a cure of the disorder, disease, or condition.
[0053] The term “contacting” or “contact” is meant to refer to bringing together of a therapeutic agent and cell or tissue such that a physiological and/or chemical effect takes place as a result of such contact. Contacting can take place in vitro, ex vivo, or in vivo. In one embodiment, a therapeutic agent is contacted with a cell in cell culture (in vitro to determine the effect of the therapeutic agent on the cell. In another embodiment, the contacting of a therapeutic agent with a cell or tissue includes the administration of a therapeutic agent to a subject having the cell or tissue to be contacted.
[0054] The term “therapeutically effective amount” or “effective amount” is meant to include the amount of a compound that, when administered, is sufficient to treat, prevent development of, or alleviate a disorder, disease, or condition, or one or more of the symptoms of the disorder, disease, or condition being treated. In some contexts, the term “therapeutically effective amount” or “effective amount” also refers to the amount of a compound that is sufficient to elicit a biological or medical response of a biological molecule (e.g., a protein, enzyme, RNA, or DNA), cell, tissue, system, animal, or human, which is being sought by a researcher, veterinarian, medical doctor, or clinician.
[0055] The term “pharmaceutically acceptable carrier,” “pharmaceutically acceptable excipient,” “physiologically acceptable carrier,” or “physiologically acceptable excipient” refers to a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, solvent, or encapsulating material. In one embodiment, each component is “pharmaceutically acceptable” in the sense of being compatible with the other ingredients of a pharmaceutical formulation, and suitable for use in contact with the tissue or organ of a subject (e.g., a human or an animal) without excessive toxicity, irritation, allergic response, immunogenicity, or other problems or complications, commensurate with a reasonable benefit/risk ratio. See, e.g., Remington: The Science and Practice of Pharmacy, 22nd ed.; Allen Ed.: Philadelphia, PA, 2012; Handbook of Pharmaceutical Excipients, 8th ed.; Sheskey etal., Eds.; The Pharmaceutical Press: 2017; Handbook of Pharmaceutical Additives, 3rd ed.; Ash and Ash Eds.; Gower Publishing Company: 2007; Pharmaceutical Preformulation and Formulation, 2nd ed.; Gibson Ed.; CRC Press LLC: Boca Raton, FL, 2009.
[0056] The term “about” or “approximately” means an acceptable error for a particular value as determined by one of ordinary skill in the art, which depends in part on how the value is measured or determined. In certain embodiments, the term “about” or “approximately” means within 1, 2, 3, or 4 standard deviations. In certain embodiments, the term “about” or “approximately” means within 50%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, or 0.05% of a given value or range.
[0057] The term “solvate” refers to a complex or aggregate formed by one or more molecules of a solute, e.g., a compound provided herein, and one or more molecules of a solvent, which are present in stoichiometric or non-stoichiometric amount. Suitable solvents include, but are not limited to, water, methanol, ethanol, ^-propanol, isopropanol, and acetic acid. In certain embodiments, the solvent is pharmaceutically acceptable. In one embodiment, the complex or aggregate is in a crystalline form. In another embodiment, the complex or aggregate is in a noncrystalline form. Where the solvent is water, the solvate is a hydrate. Examples of hydrates include, but are not limited to, a hemihydrate, monohydrate, dihydrate, trihydrate, tetrahydrate, and pentahydrate.
Pyrazole Compound
[0058] Compound A22 of this disclosure, (lr,4r)-A1-(5-chloro-4-(5-(cyclopropylmethyl)- 1 -methyl- 177-pyrazol-4-yl)pyrimidin-2-yl)cy cl ohexane-l,4-diamine, has the following structure:
A22.
[0059] Compound A22 is a CKla, CDK7, and CDK9 inhibitor. Minzel et aL, Cell 2018, 775, 1-15. The Compound can be prepared according to the procedures described in Minzel et al, Cell ' 2018, 775, 1-15; or U.S. Pat. Appl. Publ. No. 2018/0214447 Al. The entire contents of those disclosures are incorporated herein by reference. [0060] Compound A22 may also be referred to as compound A51 or BTX-A51.
[0061] In one embodiment, Compound A22 is formulated as a non-hygroscopic crystalline salt of (lr,4r)-A1-(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-U/-pyrazol-4- yl)pyrimidin-2-yl)cyclohexane-l,4-diamine with an acid, or a pharmaceutically acceptable solvate thereof. In some embodiments, Compound A22 is formulated as a non-hygroscopic crystalline tosylate salt. In some embodiments, Compound A22 is formulated as a non- hygroscopic crystalline ditosylate salt.
[0062] In another embodiment, the disclosure provides Compound A22, chloro-4-(5-(cyclopropylmethyl)- 1 -methyl- U/-pyrazol-4-yl)pyrimidin-2-yl)cyclohexane- 1 ,4- diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
[0063] In another embodiment, Compound A22 is provided as a -toluensulfonate of (lr,4r)-A1-(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-lJ7-pyrazol-4-yl)pyrimidin-2- yl)cyclohexane-l,4-diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically solvate or hydrate.
[0064] In another embodiment, Compound A22 is provided as a di- -toluensulfonate of (lr,4r)-A1-(5-chloro-4-(5-(cyclopropylmethyl)-l-methyl-lJ7-pyrazol-4-yl)pyrimidin-2- yl)cyclohexane-l,4-diamine, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically solvate or hydrate. The -toluenesulfonate and di- -toluensulfonate salts are described in U.S. Pub. No. 2022/0332702, the entire contents of which are incorporated herein by reference. The term “/?-toluenesulfonate” is used interchangeably with the term “tosylate.” Thus, in some embodiments Compound A22 is provided as a toyslate salt. In some embodiments, Compound A22 is provided as a ditosylate salt.
[0065] In certain embodiments, Compound A22 is isolated or purified. In certain embodiments, a compound described herein has a purity of at least about 50%, at least about 70%, at least about 80%, at least about 90%, at least about 95%, at least about 98%, at least about 99%, or at least about 99.5% by weight.
[0066] The compounds described herein are intended to encompass all possible stereoisomers unless a particular stereochemistry is specified. Where a compound described herein contains an alkenyl group, the compound may exist as one or mixture of geometric cisltrans (or ZZE) isomers. Where structural isomers are interconvertible, the compound may exist as a single tautomer or a mixture of tautomers. This can take the form of proton tautomerism in the compound that contains, for example, an imino, keto, or oxime group; or so-called valence tautomerism in the compound that contain an aromatic moiety. It follows that a single compound may exhibit more than one type of isomerism.
[0067] Compound A22 can be enantiomerically pure, such as a single enantiomer or a single diastereomer, or be stereoisomeric mixtures, such as a mixture of enantiomers, e.g., a racemic mixture of two enantiomers; or a mixture of two or more diastereomers. As such, one of ordinary skill in the art will recognize that administration of a compound in its (A) form is equivalent, for compounds that undergo epimerization in vivo, to administration of the compound in its (5) form. Conventional techniques for the preparation/isolation of individual enantiomers include synthesis from a suitable optically pure precursor, asymmetric synthesis from achiral starting materials, or resolution of an enantiomeric mixture, for example, chiral chromatography, recrystallization, resolution, diastereomeric salt formation, or derivatization into diastereomeric adducts followed by separation.
[0068] Compound A22 can also be provided as a pharmaceutically acceptable salt. Salts of Compound A22 are described in U.S. Pub. No. 2022/0332702, the entire contents of which are incorporated herein by reference.
[0069] Compound A22 may also be provided as a prodrug, which is a functional derivative of a compound, and is readily convertible into the parent compound in vivo. Prodrugs are often useful because, in some situations, they may be easier to administer than the parent compound. They may, for instance, be bioavailable by oral administration whereas the parent compound is not. The prodrug may also have enhanced solubility in pharmaceutical compositions over the parent compound. A prodrug may be converted into the parent drug by various mechanisms, including enzymatic processes and metabolic hydrolysis.
[0070] The compounds described herein can be prepared, isolated, or obtained by any method known to one of ordinary skill in the art, for example, by following the procedures described in U.S. Pat. No. 10,376,511. Pharmaceutical Compositions
[0071] In some embodiments, provided herein are pharmaceutical compositions, comprising Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and a pharmaceutically acceptable excipient. In some embodiments, a pharmaceutical composition comprises the -toluenesulfonate or di- -toluensulfonate salt of Compound A22.
[0072] A pharmaceutical composition provided herein can be formulated in various dosage forms, including, but not limited to, dosage forms for oral, parenteral, and topical administration. The pharmaceutical composition can also be formulated as modified release dosage forms, including delayed-, extended-, prolonged-, sustained-, pulsatile-, controlled-, accelerated-, fast-, targeted-, programmed-release, and gastric retention dosage forms. These dosage forms can be prepared according to conventional methods and techniques known to those skilled in the art. See, e.g., Remington: The Science and Practice of Pharmacy, supra, Modified-Release Drug Delivery Technology, 2nd ed.; Rathbone et al., Eds.; Drugs and the Pharmaceutical Sciences 184; CRC Press: Boca Raton, FL, 2008.
[0073] In one embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for oral administration. In some embodiments, the pharmaceutical composition is a tablet or a capsule. In another embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for parenteral administration. In yet another embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for intravenous administration. In yet another embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for intramuscular administration. In yet another embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for subcutaneous administration. In still another embodiment, a pharmaceutical composition provided herein is formulated in a dosage form for topical administration.
[0074] A pharmaceutical composition provided herein can be provided in a unit-dosage form or multiple-dosage form. A unit-dosage form, as used herein, refers to physically discrete a unit suitable for administration to a subject, and packaged individually as is known in the art. Each unit-dose contains a predetermined quantity of an active ingredient(s) (e.g., a compound provided herein) sufficient to produce the desired therapeutic effect, in association with the required pharmaceutical excipient(s). Examples of a unit-dosage form include, but are not limited to, an ampoule, syringe, and individually packaged tablet and capsule. A unitdosage form may be administered in fractions or multiples thereof. A multiple-dosage form is a plurality of identical unit-dosage forms packaged in a single container to be administered in a segregated unit-dosage form. Examples of a multiple-dosage form include, are not limited to, a vial, bottle of tablets or capsules, or bottle of pints or gallons.
[0075] A pharmaceutical composition provided herein can be administered at once or multiple times at intervals of time. It is understood that the precise dosage and duration of treatment may vary with the age, weight, and condition of the subject being treated, and may be determined empirically using known testing protocols or by extrapolation from in vivo or in vitro test or diagnostic data. It is further understood that for any particular individual, specific dosage regimens should be adjusted over time according to the subject’s need and the professional judgment of the person administering or supervising the administration of the pharmaceutical composition.
[0076] In some embodiments, a pharmaceutical composition provided herein comprises Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and sugar beads, talc, and povidone.
[0077] In some embodiments, a pharmaceutical composition provided herein comprises Compound A22, or a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof; and sugar beads, talc, and povidone.
[0078] In some embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt; and sugar beads, talc, and povidone. In some embodiments, a pharmaceutical composition comprises a tosylate salt of Compound A22. In some embodiments, a pharmaceutical composition comprises a ditosylate salt of Compound A22. In some embodiments, the pharmaceutical composition is formulated as a capsule. In some embodiments, the pharmaceutical composition is formulated as a tablet.
[0079] In some embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.1 to about 50, from about 0.2 to about 40, from about 0.5 to about 35, or from about 0.5 to about 30 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.1 to about 50 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.2 to about 35 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.5 to about 30 mg per capsule or tablet. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount ranging from about 0.5 to about 5 mg per capsule.
[0080] In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form. In certain embodiments, a pharmaceutical composition provided herein comprises Compound A22 or a pharmaceutically acceptable salt in an amount of about 0.5, about 1, or about 2 mg per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 21 mg of Compound A22 per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 30 mg of Compound A22 per capsule or tablet or unit oral dosage form. In some embodiments, a pharmaceutical composition comprises about 35 mg of Compound A22 per capsule or tablet or unit oral dosage form.
[0081] In yet another embodiment, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 or a di- -toluenesulfonate salt of Compound A22; and sugar beads, talc, and povidone. In one embodiment, the pharmaceutical composition is formulated as a capsule or tablet or unit oral dosage form.
[0082] In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50, from about 0.2 to about 20, from about 0.5 to about 10, or from about 0.5 to about 5 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.2 to about 40 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 30 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 25 mg per capsule.
[0083] In certain embodiments, a pharmaceutical composition provided herein comprises a -toluenesulfonate salt of Compound A22 in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form. In certain embodiments, a pharmaceutical composition provided herein comprises a p- toluenesulfonate salt of Compound A22 in an amount of about 21, 30, or 35 mg per capsule or tablet or unit oral dosage form.
[0084] In still another embodiment, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22; and sugar beads, talc, and povidone. In one embodiment, the pharmaceutical composition is formulated as a capsule.
[0085] In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50, from about 0.2 to about 20, from about 0.5 to about 10, or from about 0.5 to about 5 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.1 to about 50 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.2 to about 40 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 30 mg per capsule. In certain embodiments, a pharmaceutical composition provided herein comprises a di- - toluenesulfonate salt of Compound A22 in an amount ranging from about 0.5 to about 25 mg per capsule.
[0086] In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount of about 0.5, about 0.6, about 0.7, about 0.8, about 0.9, about 1, about 1.2, about 1.4, about 1.6, about 1.8, about 2, about 2.5, about 3, about 3.5, about 4, about 4.5, about 5, about 6, about 8, about 10, about 12, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, about 40 mg per capsule or tablet or unit oral dosage form. In certain embodiments, a pharmaceutical composition provided herein comprises a di- -toluenesulfonate salt of Compound A22 in an amount of about 21, 30, or 35 mg per capsule or tablet or unit oral dosage form.
[0087] In certain embodiments, a pharmaceutical composition provided herein is formulated as an immediate-release capsule with a size of, e.g., size 1.
Methods of Treatment
[0088] This disclosure provides methods of treating Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer in a subject, comprising administering to a subject in need thereof a therapeutically effective amount of Compound A22. This disclosure is based on the surprising discovery of multiple mechanisms of action of Compound A22. The methods of this disclosure are based on the discovery that Compound A22 inhibits CKla, inducing p53 activation and apoptosis, and also inhibits CDK7 and CDK9, which inhibits MDM2 expression and MCL1 expression, thereby also activating p53 and inducing apoptosis. The methods of this disclosure exploit the surprising synergistic ability of Compound A22 to effect multiple mechanisms of action. The methods of this disclosure are further based on the discovery that in patients with GATA3mt, the P53 gene is almost always wild type. Treating GATA3mt tumors in preclinical studies with MDM2 inhibitors and MDM2 siRNAs has shown that these tumors, unlike GATA3wt tumors, are sensitive to MDM2 inhibition or knockdown. Thus, GATA3 mutations are synthetic lethal with MDM2 inhibition. Despite this observation and the fact that patients with GATA3 mutations have poor survival outcomes, there are no approved agents approved for the treatment of GAT A3 mt ER+ Her2- breast cancer. A major limitation has been the lack of well tolerated treatments for inhibiting MDM2 and activating P53. This disclosure addresses the unmet need of treatment for GATA3mt ER+HER2- breast cancer by applying the discovery of the novel mechanisms of action of Compound A22 and its surprising clinical tolerability.
[0089] In some embodiments, the breast cancer is refractory. In some embodiments, the breast cancer is metastatic. In some embodiments, the breast cancer is drug-resistant. In some embodiments, prior therapy has failed to stop progression of the breast cancer.
[0090] In some embodiments, the subject has GATA3/7?/. In some embodiments, the subject has p53vi7.
[0091] In certain embodiments, the breast cancer is drug-resistant. In certain embodiments, the breast cancer is resistant to CDK4/6 inhibitors, anti-estrogen therapies including aromatase inhibitors, PI3 kinase inhibitors, mTOR inhibitors, taxane chemotherapy, fluoropyrimidine chemotherapy, anti-HER2 antibodies, and/or antibody drug conjugates targeting HER2 or TROP2.
[0092] In certain embodiments, the subject is newly diagnosed with GAT A3 mt breast cancer. In certain embodiments, the subject has failed a prior therapy. In some embodiments, the subject has failed more than one prior therapy. In some embodiments, a prior therapy has failed to stop progression of the breast cancer.
[0093] In certain embodiments, the subject is a mammal. In certain embodiments, the subject is a human.
[0094] Methods provided herein encompass treating a subject regardless of patient’s age, although some diseases are more common in certain age groups.
[0095] In certain embodiments, the therapeutically effective amount of Compound A22 is ranging from about 0.001 to about 10 mg/kg per day, from about 0.002 to about 5 mg/kg per day, from about 0.005 to about 2 mg/kg per day, from about 0.01 to about 1 mg/kg per day, or from about 0.01 to about 0.5 mg/kg per day. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.001 to about 10 mg/kg per day. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.002 to about 5 mg/kg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.005 to about 2 mg/kg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.01 to about 1 mg/kg per day. In yet another embodiment, the therapeutically effective amount of a Compound A22 ranges from about 0.01 to about 0.5 mg/kg per day. In still another embodiment, the therapeutically effective amount of Compound A22, is about 0.01, about 0.02, about 0.03, about 0.05, about 0.08, about 0.1, about 0.12, about 0.15, about 0.17, about 0.2, or about 0.25 mg/kg per day.
[0096] In certain embodiments, the therapeutically effective amount of Compound A22 ranges from about 0.1 to about 200 mg per day, from about 0.2 to about 100 mg per day, from about 0.5 to about 50 mg per day, or from about 1 mg every other day to about 20 mg per day. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.1 to about 200 mg per day. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.2 to about 100 mg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 0.5 to about 50 mg per day. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 1 to about 20 mg per day. In yet another embodiment, the therapeutically effective amount of Compound A22, is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 22, about 23, about 24, about 25, about 26, about 27, about 28, about 29, about 30, about 31, about 32, about 33, about 34, about 35, about 36, about 37, about 38, about 39, or about 40 mg per day.
[0097] In certain embodiments, the therapeutically effective amount of Compound A22 ranges from about 1 to 500, from about 2 to 250, from about 5 to about 105, from about 10 to about 90, or from about 20 to about 65 mg per week. In one embodiment, the therapeutically effective amount of Compound A22 ranges from about 1 to 500 mg per week. In another embodiment, the therapeutically effective amount of Compound A22 ranges from about 2 to 250 mg per week. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 5 to about 105 per week. In yet another embodiment, the therapeutically effective amount of Compound A22 ranges from about 10 to about 90 mg per week. In some embodiments, the therapeutically effective amount of Compound A22 ranges from about 10 to about 63 or 65 mg per week. In still another embodiment, the therapeutically effective amount of Compound A22, is about 5, about 10, about 15, about 20, about 25, about 30, about 35, about 40, about 50, about 60, about 65, about 70, about 75, about 80, about 85, about 90, about 95, about 100, about 105 mg per week.
[0098] In certain embodiments, the compound is administered at a dose of about 1 mg, about 2 mg, about 3 mg, about 4 mg, about 5 mg, about 6 mg, about 7 mg, about 8 mg, about 9 mg, about 10 mg, about 11 mg, about 12 mg, about 13 mg, about 14 mg, about 15 mg, about 16 mg, about 17 mg, about 18 mg, about 19 mg, about 20 mg, about 21 mg, about 22 mg, about 23 mg, about 24 mg, about 25 mg, about 26 mg, about 27 mg, about 28 mg, about 29 mg, about 30 mg, about 31 mg, about 32 mg, about 33 mg, about 34 mg, about 35 mg, about 36 mg, about 37 mg, about 38 mg, about 40 mg, about 42 mg, about 45 mg, about 48 mg, about 51 mg, about 52 mg, about 55 mg, about 58 mg, about 60 mg, about 62 mg, about
63mg, about 64 mg, about 65 mg, about 66 mg, about 67 mg, about 68 mg, about 69 mg, about 70 mg, about 71 mg, about 72 mg, about 73 mg, about 75 mg, about 78 mg, about 80, about 81 mg, about 84 mg, about 87 mg, about 90 mg, about 93 mg, about 96 mg, about 99 mg, about 102 mg, or about 105 mg per week. In certain embodiments, the compound is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week.
[0099] It is understood that the administered dose of Compound A22 can also be expressed in units other than mg/kg every other day. For example, doses for parenteral administration can be expressed as mg/m2 per day. One of ordinary skill in the art would readily know how to convert doses from mg/kg per day to mg/m2 per day to given either the height or weight of a subject or both. For example, a dose of 1 mg/m2 per day for a 65 kg human is approximately equal to 58 mg/kg per day.
[00100] Depending on the disease to be treated and the subject’s condition, Compound A22 may be administered by oral, parenteral (e.g., intramuscular, intraperitoneal, intravenous, CIV, intracistemal injection or infusion, subcutaneous injection, or implant), inhalation, nasal, vaginal, rectal, sublingual, or topical (e.g., transdermal or local) routes of administration.
[00101] In one embodiment, Compound A22, is administered orally. In some embodiments, Compound A22 is administered as tablet or capsule. In another embodiment, Compound A22, is administered parenterally. In yet another embodiment, Compound A22, is administered intravenously. In yet another embodiment, Compound A22, is administered intramuscularly. In yet another embodiment, Compound A22, is administered subcutaneously. In still another embodiment, Compound A22, is administered topically.
[00102] A compound described herein, e.g., Compound A22, can be delivered as a single dose such as, e.g., a single bolus injection, or oral tablets or pills; or over time such as, e.g., continuous infusion over time or divided bolus doses over time. A compound described herein, e.g., Compound A22, can be administered repetitively if necessary, for example, until the subject experiences stable disease or regression, or until the subject experiences disease progression or unacceptable toxicity. Stable disease or lack thereof is determined by a method known in the art such as evaluation of subject’s symptoms, physical examination, visualization of the cancer that has been imaged using X-ray, CAT, PET, or MRI scan and other commonly accepted evaluation modalities.
[00103] A compound described herein, e.g., Compound A22, can be administered once daily (QD), or divided into multiple daily doses such as twice daily (BID), and three times daily (TID). In addition, the administration can be continuous, i.e., every day, or intermittently. The term “intermittent” or “intermittently” as used herein is intended to mean stopping and starting at either regular or irregular intervals. For example, intermittent administration of Compound A22, is administration for one to six days per week, administration in cycles (e.g., daily administration for two to eight consecutive weeks, then a rest period with no administration for up to one week), or administration on alternate days. In one embodiment, Compound A22 is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week. In certain embodiments, Compound A22 is administered for 3 days or 5 days per week. In some embodiments, Compound A22 is administered for 3 non- consecutive days per week. In certain embodiments, Compound A22 is administered on days 1, 2, and 3 in a week. In some embodiments, Compound A22 is administered on days 1, 2, 3, 4, and 5 in a week. In some embodiments, Compound A22 is administered on days 1, 3, and 5 in a week.
[00104] It will be understood, however, that the specific dose level and frequency of dosage for any particular subject can be varied and will depend upon a variety of factors including the activity of the specific compound employed, e.g., Compound A22, the metabolic stability and length of action of the compound, the age, body weight, general health, sex, diet, mode and time of administration, rate of excretion, drug combination, the severity of the particular condition, and the host undergoing therapy. [00105] In certain embodiments, Compound A22, is cyclically administered to a subject to be treated. Cycling therapy involves the administration of the compound for a period of time, followed by a rest for a period of time, and repeating this sequential administration. Cycling therapy can reduce the development of resistance to one or more of the therapies, avoid or reduce the side effects of one of the therapies, and/or improves the efficacy of the treatment.
[00106] Consequently, in one embodiment, Compound A22, is administered for a cycle of about one week, about two weeks, about three weeks, about four weeks, about five weeks, about six weeks, about eight weeks, or about ten weeks, with a rest period of 0 days to about four weeks. In one embodiment, Compound A22, is administered for a cycle of three weeks, four weeks, five weeks, or six weeks with a rest period of 0, 1, 3, 5, 7, 9, 12, or 14 days. In some embodiments, Compound A22 is administered for a cycle of 21-42 days. In some embodiments, Compound A22 is administered for a cycle of 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, or 35 days. In some embodiments, a cycle has no rest period. In some embodiments, Compound A22 is administered for consecutive cycles with no rest period between cycles. In some embodiments, Compound A22 is administered for multiple cycles with a rest period. In certain embodiments, the rest period is 7 days. In certain embodiments, the rest period is 14 days. In certain embodiments, the rest period is a period that is sufficient for bone marrow recovery. In certain embodiments, no rest period is required between cycles. The frequency, number, and length of dosing cycles can be increased or decreased.
[00107] In one embodiment, Compound A22, is administered for three weeks in a 28-day cycle with a 7-day rest period. In one embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered every day for five days of a week. In another embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered on Days 1, 2, 3, 4, 5, 8, 9, 10, 11, 12, 15, 16, 17, 18, and 19. In one embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered every other day for three days of a week. In another embodiment, in a 28-day cycle with a 7-day rest period, Compound A22, is administered on Days 1, 3, 5, 8, 10, 12, 15, 17, and 19.
[00108] In some embodiments, Compound A22 is administered 3 days per week for four weeks in a 28-day cycle. In some embodiments, Compound A22 is administered for multiple cycles. In some embodiments, when Compound A22 is administered for multiple cycles, there is a rest period between cycles. In some embodiments when Compound A22 is administered for multiple cycles, there is no rest period between cycles. [00109] In certain embodiments, the subject is treated with Compound A22, from about 1 to about 50, from about 2 to about 20, from about 2 to 10, or from about 4 to about 8 cycles. In certain embodiments, the subject is treated with Compound A22 from about 1 to about 50 cycles. In certain embodiments, the subject is treated with Compound A22 from about 2 to about 20 cycles. In certain embodiments, the subject is treated with Compound A22 from about 2 to 10 cycles. In certain embodiments, the subject is treated with Compound A22, from about 4 to about 8 cycles. In some embodiments, Compound A22 is administered with a rest period between cycles. In some embodiments, Compound A22 is administered with no rest period between cycles.
[00110] In one embodiment, provided herein is a method of inhibiting the growth of a cell, comprising contacting the cell with an effective amount of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
[00111] In another embodiment, provided herein is a method of modulating the activity of CKla in a cell, comprising contacting the cell with a compound of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
[00112] In yet another embodiment, provided herein is a method of inducing apoptosis in a cell, comprising contacting the cell with a compound of Compound A22, or an enantiomer, a mixture of enantiomers, a diastereomer, a mixture of two or more diastereomers, a tautomer, a mixture of two or more tautomers, or an isotopic variant thereof; or a pharmaceutically acceptable salt, solvate, hydrate, or prodrug thereof.
[00113] In some embodiments, the cell is a breast cancer cell. In certain embodiments, the cell is a relapsed or refractory breast cancer cell. In certain embodiments, the cell is a relapsed breast cancer cell. In certain embodiments, the cell is a refractory breast cancer cell. In certain embodiments, the cell is a metastic breast cancer cell. In certain embodiments, the cell is a drug-resistant breast cancer cell. Embodiments
[00114] In some embodiments, the disclosures relates to one or more of the following enumerated embodiments:
1. A method of treating a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22 or a pharmaceutically acceptable salt thereof.
2. The method of embodiment 1, wherein the patient has GATA3/7?/.
3. The method of any one of embodiments 1-2, wherein the patient has p53vi7.
4. The method of any one of embodiments 1-3, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene.
5. The method of any one of embodiments 1-4, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
6. The method of any one of embodiments 1-5, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
7. The method of any one of embodiments 1-6, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
8. The method of any one of embodiments 1 to 7, wherein the breast cancer is refractory.
9. The method of any one of embodiments 1 to 7, wherein the breast cancer is metastatic.
10. The method of any one of embodiments 1 to 7, wherein the breast cancer is drugresistant.
11. The method of any one of embodiments 1 to 10, wherein the patient has failed a prior therapy. The method of any one of embodiments 1 to 11, wherein the patient is a human. The method of any one of embodiments 1 to 12, wherein the compound is formulated as a -toluenesulfonate salt or a di- -toluenesulfonate salt. The method of any one of embodiments 1 to 12, wherein the pharmaceutical composition is administered orally. The method of any one of embodiments 1 to 14, wherein the pharmaceutical composition is administered as a tablet or capsule. The method of any one of embodiments 1 to 15, wherein the therapeutically effective amount is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 25 mg, about 30 mg, or about 35 mg. The method of any one of embodiments 1 to 16, wherein the pharmaceutical composition is administered in a cycle. The method of any one of embodiments 1 to 17, wherein the pharmaceutical composition is administered in a 28-cycle. The method of any one of embodiments 1 to 18, wherein the pharmaceutical composition is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week. The method of any one of embodiments 1 to 19, wherein the pharmaceutical composition is administered for 3 days or 5 days per week. The method of any one of embodiments 1 to 20, wherein the pharmaceutical composition is administered on three non-consecutive days in a week. The method of any one of embodiments 1 to 21, wherein the pharmaceutical composition is administered in a 28-day cycle for 3 days per week. The method of any one of embodiments 1 to 22, wherein the pharmaceutical composition is administered in a 28-day cycle for 3 non-consecutive days per week. The method of any one of embodiments 1 to 20, wherein the pharmaceutical composition is administered in a 28-day cycle for 5 days per week. The method of embodiment 24, wherein the pharmaceutical composition is administered in a 28-day cycle on Days 1, 2, 3, 4, and 5 per week. The method of any one of embodiments 1 to 25, wherein the therapeutically effective amount is about 21 mg. The method of any one of embodiments 1 to 25, wherein the therapeutically effective amount is about 30 mg. The method of any one of embodiments 1 to 25, wherein the therapeutically effective amount is about 35 mg. A method of inhibiting MDM2 in a patient having ER+, HER- metastatic breast cancer and having GATA3/7?/, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22, or a pharmaceutically acceptable salt thereof. The method of embodiment 26, wherein the compound is formulated as a di- - toluenesulfonate salt. The method of any one of embodiments 26 to 27, wherein the therapeutically effective amount is about 21 mg per day, about 30 mg per day, or about 35 mg. The method of any one of embodiments 26 to 27, wherein the pharmaceutical composition is administered three times per week. The method of any one of embodiments 26 to 30, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene. The method of any one of embodiments 26 to 31, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene. The method of any one of embodiments 26 to 32, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene. The method of any one of embodiments 26 to 33, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene. A method of inhibiting CDK7 in a subject having ER+, HER- metastatic breast cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof. The method of embodiment 37, wherein the compound is formulated as a di- - toluenesulfonate salt. The method of any one of embodiments 37 to 38, wherein the breast cancer has been resistant to a CDK4/6 inhibitor. The method of any one of embodiments 37 to 39, wherein the subject has P53VI . The method of any one of embodiments 37 to 40, wherein the subject has GATA3wt. The method of any one of embodiments 37 to 40, wherein the subject has GATA3mt. The method of any one of embodiments 37 to 42, wherein the subject over-expresses MDM2 or has a >4 copy number of the MDM2 gene. The method of any one of embodiments 37 to 43, wherein the subject over-expresses MDM2 or has a >6 copy number of the MDM2 gene. The method of any one of embodiments 37 to 44, wherein the subject over-expresses MDM2 or has a >8 copy number of the MDM2 gene. The method of any one of embodiments 37 to 44, wherein the subject over-expresses MDM2 or has a >10 copy number of the MDM2 gene. The method of any one of embodiments 37 to 46, wherein Compound A22 is administered three times per week. The method of any one of embodiments 37 to 47, wherein the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg per day. A method of treating cancer in a subject having amplified expression of MDM2 or having a >4 copy number of the MDM2 gene, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof. The method of embodiment 49, wherein the compound is formulated as a di- - toluenesulfonate salt. The method of any one of embodiments 49 to 50, wherein the subject over-expresses MDM2 or has a >6 copy number of the MDM2 gene. The method of any one of embodiments 49 to 51, wherein the subject over-expresses MDM2 or has a >8 copy number of the MDM2 gene. The method of any one of embodiments 49 to 52, wherein the subject over-expresses MDM2 or has a >10 copy number of the MDM2 gene. The method of any one of embodiments 49-53, wherein administering comprises administering about 21 mg of Compound A22 three times per week. The method of any one of embodiments 46-50, wherein administering comprises administering about 30 mg of Compound A22 three times per week. The method of any one of embodiments 46-50, wherein administering comprises administering about 35 mg of Compound A22 three times per week. A method of inhibiting MDM2 expression in a subject having cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof. The method of embodiment 57, wherein the compound is formulated as a di- - toluenesulfonate salt. The method of any one of embodiments 57 to 58, wherein the subject over-expresses MDM2 or has a >4 copy number of the MDM2 gene. The method of any one of embodiments 57 to 59, wherein the subject over-expresses MDM2 or has a >6 copy number of the MDM2 gene. The method of any one of embodiments 57 to 60, wherein the subject over-expresses MDM2 or has a >8 copy number of the MDM2 gene. The method of any one of embodiments 57 to 61, wherein the subject over-expresses MDM2 or has a >10 copy number of the MDM2 gene. The method of any one of embodiments 54 to 59, wherein administering comprises administering about 21 mg of Compound A22 three times per week. The method of any one of embodiments 54 to 59, wherein administering comprises administering about 30 mg of Compound A22 three times per week. The method of any one of embodiments 54 to 59, wherein administering comprises administering about 35 mg of Compound A22 three times per week. A pharmaceutical composition comprising Compound A22, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, for use in the treatment of a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer. The pharmaceutical composition for use according to embodiment 66, wherein the patient has GAT3/7?/. 68. The pharmaceutical composition for use according to any one of embodiments 66-67, wherein the treatment comprises administration of the pharmaceutical composition to the patient on three non-consecutive days in a week.
69. The pharmaceutical composition for use according to any one of embodiments 66-68, wherein the administration occurs for one or more cycles.
70. The pharmaceutical composition of embodiment 69, wherein the cycle is 28 days.
71. The pharmaceutical composition for use according to any one of embodiments 66-70, wherein the pharmaceutical composition is orally administered to the patient.
72. The pharmaceutical composition for use according to any one of embodiments 66-71, wherein the pharmaceutical composition is formulated as an oral unit dosage form.
73. The pharmaceutical composition for use according to any one of embodiments 72, wherein the oral unit dosage form comprises about 21 mg of Compound A22.
74. The pharmaceutical composition for use according to any one of embodiments 72, wherein the oral unit dosage form comprises about 30 mg of Compound A22.
75. The pharmaceutical composition for use according to embodiment 72, wherein the oral unit dosage form comprises about 35 mg of Compound A22.
[00115] The disclosure will be further understood by the following non-limiting examples.
EXAMPLES
[00116] As used herein, the symbols and conventions used in these processes, schemes, and examples, regardless of whether a particular abbreviation is specifically defined, are consistent with those used in the contemporary scientific literature, for example, the Journal of the American Chemical Society, the Journal of Medicinal Chemistry, or the Journal of Biological Chemistry. Specifically, but without limitation, the following abbreviations may be used in the examples and throughout the specification: g (grams); mg (milligrams); mL (milliliters); mL (microliters); mM (millimolar); mM (micromolar); mmol (millimoles); h (hour or hours); and min (minutes).
Example 1
Cellular Cytotoxicity of Compound A22 and a Reference Compound, Milademetan (an MDM2 Inhibitor)
[00117] Evaluation of cellular cytotoxicity and determination of IC50 of Compound A22 was performed in two cell lines, MCF7 and RKO. Genetic features of these lines are described in the Table 2 below.
Table 2
[00118] Growth curves were determined for each cell line to determine optimal seeding for the cellular cytotoxicity assay.
[00119] Cells were seeded into 96-well plates (Corning 3904) with 100 pL media at 250, 500, 1000, 2000, 5000 and 10,000 cells/well on Day -1. Cell growth was measured by Cell Titer Glow® 2.0 Assay (Promega, G9243) on Days 0, 1, 2 and 3 (total 96 hours) measured with an Envision plate reader. The experiment was performed with 5 replicates.
[00120] The results showed that optimal seeding to achieve log-linear growth for the cytotoxicity experiment required 1,500 cells/well for MCF7 and 2,000 cells/well for RKO.
[00121] A serial titration of Compound A22 and Milademetan at ten concentrations was performed to determine the IC50 of each compound in the two cell lines. Milademetan is a direct MDM2 inhibitor. Duplicate wells were evaluated for each compound concentration and two biological replicates were performed. Cell growth was measured by Cell Titer Glow® 2.0 Assay.
[00122] Results are shown in Table 3 below:
Table 3
Summary of IC50 values (|1M) (N=l, N=2)
[00123] The data show that both Compound A22 and Milademetan take more than 24 hours for cell killing with sub-micromolar IC50 values, which is approximately the time for cell doubling.
Example 2
Evaluation of Biomarker Responses in MCF7 by Western Blot
[00124] To evaluate biomarker responses by Western blot, MCF7 cells were seeded into 10 cm cell culture dishes and cultured for 24 hours. Compound A22 or vehicle (DMSO) were added and incubated with cells for 16 hours. After incubation, the media was aspirated and cells were harvested with a trypsin-EDTA solution. Cells were transferred to tubes and washed with cold PBS.
[00125] The washed cells were lysed by the addition of 500 pL pre-chilled RIPA buffer (Boston BioProducts BP115D) with protease and phosphatase inhibitors (Roche 05892791001; Roche 0490068337001). Cell were incubated for 30 minutes on ice.
[00126] Lysates were clarified by spinning at 12000 rpm for 10 minutes at 4 degrees and the supernatant was transferred to a fresh tube. Protein concentrations were determined by BCA (Thermo Scientific) and samples were adjusted to the same final protein concentration with RIPA.
[00127] Samples were combined with a loading buffer and heated at 100°C for 10 minutes before cooling to room temperature. Samples were loaded onto a polyacrylamide gel and run for 20 minutes at 80V, followed by an increase to 120 V for 120 minutes.
[00128] Samples were electrotransferred to a nitrocellulose membrane for 90 minutes. Membranes were treated with 5% BSA for 1 hour to block non-specific binding. Primary antibody as described in Table 4 were added and incubated in blocking buffer at 4°C overnight. Table 4
[00129] The next day, membranes were washed three times (5 min each) with TBST (Millipore). Secondary antibody (anti-rabbit IgG (Licor, 926-32211) or anti-mouse IgG (LI- COR, 926-68070) was then added and incubated for 1 hour at room temperature in blocking buffer. Finally, membrane was washed three times with TBST as above and read on the LiCOR.
[00130] Results are shown in FIGS. 1 and 2 and were as follows: pRbpl S2/5 was reduced by 73% compared to DMSO control; MCL-1 was reduced by 70% compared to DMSO control; Cleaved PARP was increased by 12 fold and p53 was increased by 5 fold compared to DMSO treatment. Gamma-142 AX, a marker of DNA damage response, was increased by approximately 2-fold (data not shown).
[00131] Rb phosphyorylation was also decreased by Compound A22 treatment. Phosphorylation at S780 was decreased approximately 80% and phosphorylation at Ser807/810 was decreased by approximately 90%. Rb itself was decreased by approximately 50%. However, Actin levels were unchanged, suggesting that these changes in Rb are specific to the mechanism of action of Compound A22 rather than being non-specific effects. Example 3
Evaluation of CDK2 and Phospho-CDK2 by Immunoprecipitation
[00132] CDK7 is the catalytic component of the CAK (Cyclin Activating Kinase) protein complex. One of its roles is to phosphorylate CDK2 to enable progression from G1 into S phase in the cell cycle.
[00133] Detection of CDK2 and phospho-CDK2 in cell lysates required the use of an immunoprecipitation step prior to a Western blot analysis. Cells were seeded into culture dishes and cultivated for 24 hours as in Example 3. After treatment with 100 nM Compound A22 for 16 hours at 37°C, cells were processed using trypsin-EDTA solution, washed with cold PBS, and lysed in 0.5 mL ice cold IP -MS Cell Lysis Buffer (ThermoFisher #87787) with protease inhibitor cocktail and phosphatase inhibitor cocktail for cell lysis for 30 min on ice. Samples were clarified by centrifugation as above and the supernatant evaluated for protein concentration by BCA assay. 50 pg of the lysate was retained as input to the immunoprecipitation.
[00134] Pierce Protein A/G magnetic beads (100 uL per IP) were prepared by washing 3 times in TBST followed by incubation with the anti-Cdk2 antibody (Cell Signaling Technology #2546) for 1 hour at room temperature. The immunoprecipitation was carried out by adding the antibody conjugated magnetic beads to 1 mg of cell lysate. Samples were incubated overnight at 4°C with rotation on a ThermoMixer at 300 rpm. The beads were recovered magnetically and washed 3 times with TBST.
[00135] For Western analysis, 24 pL of sample was mixed with 6 pL 5X SDS-PAGE loading buffer (Beyotime, ST628), mixed and incubated at 95°C for 10 min. 15 pL of IP supernatant was loaded on a gel, run for 30 minutes at 80V and then 1.5 hours at 120V, followed by electro transfer to a nitrocellulose membrane as described in Example 3. Membrane was blocked with LLCOR blocking buffer (LLCOR, 927-60001) for 30 minutes, washed three times with TBST, and incubated with primary antibody in LLCOR blocking buffer. After washing the membrane three times with TBST, they were incubated with secondary antibody for 1 hour at room temperature and read on the Li-COR.
[00136] Results for CDK2 and phospho-CDK2 are shown in FIG. 3. Phospho-CDK2 (pCDK2-T160; Cell Signaling, 2561 S) decreased approximately 2-fold while CDK2 (78B2; Cell Signaling, 2546S) decreased approximately 25% after Compound A22 (BTX-A51) treatment.
Example 4
Open Label, Escalating Multiple Dose Study to Evaluate the Safety, Toxicity, and Pharmacokinetics of Compound A22 Capsules in Subjects with Advanced Solid Tumors and Estrogen Receptor Positive, Human Epidermal Growth Factor Receptor 2 Negative Breast Cancer
[00137] This is a multicenter, open label, nonrandomized, sequential dose escalation/cohort expansion, multiple dose study designed to evaluate the safety, toxicity, and PK as well as preliminary efficacy of Compound A22 in subjects with advanced solid tumors. The study has two phases: Phases la and lb. Throughout the study, safety is evaluated by a Dose Escalation Committee (DEC) that includes the Principal Investigator(s), the Sponsor’ s physician (in consultation with the Sponsor’s pharmacologist/ pharmacokineticist as needed), and/or independent experts. The DEC reviews all cumulative available data and authorizes plans for the dosing of each subsequent cohort. Treatment decisions for individual subjects is the responsibility of the Investigators.
[00138] Phase la is designed to determine the dose limiting toxi cities (DLTs), maximum tolerated dose (MTD), and recommended Phase 2 dose (RP2D) of orally administered Compound A22. The Compound A22 starting dose for Cohort 1 is <21 mg and does not exceed the highest dose deemed safe as determined in the BTX-A51-001 study of Compound A22 in subjects with acute myeloid leukemia and myelodysplastic syndrome. Compound A22 is administered once daily on a weekly schedule of 5 days on/2 days off. Dose escalation proceeds according to a modified 3+3 design. Each cycle consists of 28 days (4 weeks), and the DLT observation period is the first cycle (i.e., 28 days after initiation of dosing). A DLT may be observed in no more than 0 out of 3 or 1 out of 6 subjects who have completed the DLT observation period before the next cohort initiates accrual. Barring DLT, sequential dose escalation of Compound A22 proceeds with up to a total of 6 dose levels to a maximum of 200 mg; on the basis of these an MTD is identified. The MTD is defined as the highest dose level with a subject incidence of DLTs of 0 or 1 out of 6 during the first 28 days of study drug dosing. A minimum of 6 subjects are treated at a dose level before this dose level can be deemed as the MTD. Subjects who receive less than 75% of planned doses due to reasons other than a DLT are replaced for the determination of the MTD but remain part of the safety population if they received at least one dose of study drug. At dose levels equal to or below the MTD, up to 6 additional subjects per dose level cohort may be enrolled to obtain additional safety, PK, and pharmacodynamic (PD) data.
[00139] Toxicity severity is graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events (NCI CTCAE) Version 5.0. For purposes of dose escalation, the totality of accrued safety information across all cycles completed at the time of DEC data review is taken into consideration. A DLT is defined as a severe or clinically significant AE or abnormal laboratory value (Grade 3 or greater, unless otherwise specified), unless it is clearly related to disease progression, intercurrent illness, preexisting condition, or concomitant medications.
[00140] In Phase la, the RP2D has been determined to be 21 mg administered 3x/week based on a comprehensive analysis of available safety, tolerability, PK/PD, and preliminary efficacy as well as the MTD from the dose escalation phase.
[00141] In Phase lb, up to 40 additional subjects are enrolled to evaluate safety and preliminary efficacy of Compound A22 in subjects with estrogen receptor positive (ER+), human epidermal growth factor receptor 2 negative (HER2-), GATA3 mutant (mt) and wildtype (wt) metastatic breast cancer (mBC). Based on the Phase la results, the RP2D for Compound A22 was determined to be 21 mg administered orally 3-times-weekly (3x/week) on a Monday /Wednesday /Friday (MWF) schedule for 4 weeks in a 28 day cycle; this dose and schedule is evaluated in Phase lb.
[00142] Phase lb is conducted in a staged manner with 22 subjects enrolled to Stage 1. If an objective response (i.e., PR or CR) is observed in >2 subjects of the first 22 enrolled, then an additional 18 subjects are enrolled. Of the first 22 subjects enrolled, a minimum of 13 subjects should have a pathogenic GAT A3 mutation based on next generation sequencing (NGS) of tumor tissue. Of the 40 subjects enrolled to Stages 1 and 2, a minimum of 24 subjects should have a GAT A3 mutation.
[00143] Dosing in this phase of the study consists of the first cycle of therapy (i.e., 28 days). The DEC reviews cumulative safety and available PK data in subjects treated in Phase lb for DLTs, with DEC reviews scheduled after the 6th, 12th, 22nd (interim analysis), 30th, and 40th subjects complete a cycle of Compound A22. [00144] Subjects who complete one cycle of Compound A22 in either Phase la or Phase lb are offered continued access to study drug until disease progression or unacceptable toxicity. The DEC continues to review accruing safety/PK data, inclusive of all cycles, for subjects who continue with treatment.
[00145] Once treatment has completed, subjects are contacted by telephone every 3 months for survival status and anticancer therapy; the cause of death is documented. Individual subjects are considered to have completed the study 2 years after their last treatment or upon death, whichever occurs first.
[00146] The study ends after a minimum of 70% of subjects have died or 5 years after the last subject is enrolled, whichever occurs first.
[00147] The duration of the treatment phases of this study is approximately 38 months. The Follow up Phase is 5 years after the last subject is enrolled.
[00148] The Phase lb includes a cohort of up to approximately 40 subjects with advanced ER+, HER2-, GATA3 mt and wt mBC to further characterize the safety, tolerability, and preliminary efficacy when administered at the recommended dose (21 mg 3x/week) identified in the Dose-Escalation Phase la.
[00149] Eligible subjects for the study demonstrate an understanding by voluntarily signing an informed consent form and are 18 years old or older with histologically or cytologically documented, incurable or metastatic solid tumor that is refractory to or intolerant of all standard therapy or for which no standard therapy is available. To be eligible for Phase lb, subjects have histologically confirmed diagnosis of ER+, HER2- mBC not amenable to resection or radiation therapy with curative intent. Further, the subject has (i) adequate tumor tissue sample from locally recurrent or metastatic site; (ii) documented evidence of ER+ mBC confirmed with the most recently available tumor biopsy from a locally recurrent or metastatic site. ER should be more than 10% ER positive or Allred >5 by local laboratory testing; (iii) documented evidence of HER2- status defined as a negative in situ hybridization test or an IHC status of 0, 1+ or 2+. If IHC is 2+, a negative in situ hybridization (FISH, CISH, or SISH) test is required by local laboratory testing and based on the most recently analyzed tissue sample; and (iv) GATA3 and p53 mutational status based on NGS data obtained prior to or during screening. [00150] Participants in the Phase lb expansion phase have progressed on or been intolerant of standard therapy (at least 2 prior lines of endocrine therapy and up to 3 lines of chemotherapy). Participants that qualify for PARP inhibitors must have progressed on or been intolerant of these agents. For participants who experienced metastatic disease recurrence within 12 months of curative-intent chemotherapy, the (neo)adjuvant chemotherapy would count as one line of chemotherapy in the metastatic setting. For participants whose disease recurs within 12 months of completing adjuvant endocrine therapy, this will count as one line of endocrine therapy for eligibility requirements.
[00151] Disease is measurable per RECIST vl.l.
[00152] Adequate organ function as defined by the following criteria; patients should not have had a transfusion within 2 weeks prior to hematology evaluation for eligibility: absolute neutrophil count (ANC) >1000 cells/pL; white blood cell (WBC) count >1500/pL; platelet count >100,000/pL; hemoglobin >9.0 g/dL; serum AST and serum ALT <3.0 x ULN (<5.0 x ULN in subjects with documented liver involvement); total bilirubin <1.5 x ULN or < 3.0 x ULN and primarily unconjugated bilirubin (per contemporaneous evaluation of both total and direct bilirubin) if patient has a documented history of Gilbert's syndrome or genetic equivalent; creatinine clearance of >50 mL/min; international normalized ratio (INR) and activated partial thromboplastin time (aPTT) <1.5 x ULN; subjects on full-dose oral anti coagulation must be on a stable dose (minimum duration 14 days); if receiving warfarin, the subject must have an INR <3.0 and no active bleeding (i.e., no bleeding within 14 days prior to first dose of study drug); subjects on low molecular weight heparin are allowed.
[00153] Subjects must have an Eastern Cooperative Oncology Group (ECOG) performance status of 0 or 1.
[00154] Female participants of childbearing potential may participate provided they have a negative serum pregnancy test at screening and a negative serum pregnancy test within 72 hours prior to starting on treatment; they also must agree to either abstain from sexual intercourse or use a highly effective method of contraception (for up to 3 months after the last dose of study drug) such as: established use of oral or long acting injected or implanted hormonal methods of contraception for at least 90 days prior to Cycle 1 Dose 1 (participants in Part lb should not be on any form of hormonal contraception); placement of an intrauterine device or intrauterine system; barrier methods: condom, diaphragm, or cervical/vault cap with spermicidal foam/gel/film/cream/suppository; male partner sterilization (with the appropriate post vasectomy documentation of the absence of spermatozoa in the ejaculate [the vasectomized partner should be the sole partner for that subject]); or true abstinence (when this is in line with the preferred and usual lifestyle of the subject).
[00155] Male participants sexually active with a woman of childbearing potential (even those who have had a vasectomy) must agree to use a barrier method of birth control (see above) during and after the study (up to 3 months after the last dose of study drug); all men must not donate sperm during the study and for 3 months after receiving the last dose of study drug.
[00156] The study excludes those with life expectancy <3 months. In the Phase lb expansion phase, the study excludes subjects with inflammatory breast cancer. The study also excludes subjects undergoing treatment with any local or systemic antineoplastic therapy (including chemotherapy, hormonal therapy, or radiation) within 3 weeks prior to first dose of Compound A22, with the following exceptions: hormonal therapy with gonadotropinreleasing hormone (GnRH) agonists or antagonists for prostate cancer; hormone-replacement therapy or oral contraceptives; herbal therapy intended as anti -cancer >1 week prior to first dose of Compound A22; palliative radiotherapy for bone metastases >2 weeks prior to first dose of Compound A22. The study excludes those who have engaged in chronic use of corticosteroids in excess of 10 mg daily of prednisone or equivalent within 4 weeks prior to first dose of Compound A22 (replacement doses of corticosteroids, e.g., prednisone 5-7.5 mg daily are acceptable). The study excludes those who have had major trauma or major surgery within 4 weeks prior to first dose of Compound A22. The study also excludes subjects who experienced adverse events from prior anti-cancer therapy that have not resolved to Grade <1 except for alopecia or Grade <2 immunotherapy related thyroid toxicity. The study excludes subjects with active central nervous system (CNS) disease involvement, or prior history of NCI CTCAE Grade >3 drug-related CNS toxicity. Subject with known CNS metastases that are treated and stable (without evidence of CNS toxicity) and are not requiring systemic steroids are allowed to be enrolled. This study excludes subjects with active or prior history of leptomeningeal disease. The study also excludes subjects with clinically significant cardiac disease, such as: myocardial infarction within 6 months before Cycle 1, Day 1 or unstable or uncontrolled disease/ condition related to or affecting cardiac function (e.g., unstable angina, congestive heart failure [New York Heart Association Class III or IV]); screening 12-lead electrocardiogram (ECG) showing a Baseline average QT interval as corrected by Fridericia's formula (QTcF) >470 msec. Subjects with left or right bundle branch block, where QT cannot be accurately assessed, will be allowed into the study with the approval of the Medical Monitor; LVEF <40%.
[00157] This study excludes subjects with active uncontrolled systemic fungal, bacterial, mycobacterial, or viral infection (defined as ongoing signs/symptoms related to the infection without improvement despite appropriate antibiotics, antiviral therapy, and/or other treatment); prophylactic therapy according to institutional protocols is acceptable. This study excludes subjects with known positive test result for human immunodeficiency virus (HIV) or acquired immune deficiency syndrome (AIDS). This study excludes subjects with active hepatitis C virus (HCV) or hepatitis B virus (HBV); subjects who are positive for hepatitis B core antibody, hepatitis B surface antigen, or hepatitis C antibody must have a negative PCR result before enrollment; those who are PCR positive are excluded. This study excludes subjects with second primary malignancy that has not been in remission for greater than 3 years; exceptions that do not require a 3-year remission include: non-melanoma skin cancer, cervical carcinoma in situ on biopsy or squamous intraepithelial lesion on Papanicolaou (PAP) smear, localized prostate cancer (Gleason score <6), or resected melanoma in situ; other localized, solid tumors in situ or other low risk cancers may also be exempt after discussion with the Sponsor Medical Monitor. This study excludes subjects with any serious underlying medical (e.g., pulmonary, renal, hepatic, gastrointestinal, or neurological) or psychiatric condition (e.g., alcohol or drug abuse, dementia or altered mental status) or any issue that would limit compliance with study requirements, impair the ability of the subject to understand informed consent, or that in the opinion of the investigator would contraindicate the subject's participation in the study or confound the results of the study. This study excludes subjects who are pregnant, lactating, or breastfeeding. This study excludes subjects who participate or plan to participate in another interventional clinical study while taking part in this protocol; participation in an observational study is acceptable.
[00158] The study drug in phases la and lb is orally administered, immediate-release capsules of 1.0 mg, 2.0 mg, and 7.0 mg. The capsules contain Compound A22 as the active moiety as well as the following inactive compendial excipients: Suglets (Sugar beads), Talc (USP/EP), and Povidone (USP). On Day 1 of each applicable cycle, the study drug is dispensed in separate bottles for each capsule strength, including a sufficient amount to complete the treatment cycle (with the exception that the 0.5-mg and 1.0-mg strengths are not be dispensed together). The 1.0 mg and 2.0 mg capsules are visually distinguishable from one another with the 1.0 mg capsules being white, and the 2.0 mg capsules being orange. The 7 mg capsules are also white but are much bigger. The 7 mg is a size 000 capsule, whereas the 1 mg is a size 1. Note that the capsules strengths on the study drug bottle label are highlighted in different colors: 1 mg - blue, 2 mg - red, and 7 mg - green.
[00159] The six dosing levels in Phase la are listed in Table 1. The numbers of subjects and doses administered may be altered in response to toxicity or tolerability.
Table 1 : Dosing Levels in Phase la
Abbreviation: TBD=to be determined. a De-escalation in dose, if required in response to DLT, will be to a mid-dose between the current dose and the previous lower dose with DLT subject incidence of <33%. b Dose level increases will be 2-fold or less. c Available for dose modification (de-escalation or intermediate dose) that deviates from the planned treatment groups.
[00160] Dosing in Phase lb is at the RP2D, which is 21 mg administered 3x/week. To the greatest extent practicable, Day 1 (and thus the first dose) of each cycle is scheduled to occur preferably on a Monday. For a given subject, one or more doses may be held as needed to manage toxicity. In some cases, the dose and/or schedule is lowered in subjects in response to toxicity.
[00161] Subjects are instructed to take their whole daily dose at approximately the same time each day and to swallow capsules whole. Subjects are instructed to open the Compound A22 packaging as close as possible to when they are going to take Compound A22. Subjects may take study drugs with or without food. Subjects are advised that if study drug is taken with food, they should avoid consuming a high fat meal.
[00162] Dosing may be interrupted in response to toxi cities or tolerability issues not amounting to a DLT. In such subjects, dosing will resume at the original dose; no dose reductions are permitted for non-DLT AE. In the event of a dose interruption due to a DLT, treatment is recommenced at a lower dose or schedule following appropriate resolution of toxicity. Subjects who experience any Grade > 3 skin drug-related AE or suspected Stevens Johnson syndrome (SJS) or toxic epidermal necrolysis (TEN) are withdrawn.
[00163] Treatment is discontinued in the event of disease progression, unacceptable toxicity or prolonged time required for recovery from toxicity, confirmed pregnancy, withdrawal of consent, loss to follow-up or noncompliance with protocol, or if the Investigator determines removal from the study is in the subject’s best interest.
[00164] Following provision of written informed consent, eligibility for participation is assessed during a screening period occurring up to 28 days before the start of study drug. The study follows the below schedule of assessments.
Schedule of Assessments
[00165] Abbreviations: AE=adverse event; C=cycle; CR=complete response; CT=computed tomography; D=Day; DEC=Dose Escalation Committee; DNA= deoxyribonucleic acid; ECG=electrocardiogram; ECHO=echocardiogram; ECOG=Eastem Cooperative Oncology Group; EOT=End of Treatment; GLS=global longitudinal strain; HBsAg=hepatitis B surface antigen; HBV=hepatitis B virus; HCV=hepatitis C virus;
HIV=human immunodeficiency virus; LVEF=left ventricular ejection fraction; MRI=magnetic resonance imaging; PBMC=peripheral blood mononuclear cells; PE=physical examination; PK=pharmacokinetics; SAE=serious adverse event.
[00166] Subjects return to clinic for the EOT visit 14 to 28 days after the last dose of study drug, to assess AEs and SAEs. In addition, a 30-day Safety follow-up is conducted by phone 30 days (±7 days) after the last dose to assess AEs.
[00167] Medical history includes significant past medical events (e.g., prior hospitalizations or surgeries), a review of the disease under study, prior anti-cancer therapies, and any concurrent medical illnesses.
[00168] Screening and end of treatment physical examinations include a complete physical examination. Beginning on CID 1, a modified physical examination is performed to monitor for any changes (e.g., lymph nodes, lung, cardiac, abdomen, skin, neurologic, and any systems, as clinically indicated). Weight (without shoes) is measured at each physical examination. Height (without shoes) is measured at screening only.
[00169] Vital signs are taken within 15 min pre-Compound A22 dose. On CID 1 and C1D5, vitals are collected at 2 and 4 hours postdose (±15 min); at all other visits, vital signs are taken predose only. [00170] A triplicate 12-lead ECG is collected at screening. Single 12-lead ECGs are collected for the remaining timepoints specified in the table.
[00171] Screening chemistry, hematology, and coagulation is collected within 10 days of Study Day 1/C ID 1.
[00172] Serum pregnancy is conducted at screening for female subjects of childbearing potential. Urine pregnancy is conducted predose on Day 1 of each cycle and at EOT.
[00173] HIV, HBsAg, and HCV antibody viral testing in subjects with current or prior history of HIV, HBV, and HCV infection, respectively, is conducted at screening.
[00174] Plasma samples for Compound A22 PK are collected prior to the dose administered in the clinic on that day in all patients that are not having intensive PK collection. Blood samples are collected from each subject for measurement of plasma concentrations of Compound A22 (and its demethylated metabolite, if applicable) on the days indicated in the Schedule of Assessments.
[00175] Intensive PK is collected in all subjects in Phase la dose escalation cohorts and in the first 10 subjects in the Phase lb dose expansion. Plasma samples for intensive PK are collected on C1D1 and C1D5 at pre-dose (within -2 hrs) and 1 (±10 min), 2 (±15 min), 3 (±20 min), 6 (±30 min), and 24 (±1 hr) hours post dose. A 48 (± 3hr) hour post dose sample is collected prior to the dose scheduled for Day 3. The Day 8 sample (±1 day) is collected prior to the dose scheduled for Day 8. Actual sample collection time is recorded. In response to a DLT or any significant safety concern, PK blood samples are collected from the affected subject in order to measure levels of Compound A22.
[00176] Exploratory biomarker blood samples are collected for future analyses, in efforts to identify novel biomarkers of response in breast cancer and other solid tumors and to better understand the biological effect of Compound A22. This includes PBMCs to evaluate MCL- 1, MDM2, and p53 protein expression and serum-based biomarkers (including but not limited to MIC-1, Macrophage Inhibitory Cytokine-1) as a surrogate of p53 since it is a p53- regulated secreted cytokine. Exploratory biomarker samples are collected: pre-dose on Day 1, on Day 2 (24-hours post-dose), 4 hours post-dose on Day 5 of Cycle 1, and 4 hours postdose on Day 1 at the beginning of Cycles 2 and 3. Collection timepoints may be adjusted in response to emerging data. [00177] Biological specimens are collected for the evaluation of exploratory systemic and/or tissue specific biomarkers, to explore association with study drug response, including efficacy and/or AEs, and to increase knowledge and understanding of the biology of Compound A22. Samples for pharmacodynamic/biomarker assessments are collected predose on Day 1, on Day 2 (24 hours post-dose), 4 hours post-dose on Day 5 of Cycle 1, and 4 hours post-dose on Day 1 at the beginning of Cycles 2 and 3. Collection timepoints may be adjusted in response to emerging data. The specific analyses include, but are not limited to, the biomarkers and assays described in the following assessments: serum biomarkers (including but not limited to MIC-1, Macrophage Inhibitory Cytokine- 1) as a surrogate of p53 since it is a p53 -regulated secreted cytokine; plasma biomarkers; whole blood genomic DNA; whole blood RNA; circulating tumor DNA; exploratory peripheral blood mononuclear cells (PBMC) to evaluate MCL-1, MDM2, and p53 protein expression; cytogenetics and mutation panel may including: gene expression levels of target SE genes (i.e., Mell, MYC, MYB, and MDM2) and gene mutation analysis by NGS.
[00178] Genomic DNA is collected predose on CID 1 or may be collected at any time during the course of the study.
[00179] Archival tumor tissue for biomarker analysis is collected. The tissue is obtained either at the time of or after the diagnosis of advanced disease has been made and from a site not previously irradiated. Tissue is collected between screening and Cycle 2.
[00180] For patients enrolled in Phase la dose escalation, an ECHO, to be performed for the evaluation of LVEF and GLS, is performed within the 14 days prior to starting study treatment. ECHO testing is also conducted during Cycles 2, 4, and 6 (no earlier than the Day 15 Visit) and at the end of Cycle 8 (no earlier than the Day 22 Visit), or more frequently if clinically indicated. It is acceptable for ECHO results to be obtained within 7 days prior to the next cycle’s Day 1 Visit; however, results and interpretation must be received prior to the start of the next cycle (Cycles 3, 5, and 7). For patients enrolled in Phase lb dose expansion, ECHO is performed within the 14 days prior to the start of treatment, i.e., Day 1 of Cycle 1. If troponin levels are elevated, post-treatment ECHO is conducted at the time of the elevation and every 2 months thereafter until troponin levels have returned to baseline.
[00181] Tumor evaluation by CT/MRI is performed during screening (within 28 days of
Day 1) and every 8 weeks on Day 1 of each odd numbered cycle. A baseline scan within 6 weeks of Day 1 is acceptable. C3D1 and C5D1 scans have a -7-day window. The same radiographic procedure used to define measurable lesions is used throughout the study for each subject. For subjects in Phase lb, a baseline CT/MRI of the head is performed during screening.
[00182] Primary analysis of efficacy of Compound A22 is based on the investigator’s assessment of response. For solid tumors, responses are evaluated using RECIST vl.l. For subjects with solid tumors, tumor evaluation by CT/MRI is performed during screening (within 28 days of Day 1) and every 8 weeks on Day 1 of each odd numbered cycle. A baseline scan within 6 weeks of Day 1 is acceptable. Responses are evaluated using RECIST vl.l, or the appropriate response evaluation for the corresponding malignancy, which may include a bone marrow biopsy. The same radiographic procedure used to define measurable lesions is used throughout the study for each subject. Response to disease (CR or PR) is confirmed by the same imaging technique no less than 4 weeks after the criteria for response are first met.
[00183] For subjects in Phase lb, a baseline CT/MRI of the head is performed during screening.
[00184] The End of Treatment (EOT) Visit is conducted within 14 to 28 days after the last dose of study drug is administered in a given living subject, regardless of the reason for discontinuation. Adverse events > Grade 2 ongoing at the EOT Visit are followed until the event resolves to < Grade 1, stabilizes, subjects start alternate therapy, returns to a status that is clinically acceptable in the judgment of Investigator, is lost to follow-up, or terminates with the subject death. After discontinuation of Compound A22, all surviving subjects are contacted by telephone once every 3 months thereafter for up to 2 years or until death, withdrawal of consent, loss to follow-up, or end of study. Subjects with continued response (i.e., beyond the 2-year survival follow-up period) are to be followed until this study is closed.
Example 5
A 59 year old white female with ER+, HER2-, GATA3 mutant (P409fs) breast cancer received 21 mg Compound A22 on a 3x/week schedule (MWF)
[00185] A 59 year old white female with ER+, HER2-, GAT A3 mutant (P409fs) breast cancer received 21 mg Compound A22 on a 3x/week schedule (Monday, Wednesday, Friday). Prior to receiving Compound A22, the patient was initially diagnosed with Stage 1 intraductal carcinoma of the left breast and underwent lumpectomy followed by radiation therapy for approximately 2 months. The patient was treated with tamoxifen therapy for 4 years but was discontinued due to uterine thickening/polyps and Grade 3 hot flashes. Two years after discontinuation of tamoxifen, the patient was diagnosed with recurrence of breast cancer in her lungs and pleural space. The patient was treated with two cycles of Ibrance/letrozole and 1 cycle of ribociclib but showed disease progression. The patient also received Enhertu for 3 cycles but developed shortness of breath associated with disease progression. Biopsy of a metastatic lesion in the right lung showed breast cancer origin and GATA3 mutation (P409fs) with an allele frequency of 43%.
[00186] The patient was treated with 21 mg Compound A22 on a 3x/week schedule (Monday-Wednesday-Friday). At baseline, the target lesion linear diameter measured 6.4 cm. A scan conducted after 2 cycles of Compound A22 treatment showed a reduction of target lesion linear diameter to 5.5 cm, a -14% reduction in the lesion.
[00187] The examples set forth above are provided to give those of ordinary skill in the art with a complete disclosure and description of how to make and use the claimed embodiments and are not intended to limit the scope of what is disclosed herein. Modifications that are obvious to persons of skill in the art are intended to be within the scope of the following claims. All publications, patents, and patent applications cited in this specification are incorporated herein by reference as if each such publication, patent or patent application were specifically and individually indicated to be incorporated herein by reference.

Claims

What is claimed is:
1. A method of treating a patient having with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22. or a pharmaceutically acceptable salt thereof.
2. The method of claim 1, wherein the patient has GATA3/7?/.
3. The method of any one of claims 1-2, wherein the patient has p53vi7.
4. The method of any one of claims 1-3, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene.
5. The method of any one of claims 1-4, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
6. The method of any one of claims 1-5, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
7. The method of any one of claims 1-6, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
8. The method of any one of claims 1 to 7, wherein the breast cancer is refractory.
9. The method of any one of claims 1 to 7, wherein the breast cancer is metastatic.
10. The method of any one of claims 1 to 7, wherein the breast cancer is drug-resistant.
11. The method of any one of claims 1 to 10, wherein the patient has failed a prior therapy.
12. The method of any one of claims 1 to 11, wherein the patient is a human.
13. The method of any one of claims 1 to 12, wherein the compound is formulated as a p- toluenesulfonate salt or a di- -toluenesulfonate salt.
14. The method of any one of claims 1 to 13, wherein the pharmaceutical composition is administered orally.
15. The method of any one of claims 1 to 14, wherein the pharmaceutical composition is administered as a tablet or capsule.
16. The method of any one of claims 1 to 15, wherein the therapeutically effective amount is about 1, about 2, about 3, about 5, about 8, about 10, about 11, about 14, about 15, about 17, about 20, about 21, about 25 mg, about 30 mg, or about 35 mg.
17. The method of any one of claims 1 to 16, wherein the pharmaceutical composition is administered in a cycle.
18. The method of any one of claims 1 to 17, wherein the pharmaceutical composition is administered in a 28-day cycle.
19. The method of any one of claims 1 to 18, wherein the pharmaceutical composition is administered for 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, or 7 days per week.
20. The method of any one of claims 1 to 19, wherein the pharmaceutical composition is administered for 3 days or 5 days per week.
21. The method of any one of claims 1 to 20, wherein the pharmaceutical composition is administered on three non-consecutive days in a week.
22. The method of any one of claims 1 21, wherein the pharmaceutical composition is administered in a 28-day cycle for 3 days per week.
23. The method of any one of claims 1 to 22, wherein the pharmaceutical composition is administered in a 28-day cycle for 3 non-consecutive days per week.
24. The method of any one of claims 1 to 20, wherein the pharmaceutical composition is administered in a 28-day cycle for 5 days per week.
25. The method of claim 24, wherein the pharmaceutical composition is administered in a 28-day cycle on Days 1, 2, 3, 4, and 5 per week.
26. The method of any one of claims 1 to 25, wherein the therapeutically effective amount is about 21 mg.
27. The method of any one of claims 1 to 25, wherein the therapeutically effective amount is about 30 mg.
28. The method of any one of claims 1 to 25, wherein the therapeutically effective amount is about 35 mg.
29. A method of inhibiting MDM2 in a patient having ER+, HER- metastatic breast cancer and having GATA3/7?/, comprising administering to the patient a pharmaceutical composition comprising a therapeutically effective amount of Compound A22,
A22, or a pharmaceutically acceptable salt thereof.
30. The method of claim 29, wherein the compound is formulated as a di- - toluenesulfonate salt.
31. The method of any one of claims 29 to 30, wherein the therapeutically effective amount is about 21 mg per day, about 30 mg per day, or about 35 mg.
32. The method of any one of claims 29 to 31, wherein the pharmaceutical composition is administered three times per week.
33. The method of any one of claims 29 to 32, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene.
34. The method of any one of claims 29 to 33, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
35. The method of any one of claims 29 to 34, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
36. The method of any one of claims 29 to 35, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
37. A method of inhibiting CDK7 in a patient having ER+, HER- metastatic breast cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
38. The method of claim 37, wherein the compound is formulated as a di- - toluenesulfonate salt.
39. The method of any one of claims 39 to 38, wherein the breast cancer has been resistant to a CDK4/6 inhibitor.
40. The method of any one of claims 39 to 38, wherein the patient has P53VI .
41. The method of any one of claims 39 to 40, wherein the patient has GATA3wt.
42. The method of any one of claims 39 to 40, wherein the patient has GATA3mt.
43. The method of any one of claims 39 to 42, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene.
44. The method of any one of claims 39 to 43, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
45. The method of any one of claims 39 to 44, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
46. The method of any one of claims 39 to 45, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
47. The method of any one of claims 37 to 46, wherein Compound A22 is administered three times per week.
48. The method of any one of claims 37 to 47, wherein the therapeutically effective amount is about 21 mg, about 30 mg, or about 35 mg per day.
49. A method of treating cancer in a patient having amplified expression of MDM2 or having a >4 copy number of the MDM2 gene, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
50. The method of claim 49, wherein the compound is formulated as a di- - toluenesulfonate salt.
51. The method of any one of claims 51 to 50, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
52. The method of any one of claims 51 to 51, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
53. The method of any one of claims 51 to 52, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
54. The method of any one of claims 49-53, wherein administering comprises administering about 21 mg of Compound A22 three times per week.
55. The method of any one of claims 49-53, wherein administering comprises administering about 30 mg of Compound A22 three times per week.
56. The method of any one of claims 49-53, wherein administering comprises administering about 35 mg of Compound A22 three times per week.
57. A method of inhibiting MDM2 expression in a patient having cancer, comprising administering to the patient a therapeutically effective amount of Compound A22, or a pharmaceutically acceptable salt thereof.
58. The method of claim 57, wherein the compound is formulated as a di- - toluenesulfonate salt.
59. The method of any one of claims 61 to 58, wherein the patient over-expresses MDM2 or has a >4 copy number of the MDM2 gene.
60. The method of any one of claims 61 to 59, wherein the patient over-expresses MDM2 or has a >6 copy number of the MDM2 gene.
61. The method of any one of claims 61 to 60, wherein the patient over-expresses MDM2 or has a >8 copy number of the MDM2 gene.
62. The method of any one of claims 61 to 61, wherein the patient over-expresses MDM2 or has a >10 copy number of the MDM2 gene.
63. The method of any one of claims 57 to 62, wherein administering comprises administering about 21 mg of Compound A22 three times per week.
64. The method of any one of claims 57 to 62, wherein administering comprises administering about 30 mg of Compound A22 three times per week.
65. The method of any one of claims 57 to 62, wherein administering comprises administering about 35 mg of Compound A22 three times per week.
66. A pharmaceutical composition comprising Compound A22, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier, for use in the treatment of a patient diagnosed with Estrogen Receptor positive (ER+) Human Epidermal Growth Factor Receptor 2 negative (HER2-) breast cancer.
67. The pharmaceutical composition for use according to claim 66, wherein the patient has GAT3/7?/.
68. The pharmaceutical composition for use according to any one of claims 66 to 67, wherein the treatment comprises administration of the pharmaceutical composition to the patient on three non-consecutive days in a week.
69. The pharmaceutical composition for use according to any one of claims 66 to 68, wherein the administration occurs for one or more cycles.
70. The pharmaceutical composition of claim 69, wherein the cycle is 28 days.
71. The pharmaceutical composition for use according to any one of claims 66-70, wherein the pharmaceutical composition is orally administered to the patient.
72. The pharmaceutical composition for use according to any one of claims 66-71, wherein the pharmaceutical composition is formulated as an oral unit dosage form.
73. The pharmaceutical composition for use according to claim 72, wherein the oral unit dosage form comprises about 21 mg of Compound A22.
74. The pharmaceutical composition for use according to claim 72, wherein the oral unit dosage form comprises about 30 mg of Compound A22.
75. The pharmaceutical composition for use according to claim 72, wherein the oral unit dosage form comprises about 35 mg of Compound A22.
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