Combination therapy for treating cancer

Abstract

This disclosure provides compounds of formula (I) and pharmaceutically acceptable salts thereof that inhibit phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3K) isoform alpha (PI3Kα), for use in combination with additional therapeutic agents for treating a condition, disease, or disorder (e.g., cancer) in which increased (e.g., excessive) PI3Kα activation contributes to the pathology and / or symptoms and / or progression of the condition, disease, or disorder in a subject.

Description

[Technical Field]

[0001] (Cross-reference of related applications) This application claims the benefits as of the filing dates of U.S. Provisional Patent Application No. 63 / 421,082, filed October 31, 2022; U.S. Provisional Patent Application No. 63 / 423,383, filed November 7, 2022; U.S. Provisional Patent Application No. 63 / 488,674, filed March 6, 2023; and U.S. Provisional Patent Application No. 63 / 531,990, filed August 10, 2023. The contents of each application are incorporated herein by reference in their entirety.

[0002] (Sequence Listing) This application includes a sequence listing submitted electronically as an XML file named 50006-0099WO1_ST26_SL.XML. The XML file, created on October 30, 2023, is 2,946 bytes in size. The material within the XML file is incorporated herein by reference in its entirety.

[0003] This disclosure provides compounds of formula (I) and pharmaceutically acceptable salts thereof that inhibit phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3K) isoform alpha (PI3Kα), for use in combination with additional therapeutic agents for treating a condition, disease, or disorder (e.g., cancer) in which increased (e.g., excessive) PI3Kα activation contributes to the pathology and / or symptoms and / or progression of the condition, disease, or disorder in a subject. [Background technology]

[0004] The phosphatidylinositol 4,5-bisphosphate 3-kinase (PI3K) isoform alpha (PI3Kα), encoded by the PIK3CA gene, is part of the PI3K / AKT / TOR signaling network and is altered in several human cancers. Several researchers have demonstrated that the role of PI3K / AKT signaling is involved in physiological and pathophysiological functions that drive tumor progression, including metabolism, cell growth, proliferation, angiogenesis, and metastasis. See Fruman, DACell 2017, 170, 605-635 and Janku, F. et al., Nat. Rev. Clin. Oncol. 2018, 15, 273-291. Pharmacological or genetic suppression of PI3K / AKT / TOR signaling can induce cancer cell death and regression of tumor growth.

[0005] The PI3K pathway can be activated, for example, through point mutations in the PIK3CA gene or through inactivation of phosphatase and tensin homolog (PTEN) genes. Activation of this pathway occurs in approximately 30–50% of human cancers and contributes to resistance to various anticancer therapies. See Martini, M. et al., Ann. Med. 2014, 46, 372–383 and Bauer, TM et al., Pharmacol. Ther. 2015, 146, 53–60. [Overview of the project]

[0006] A method for treating cancer in a subject requiring treatment for cancer is provided herein, comprising administering to the subject (a) compound 1 or a pharmaceutically acceptable salt thereof, and (b) one or more additional therapeutic agents.

[0007] A method for treating cancer in a subject requiring treatment for cancer is provided herein, comprising administering to the subject (a) compound 1 or a pharmaceutically acceptable salt thereof, and (b) one or more independently selected additional therapeutic agents selected from the group consisting of selective estrogen receptor modulators (SERMs) / selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, and RAF inhibitors, PIM (Provirus Integration site for Moloney leukemia virus kinase, e.g., PIM1, PIM2, and PIM3) inhibitors, or any combination thereof.

[0008] Furthermore, pharmaceutical compositions comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof, one or more pharmaceutically acceptable excipients, and one or two independently selected additional therapeutic agents are also provided herein.

[0009] The disclosure also provides a method for inhibiting PI3Kα in mammalian cells, comprising contacting mammalian cells with a compound of formula (I) or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents. In some embodiments, the one or more additional therapeutic agents are selective estrogen receptor modulators (SERMs) / selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, RAF inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, or any combination thereof.

[0010] Other embodiments include those described in the detailed description and / or claims.

[0011] Additional definitions To facilitate understanding of the disclosures contained herein, several additional terms are defined below. In general, the nomenclature used herein, as well as the experimental procedures in organic chemistry, medicinal chemistry, and pharmacology described herein, are well known and commonly used in the art. Unless otherwise defined, all technical and scientific terms used herein generally have the same meaning as commonly understood by those skilled in the art to which this disclosure belongs. Each of the patents, applications, published applications, and other publications referenced throughout this specification and its appendices is incorporated herein by reference in their entirety.

[0012] When referring to a number or numerical range, the term "approximately" means that the number or numerical range referred to is an approximation within, for example, experimental variability and / or statistical experimental error, and therefore the number or numerical range may vary by up to ±10% of the stated number or numerical range.

[0013] When used herein, the term “acceptable” with respect to a formulation, composition, or component means that it does not have any lasting adverse effects on the overall health of the subject being treated.

[0014] The term "therapeutic dose" means an amount of a compound that, when administered to a subject in need of such treatment, is sufficient to (i) treat a PI3Kα protein-related disease or disorder, (ii) reduce, improve or eliminate one or more symptoms of a particular disease, condition, or disorder, or (iii) delay the onset of one or more symptoms of a particular disease, condition, or disorder as described herein. When used in relation to treatment with two or more therapeutic agents, each agent may be administered independently in a therapeutic dose (e.g., an amount that may be therapeutically effective as monotherapy), or one or more therapeutic agents may together constitute a therapeutic dose for treating the indicated disease or disorder (e.g., a therapeutic dose of combination therapy). In other words, the amount of each component in a therapeutic dose of combination therapy may be administered (in combination) at a level less than the therapeutic dose when administered independently as monotherapy.

[0015] The term "pharmaceutically acceptable excipient" means a pharmaceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, carrier, solvent, or encapsulating material. In one embodiment, each component is "pharmaceutically acceptable" in the sense that it is compatible with other components of a pharmaceutical formulation, suitable for use in contact with human and animal tissues or organs without excessive toxicity, irritation, allergic reactions, immunogenicity, or other problems or complications, and is commensurate with a reasonable benefit / risk ratio. For example, Remington: The Science and Practice of Pharmacy, 21st ed.; Lippincott Williams & Wilkins: Philadelphia, PA, 2005, Handbook of Pharmaceutical Excipients, 6th ed.; Rowe et al., Eds.; The Pharmaceutical Press and American Pharmaceutical Association: 2009, 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.

[0016] The term "pharmaceutically acceptable salt" refers to a formulation of a compound that does not cause significant irritation to the organism to which it is administered and does not inhibit the biological activity and properties of the compound. In certain cases, pharmaceutically acceptable salts are obtained by reacting the compounds described herein with acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. In some cases, pharmaceutically acceptable salts are obtained by reacting the acidic compounds described herein with a base to form salts such as ammonium salts, alkali metal salts such as sodium or potassium salts, alkaline earth metal salts such as calcium or magnesium salts, salts of organic bases such as dicyclohexylamine, N-methyl-D-glucamine, and tris(hydroxymethyl)methylamine, and salts with amino acids such as arginine and lysine, or by other methods previously determined. Pharmacologically acceptable salts are not particularly limited as long as they can be used pharmacopoeia. Examples of salts formed by the compounds described herein with bases include: salts having inorganic bases such as sodium, potassium, magnesium, calcium, and aluminum; salts having organic bases such as methylamine, ethylamine, and ethanolamine; salts having basic amino acids such as lysine and ornithine; and ammonium salts. The salts may also be acid addition salts, which specifically include: mineral acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, nitric acid, and phosphoric acid; organic acids such as formic acid, acetic acid, propionic acid, oxalic acid, malonic acid, succinic acid, fumaric acid, maleic acid, lactic acid, malic acid, tartaric acid, citric acid, methanesulfonic acid, and ethanesulfonic acid; and acidic amino acids such as aspartic acid and glutamic acid.

[0017] As used herein, “Subject” refers to any animal, including primates (e.g., humans), mice, rats, other rodents, rabbits, dogs, cats, pigs, cattle, sheep, horses, primates, and mammals such as humans. In some embodiments, the subject is human. In some embodiments, the subject is experiencing and / or exhibiting at least one symptom of the cancer being treated.

[0018] The “treatment” or “therapy” of a subject means any type of intervention or process performed on the subject, or administration of an activator to the subject, for the purpose of reversing, mitigating, improving, inhibiting, slowing, or preventing the onset, progression, development, severity, or recurrence of symptoms, complications, conditions, or biochemical signs associated with the disease. In some embodiments, the disease is cancer. As used herein, the terms “treatment” and “treating” are not intended to be absolute terms, for example, when referring to the treatment of cancer. For example, “treatment of cancer” and “treating cancer,” when used in a clinical setting, are intended to include obtaining a beneficial or desired clinical outcome and may include improvement of the condition of a subject having cancer. Beneficial or desired clinical outcomes include, but are not limited to, one or more of the following: reduced proliferation (or destruction) of neoplastic or cancer cells; inhibition of metastasis of neoplastic cells; reduced metastasis in the subject; reduction or decrease in tumor size; alteration of the growth rate of one or more tumors in the subject; increased duration of remission (partial or complete) for the subject (e.g., compared to one or more metrics in a subject with a similar cancer that is untreated or treated differently, or compared to one or more metrics in the same subject before treatment); reduced symptoms resulting from the disease; improved quality of life for the person with the disease (e.g., assessed using FACT-G or EORTC-QLQC30); reduced doses of other medications required to treat the disease; delayed disease progression; and / or extended survival in the subject with the disease. "Treatment" may also mean an extension of survival compared to the survival expected without treatment.

[0019] The term "metastasis" is a well-known term in the art, referring to the spread of cancer cells from the site where they were first formed (primary site) to one or more other sites in the body (one or more secondary sites). In metastasis, cancer cells detach from the original (primary) tumor and travel through the blood or lymphatic system to form a new tumor (metastatic tumor) in another organ or tissue of the body. The new metastatic tumor contains cancer cells that are the same as or similar to those in the primary tumor. At the secondary site, tumor cells proliferate and may initiate the growth or colonization of a secondary tumor at this distal site.

[0020] As used herein, the term “metastatic cancer” (also known as “secondary cancer”) refers to a type of cancer that originates from one histological type but subsequently spreads to one or more tissues other than the (primary) cancer of origin. Metastatic brain cancer refers to cancer within the brain, i.e., cancer that originates from tissues other than the brain and has metastasized to the brain. In some embodiments described herein, the subjects have metastatic brain cancer and / or metastatic spinal cancer.

[0021] The term "tumor growth inhibition (TGI) index" refers to a value used to express the degree to which a drug (e.g., Compound 1 or a pharmaceutically acceptable salt thereof, either alone or in combination with one or more additional therapeutic agents described herein) inhibits tumor growth compared to an untreated control or a control treated with monotherapy as part of a combination therapy. The TGI index is calculated for a specific point in time (e.g., a specific number of days in an experiment or clinical trial) according to the following formula:

[0022]

number

[0023] The compounds provided herein may encompass a variety of stereochemical forms. The compounds also include enantiomers (e.g., R and S isomers), diastereomers, and mixtures of enantiomers (e.g., R and S isomers), including racemic mixtures and mixtures of diastereomers, as well as individual enantiomers and diastereomers resulting from structural asymmetry in particular compounds. Unless otherwise indicated, if a disclosed compound is named or described by its structure (e.g., a "flat" structure) without specifying its stereochemistry and has one or more chiral centers, it is understood to represent all possible stereoisomers of the compound. Similarly, unless otherwise indicated, if a disclosed compound is named or described by a structure that specifies its stereochemistry (e.g., a structure with "wedge" and / or "dashed" bonds) and has one or more chiral centers, it is understood to represent the indicated stereoisomer of the compound.

[0024] The compounds of formula (I) or their pharmaceutically acceptable salts are as follows:

[0025] [ka] or a pharmaceutically acceptable salt thereof, in the formula, Z is O or NR x And, Rx is hydrogen, C1-C6 alkyl, or C3-C6 cycloalkyl, each R 1 is independently selected from halogen, hydroxyl, cyano, C1-C6 alkyl optionally substituted with hydroxyl, and C3-C6 cycloalkyl, m is 0, 1, 2, or 3, R 2 is halogen, hydroxyl, C1-C6 alkyl optionally substituted with hydroxyl, C1-C6 haloalkyl, C3-C6 cycloalkyl optionally substituted with 1 or 2 fluorines, R 3 is C1-C6 alkyl, C1-C6 haloalkyl, or C3-C6 cycloalkyl substituted with 1 or 2 substituents independently selected from fluorine and C1-C6 alkyl, Ring A is 6-10 member aryl, C3-C8 cycloalkyl, 5-10 member heteroaryl, or 4-10 member heterocyclyl, each R 4 is independently (i) halogen, (ii) C1-C6 alkyl optionally substituted with 1 or 2 hydroxyls or -NR A R B ; (iii) C1-C6 alkoxy optionally substituted with 1-2 substituents independently selected from hydroxyl and C3-C6 cycloalkyl, (iv) C1-C6 haloalkyl, (v) hydroxyl, (vi) cyano, (vii) -CO2H, (viii) -NR A R B , (ix) =NR A2 , (x) -C(=O)NR C R D , (xi) -SO2(NR E R F ), (xii)-SO2(C1-C6 alkyl), (xiii)-S(=O)(=NH)(C1-C6 alkyl), (xiv)-C(=O)(C1-C6 alkyl), (xv)-CO2(C1-C6 alkyl), (xvi) 5-6 member heteroaryls optionally substituted with C1-C6 alkyl groups, (xvii) One or two independently selected R G A 3- to 9-membered heterocycline that is substituted by, and (xviii) One or two independently selected R G Selected from the group consisting of 3- to 6-membered cycloalkyl groups that are substituted with, n is 0, 1, or 2. Each R A , R A1 , R B , R B1 , R C , R C1 , R D , R D1 , R E , and R F Independently, (i) Hydrogen, (ii) Hydroxyl, (iii) 4-6 member heterocycline, (iv) C1-C6 haloalkyl, (v)-C(=O)(C1-C6 alkyl), (vi)-C(=O)O(C1-C6 alkyl), (vii)-SO2(C1-C6 alkyl), (viii) 3-6 member cycloalkyls optionally substituted with hydroxyl, (ix) Optionally hydroxyl, -C(=O)NR B2 R C2 , a C1-C6 alkyl substituted with one or two substituents independently selected from 5-6 member heteroaryl, 3-6 member cycloalkyl, -SO2(C1-C6 alkyl), -CO2H, and -SO2(NH2), or R C and RD These, together with the nitrogen atom to which they are bonded, can selectively form hydroxyl, halogen, and -C(=O)NR B1 R C1 , -SO2(C1-C6 alkyl), -CO2H, C1-C6 alkyl, C1-C6 alkoxy, and C1-C6 haloalkoxy are optionally substituted with hydroxyl molecules to form 4-10 membered heterocyclines substituted with 1-2 substituents independently selected from these groups. Each R A2 , R B2 , and R C2 These are independently hydrogen or a C1-C6 alkyl group. Each R G These are independently substituted with fluoro, cyano, hydroxyl, optionally hydroxyl-substituted C1-C6 alkyl, C1-C6 alkoxy, and -NR. A1 R B1 ,=NR A2 -C(=O)NR C1 R D1 This refers to a compound of (I) or a pharmaceutically acceptable salt thereof, selected from the group consisting of -CO2(C1-C6 alkyl), C1-C6 haloalkyl, C3-C6 cycloalkyl, C1-C6 haloalkoxy, -SO2(C1-C6 alkyl), and -CO2H.

[0026] "Compound 1" or a pharmaceutically acceptable salt thereof refers to (R)-1-(2-aminopyrimidine-5-yl)-3-(1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethyl)urea or a pharmaceutically acceptable salt thereof having the following structure.

[0027] [ka] Or it refers to a pharmaceutically acceptable salt thereof.

[0028] Details of one or more embodiments of this disclosure are described in the accompanying drawings and the following description. Other features and advantages of this disclosure will be apparent from the description and drawings, as well as from the claims. [Brief explanation of the drawing]

[0029] [Figure 1A] The isobolograms of fulvestrant versus compound 1 are shown. [Figure 1B] The isobologram of lapatinib versus compound 1 is shown. [Figure 1C] The isobologram of abemaciclib versus compound 1 is shown. [Figure 1D] This is an overview of potential inhibitors of the RAS pathway and MEK1 / 2 pathway genes. [Figure 1E] The results of the Profiling Relative Inhibition Simultaneously in Mixture (PRISM) screening for drug profiling against wild-type MAPK and mutant MAPK are shown. [Figure 1F] The isobologram of trametinib versus compound 1 is shown. [Figure 1G] This shows inhibition of MSI-H colon cancer cell proliferation upon treatment with compound 1 and / or binimetinib. [Figure 2A] This study demonstrates that compound 1 reduces tumor volume in the PDX model, and that this reduction is more pronounced when compound 1 is administered in combination with fulvestrant. PDX model 1 measures PIK3CAH1047R, ER+ tumor volume, with circles representing the vehicle, diamonds representing fulvestrant 5 mg SC QW, triangles representing compound 1 at 100 mg / kg PO QD, and squares representing compound 1 + fulvestrant at 100 mg / kg. [Figure 2B] The tumor volume before and after treatment with compound 1 + fulvestrant at a dose of 100 mg / kg is shown. [Figure 2C] The tumor volume before and after treatment with compound 1 at 100 mg / kg PO QD is shown. [Figure 3A]This study demonstrates that compound 1 reduces tumor volume in the PDX model, and that this reduction is more pronounced when compound 1 is administered in combination with palbociclib. PDX model 2 measures PIK3CAH1047R / R108H, ER+ / HER2+ tumor volume, with circles representing the vehicle, triangles representing compound 1 at 100 mg / kg PO QD, green circles representing palbociclib PO QD, and squares representing compound 1 at 100 mg / kg + palbociclib at 50 mg / kg. [Figure 3B] This shows that compound 1 reduces tumor volume in the PDX model, and that the reduction in tumor volume is more pronounced when compound 1 is administered in combination with palbociclib. Figure 3B shows the volume changes of PDX model 3: PIK3CAH1047R, ER+ / HER2+ tumor, where circles represent vehicles, triangles represent compound 1 at 100 mg / kg PO QD, green circles represent palbociclib PO QD, and squares represent compound 1 at 100 mg / kg + palbociclib at 50 mg / kg. [Figure 4A] This shows combination therapy with compound 1, fulvestrant and / or palbociclib, in an ER+ breast cancer model. Figure 4A shows established T47D xenograft tumors in female NSG mice. Mice were randomized and treated as shown when the tumors reached approximately 200 mm3. Data represent the percentage change in tumor volume of individual tumors at day 20 compared to randomization. [Figure 4B]This shows combination therapy of compound 1 with fulvestrant and / or palbociclib in an ER+ breast cancer model. Figures 4B and 4C show that NSG mice harboring T47D tumors were administered a single dose of the vehicle or compound as shown. Tumors were collected 4 hours after treatment with compound 1, and (4B) Western blots of pAKT(S473) and S6(S240 / 244), and (4C) 24 hours after treatment with fulvestrant for IHC of AKT and S6. The pS6 and pAKT(S473) groups were blotted and analyzed together. All samples for a given protein were performed on the same gel. Vinculin analysis is shown. For clarity, images were cropped to remove other compounds. 20X IHC images are shown with a 200 μm scale bar. [Figure 4C] This shows combination therapy of compound 1 with fulvestrant and / or palbociclib in an ER+ breast cancer model. Figures 4B and 4C show that NSG mice harboring T47D tumors were administered a single dose of the vehicle or compound as shown. Tumors were collected 4 hours after treatment with compound 1, and (4B) Western blots of pAKT(S473) and S6(S240 / 244), and (4C) 24 hours after treatment with fulvestrant for IHC of AKT and S6. The pS6 and pAKT(S473) groups were blotted and analyzed together. All samples for a given protein were performed on the same gel. Vinculin analysis is shown. For clarity, images were cropped to remove other compounds. 20X IHC images are shown with a 200 μm scale bar. [Figure 4D] This shows combination therapy with compound 1, fulvestrant and / or palbociclib, in an ER+ breast cancer model. Figure 4D shows established ST1056 xenograft tumors in BALB / c nude mice. Mice were randomized to treatment groups as shown, approximately 260 mm3. Mice were treated for 94 days or until they were excluded from the study due to tumor volume. After 94 days, treatment was discontinued and tumors were monitored for regrowth. [Figure 4E]This shows combination therapy with compound 1, fulvestrant and / or palbociclib, in an ER+ breast cancer model. Figure 4E shows the percentage change in body weight in Figure 4D. [Figure 4F] This shows combination therapy with compound 1, fulvestrant and / or palbociclib, in an ER+ breast cancer model. Figures 4F and 4G show the percentage changes in body weight (upper) and tumor volume measurements (lower) over time from the T47D efficacy shown in Figure 4A (N=9). [Figure 4G] This shows combination therapy with compound 1, fulvestrant and / or palbociclib, in an ER+ breast cancer model. Figures 4F and 4G show the percentage changes in body weight (upper) and tumor volume measurements (lower) over time from the T47D efficacy shown in Figure 4A (N=9). [Figure 5A] Alpericib and compound 1 inhibit the enzymatic activity of WT PI3Kα and kinase domain and helical main mutant proteins, and the geometric mean and standard deviation are shown. [Figure 5B] The table below the sensorgrams shows surface plasmon resonance (SPR) sensorgrams for compound 1 (upper graph) and duvarisive (lower graph) that bind to WT, H1047R, and E545K mutant proteins. [Figure 5C] The 2.9 Å X-ray structure of PI3Kα (p110=purple, p85=orange) is shown, with GDC-0077 (red sphere) bound to the ATP binding site and compound 1 (green sphere) bound to the allosteric site. [Figure 5D] Arrows indicate Cα (sphere) movement of ≥3 Å between the aligned H1047R (teal=PDB 3HHm) and the compound 1-bond structure (purple). 3HHM lacks residues 941-952 for direct comparison. [Figure 5E] Detailed diagrams of the bound compound 1 and the label residues that significantly contribute to compound binding are shown. [Figure 5F] The molecular structure of compound 1 is shown. [Figure 5G]This demonstrates that compound 1 possesses broad quinome selectivity. The upper panel is a table summarizing the quinome profiling of compound 1 compared to publicly available alpericib data. AurB kinase was the only off-target with an IC50 < 10 mM (approximately 1658 nM). The lower panel shows dose-dependent inhibition of phospho-H3 (Ser10) observed 1 hour after treatment with the AurB kinase inhibitor, valacertib. No significant inhibition of AurB was observed with treatment with compound 1. [Figure 6A] This shows the PI3Kα-muta selectivity profiling of compound 1 in cell assays. Figure 6A shows correlation plots of alpericib and compound 1 comparing pAKT IC50 at 1 hour and viability (CTGlo)GI50 at 72 hours across a panel of cell lines. [Figure 6B] Figure 6B shows the PI3Kα-muta selectivity profiling of compound 1 in a cell assay. Figure 6B shows the pAKT inhibition dose-response curve (HTRF assay). [Figure 6C] This shows the PI3Kα-mutatric selectivity profiling of compound 1 in cell assays. Figure 6C shows correlation plots comparing the potency of alpelisib and compound 1 (pAKT HTRF assay) in panels of kinase domain mutant cell lines (orange dots) and WT PI3Kα SKBR3 (black dots). [Figure 6D] This shows the PI3Kα-mutatric selectivity profiling of compound 1 in cell assays. Figure 6D shows the AUC of compound 1 and alpericib from the Broad Institute's PRISM screening (theprismlab.org) across CCLE, grouped by PIK3CA mutation status. [Figure 6E] This shows the PI3Kα-mutant selectivity profiling of compound 1 in a cell assay. Figure 6E shows the 2-hour glucose uptake in primary human adipocytes (percent vehicle response), shown as a bar graph. For comparison, 72-hour survival data for the H1047R-PI3Kα-mutant T47D cell line are superimposed on the orange dose-response curve. [Figure 6F]This shows the PI3Kα-muta selectivity profiling of compound 1 in a cell assay. [Figure 7A] The effects of compound 1 and alpelisib on glucose homeostasis are shown. Figure 7A shows that tumor-naive female BALB / c mice were administered the compounds for 5 days as shown (n=5 / group) and subjected to an insulin tolerance test (ITT). On day 5, the animals were fasted for 6 hours and administered the compounds as shown 1 hour before the end of fasting (-1 hour). At T=0, the animals were administered 0.75 U / kg of intraperitoneal insulin. Blood glucose levels were monitored over time, and the group mean and standard error mean are shown. [Figure 7B] The effects of compound 1 and alperisib on glucose homeostasis are shown. Figure 7B shows the AUC calculated from Figure 7A. The AUC of each treatment group was compared with the vehicle group using standard one-way ANOVA and Dunnett's multiple comparison test. [Figure 7C] The effects of compound 1 and alpelisib on glucose homeostasis are shown. Figure 7C shows an oral glucose tolerance test conducted in the same manner as the ITT in Figure 7A, except that at T=0, mice were administered 2 g / kg glucose, blood glucose levels were monitored over time, and the group mean and standard error mean are shown. [Figure 7D] The effects of compound 1 and alperisib on glucose homeostasis are shown. Figure 7D shows the AUC calculated from the data shown in Figure 7C (calculated as analyzed in Figure 7B). [Figure 8A] The efficacy and pharmacodynamic profiling of compound 1-alperisib in CAL33 xenograft tumors are shown. Figures 8A and 8B show that Cal33 xenograft tumors were established in female BALB / c nude mice. Animals were randomized to treatment groups (n=6) in approximately 160 mm3 and treated as shown. Tumors (8A) and BW (8B) were measured twice weekly, and group mean and standard error mean are shown. [Figure 8B]The efficacy and pharmacodynamic profiling of compound 1-alperisib in CAL33 xenograft tumors are shown. Figures 8A and 8B show that Cal33 xenograft tumors were established in female BALB / c nude mice. Animals were randomized to treatment groups (n=6) in approximately 160 mm3 and treated as shown. Tumors (8A) and BW (8B) were measured twice weekly, and group mean and standard error mean are shown. [Figure 8C] The efficacy and pharmacodynamic profiling of compound 1-pair alpelisib in CAL33 xenograft tumors are shown. Figures 8C–8D show that mice carrying Cal33 tumors were treated for 3 days as shown, and serum insulin (8C) and blood glucose levels (8D) were measured using standard one-way ANOVA and Dadanet's multiple comparison tests, with the group mean and standard deviation mean shown for each treatment group compared to the vehicle group. [Figure 8D] The efficacy and pharmacodynamic profiling of compound 1-pair alpelisib in CAL33 xenograft tumors are shown. Figures 8C–8D show that mice carrying Cal33 tumors were treated for 3 days as shown, and serum insulin (8C) and blood glucose levels (8D) were measured using standard one-way ANOVA and Dadanet's multiple comparison tests, with the group mean and standard deviation mean shown for each treatment group compared to the vehicle group. [Figure 8E] The efficacy and pharmacodynamic profiling of compound 1-pair alpelisib in CAL33 xenograft tumors are shown. Figure 8E shows pAKT(S473) measured by Western blotting in all Cal33 tumors from mice in a 28-day efficacy study (30 and 100 mg dose groups) and a 3-day PK / PD study (30, 100, and 300 mg / kg dose groups). pAKT(S473) and total AKT were measured by Western blotting in Cal33 tumors. Normalized pAKT(S473) levels are plotted against unbound concentration (blue). In vitro pAKT dose-response in Cal33 is included for reference. [Figure 8F]This shows the efficacy and pharmacodynamic profiling of compound 1 versus alpelisib in CAL33 xenograft tumors. Figure 8F shows the pAKT levels in Cal33 tumors showing compound 1 (100 mg / kg) and alpelisib (50 mg / kg) alone over time. [Figure 8G] The efficacy and pharmacodynamic profiling of compound 1-alpelisib in CAL33 xenograft tumors are shown. Figure 8G shows pAKT(S473) measured by Western blotting from gastrocnemius muscle of mice from Figures 8E-8F. Significance was measured using standard one-way ANOVA and Dunnett's multiple comparison test. [Figure 8H] The efficacy and pharmacodynamic profiling of compound 1 versus alpelisib in CAL33 xenograft tumors are shown. Figure 8H shows N=4 mice fasted for 4 hours, then administered either vehicle, compound 1 (100 mg / kg), or alpelisib (50 mg / kg). One hour later, mice were orally administered [U-13C]-glucose. Thirty minutes later, tissues were collected and analyzed by MS. The abundance of labeled [M+4]succinate (S), fumarate (F), and malate (M) in the tumors and gastrocnemius muscle is shown. Individual values, group mean, and standard deviation mean are shown along with each treatment group compared to the vehicle group using two-way ANOVA and the Holm-Sidak post-hoc test. [Figure 8I] This report presents the efficacy and pharmacodynamic profiling of compound 1-alpelisib in CAL33 xenograft tumors. Figure 8I shows plasma insulin measurements immediately before and 30 minutes after administration of labeled glucose. [Figure 8J] The efficacy and pharmacodynamic profiling of compound 1-alpelisib in CAL33 xenograft tumors are shown. Figure 8J shows measurements from plasma of mice shown in Figure 8H. The isotopologic state represents the count of labeled U-13C carbons on glucose. Animals from the 0-hour group were not administered glucose. Samples were collected at the time of glucose administration to mice collected 30 minutes after [U-13C]-glucose. Bars represent standard deviation (SD). [Figure 9]This report shows that mice carrying Cal33 tumors were treated for 3 days as shown, and serum insulin was measured using the individual values, group mean, and standard deviation mean for each treatment group compared to the vehicle group using standard one-way ANOVA and Dadanet's multiple comparison tests. [Figure 10] p-AKT and AKT WB analysis in the efficacy group. Tumor samples were analyzed for total AKT and p-AKT by Western blotting. For all treatment groups analyzed, corresponding vehicle control samples were processed in parallel and performed on the same gel and WB to minimize variability. The p-AKT:AKT ratio for the time-matched vehicle control was defined as 100%, and all treated samples were represented against it, shown in the corresponding scatter plots with the indicated mean and SEM. [Figure 11A] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11A–11C show tumor volume over time from PDX models (N=3 / group) carrying PI3Kα mutations in the kinase domain (ST1056-H1047R; 11C), kinase and helical domain (ST1799-E542K / H1065L; 11B), and helical domain (ST2652-E545K; 11A) (treated with either vehicle, compound 1, or alpelisib). Statistical significance was calculated using two-way ANOVA and Dunnett's multiple comparison test. At the end of the study, tumors were collected 4 hours after the final dose, and pAKT(S473) was analyzed by Western blotting. To determine that the E542K / H1065L PI3Kα mutation is cis by IsoSeq, one of three tumors from the model ST1799 was used. AKT and pAKT(S473) were blotted separately, and vinculin analysis is shown. [Figure 11B]This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11A–11C show tumor volume over time from PDX models (N=3 / group) carrying PI3Kα mutations in the kinase domain (ST1056-H1047R; 11C), kinase and helical domain (ST1799-E542K / H1065L; 11B), and helical domain (ST2652-E545K; 11A) (treated with either vehicle, compound 1, or alpelisib). Statistical significance was calculated using two-way ANOVA and Dunnett's multiple comparison test. At the end of the study, tumors were collected 4 hours after the final dose, and pAKT(S473) was analyzed by Western blotting. To determine that the E542K / H1065L PI3Kα mutation is cis by IsoSeq, one of three tumors from the model ST1799 was used. AKT and pAKT(S473) were blotted separately, and vinculin analysis is shown. [Figure 11C] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11A–11C show tumor volume over time from PDX models (N=3 / group) carrying PI3Kα mutations in the kinase domain (ST1056-H1047R; 11C), kinase and helical domain (ST1799-E542K / H1065L; 11B), and helical domain (ST2652-E545K; 11A) (treated with either vehicle, compound 1, or alpelisib). Statistical significance was calculated using two-way ANOVA and Dunnett's multiple comparison test. At the end of the study, tumors were collected 4 hours after the final dose, and pAKT(S473) was analyzed by Western blotting. To determine that the E542K / H1065L PI3Kα mutation is cis by IsoSeq, one of three tumors from the model ST1799 was used. AKT and pAKT(S473) were blotted separately, and vinculin analysis is shown. [Figure 11D]This shows that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figure 11D shows N=9 for the NCI-H1048, Detroit562, and GP2D models, and n=6 for the HCC1954 model. Figures 11A-11C show the percentage change in body weight in the PDX efficacy trial. N=3 for each model. Bars represent SEM. [Figure 11E] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figure 11E shows the final tumor volume percentage change for GP2D, Detroit562, NCI-H1048, and HCC 1954 CDX tumors. [Figure 11F] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11F–11I show tumor volume measurements over time (left) and percentage change in body weight (right) from the CDX efficacy study in Figure 11E. [Figure 11G] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11F–11I show tumor volume measurements over time (left) and percentage change in body weight (right) from the CDX efficacy study in Figure 11E. [Figure 11H] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11F–11I show tumor volume measurements over time (left) and percentage change in body weight (right) from the CDX efficacy study in Figure 11E. [Figure 11I] This demonstrates that a metabolically reduced dose of compound 1 is as effective as high-dose alpelisib across PI3Kα-mutated tumor xenografts. Figures 11F–11I show tumor volume measurements over time (left) and percentage change in body weight (right) from the CDX efficacy study in Figure 11E. [Figure 12]This shows the relationship between exposure to compound 1 versus time after a single oral administration of 30, 100, or 300 mg / kg of compound 1 in CD1 mice. Total exposure and IC80 efficacy are corrected for both mouse PPB (fu=0.033) and assay medium-bound (10% FBS fu=0.26), respectively. [Modes for carrying out the invention]

[0030] This disclosure provides compounds of formula (I) that inhibit PI3Kα and pharmaceutically acceptable salts thereof, for use in combination with additional therapeutic agents for treating a condition, disease, or disorder (e.g., cancer) in which increased PI3Kα activation contributes to the pathology and / or symptoms and / or progression of the condition, disease, or disorder in a subject.

[0031] As described herein, selective targeting of mutant PI3Kα by compound 1 lacked the systemic metabolic dysfunction caused by alpelisib. Compound 1 conserved glucose uptake in human adipocytes and did not induce systemic insulin resistance in vivo, as demonstrated by insulin resistance tests, which include the absence of insulin spikes after chronic compound 1 administration in six independent CDX and OGTTs, compared to alpelisib. Selective targeting of mutant PI3Kα was further demonstrated by the differential effect of compound 1 on pAKT / AKT suppression in tumors and muscles, and by evaluating glucose oxidation in these tissues in isotopic-labeled glucose OGTTs. These data demonstrate a superior metabolic safety profile of compound 1 compared to alpelisib, which may enhance efficacy as a result of blunting the counterregulatory insulin spikes that reduce efficacy in preclinical studies (Hopkins et al. Nature 2018;560(7719):499-503).

[0032] Compound 1 monotherapy was tested in one case each for colon cancer and lung cancer in a panel of 10 CDX and PDX cancers, primarily breast cancer and HNSCC cancer. This panel represented the dominant PI3Kα mutations in cancer types where there is a high unmet need for improved treatment options. Compound 1 100 mg / kg QD demonstrated robust efficacy comparable to or better than high-dose alpelisib in xenograft models, with mouse alpelisib exposure being approximately twice as high as patient exposure (AUC at 50 mg dose). 24時間 (Based on [the above]). Compound 1 was equally effective in GP2D mutant colorectal cancer xenografts harboring the H1047L variant, the second most dominant kinase domain mutation after H1047R. Importantly, the treatment was highly effective in ST2652 and ST1799 PDX tumors harboring the E545K and E542K mutations, which are the second and third most common hotspots, respectively. Finally, both NCIH1048 and ST1799 tumors harbor a secondary PI3Kα mutation, which occurs in approximately 10% of primary tumor samples, and both tumors responded favorably to treatment with Compound 1.

[0033] CDK4 / 6 inhibitors and anti-estrogen therapy are used in ER. + It is an important standard of treatment for breast cancer. The clear superiority of alpelisib and fulvestrant combination therapy over fulvestrant monotherapy highlights the importance of testing PI3Kα inhibitors in combination in preclinical models. Benchmark T47D ER + HER2 -In the BrCa CDX model, fulvestrant monotherapy provided a moderate level of tumor growth control, while low-dose compound 1 monotherapy and high-dose alpericib resulted in tumor quiescence. The combination of fulvestrant and low dose was superior to low-dose compound 1 monotherapy and resulted in regression in the majority of animals. Higher doses of compound 1 resulted in deep tumor regression, with or without fulvestrant combination therapy. All combinations, including monotherapy of palbociclib, fulvestrant, and compound 1, as well as triple therapy in the invasive BrCa PDX model (ST1056), were tested. Compound 1 monotherapy provided a much more robust and sustained response in this model than fulvestrant or palbociclib monotherapy or any combination thereof. However, the combination of fulvestrant and compound 1 provided exceptional tumor growth control, and regression persisted in all animals over 90 days of treatment and was preserved for several weeks after discontinuation of medication until the end of the study. Palbociclib showed limited efficacy as monotherapy or in combination with compound 1 in this PDX model, while triple therapy with compound 1 and fulvestrant was well tolerated in mice over 90 days of treatment. In contrast, triple therapy with alpelisib, ribociclib, and fulvestrant resulted in increased hepatobiliary toxicity and a higher incidence and severity of rash, and was untolerable in patients (Tolaney SM, et al. Clin Cancer Res 2021;27(2):418-28). Notably, the toxicity from the triple therapy was not due to intolerance to the combined mechanism of action, but rather to interference with drug metabolism, given that another non-mutant selective PI3Kα inhibitor, inabolisi, appears to be intolerant in the triple therapy (Bedard PL, et al. J Clin Oncol 2022;40(16_suppl):1052), and is progressing in Phase 3 trials with a CDK4 / 6 inhibitor and fulvestrant (NCT04191499).

[0034] Treatment method Indications Methods for treating or preventing diseases or disorders associated with dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or any of them (i.e., PI3Kα-related diseases or disorders), such as PIK3CA-related overgrowth syndromes (PROS), see, e.g., Venot, et al., Nature, 558, 540-546 (2018), brain disorders (e.g., macrocapillary malformation (MCAP) and hemimegalencephaly), congenital lipomas (e.g., abnormal proliferation of vascular malformations), epidermal nevi and skeletal / spinal cord abnormalities (Cloves syndrome) and fibroadipose hyperplasia (FH), or cancer (e.g., PI3Kα-related cancer).

[0035] In some embodiments, the compounds provided herein may exhibit potent and selective inhibition of PI3Kα. For example, the compounds provided herein can bind to the helical phosphatidylinositol kinase homologous domain catalytic domain of PI3Kα. In some embodiments, the compounds provided herein may exhibit nanomolar potency against PI3Kα kinases containing one or more mutations, for example, the mutations in Tables 1 and 2.

[0036] In some embodiments, the compounds provided herein may exhibit potent and selective inhibition of mutant PI3Kα. For example, the compounds provided herein can bind to allosteric sites within the kinase domain. In some embodiments, the compounds provided herein may exhibit nanomolar potency against PI3Kα proteins containing activating mutations with minimal activity against the relevant kinase (e.g., wild-type PI3Kα). Inhibition of wild-type PI3Kα may cause undesirable side effects (e.g., hyperglycemia and rash) that can affect quality of life and compliance. In some cases, inhibition of wild-type PI3Kα may result in dose-limiting toxicity. See, for example, Hanker, et al., Cancer Disc. 2019, 9, 4, 482-491. Mutaselective inhibitors may reduce the risk of such dose-limiting toxicity, including hyperglycemia, observed with inhibitors of wild-type PI3Kα.

[0037] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof selectively targets PI3Kα. For example, compound 1 or a pharmaceutically acceptable salt thereof selectively targets PI3Kα more than another kinase or non-kinase target.

[0038] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof may exhibit greater inhibition of PI3Kα containing one or more mutations described herein (e.g., one or more mutations described in Table 1 or Table 2) compared to inhibition of wild-type PI3Kα. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof may exhibit inhibition of PI3Kα containing one or more mutations described herein that is at least 2-fold, 3-fold, 5-fold, 10-fold, 25-fold, 50-fold, or 100-fold greater than inhibition of wild-type PI3Kα. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof may exhibit inhibition of PI3Kα containing one or more mutations described herein that is up to 1000-fold greater than inhibition of wild-type PI3Kα. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof may exhibit inhibition of PI3Kα having combinations of mutations described herein that are up to 10000-fold greater than inhibition of wild-type PI3Kα.

[0039] In some embodiments, compound 1 or its pharmaceutically acceptable salt is mutant PI3Kα (e.g., PI3Kα) rather than wild-type PI3Kα. H1047R ) selectively targets.

[0040] Compound 1 or its pharmaceutically acceptable salts are useful for treating PI3Kα-related diseases and disorders, such as PIK3CA-associated proliferative disorders (PROS), and proliferative disorders such as cancers including hematological malignancies and solid tumors (e.g., progressive or metastatic solid tumors), which can be treated with PI3Kα inhibitors.

[0041] In some embodiments, subjects have been identified or diagnosed with a cancer (PI3Kα-related cancer) that has dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof (determined, for example, using a regulatory-approved, e.g., FDA-approved assay or kit). In some embodiments, subjects have a tumor that is positive for dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof (determined, for example, using a regulatory-approved assay or kit). For example, a subject has a tumor that is positive for mutations such as those listed in Table 1 or Table 2. A subject may have a tumor that is positive for dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof (determined, for example, by a regulatory-approved, e.g., FDA-approved assay or kit). A subject may have a tumor that has dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof (for example, if the tumor is identified as such using a regulatory-approved, e.g., FDA-approved assay or kit). In some embodiments, the subject is suspected of having a PI3Kα-related cancer. In some embodiments, the subject has a clinical record indicating that the subject has a tumor with dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof (and optionally, the clinical record indicates that the subject should be treated with any of the compositions provided herein).

[0042] In some embodiments, the subjects are children. See, for example, Berhman RE, et al., Textbook of Pediatrics, 15th Ed. Philadelphia: WBSaunders Company, 1996; Rudolph AM, et al., Rudolph's Pediatrics, 21st Ed. New York: McGraw-Hill, 2002; and Avery and First, Pediatric Medicine, 2nd Ed. Baltimore: Williams & Wilkins; 1994.

[0043] In certain embodiments, compounds of formula (I) or pharmaceutically acceptable salts thereof are useful for preventing diseases and disorders as defined herein (e.g., PIK3CA-associated overgrowth syndrome (PROS) and cancer). As used herein, the term “prevent” means delaying the onset, recurrence, or progression of all or part of the disease or condition or symptoms thereof described herein.

[0044] As used herein, the term “PI3Kα-related disease or disorder” refers to a disease or disorder that is related to, or has a dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any one of them (e.g., one or more of them) (e.g., any kind of dysregulation of the expression, activity, or level of the PIK3CA gene, or the PI3Kα protein, or any one of them). Non-exclusive examples of PI3Kα-related diseases or disorders include, for example, PIK3CA-related hyperproliferative syndromes (PROS), brain disorders (e.g., macrocapillary malformations (MCAP) and hemimegalencephaly), congenital lipomas (e.g., abnormal proliferation of vascular malformations), epidermal nevi and skeletal / spinal cord abnormalities (e.g., CLOVES syndrome) and fibrolipid hyperplasia (FH), or cancer (e.g., PI3Kα-related cancer).

[0045] As used herein, the term “PI3Kα-related cancer” refers to cancers that are related to, or have, dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any of them. Non-exclusive examples of PI3Kα-related cancers are described herein.

[0046] The phrase "dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or any of them" refers to increased expression (e.g., increased levels) of wild-type PI3Kα in mammalian cells due to gene mutations (e.g., mutations in the PIK3CA gene resulting in the expression of PI3Kα having at least one amino acid deletion compared to wild-type PI3Kα, mutations in the PIK3CA gene resulting in the expression of PI3Kα having one or more point mutations compared to wild-type PI3Kα, mutations in the PIK3CA gene resulting in the expression of PI3Kα having at least one inserted amino acid compared to wild-type PI3Kα, gene duplication resulting in increased levels of PI3Kα in cells, or mutations in regulatory sequences (e.g., promoters and / or enhancers) resulting in increased levels of PI3Kα in cells), alternative splicing versions of PI3Kα mRNA resulting in PI3Kα having at least one amino acid deletion compared to wild-type PI3Kα, or increased expression (e.g., increased levels) of wild-type PI3Kα in mammalian cells due to abnormal cellular signaling and / or dysregulation of autocrine / paracrine signaling (e.g., compared to control non-cancerous cells). As another example, dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either of them may be due to a mutation in the PIK3CA gene encoding a PI3Kα protein that is constitutively active or has increased activity compared to the protein encoded by the unmutated PIK3CA gene. Non-limiting examples of point mutations / substitutions / insertions / deletions of PI3Kα are listed in Tables 1 and 2.

[0047] The term "activating mutation" with respect to PI3Kα refers to a mutation in the PIK3CA gene that results in the expression of PI3Kα with increased kinase activity, for example, compared to wild-type PI3Kα, when assayed under identical conditions. In another example, an activating mutation may be a mutation in the PIK3CA gene that results in the expression of PI3Kα having one or more amino acid substitutions (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10) (e.g., any combination of amino acid substitutions described herein) with increased kinase activity, for example, compared to wild-type PI3Kα, when assayed under identical conditions. In yet another example, an activating mutation may be a mutation in PIK3CA that results in the expression of PI3Kα having one or more amino acid deletions (e.g., 2, 3, 4, 5, 6, 7, 8, 9, or 10) (e.g., compared to wild-type PI3Kα), when assayed under identical conditions. In another example, an activating mutation could be a mutation in the PIK3CA gene that, when assayed under identical conditions, results in the expression of PI3Kα having at least one (e.g., at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 12, at least 14, at least 16, at least 18, or at least 20) amino acid insertions compared to wild-type PI3Kα, for example, the exemplary wild-type PI3Kα described herein. Examples of additional activating mutations are known in the art.

[0048] The term "wild type" refers to a nucleic acid (e.g., PIK3CA gene or PI3Kα mRNA) or protein (e.g., PI3Kα) sequence typically found in subjects without disease or disorder related to a reference nucleic acid or protein.

[0049] The term "wild-type PI3Kα" or "wild-type PI3Kα" refers to the normal PI3Kα nucleic acid (e.g., PIK3CA or PI3Kα mRNA) or protein found in subjects that do not have PI3Kα-related diseases, such as PI3Kα-related cancer (and, optionally, those that do not have an increased risk of developing PI3Kα-related diseases and / or are not suspected of having PI3Kα-related diseases), or in cells or tissues from subjects with PI3Kα-related diseases, such as PI3Kα-related cancer (and, optionally, those that do not have an increased risk of developing PI3Kα-related diseases and / or are not suspected of having PI3Kα-related diseases).

[0050] A method for treating cancer (e.g., PI3Kα-associated cancer) in a subject requiring treatment of cancer is provided herein, comprising administering to the subject a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof), or a pharmaceutical composition thereof, and one or more independently selected additional therapeutic agents described herein. For example, a method for treating PI3Kα-associated cancer in a subject requiring treatment of PI3Kα-associated cancer is provided herein, comprising: a) detecting dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof, in a sample from the subject; and b) administering compound 1 or a pharmaceutically acceptable salt thereof, and one or more independently selected additional therapeutic agents. In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof includes substitutions / point mutations / insertions of one or more PI3Kα proteins. Non-limiting examples of PI3Kα protein substitutions / insertions / deletions are given in Tables 1 and 2.

[0051] Some embodiments provide a method for treating cancer in a subject requiring cancer treatment, comprising administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0052] Some embodiments provide a method for treating cancer in a subject requiring cancer treatment, comprising administering to the subject a combination therapy comprising compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0053] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) Testing or having tested a subject in order to determine whether the subject has PI3Kα-related cancer, (b) A method is provided comprising administering to a subject compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0054] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) The subject is determined to have PI3Kα-related cancer, (b) A method is provided comprising administering to a subject compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0055] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) The subject is determined to have PI3Kα-related cancer, (b) A method is provided comprising administering to a subject a combination therapy comprising compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0056] Some embodiments provide a method for treating cancer in a subject previously determined to have PI3Kα-associated cancer, comprising administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or more independently selected additional therapeutic agents.

[0057] Some embodiments provide a method for treating cancer in a subject previously determined to have PI3Kα-associated cancer, comprising administering to the subject a combination therapy comprising compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents.

[0058] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, and the subjects (a) Compound 1 or a pharmaceutically acceptable salt thereof, (b) A method is provided comprising administering one or more independently selected additional therapeutic agents selected from the group consisting of selective estrogen receptor modulators (SERMs), selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, and RAF inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0059] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, and the subjects (a) Compound 1 or a pharmaceutically acceptable salt thereof, (b) A method is provided comprising administering a combination therapy comprising one or more independently selected additional therapeutic agents selected from the group consisting of selective estrogen receptor modulators (SERMs), selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, and RAF inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0060] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) Testing or having tested a subject in order to determine whether the subject has PI3Kα-related cancer, (b) A method is provided comprising administering to a subject compound 1 or a pharmaceutically acceptable salt thereof and a selective estrogen receptor modulator (SERM) / selective estrogen receptor degrader (SERD), CDK4 / 6 inhibitor, HER2 inhibitor, EGFR inhibitor, immune checkpoint inhibitor, MEK inhibitor, RAS inhibitor, RAF inhibitor, PIM (e.g., PIM1 and PIM3) inhibitor, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0061] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) The subject is determined to have PI3Kα-related cancer, (b) A method is provided comprising administering to a subject compound 1 or a pharmaceutically acceptable salt thereof and a selective estrogen receptor modulator (SERM) / selective estrogen receptor degrader (SERD), CDK4 / 6 inhibitor, HER2 inhibitor, EGFR inhibitor, immune checkpoint inhibitor, MEK inhibitor, RAS inhibitor, RAF inhibitor, PIM (e.g., PIM1 and PIM3) inhibitor, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0062] Some embodiments are methods for treating cancer in subjects requiring cancer treatment, (a) The subject is determined to have PI3Kα-related cancer, (b) A method is provided comprising administering to a subject a combination therapy comprising compound 1 or a pharmaceutically acceptable salt thereof and a selective estrogen receptor modulator (SERM) / selective estrogen receptor degrader (SERD), CDK4 / 6 inhibitor, HER2 inhibitor, EGFR inhibitor, immune checkpoint inhibitor, MEK inhibitor, RAS inhibitor, RAF inhibitor, PIM (e.g., PIM1 and PIM3) inhibitor, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0063] Some embodiments are methods for treating cancer in subjects previously determined to have PI3Kα-related cancer, wherein the subjects (a) Compound 1 or a pharmaceutically acceptable salt thereof, (b) A method is provided comprising administering one or more independently selected additional therapeutic agents selected from the group consisting of selective estrogen receptor modulators (SERMs), selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, and RAF inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0064] Several embodiments provide a method for treating cancer in a subject previously determined to have PI3Kα-associated cancer, comprising administering to the subject a combination therapy comprising compound 1 or a pharmaceutically acceptable salt thereof and a selective estrogen receptor modulator (SERM) / selective estrogen receptor degrader (SERD), CDK4 / 6 inhibitor, HER2 inhibitor, EGFR inhibitor, immune checkpoint inhibitor, MEK inhibitor, RAS inhibitor, RAF inhibitor, PIM (e.g., PIM1 and PIM3) inhibitor, or any combination thereof. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with one additional therapeutic agent. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered together with two independently selected additional therapeutic agents.

[0065] In some embodiments, substitutions / insertions / deletions of one or more PI3Kα proteins are selected from the group consisting of E542A, E542G, E542K, E542Q, E542V, E545A, E545D, E545G, E545K, E545Q, M1043I, M1043L, M1043T, M1043V, H1047L, H1047Q, H1047R, H1047Y, G1049R, and combinations thereof. In some embodiments, the substitution / insertion / deletion of the PI3Kα protein is H1047X, where X is any amino acid. In some embodiments, the substitution / insertion / deletion of the PI3Kα protein is H1047R. In some embodiments, the substitution / insertion / deletion of one or more PI3Kα proteins is selected from the group consisting of E542A, E542G, E542K, E542Q, E542V, E545A, E545D, E545G, E545K, E545Q, and H1047R.

[0066] In some embodiments, the cancer (e.g., PI3Kα-related cancer) is selected from hematological cancers and solid tumors.

[0067] In some embodiments, the cancer was refractory to one or more prior therapies. In some embodiments, the subjects experienced dose-limiting toxicity to one or more prior therapies. In some embodiments, one or more prior therapies included PI3K inhibitors.

[0068] In some embodiments, the cancer is unresectable and / or metastatic.

[0069] In some embodiments, the cancer is locally advanced. In some embodiments, the cancer is unresectable. In some embodiments, the cancer is metastatic. In some embodiments, the cancer is metastatic brain cancer as described herein. In some embodiments, the cancer is metastatic spinal cancer as described herein.

[0070] In some embodiments, cancer is breast cancer (HER2 + and HER2 -Both breast cancers, ER + The following are selected from breast cancer (including triple-negative breast cancer), endometrial cancer, lung cancer (including lung adenocarcinoma and squamous cell carcinoma), esophageal cancer (including esophageal squamous cell carcinoma), ovarian cancer, colorectal cancer, esophageal and gastric adenocarcinoma, stomach cancer, bladder cancer, head and neck cancer (including head and neck squamous cell carcinoma such as oropharyngeal squamous cell carcinoma), thyroid cancer, glioma, cervical cancer, lymphangioma, meningioma, melanoma (including uveal melanoma), prostate cancer, kidney cancer, pancreatic neuroendocrine neoplasm (pNET), stomach cancer, esophageal cancer, acute myeloid leukemia, relapsed and refractory multiple myeloma, and pancreatic cancer.

[0071] In some embodiments, cancer is breast cancer (HER2 + and HER2 - Both breast cancers, ER + The following cancers are selected from breast cancer (including triple-negative breast cancer), colon cancer, rectal cancer, colorectal cancer, ovarian cancer, lymphangioma, meningioma, squamous cell carcinoma of the head and neck (including oropharyngeal squamous cell carcinoma), melanoma (including uveal melanoma), kidney cancer, pancreatic neuroendocrine major (pNET), gastric cancer, esophageal cancer, acute myeloid leukemia, relapsed and refractory multiple myeloma, pancreatic cancer, lung cancer (including lung adenocarcinoma and squamous cell carcinoma), and endometrial cancer.

[0072] In some embodiments, the cancer is a gynecological cancer. In some embodiments, the gynecological cancer is endometrial cancer, ovarian cancer, or cervical cancer.

[0073] In some embodiments, the cancer is endometrial cancer that does not have a deficiency in DNA mismatch repair (dMMR), i.e., endometrial cancer is dMMR-free.

[0074] In some embodiments, the cancer is selected from breast cancer, lung cancer, endometrial cancer, esophageal cancer, stomach cancer, ovarian cancer, colorectal cancer, bladder cancer, head and neck cancer, thyroid cancer, prostate cancer, glioma, and cervical cancer.

[0075] In some embodiments, the cancer is breast cancer.

[0076] In some embodiments, the cancer is lung cancer.

[0077] In some embodiments, the cancer is endometrial cancer.

[0078] In some embodiments, the cancer is esophageal cancer.

[0079] In some embodiments, the cancer is stomach cancer.

[0080] In some embodiments, the cancer is ovarian cancer.

[0081] In some embodiments, the cancer is colorectal cancer.

[0082] In some embodiments, the cancer is bladder cancer.

[0083] In some embodiments, the cancer is head and neck cancer.

[0084] In some embodiments, the cancer is thyroid cancer.

[0085] In some embodiments, the cancer is prostate cancer.

[0086] In some embodiments, the cancer is a glioma.

[0087] In some embodiments, the cancer is cervical cancer.

[0088] In some embodiments, the cancer described herein is HER2 + It is cancer. In some embodiments, the cancer described herein is HER2 - It's cancer.

[0089] In some embodiments, the cancer described herein is a HER2-low-expressing cancer. In some embodiments, the cancer has a HER2 score of 0, +1, or +2 (e.g., an IHC score). In some embodiments, the cancer has a HER2 score of 0 or +1 (e.g., an IHC score). In some embodiments, the cancer has a HER2 score of 0. In some embodiments, the cancer has a HER2 score of +1. In some embodiments, the cancer has a HER2 score of +2. In some embodiments, the cancer has a HER2 score of +2 or +3. In some embodiments, the cancer has a HER2 score of +3.

[0090] In some embodiments, the cancers described herein are hormone receptor-positive (HR + ) is cancer. In some embodiments, the cancer described herein is HER2 - and HR + It is cancer. In some embodiments, the cancer described herein is ER + In some embodiments, the cancer described herein is PR + In some embodiments, cancer is HR + / ER - I have breast cancer.

[0091] In some embodiments, the cancer described herein is HR+ and has a HER2 score of 0 or +1. In some embodiments, the cancer described herein is HR+ and has a HER2 score of 0. In some embodiments, the cancer described herein is HR+ and has a HER2 score of +1.

[0092] In some embodiments, the cancer described herein is HR+ and has a HER2 score of +2 or +3. In some embodiments, the cancer described herein is HR+ and has a HER2 score of +2. In some embodiments, the cancer described herein is HR+ and has a HER2 score of +3.

[0093] In some embodiments, the subject has been previously identified or determined not to have activating mutations in AKT and / or PTEN.

[0094] In some embodiments, PI3Kα-related cancers are selected from the cancers listed in Tables 1 and 2.

[0095] Table 1. Amino acid substitutions / insertions / deletions of PI3Kα protein A [Table 1-1] TIFF2026509320000006.tif68169

[0096] (Continued from Table 1) [Table 1-2] TIFF2026509320000008.tif116169

[0097] (Continued from Table 1) [Table 1-3]

[0098] (Continued from Table 1) [Table 1-4] TIFF2026509320000011.tif20169

[0099] (Continued from Table 1) [Table 1-5] TIFF2026509320000013.tif58169

[0100] (Continued from Table 1) [Table 1-6] TIFF2026509320000015.tif26169

[0101] (Continued from Table 1) [Table 1-7] TIFF2026509320000017.tif24169

[0102] (Continued from Table 1) [Table 1-8] TIFF2026509320000019.tif77169

[0103] (Continued from Table 1) [Table 1-9] TIFF2026509320000021.tif69169

[0104] (Continued from Table 1) [Table 1-10] TIFF2026509320000023.tif58169

[0105] (Continued from Table 1) [Table 1-11] TIFF2026509320000025.tif49169

[0106] (Continued from Table 1) [Table 1-12]

[0107] (Continued from Table 1) [Table 1-13] TIFF2026509320000028.tif62169 A Unless otherwise noted, the mutations in Table 1 are found in the cBioPortal database, derived from Cerami et al. The cBio Cancer Genomics Portal: An Open Platform for Exploring Multidimensional Cancer Genomics Data. Cancer Discovery. May 2012 2;401, and Gao et al. Integrative analysis of complex cancer genomics and clinical profiles using the cBioPortal. Sci.Signal.6,pl1 (2013). † Velho S, Oliveira C, Ferreira A, Ferreira AC, Suriano G, Schwartz S Jr, Duval A, Carneiro F, Machado JC, Hamelin R, Seruca R. The prevalence of PIK3CA mutations in gastric and colon cancer.Eur J Cancer.2005 Jul;41(11):1649-54.doi:10.1016 / j.ejca.2005.04.022.PMID:15994075.

[0108] Table 2. Additional amino acid substitutions / insertions / deletions of PI3Kα protein A [Table 2-1] TIFF2026509320000030.tif188170

[0109] (Continued from Table 2) [Table 2-2] A Unless otherwise noted, the mutations in Table 2 are found in the cBioPortal database, derived from Cerami et al. The cBio Cancer Genomics Portal: An Open Platform for Exploring Multidimensional Cancer Genomics Data. Cancer Discovery. May 2012 2;401, and Gao et al. Integrative analysis of complex cancer genomics and clinical profiles using the cBioPortal. Sci.Signal.6,pl1 (2013). † Velho S, Oliveira C, Ferreira A, Ferreira AC, Suriano G, Schwartz S Jr, Duval A, Carneiro F, Machado JC, Hamelin R, Seruca R. The prevalence of PIK3CA mutations in gastric and colon cancer.Eur J Cancer.2005 Jul;41(11):1649-54.doi:10.1016 / j.ejca.2005.04.022.PMID:15994075.

[0110] In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any of them includes splice variations in PI3Kα mRNA, resulting in an expressed protein that is an alternative splicing variant of PI3Kα that has at least one residue deleted (compared to the wild-type PI3Kα protein) and results in constitutive activity of the PI3Kα protein domain.

[0111] In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or any of them includes at least one point mutation in the PIK3CA gene resulting in the production of a PI3Kα protein having one or more amino acid substitutions, insertions, or deletions in the PIK3CA gene, which results in the production of a PI3Kα protein in which one or more amino acids are inserted or removed compared to the wild-type PI3Kα protein. In some cases, the resulting mutant PI3Kα protein has increased activity compared to the wild-type PI3Kα protein or a PI3Kα protein that does not contain the same mutation. In some embodiments, the compounds described herein selectively inhibit the resulting mutant PI3Kα protein compared to the wild-type PI3Kα protein or a PI3Kα protein that does not contain the same mutation.

[0112] Exemplary sequence (SEQ ID NO: 1) of human phosphatidylinositol 4,5-bisphosphate 3-kinase isoform alpha (UniProtKB entry P42336)

[0113] [Table 3]

[0114] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is useful for treating cancers identified as having one or more PI3Kα mutations. Accordingly, a method for treating a subject diagnosed with (or identified as having) cancer is provided herein, comprising administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof can cross the blood-brain barrier (BBB) ​​and inhibit mutant PI3Kα in brain and / or other central nervous system (CNS) structures. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof can cross the BBB in therapeutically effective doses.

[0115] Also provided herein are methods for treating subjects identified or diagnosed with PI3Kα-related cancer, comprising administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents. In some embodiments, subjects are identified or diagnosed with PI3Kα-related cancer by using regulatory authority-approved tests or assays, such as FDA-approved tests or assays, or by performing any non-limiting example of assays described herein, to identify dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or any of them in the subject or a biopsy sample from the subject. In some embodiments, the tests or assays are provided as kits. In some embodiments, the cancer is PI3Kα-related cancer.

[0116] The term "regulatory authority" refers to a national agency that approves the medical use of drugs in a given country. For example, a non-specific example of a regulatory agency is the U.S. Food and Drug Administration (FDA).

[0117] Also provided is a method for treating cancer in a subject requiring cancer treatment, comprising (a) detecting PI3Kα-related cancer in the subject, and (b) administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents. In some embodiments, the subject has previously been treated with another anti-cancer treatment, e.g., at least partial resection of the tumor or radiotherapy. In some embodiments, the subject is determined to have PI3Kα-related cancer in the subject or a biopsy sample from the subject by using a regulatory authority-approved test or assay, e.g., an FDA-approved test or assay, or by performing any non-limiting example of an assay described herein, to identify dysregulation of the expression, activity or level of the PIK3CA gene, the PI3Kα protein, or either thereof. In some embodiments, the test or assay is provided as a kit. In some embodiments, the cancer is PI3Kα-related cancer.

[0118] Also provided is a method for treating a subject, comprising: performing an assay on a sample obtained from the subject to determine whether the subject has dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof; and administering (e.g., specifically or selectively) compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents to the subject determined to have dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof. In some embodiments of these methods, the subject has been previously treated with another anti-cancer treatment, such as at least partial resection of a tumor or radiotherapy. In some embodiments, the subject is a subject suspected of having PI3Kα-related cancer, a subject exhibiting one or more symptoms of PI3Kα-related cancer, or a subject at high risk of developing PI3Kα-related cancer. In some embodiments, the assay utilizes next-generation sequencing, pyrosequencing, immunohistochemistry, or break-apart FISH analysis. In some embodiments, the assay is a regulatory-approved assay, such as an FDA-approved kit. In some embodiments, the assay is a liquid biopsy. Additional non-limiting assays that may be used in these methods are described herein. Additional assays are also known in the art.

[0119] Furthermore, to determine whether a subject has dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof, the present invention provides compound 1 or a pharmaceutically acceptable salt thereof, along with one or more additional therapeutic agents, for use in treating PI3Kα-related cancer in subjects identified or diagnosed with PI3Kα-related cancer through a step of performing an assay (e.g., an in vitro assay) on a sample obtained from the subject, wherein the presence of dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof identifies the subject as having PI3Kα-related cancer. Furthermore, to determine whether a subject has dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof, a step of performing an assay on a sample obtained from the subject is provided, which provides compound 1 or a pharmaceutically acceptable salt thereof, and one or more additional therapeutic agents for the manufacture of a pharmacopoeia for the treatment of PI3Kα-related cancer in a subject identified or diagnosed with PI3Kα-related cancer, wherein the presence of dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof identifies the subject as having PI3Kα-related cancer. Some embodiments of any of the methods or uses described herein further include recording in the subject's clinical record (e.g., on a computer-readable medium) that the subject has been determined, through the performance of the assay, to have dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof, and that the subject needs to be administered compound 1 or a pharmaceutically acceptable salt thereof, and one or more additional therapeutic agents. In some embodiments, the assay utilizes next-generation sequencing, pyrosequencing, immunohistochemistry, or break-apart FISH analysis. In some embodiments, the assay is a regulatory-approved assay, such as an FDA-approved kit. In some embodiments, the assay is a liquid biopsy.

[0120] Furthermore, compound 1 or a pharmaceutically acceptable salt thereof, along with one or more additional therapeutic agents, are provided for use in the treatment of cancer in subjects requiring treatment for cancer or subjects identified or diagnosed with PI3Kα-related cancer. Also provided are compound 1 or a pharmaceutically acceptable salt thereof, along with one or more additional therapeutic agents, for the manufacture of pharmaceuticals for the treatment of cancer in subjects identified or diagnosed with PI3Kα-related cancer. In some embodiments, subjects are identified or diagnosed with PI3Kα-related cancer through the use of a regulatory-approved, e.g., FDA-approved kit for identifying dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or any of them, in the subject or a biopsy sample from the subject. As provided herein, PI3Kα-related cancers include those described herein and those known in the art.

[0121] In some embodiments, the subject is identified or diagnosed with a cancer involving dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either of them. In some embodiments, the subject has a tumor that is positive for dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either of them. In some embodiments, the subject may have a tumor that is positive for dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either of them. In some embodiments, the subject may have a tumor whose tumor has dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either of them. In some embodiments, the subject is suspected of having a PI3Kα-related cancer. In some embodiments, a method for treating PI3Kα-associated cancer in a subject requiring treatment for PI3Kα-associated cancer is provided herein, comprising: a) detecting dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any of them in a sample from the subject; and b) administering compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents. In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any of them includes one or more point mutations / insertions / deletions of the PI3Kα protein. Non-limiting examples of point mutations / insertions / deletions of the PI3Kα protein are listed in Tables 1 and 2. In some embodiments, the point mutation / insertion / deletion of the PI3Kα protein is H1047X, where X is any amino acid. In some embodiments, point mutations / insertions / deletions of the PI3Kα protein are selected from the group consisting of E542A, E542G, E542K, E542Q, E542V, E545A, E545D, E545G, E545K, E545Q, M1043I, M1043L, M1043T, M1043V, H1047L, H1047Q, H1047R, H1047Y, and G1049R.In some embodiments, tumors with dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof are determined using a regulatory-approved assay or kit, such as an FDA-approved one.

[0122] In some embodiments, a subject has a clinical record indicating that the subject has a tumor with dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof. Also provided is a method of treating a subject having a clinical record indicating that the subject has dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof, comprising administering compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents to the subject.

[0123] In some embodiments, the methods described herein include performing an assay on a sample obtained from a subject to determine whether the subject has dysregulation of the expression or level of the PIK3CA gene, the PI3Kα protein, or either thereof. In some such embodiments, the methods also include administering compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents to a subject determined to have dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof. In some embodiments, the methods include determining whether a subject has dysregulation of the expression or level of the PIK3CA gene, the PI3Kα protein, or either thereof via an assay performed on a sample obtained from a subject. In some such embodiments, the methods also include administering compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents to the subject. In some embodiments, the dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or either thereof is one or more point mutations in the PIK3CA gene (e.g., one or more of the PI3Kα point mutations described herein). One or more point mutations in the PIK3CA gene may result in the translation of the PI3Kα protein having one or more amino acid substitutions, deletions, and insertions of, for example, E542A, E542G, E542K, E542Q, E542V, E545A, E545D, E545G, E545K, E545Q, M1043I, M1043L, M1043T, M1043V, H1047L, H1047Q, H1047R, H1047Y, and G1049R. One or more mutations in the PIK3CA gene may result in the translation of the PI3Kα protein having one or more of the following amino acids: 542, 545, 1043, and 1047 and 1049. In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, the PI3Kα protein, or any of them is due to one or more PI3Kα amino acid substitutions (e.g., any of the PI3Kα amino acid substitutions described herein).Some embodiments of these methods further include administering another anticancer drug (e.g., immunotherapy) to the subject.

[0124] In some embodiments, assays used to determine whether a subject has dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof, using a sample from the subject, may include, for example, next-generation sequencing, immunohistochemistry, fluorescence microscopy, break-apart FISH analysis, Southern blotting, Western blotting, FACS analysis, Northern blotting, and PCR-based amplification (e.g., RT-PCR and quantitative real-time RT-PCR). As is well known in the art, the assay is typically carried out using, for example, at least one labeled nucleic acid probe or at least one labeled antibody or its antigen-binding fragment. The assay may utilize other detection methods known in the art for detecting dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof (see, for example, the references cited herein). In some embodiments, the sample is a biological or biopsy sample from the subject (e.g., a paraffin-embedded biopsy sample). In some embodiments, the subjects are those suspected of having PI3Kα-related cancer, those having one or more symptoms of PI3Kα-related cancer, and / or those at increased risk of developing PI3Kα-related cancer.

[0125] In some embodiments, dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof can be identified using liquid biopsy (variously referred to as fluid biopsy or fluid-phase biopsy, etc.). See, for example, Karachialiou et al. Ann. Transl. Med., 3(3):36, 2016. Liquid biopsy can be used to detect dysregulation of total tumor volume and / or the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof. Liquid biopsy can be performed on biological samples obtained relatively easily from the subject (e.g., via simple blood sampling) and is generally less invasive than conventional methods used to detect dysregulation of tumor volume and / or the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof. In some embodiments, liquid biopsy can be used to detect the presence of dysregulation of the expression, activity, or level of the PIK3CA gene, PI3Kα protein, or either thereof at an earlier stage than conventional methods. In some embodiments, the biological samples used in liquid biopsy may include blood, plasma, urine, cerebrospinal fluid, saliva, sputum, bronchoalveolar lavage fluid, bile, lymph, cystic fluid, feces, ascites, and combinations thereof. In some embodiments, liquid biopsy can be used to detect circulating tumor cells (CTCs). In some embodiments, liquid biopsy can be used to detect cell-free DNA. In some embodiments, the cell-free DNA detected using liquid biopsy is circulating tumor DNA (ctDNA) derived from tumor cells. Analysis of ctDNA (e.g., using highly sensitive detection techniques such as next-generation sequencing (NGS), conventional PCR, digital PCR, or microarray analysis, etc.) can be used to identify dysregulation of the expression, activity, or levels of PIK3CA, PI3Kα proteins, or either of them.

[0126] Also provided herein is a method for inhibiting cell proliferation, comprising contacting cells with compound 1 and one or two additional independently selected therapeutic agents as defined herein.

[0127] A method for increasing cell death is further provided herein, comprising contacting cells with compound 1 and one or two additional independently selected therapeutic agents as defined herein.

[0128] In some embodiments, the contact is in vitro. In some embodiments, the contact is in vivo. In some embodiments, the contact is in vivo, and the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof and one or two independently selected additional therapeutic agents as defined herein to a subject having cells having abnormal PI3Kα activity. In some embodiments, the cells are cancer cells. In some embodiments, the cancer cells are any cancer as described herein. In some embodiments, the cancer cells are PI3Kα-associated cancer cells.

[0129] As used herein, the term “contact” means bringing together the indicated portions in an in vitro or in vivo system. For example, “contact” the PI3Kα protein with the compound provided herein includes administering the compound provided herein to an individual or subject, such as a human, having the PI3Kα protein, and introducing the compound provided herein into a sample containing cells or a purified preparation containing the PI3Kα protein, for example.

[0130] Also provided herein is a method for increasing tumor cell death in a subject, comprising administering to the subject compound 1 or a pharmaceutically acceptable salt thereof and one or two additional independently selected therapeutic agents as defined herein.

[0131] combination The methods described herein relate, in particular, to the treatment of cancer (e.g., cancers described herein, such as breast cancer) using compound 1 or a pharmaceutically acceptable salt thereof and one or more additional therapeutic agents.

[0132] Such additional therapeutic agents include additional therapeutic molecules (e.g., small molecules or antibodies), as well as radiotherapy (or radiotherapy) and surgical procedures, such as open surgery or minimally invasive surgery. Therefore, compounds of formula I, such as compound 1, or pharmaceutically acceptable salts thereof, may also be useful as adjuvants in cancer treatment; that is, they can be used in combination with one or more additional therapies or therapeutic agents, such as chemotherapeutic agents acting by the same or different mechanisms of action.

[0133] In some embodiments, a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof) can be used before administering one or more independently selected additional therapeutic agents or additional therapies. For example, one or more doses of a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof) can be administered to a subject in need for a period of time, followed by at least partial resection of the tumor. In some embodiments, treatment with one or more doses of a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof) reduces the size of the tumor (e.g., tumor volume) before at least partial resection of the tumor. In some embodiments, one or more doses of a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof) can be administered to a subject in need for a period of time under one or more radiotherapy sessions. In some embodiments, treatment with one or more doses of a compound of formula (I) or a pharmaceutically acceptable salt thereof (e.g., compound 1 or a pharmaceutically acceptable salt thereof) reduces the size of the tumor (e.g., tumor volume) before one or more radiotherapy sessions.

[0134] In some embodiments, the subjects have cancer (e.g., locally advanced or metastatic tumors) that is refractory or intolerant to standard therapy (e.g., chemotherapy agents such as multi-kinase inhibitors, immunotherapy, or radiation (e.g., radioactive iodine)). In some embodiments, the subjects have cancer (e.g., locally advanced or metastatic tumors) that is refractory or intolerant to prior therapy (e.g., chemotherapy agents such as multi-kinase inhibitors, immunotherapy, or radiation (e.g., radioactive iodine)). In some embodiments, the subjects have cancer (e.g., locally advanced or metastatic tumors) for which there is no standard therapy. In some embodiments, the subjects are PI3Kα inhibitor naive. For example, the subjects are naive to treatment with selective PI3Kα inhibitors. In some embodiments, the subjects are not PI3Kα inhibitor naive. In some embodiments, the subjects are kinase inhibitor naive. In some embodiments, the subjects are not kinase inhibitor naive. In some embodiments, the subjects have received prior therapy.For example, multi-kinase inhibitors (MKIs) or other PI3K inhibitors, such as buparlisib (BKM120), alpelisib (BYL719), WX-037, idelalisib, duvelisib, copanlisib, umbralisib (ALIQOPATM, BAY80-6946), dactricib (NVP-BEZ235, BEZ-235), taselicib (GDC-0032, RG7604), sonolicib (PX-866), fimepinostat (CUDC-907), vimiralisib (PQR309), ZSTK474, SF112. 6, AZD8835, Inaborishi (GDC-0077), ASN003, Pictilisib (GDC-0941), Pilaralisib (XL147, SAR245408), Gedatricib (PF-05212384, PKI-587), Ceravelisib (TAK-117, MLN1117, INK1117), BGT-226 (NVP-BGT226), PF-04691502, Apitricib (GDC-0980), Omiparisib (GSK2126458, GSK458), Boxtalisib (XL756, SAR245409), AMG Treatment using 511, CH5132799, GSK1059615, paxalisib (GDC-0084 (RG7666), VS-5584 (SB2343), PKI-402, wartmannin, LY294002, PI-103, ligosertib (ON-01910 sodium salt), boxalisib (XL-765), LY2023414, SAR260301, KIN-193 (AZD-6428), acalisib (GS-9820), AMG319, or GSK2636771.

[0135] In some embodiments of any method described herein, the compound of formula I (or a pharmaceutically acceptable salt thereof) is administered in combination with at least one additional therapeutic agent selected from one or more additional therapeutic or therapeutic (e.g., chemotherapy) agents in a therapeutically effective amount.

[0136] Non-limiting examples of additional therapeutic agents include other PI3Kα-targeting therapies (i.e., other PI3Kα inhibitors), EGFR inhibitors, VEGFR / VEGF inhibitors, HER2 inhibitors, MEK pathway-targeting therapies (including RAS pathway-targeting therapies, including mTOR modulators / inhibitors as described herein), SHP2 inhibitors, ULK inhibitors, CDK4 / 6 inhibitors, NTRK / ROS inhibitors, ALK inhibitors, RET inhibitors, MET inhibitors, PARP inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, other kinase inhibitors (e.g., Trk inhibitors or multi-kinase inhibitors), KAT6A inhibitors, farnesyltransferase inhibitors, and aromatizers. Examples include ester inhibitors, selective estrogen receptor modulators or degraders (including SERM / SERD, ERα inhibitors or ERα degraders), vinca alkaloids, antimetabolites, antiandrogens (e.g., androgen receptor (AR) antagonists, AR degraders, AR modulators), alkylating agents, checkpoint inhibitors, apoptotic pathway modulators, cytotoxic chemotherapeutic agents (also called antineoplastic chemotherapeutic agents), angiogenesis-targeted therapies (e.g., angiogenesis inhibitors), immunotargeted agents (including immunotherapy), radiotherapy, glucocorticoids, antidiarrheals (such as loperamide and diphenoxylate-atropine), antihistamines, and retinoic acids.

[0137] As used herein, PIM inhibitors are inhibitors of the provirus integration site for Moloney leukemia virus kinase (sometimes called the proviral insertion site in Murine leukemia virus protein kinase), e.g., PIM1, PIM2, and PIM3, and any of their isoforms (e.g., PIM-1L (molecular weight 44 kDa) and PIM-1S (molecular weight 33 kDa)). PIM kinases regulate cell proliferation, survival, metabolism, cell transport, and signaling, and are overexpressed in several human cancers. Non-exclusive examples of PIM1 inhibitors include A47, abemaciclib (Verzenio:NCT03905889), AZD1208 (NCT01588548), AZD1897, ETH-155008, ETP-390101, ETP-45299, ETP-47551, INCB053914 (uzansertib), JP11646, K00135, K00486, LGB321, LGH447 (PIM447), PIM447, SEL24 / MEN1703 (SEL24-B489), SGI-1776, and TP-3654 (Belon and Nicot (2023) Mol. Cancer 22(1):18; Mahata S., et al.). a;.(2022)Med.Oncol.39(5):74, Asasti V.,et al.(2019)Eur J Med Chem.172:95-108,Le X.,et al.(2016)Cancer Discov.6(10):1134-47,Keeton EK,et al.(2014)Blood 123:905-13, Garcia P., et al. (2013) ASH 122:1666, Grundler R, et al. (2009) J Exp Med.206(9):1957-70, Pogacic V., et al., (2007) Cancer Res.67(14):6916-6924).

[0138] Examples of SHP2 inhibitors include JAB-3312, SHP099, SHP099 hydrochloride, SHP504, RMC-3943, AS1949490, SHP394, SHP389, and RMC-4630.

[0139] In some embodiments, the SHP2 inhibitor is RMC-4630.

[0140] Examples of ULK inhibitors include ULK-101, SBP-7455, SBI-0206965, ULK1-IN-2, MRT68921, MRT68921 dihydrochloride, MRT67307, MRT67307 hydrochloride, XST-14, and GW406108X (CW108X).

[0141] Examples of NTRK / ROS inhibitors include entrectinib (NMS-E628, RXDX-101, ROZLYTREK®), taretrectinib (DS-6051b, AB-106), or repotrectinib (TPX-0005).

[0142] Examples of ALK inhibitors include crizotinib (XALKORI®, PF-02341066), ceritinib (ZYKADIA®, LDK-378), alectinib (ALECENSA®, CH5424802, RO5424802, AF802), brigatinib (ALUNBRIG®, AP-26113), lorlatinib (LORBRENA®, PF-06463922), entrectinib (NMS-E628, RXDX-101, ROZLYTREK®), ASP3026, TSR-011, PF-06463922, enzalutinib (X-396), or CEP-37440.

[0143] Examples of RET inhibitors include serpercatinib (RETEVMO®, LOXO-292), zeteretinib (BOS-172738, DS-5010), GSK3179106, amivatinib hydrochloride (MP470 hydrochloride, HPK 56 hydrochloride), TPX-0046, or pralcetinib (GAVRETO®, BLU-667).

[0144] Examples of MET inhibitors include capmatinib (TABRECTA®, INC280, INCB28060), tepotinib (TEPMETKO®), tivantinib (ARQ197), saboltinib (ORPATHYS®, vortinib, HMPL-504, AZD-6094), forretinib (XL880, GSK1363089, GSK089, EXEL-2880), pamfetinib (TAS-115), c-Met-IN-2, PHA-665752, SU11274, SYN1143, or amvatinib hydrochloride (MP470 hydrochloride, HPK 56 hydrochloride).

[0145] Examples of TRK or multitarget kinase inhibitors include altiratinib (DCC-2701), CH7057288, larotrectinib (VITRAKVI®), entrectinib, ANA-12, repotrectinib (TPX-0005), citravatinib (MGCD516, MG-516), restaurtinib (CEP-701, KT-5555), tilhostin AG 879 (AG 879), and ceritrectinib (LOXO-195).

[0146] Examples of tyrosine kinase inhibitors include axitinib (INLYTA®), dasatinib (SPRYCEL®), erlotinib (TARCEVA®), imatinib (GLIVEC), nilotinib (TASIGNA®), pazopanib (VOTRIENT®), sunitinib (SUTENT®), and vemurafenib.

[0147] Similar vinca alkaloids include vinorelbine, vinblastine, vincristine, vindesine, and vinflunin.

[0148] Examples of antimetabolites include methotrexate, 5-fluorouracil (5-FU), 6-mercaptopurine (6-MP), capecitabine (XELODA®), phloxuridine, cytarabine (ARA-C®), fludarabine, gemcitabine (GEMZAR®), hydroxycarbamide, pemetrexed (ALIMTA®), phototrexate, decitabine, Vidaza, DFP-11207, and RX-3117 and TAS-114.

[0149] Examples of alkylating agents include cyclophosphamide, lomustine, carmustine, streptozocin, bendamustine, chlorambucil, cyclophosphamide, ifosfamide, mechloretamine, melphalan, chlorambucil, melphalan, busulfan, dacarbazine, temozolomide, altoretamine, thiotepa, carboplatin, cisplatin, lurubinectedin, trabectedin, carmustine, oxaliplatin, and oxaliplatin.

[0150] Exemplary checkpoint inhibitors include nivolumab, pembrolizumab, semipramab, atezolizumab, durvalumab, avelumab, ipilimumab, LAG525 (IMP701), REGN3767 (R3767), BI754091, tebotelimab (MGD013), efalizumab alpha (IMP321), FS118, MBG453, Sym023, TSR-022, MGC018, FPA150, EOS100850, AB928, CPI-006, monalizumab, COM701, CM24, NEO-201, defactinib, PF-04136309, MSC-1, Hu5F9-G4 (5F9), ALX148, TTI-662, RRx-001, lacnotuzumab (MCS110), LY3022855, SNDX-6352, ematuzumab (RG7155),pexidartinib (PLX3397), CAN04, canakinumab (ACZ885), BMS-986253, pepinemab (VX-15 / 2503), trebananib, FP-1305, enapotamab vedotin (EnaV), and babiximab.

[0151] Exemplary modulators of the apoptosis pathway include obatoclax, oblimersen, ABT-737, navitoclax (ABT-263), venetoclax (ABT-199), Z-VAD-FMK, emricasan, Q-VD-Oph, Z-VAD(OH)-FMK, bernasane (caspase-1), Z-DEVD-FMK (caspase-3), Q-VD-OphZ-IETD-FMK (caspase-8), PAC-1 (procaspase-3), nutlin-3, nutlin-3a, idasanutlin, HDM201, APR-246, CBL0137, pifithrin-alpha, pifithrin-mu, Z-VAD-FMK, emricasan, Q-VD-Oph, Z-VAD(OH)-FMK, pomalidomide, lenalidomide, YM155, venetoclax (Bcl-2), S63845 (MCL-1), and A-1`331852 (BCL-XL).

[0152] Exemplary cytotoxic chemotherapeutic agents include 5-fluorouracil, gemcitabine, methotrexate, NB1011, cyclophosphamide, dacarbazine, melphalan, trabectedin, temozolomide, doxorubicin, daunorubicin, mitozantrone, vinblastine, paclitaxel, docetaxel, irinotecan, etoposide, and platinum agents such as carboplatin, cisplatin, and oxaliplatin.

[0153] Exemplary topoisomerase inhibitors include etoposide, irinotecan, camptothecin (CPT), topotecan (TPT), irinotecan, verotecan, indenoisoquinoline, phenanthridine, and indolocarbazole. Further examples of topoisomerase inhibitors include aminocamptothecin, CT-2106, crisnatol mesylate, DE-310, elanifide, lucanthone, MLN576, and miindomid.

[0154] Exemplary angiogenesis target therapies include bevacizumab, itraconazole, carboxyamidotriazole, TNP-470, CM101, IFN-α, IL-12, platelet factor-4, suramin, SU5416 thrombospondin, angiostatin, endostatin, 2-methoxyestradiol, tecogalan, tetrathiomolybdate, thalidomide, thrombospondin, prolactin, lenomid, ramucirumab, tasquinimod, ranibizumab, sorafenib, sunitinib, pazopanib, everolimus, lenalidomide (e.g., REVLIMID®), and pomalidomide (e.g., POMALYST® or Imnovid®), marimistat, 2-methoxyestradiol (PANZEM), SU5415, SU6668, pemaxanib, sunitinib, vandetanib, vitaxin, YM598, ZD6126, and aflibercept.

[0155] In some embodiments, the angiogenesis target therapy is lenalidomide.

[0156] In some embodiments, the angiogenesis-targeted therapy is pomalidomide.

[0157] In some embodiments, the EGFR inhibitor is osimertinib (AZD9291, merelectinib, TAGRISSO®), erlotinib (TARCEVA®), gefitinib (IRESSA®), cetuximab (ERBITUX®), necitumumab (PORTRAZZA®, IMC-11F8), neratinib (HKI-272, NERLYNX®), panitum Mab (ABX-EGF, VECTIBIX®), Vandetanib (CAPRELSA®), Rosiretinib (CO-1686), Olumutinib (OLITA®, HM61713, BI-1482694), Nacotinib (ASP8273), Nazartinib (EGF816, NVS-816), Mabellerutinib (PF-06747775), Icotinib (BPI-2009H), Afatinib (BIBW) 2992, GILOTRIF(registered trademark), dacomitinib (PF-00299804, PF-804, PF-299, PF-299804), abitinib (AC0010), AC0010MA, EAI045, matsuzumab (EMD-7200), nimotuzumab (h-R3, BIOMAb EGFR(registered trademark)), salzumab, MDX447, depatuximab (humanized mAb 806, ABT-806), depatuximab mafodotin (ABT-414), ABT-806, mAb 806, canertinib (CI-1033), shikonin, shikonin derivatives (e.g., deoxyshikonin, isobutyrylshikonin, acetylshikonin, β,β-dimethylacryliconine and acetylalkanin), poziotinib (NOV120101, HM781-36B), AV-412, ibrutinib, WZ4002, brigatinib (AP26113, ALUNBRIG®), peritinib (EKB-569), tarloxotini (TH-4000, PR610), BPI-15086, Hemay022, ZN-e4, tesevatinib (KD019, XL647), ibrutinib (YH25448), epitinib (HMPL-813), olafertinib (CK-101, RX518), MM-151, zolife Rutinib (AZD3759), Vandetanib (ZD6474), PF-06459988, Baritinib (ASLAN001, ARRY-334543), Modotuximab (AP32788, TAK-788), Pimulutamab (HLX07), Befotertinib (D-0316), AEE788 (NVP-AEE788), Amoreltinib (formerly Almonertinib, HS-10296), Abitinib, Lapatinib (GW572016), Pilotinib (SHR1258), SCT200, CPGJ602, Sym004 (combination of futuximab and modotuximab), EMD These include 55900 (MAb-425), modotuximab (TAB-H49), futuximab (992 DS), saltumumab, RO5083945, laprituximab emtansine (IMGN289), amivantamab (RYBREVANT® JNJ-61186372), LY3164530, pan-HER (Sym013), AMG 595, tuxovertinib (BDTX-189), abatinib, disulptin, CL-387785 (EKI-785, WAY-EKI 785), EGFRBi-armed autologous T cells, and EGFR CAR-T therapy. In some embodiments, EGFR-targeted therapeutic agents are selected from gefitinib, erlotinib, afatinib, lapatinib, neratinib, osimertinib (AZD-9291, e.g., TAGRISSO®), CL-387785 (EKI-785, WAY-EKI 785), rosiletinib (CO-1686), WZ4002, OMP-305B83, trastuzumab (e.g., TRAZIMERA®, HERCEPTIN®), RG-7597, and amivantanab.

[0158] In some embodiments, the EGFR inhibitor is razertinib. In some embodiments, the EGFR inhibitor is amivantanab. In some embodiments, the EGFR inhibitor is trastuzumab.

[0159] Examples of HER2 inhibitors include trastuzumab (e.g., TRAZIMERA®, HERCEPTIN®), pertuzumab (e.g., PERJETA®), trastuzumab emtansine (T-DM1 or ad-trastuzumab emtansine, e.g., KADCYLA®), famtrastuzumab deruxtecan (ENHERTU®), lapatinib, KU004, neratinib (e.g., NERLYNX®), and emvatinib. 57 Govitecan-hziy (TRODELVY®), Dacomitinib (e.g., VIZIMPRO®), Afatinib (GILOTRIF®), Tucatinib (Irbinitinib, ONT-380, ARRY-380, e.g., TUKYSA®), Erlotinib (e.g., TARCEVA®), Pilotinib, Pozotinib, C Examples include P-724714, CUDC-101, sapitinib (AZD8931), tanespimycin (17-AAG), IPI-504, dacomitinib (PF299804, PF299), peritinib, margetuximab, AEE-788 (NVP-AEE788), enfortumab vedotin (PADCEV®), and datopotamab deruxtecan.

[0160] In some embodiments, the HER2 inhibitor is famtrastuzumab deruxtecan. In some embodiments, the HER2 inhibitor is 58-emvatinib 58-govitecan-hziy. In some embodiments, the HER2 inhibitor is datopotamab deruxtecan.

[0161] Examples of VEGFR / VEGF inhibitors include pazopanib, sunitinib, 58-emvatinib 58, cabozantinib, sorafenib, regorafenib, ponatinib, 58-emvatinib axitinib, ziv-aflibercept, vandetanib, tivozanib, batalanib, AZD-2932, aflibercept, vanucizumab, BI836880, double antiangiogenic protein (DAAP), and ramucirumab.

[0162] As used herein, “MEK pathway-targeted therapeutic agents” include any compound that exhibits inactivating activity (e.g., kinase inhibition, allosteric inhibition, dimerization inhibition, and degradation induction) of any protein in the MEK pathway, including any protein in the RAS pathway and the RAF pathway. As used herein, “RAS pathway-targeted therapeutic agents” include any compound that exhibits inactivating activity (e.g., kinase inhibition, allosteric inhibition, dimerization inhibition, and degradation induction) of any protein in the RAS pathway. Non-limiting examples of proteins in the RAS pathway include any one of the proteins in the RAS-RAF-MAPK pathway or PI3K / AKT pathway, such as RAS (e.g., KRAS, HRAS, and NRAS), RAF (ARAF, BRAF, CRAF), MEK, ERK, PI3K, AKT, and mTOR. In some embodiments, the RAS pathway modulator may be selective for proteins in the RAS pathway; for example, the RAS pathway modulator may be selective for RAS (also referred to as the RAS modulator). In some embodiments, the RAS modulator is a covalent inhibitor. In some embodiments, the RAS pathway-targeted therapeutic agent is a "KRAS pathway modulator." A KRAS pathway modulator includes any compound that exhibits inactivating activity (e.g., kinase inhibition, allosteric inhibition, dimerization inhibition, and degradation induction) of any protein in the KRAS pathway. Non-limiting examples of proteins in the KRAS pathway include any one of the proteins in the KRAS-RAF-MAPK pathway or the PI3K / AKT pathway, e.g., KRAS, RAF, BRAF, MEK, ERK, PI3K (i.e., other PI3K inhibitors described herein), AKT, and mTOR. In some embodiments, the KRAS pathway modulator may be selective for proteins in the RAS pathway; for example, a KRAS pathway modulator may be selective for KRAS (also referred to as the KRAS modulator). In some embodiments, the KRAS modulator is a covalent inhibitor.

[0163] Non-limiting examples of RAS-targeted therapies include sotrasib (AMG 510, Lumakras®), adaglasib (MRTX849), tipifarnib (R115777, Zarnestra), cismethinil, UCM-1336, delta-lazine, NHTD, RM007, RM008, gefitib, apatinib, onkurasin-1, bismodegib (GDC-0449), N-(1-acryloylazetidine-3-yl)-2-(5-bromo-3-(5-methoxy-1,2,3,4-tetrahydroisoquinoline-2-carbonyl)-1H-indole-1-yl)acetamide, 2-((4-((1-(2-(2,4-dichloro Phenoxy)acetyl)piperidine-4-yl)amino)-4-oxobutyl)disulfane)-N,N-dimethylethane-1-aminium, ARS-1620, ARS-853, vemcentinib (BGB324), ABT-737, selumetinib (AZD6244), dactricib (NVP-BEZ235), PPIN-1, PPIN-2, pan-RAS inhibitor 3144 (RAS-IN-3144), deltalacin, SML-8-73-1, SML-10-70-1, 1-(2-hydroxyethyl)-4-(2-methyl-3,5-diphenylpyrazolo[1,5-a] Pyrimidine-7-yl)piperazine-1-ium, (2R,4aR)-3-acryloyl-11-chloro-9-fluoro-10-(6-fluoro-2-hydroxycyclohexa-2,4-dien-1-yl)-2,6-dimethyl-2,3,4,4a-tetrahydro-1H-pyrazino[1',2':4,5)pyrazino[2,3-c]quinoline-5(6H)-one, NHTD, PD98059, Waltmannin, Tarniflumate, Gefitib, C Examples include PD-0857, KY1022, KYA1797K (ab229170), 0375-0604 (DUN09716), 7773, NSC-658497, JNJ-74699157, PKF115-584 (Carphostine C), Kobe0065, Kobe2602, Salilasib, 3,3'-(ethylazandiyl)bis(N-phenylpropanamide), ML264, GDC-6036, LY3499446, and D-1553.

[0164] Non-exclusive examples of KRAS-targeted therapies (e.g., KRAS inhibitors) include BI 1701963, sotrasib (AMG 510), ARS-3248 (JNJ-74699157), ARS1620, AZD4785 (ION651987), SML-8-73-1, SML-10-70-1, VSA9, AA12, adagrasib (MRTX-849), LY3499446, ARS853, and siG12D LODER.

[0165] A non-limiting example of an HRAS-targeted therapeutic agent (e.g., an HRAS inhibitor) is tipifarnib (ZARNESTRA®). Further non-limiting examples of HRAS-targeted therapeutic agents include BRAF inhibitors, MEK inhibitors, ERK inhibitors, PI3K inhibitors, AKT inhibitors, and mTOR inhibitors. In some embodiments, In some embodiments, the MEK inhibitor is trametinib (MEKINIST®, GSK1120212), cobimetinib (COTELLIC®), binimetinib (MEKTOVI®, MEK162), selumetinib (AZD6244), mildametinib (PD0325901), pimacertib (MSC1936369B), SHR7390, TAK-733, RO5126766 (CH5126766), CS3006, WX-554, PD98059, CI1040 (PD184352), hypothemycin, or a combination thereof.

[0166] In some embodiments, the ERK inhibitor is Ameriolex (FRI-20, ON-01060), VTX-11e, 25-OH-D3-3-BE (B3CD, bromoacetoxycalcidiol), FR-180204, AEZ-131 (AEZS-131), AEZS-136, AZ-13767370, BL-EI-001, Temterquib (LY -3214996), lineterquib (LTT-462), KO-947, MK-8353 (SCH900353), SCH772984, ulixetinib (BVD-523), CC-90003, laboxerutinib (GDC-0994, RG-7482), ASN007, 5Z-7-oxozeaenol (FR148083, L783279, LL-Z 1640-2), 5-iodotubercidin (NSC 113939), ONC201 (TIC10), or combinations thereof.

[0167] In some embodiments, the antiandrogens are leuprolide (LUPRON®, ELIGARD®), goserelin (ZOLDEX®), triptorelin (TRELSTAR®), leuprolide mesylate (CAMCEVI®), flutamide (EULEXIN®), bicalutamide (CASODEX®), nilutamide (NILANDRON®), degarelix (FIRMAGON®), relugolix (ORGOVYX®), enzalutamide (MDV3100, XTANDI®), abiraterone (ZYTIGA®), flutamide (EULEXIN®), AR inhibitor EPI-506, apalutamide (ERLEADA®), and dalotamide (NUBEQA®).

[0168] In some embodiments, the other PI3K inhibitor is another PI3Kα inhibitor. In some embodiments, the other PI3K inhibitor is a pan-PI3K inhibitor. In some embodiments, the other PI3K inhibitor is buparlisib (BKM120), alpelisib (BYL719, PIQRAY®), idelalisib, duvelisib, umbralicib, WX-037, copanlisib (ALIQOPA®, BAY80-6946), dactricib (NVP-BEZ235, BEZ-235), taselicib (GDC-0032, RG7604), sonolicib (PX-866), fimepinostat (CUDC-907), vimiralisib (PQR309), ZSTK474, SF1 126, AZD8835, Inaborishi (GDC-0077), ASN003, Pictilisib (GDC-0941), Pilaralisib (XL147, SAR245408), Gedatricib (PF-05212384, PKI-587), Ceravelisib (TAK-117, MLN1117, INK1117), BGT-226 (NVP-BGT226), PF-04691502, Apitricib (GDC-0980), Omiparisib (GSK2126458, GSK458), Boxtalisib (XL756, SAR245409), AMG Selected from 511, CH5132799, GSK1059615, paxalisib (GDC-0084, RG7666), VS-5584 (SB2343), PKI-402, wartmannin, LY294002, PI-103, ligosertib (ON-01910 sodium salt), boxalisib (XL-765), LY2023414, SAR260301, KIN-193 (AZD-6428), acalisib (GS-9820), AMG319, GSK2636771, or a combination thereof.

[0169] In some embodiments, the AKT inhibitor is miltefosine (IMPADIVO®), wartmannin, NL-71-101, H-89, GSK690693, CCT128930, capivacertib (AZD5363), ipatasertib (GDC-0068, RG7440), A-674563, A-443654, AT7867, AT13148, uprosertib (GSK2141795), afresertib (GSK2110183), DC120, 2-[4-(2-aminoprop-2-yl)phenyl]-3-phenylquinoxali N, MK-2206, Edelfosine, Miltefosine, Perifosine (KRX-0401), Elsylphosphocholine, Elfosine, SR13668, OSU-A9, PH-316, PHT-427 (CS-0223), PIT-1, DM-PIT-1, Trisilibine (Tricilibine phosphate monohydrate), API-1, N-(4-(5-(3-acetamidophenyl)-2-(2-aminopyridine-3-yl)-3H-imidazo[4,5-b]pyridine-3-yl)benzyl)-3-fluorobenzamide, Miransertib (ARQ092), BAY Selected from 1125976, 3-oxo-cylic acid, lactokinomycin, boc-Phe-vinyl ketone, perifosine (D-21266), TCN, TCN-P, ONC201 (TIC10), and TAS117.

[0170] In some embodiments, the AKT inhibitor is capivacertib.

[0171] In some embodiments, the mTOR inhibitor is an analog of rapamycin. Examples of rapamycin analogs include sapatinib (MLN0128), vistusertib (AZD-2014), onatasertib (CC-223), CC-115, everolimus (RAD001), temsirolimus (CCI-779), ridafolimus (AP-23573), sirolimus (rapamycin), ridafolimus (MK-8669), everolimus (RAD001, e.g., AFINITOR® or ZORTRESS®), umirolimus, zotarolimus, and RMC-5552. In some embodiments, the mTOR inhibitor is an ATP-competitive mTOR kinase inhibitor that competes with ATP at the catalytic site of mTOR. Examples of ATP-competitive mTOR kinase inhibitors include torin-1, torin-2, and vistusertib. Types of ATP-competitive mTOR kinase inhibitors can include mTOR / PI3K dual inhibitors and mTORC1 / mTORC2 dual inhibitors (also called TORCdIs). Examples of mTOR / PI3K dual inhibitors include dactolisib, buparlisib, BGT226, SF1126, PKI-587, and NVPBE235. Examples of mTORC1 / mTORC2 dual inhibitors include sapatinib (code name INK128), AZD8055, and AZD2014.

[0172] Non-limiting examples of farnesyltransferase inhibitors include lonafarnib, tipifarnib, BMS-214662, L778123, L7,448,32, and FTI-277.

[0173] In some embodiments, chemotherapeutic agents include anthracyclines, alkylating agents, taxanes, platinum-based drugs, mitomycin, gemcitabine, pemetrexed, eribulin (HALAVEN™), or combinations thereof.

[0174] Non-exclusive examples of taxanes include paclitaxel, docetaxel, abraxane, and taxotere.

[0175] In some embodiments, the anthracycline is selected from daunorubicin, doxorubicin, epirubicin, and idarubicin.

[0176] In some embodiments, the platinum-based drug is selected from carboplatin, cisplatin, oxaliplatin, nedoplatin, triplatin tetranitrate, phenantriplatin, picoplatin, satraplatin, and lovaplatin. Any of the platinum-based drugs can be conjugated to a nanocarrier such as gold nanoclusters, gold nanoparticles, or superparamagnetic iron oxide nanoparticles. See, for example, Zhang et al. 2022. Theranostics. 12(5):2115-2132.

[0177] Non-exclusive examples of PARP inhibitors include olaparib (LYNPARZA®), talazoparib, lucaparib, niraparib, veliparib, BGB-290 (pamiparib), CEP-9883, CEP 9722, E7016 (GPI 21016), iniparib, senaparib (IMP4297), benadaparib-Idience (NOV1401, IDX-1197), stenoparib (2X-121), ABT-767, atamparib (RBN-2397), and talazoparib (BMN Examples include 673), olaparib (KU-0059436, AZD2281, e.g., LYNPARZA®), iniparib (BSI-201, SAR240550), lucaparib (AG-014699, PF-01367338), INO-1001, and amerparib (JPI-289).

[0178] Non-exclusive examples of aromatase inhibitors include aminoglutethimide, testactone, anastrozole, letrozole, exemestane, volozol, formestan, and fadrozol.

[0179] Non-exclusive examples of selective estrogen receptor modulators or degraders (SERMs / SERDs) include clomiphene, cyclophenyl, anoldrin, propalestrol, napoxidine, olmeroxifene, raloxifene, toremifene, rasofoxifene, bazedoxifene, ospemifene, afimoxifene, enclomifene, cerophene, alzoxifene, tamoxifene, etaxtil (GW-5638, DPC974), fulvestrant ( Examples include FASLODEX®, brillanerastine, eracestatin (ORSERDU®), ziledetrant, amsenestrant (SAR439859), camizestrant (AZD9833), lintodestrant, imurunestrant, LSZ102, LY3484356, ZN-c5, taragastrant (D-0502), AZD9496, clotrimazole, fenticonazole, SHR9549, and parazestrant (OP-1250).

[0180] In some embodiments, SERM / SERD is a parazestrant. In some embodiments, SERM / SERD is an erasestrant. In some embodiments, SERM / SERD is a kamizestrant.

[0181] Non-exclusive examples of glucocorticoids include dexamethasone, beclomethasone, betamethasone, budesonide, cortisone, hydrocortisone, methylprednisolone, prednisolone, prednison, and triamcinolone.

[0182] In some embodiments, the glucocorticoid is dexamethasone.

[0183] In some embodiments, the additional therapeutic agent is retinoic acid.

[0184] Non-exclusive examples of epigenetic agents include EZH2 inhibitors (e.g., TAZVERIK®, tazemetostat such as 3-deazanepranosin A (DZNep or C-c3Ado), EPZ005687, EI1, GSK126, and UNC1999) and HDAC inhibitors (e.g., vorinostat (SAHA) and panobinostat (LBH589)). Non-exclusive examples of HDAC inhibitors include hydroxamic acids (or hydroxamates), e.g., trichostatin A, vorinostat (SAHA), bellinostat (PXD101), LAQ824, and panobinostat (LBH589); cyclic tetrapeptides, e.g., trapoxin B and depsipeptide; benzamides, e.g., entinostat (MS-275), tasejinarin (CI994), and mosetinostat (MGCD0103); electrophilic ketones; and fatty acid compounds, e.g., phenylbutyrate and valproic acid.

[0185] In some embodiments, the epigenetic agent is an EZH2 inhibitor. In some embodiments, the epigenetic agent is tazemettostat. In some embodiments, the EZH2 inhibitor is tazemettostat.

[0186] In some embodiments, the epigenetic agent is an HDAC inhibitor. In some embodiments, the epigenetic agent is vorinostat. In some embodiments, the epigenetic agent is panobinostat. In some embodiments, the HDAC inhibitor is vorinostat. In some embodiments, the HDAC inhibitor is panobinostat.

[0187] Non-limiting examples of KAT6A inhibitors include WM-8014, PF-07248144, CTx-648 (PF-9363), and CTX-0124143. In some embodiments, the KAT6A inhibitor is WM-8014. In some embodiments, the KAT6A inhibitor is PF-07248144. In some embodiments, the KAT6A inhibitor is CTx-648. In some embodiments, the KAT6A inhibitor is CTX-0124143.

[0188] Non-exclusive examples of immunotherapy include immune checkpoint therapy, atezolizumab (TECENTRIQ®), and albumin-conjugated paclitaxel. Non-exclusive examples of immune checkpoint therapy include inhibitors targeting CTLA-4, PD-1, PD-L1, BTLA, LAG-3, A2AR, TIM-3, B7-H3, VISTA, IDO, and combinations thereof. In some embodiments, the CTLA-4 inhibitor is ipilimumab (YERVOY®). In some embodiments, the PD-1 inhibitor is nivolumab OPDIVO®, pembrolizumab KEYTRUDA®, semiprimab LIBTAYO®, atezolizumab TECENTRIQ®, durvalumab IMFINZI®, avelumab BAVENCIO®, dostallimab (JEMPERLI®), retifanlimab (ZYNYZ®), voplaterimab (JTX-4014), spartalizumab (PDR001), camrelizumab (SHR1210), cintilimab (IBI308), tislerizumab (BGB-A317), tripalimab (JS The inhibitor is selected from 001), INCMGA00012 (MGA012), AMP-224, AMP-514 (MEDI0680), or acrixolimab (YBL-006). In some embodiments, the PD-1 inhibitor is selected from pembrolizumab (KEYTRUDA®), nivolumab (OPDIVO®), semiprimab (LIBTAYO®), or a combination thereof. In some embodiments, the PD-L1 inhibitor is selected from atezolizumab (TECENTRIQ®), avelumab (BAVENCIO®), durvalumab (IMFINZI®), or a combination thereof. In some embodiments, the LAG-3 inhibitor is relamirimab (IMP701, LAG525). In some embodiments, the A2AR inhibitor is siforadenanth (CPI-444). In some embodiments, the TIM-3 inhibitor is sabatrimab (MBG453). In some embodiments, the B7-H3 inhibitor is enobrituzumab.In some embodiments, the VISTA inhibitor is ombatirimab (JNJ-61610588). In some embodiments, the IDO inhibitor is indoximod. See, for example, Table 1 of Marin-Acevedo, et al., J Hematol Oncol. 11:39 (2018), which is incorporated herein by reference in its entirety.

[0189] In some embodiments, the CDK4 / 6 inhibitor is palbociclib (IBRANCE®, TQB3616, PD-0332991), ribociclib (KISQALI®), abemaciclib (VERZENIO®), boriclib (P1446A-05), trilaciclib, darpicilib (SHR6390), roniciclib (BAY1000394), dinacilib, flavopyridol (arbocidib, L868275, HMR-1275), roscovitine (R-roscovitine, CYC202, sericiclib), ribicib (P276-00, P276), AT7519, TG02 (SB1317), RGB-286638, dinacilib (SCH Selected from 727965), PHA-793887, ZK-304709, xytosidine, SNS032 (BMS-387032), R547 (R04584820), RGB286147, Pulvaranol A (NG60), Meliolin 3, JNJ7706621, Indirubin, AZD-5438, 10Z-Himennialdisine, AGO24322, PF-06873600, and KIN-8741.

[0190] In some embodiments, the CDK4 / 6 inhibitor is KIN-8741. In some embodiments, the CDK4 / 6 inhibitor is palbociclib. In some embodiments, the CDK4 / 6 inhibitor is ribociclib. In some embodiments, the CDK4 / 6 inhibitor is trilaciclib. In some embodiments, the CDK4 / 6 inhibitor is darupiciclib. In some embodiments, the CDK4 / 6 inhibitor is palbociclib. In some embodiments, the CDK4 / 6 inhibitor is roniciclib. In some embodiments, the CDK4 / 6 inhibitor is dinacilib A.

[0191] In some forms, additional therapies or medications include fulvestrant, capecitabine, trastuzumab, ado-trastuzumab emtansine, pertuzumab, paclitaxel, nab-paclitaxel, enzalutamide, olaparib, pegylated liposomal doxorubicin (PLD), trametinib, palbociclib (IBRANCE®), buparlisib, sotrastaurin (AEB071), everolimus, exemestane, cisplatin, letrozole, and ganitumab (AMG). The following are selected from 479), LSZ102, ribociclib (LEE011), cetuximab, luminespib (NVP-AUY922, AUY922), infigalatinib (BGJ398), binimetinib (MEK162, ARRY-162, ARRY-438162), LJM716, PIM447 (LGH447, LGB321), imatinib, gemcitabine, encorafenib (LGX818), and amsenestrant.

[0192] In some embodiments, the additional therapeutic agent is everolimus. In some embodiments, additional therapeutic agents, such as opioids and corticosteroids, may be administered to treat potential side effects of certain anticancer therapies and / or as palliative care. In some embodiments, the additional therapies or therapeutic agents described herein are selected from the group consisting of glucagon-like peptide-1 (GLP-1) receptor agonists, sodium-glucose transporter protein 2 (SGLT-2) inhibitors, dipeptidyl peptidase 4 (DPP-4) inhibitors, metformin, and combinations thereof.

[0193] Non-exclusive examples of GLP-1 receptor agonists include liraglutide NN2211 (e.g., VICTOZA®), dulaglutide (LY2189265, e.g., TRULICITY®), exenatide (e.g., BYETTA®, BYDUREON®, exendin-4), taspoglutide, lixisenatide (e.g., LYXUMIA®), albiglutide (e.g., TANZEUM®), semaglutide (e.g., OZEMPIC®, RYBELSUS®), ZP2929, NNC0113-0987 (QBR110395), BPI-3016, and TT401.

[0194] Non-exclusive examples of SGLT-2 inhibitors include bexagliflozin, canagliflozin (e.g., INVOKANA®), dapagliflozin (e.g., FARXIGA®), empagliflozin (e.g., JARDIANCE®), ertugliflozin (e.g., STEGLATRO®), ipragliflozin (e.g., SUGLAT®), luseogliflozin (e.g., LUSEFI®), remogliflozin, selfliflozin, lycofliglozin, sotagliflozin (e.g., ZYNQUISTA®), and tofogliflozin.

[0195] Non-exclusive examples of DPP-4 inhibitors include sitagliptin (e.g., JANUVIA®), vildagliptin, saxagliptin (e.g., ONGLYZA®), linagliptin (e.g., TRADJENDA®), gemigliptin, anagliptin, teneligliptin, alogliptin, trelagliptin (e.g., NESINA®), omaligliptin, evogliptin, and dutogliptin.

[0196] In some embodiments, the additional therapeutic agent is metformin. In some embodiments, the method described herein further comprises administering a therapeutically effective dose of metformin to the subject.

[0197] In some embodiments, the method involves compound 1 or a pharmaceutically acceptable salt thereof, and one additional therapeutic agent, such as an aromatase inhibitor, a CDK4 / 6 inhibitor, a SERM / SERD, radiotherapy, an anti-HER2 antibody or its antibody-drug conjugate (ADC), immunotherapy, or a checkpoint inhibitor (e.g., an anti-PD-1 or PD-L1 antibody, an anti-CLTA). Therapies include administering 4 antibodies, VEGFR inhibitors / VEGF inhibitors, KAT6A inhibitors (also known as MOZ inhibitors or MYST3 inhibitors), PI3Kα inhibitors, MEK pathway targeting therapies (including RAS pathway targeting therapies, including mTOR inhibitors as described herein), SHP2 inhibitors, ULK inhibitors, NTRK / ROS inhibitors, ALK inhibitors, RET inhibitors, MET inhibitors, PARP inhibitors, PIM (e.g., PIM1 and PIM3) inhibitors, other kinase inhibitors (e.g., Trk inhibitors or multi-kinase inhibitors), farnesyltransferase inhibitors, vinca alkaloids, antimetabolites, antiandrogens, alkylating agents, checkpoint inhibitors, apoptotic pathway modulators; cytotoxic chemotherapeutic agents, angiogenesis-targeted therapies, immunotherapy-targeted agents, or anti-EGFR antibodies.

[0198] In some embodiments, the additional therapeutic agent is an antibody or ADC described herein. In some embodiments, the antibody is daratumumab (e.g., DARZALEX®).

[0199] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with one additional therapeutic agent, such as a HER2 inhibitor, SERM / SERD, CDK4 / 6 inhibitor, MEK inhibitor, checkpoint inhibitor (e.g., anti-PD-1 or PD-L1 antibody, anti-CLTA 4 antibody), multi-kinase inhibitor, and PI3K inhibitor.

[0200] In some embodiments, the method involves compound 1 or a pharmaceutically acceptable salt thereof and one additional therapeutic agent, such as trastuzumab, pertuzumab, trastuzumab emtansine, famtrastuzumab deruxtecan, lapatinib, neratinib, dacomitinib, afatinib, tucatinib, erlotinib, pirotinib, tanespimycin, dacomitinib, peritinib, margetuximab, clomiphene, cyclophenyl, propalestrol, olmeroxifene, raloxifene, toremifene, rasofoxifene, bazedoxifene, ospemifene, enclomifene, cerophene, tamoxifene, f This includes administering rubestrant, elastrant, camizestrant, lintodestrant, clotrimazole, fenticonazole, nivolumab, pembrolizumab, semiprimab, atezolizumab, durvalumab, avelumab, ipilimumab, palbociclib, ribociclib, abemaciclib, trilaciclib, darpiciclib, trametinib, cobimetinib, binimetinib, selumetinib, mildametinib, pimacertib, alpelisib, idelalisib, duvelisib, copanlisib, umbralicib, ribociclib, trilaciclib, darpiciclib, boriclib, and roniciclib.

[0201] In some embodiments, the additional therapeutic agent is fulvestrant.

[0202] In some embodiments, an additional therapeutic agent is lapatinib.

[0203] In some embodiments, the additional therapeutic agent is abemaciclib.

[0204] In some embodiments, an additional therapeutic agent is trametinib.

[0205] In some embodiments, an additional therapeutic agent is binimetinib.

[0206] In some embodiments, the additional therapeutic agent is alpelisib.

[0207] In some embodiments, the additional therapeutic agent is palbociclib.

[0208] In some embodiments, the additional therapeutic agent is ribociclib.

[0209] In some embodiments, the additional therapeutic agent is trilasiclib.

[0210] In some embodiments, the additional therapeutic agent is darpiciclib.

[0211] In some embodiments, the additional therapeutic agent is borsicrib.

[0212] In some embodiments, the additional therapeutic agent is roniciclib.

[0213] In some embodiments, the additional therapeutic agent is dinaciclib.

[0214] In some embodiments, additional therapeutic agents are palbociclib and clomiphene.

[0215] In some embodiments, additional therapeutic agents are palbociclib and cyclofenil.

[0216] In some embodiments, additional therapeutic agents are palbociclib and anodrin.

[0217] In some embodiments, additional therapeutic agents are palbociclib and propalestrol.

[0218] In some embodiments, additional therapeutic agents are palbociclib and napoxidine.

[0219] In some embodiments, additional therapeutic agents are palbociclib and olmeroxifen.

[0220] In some embodiments, additional therapeutic agents are palbociclib and raloxifene.

[0221] In some embodiments, additional therapeutic agents are palbociclib and toremifene.

[0222] In some embodiments, additional therapeutic agents are palbociclib and rasofoxifen.

[0223] In some embodiments, additional therapeutic agents are palbociclib and bazedoxifene.

[0224] In some embodiments, additional therapeutic agents are palbociclib and ospemifene.

[0225] In some embodiments, additional therapeutic agents are palbociclib and afimoxifen.

[0226] In some embodiments, additional therapeutic agents are palbociclib and enclomifene.

[0227] In some embodiments, additional therapeutic agents are palbociclib and cerophene.

[0228] In some embodiments, additional therapeutic agents are palbociclib and alzoxifen.

[0229] In some embodiments, additional therapeutic agents are palbociclib and tamoxifen.

[0230] In some embodiments, additional therapeutic agents are palbociclib and etactyl.

[0231] In some embodiments, additional therapeutic agents are palbociclib and fulvestrant.

[0232] In some embodiments, additional therapeutic agents are palbociclib and brillanestran.

[0233] In some embodiments, additional therapeutic agents include palbociclib and elastrant.

[0234] In some embodiments, additional therapeutic agents include palbociclib and ziledetrant.

[0235] In some embodiments, additional therapeutic agents include palbociclib and amsenestrant.

[0236] In some embodiments, additional therapeutic agents are palbociclib and camizestrant.

[0237] In some embodiments, additional therapeutic agents include palbociclib and lintodestrant.

[0238] In some embodiments, additional therapeutic agents are palbociclib and imurunestrant.

[0239] In some embodiments, additional therapeutic agents include palbociclib and LSZ102.

[0240] In some embodiments, additional therapeutic agents include palbociclib and LY3484356.

[0241] In some embodiments, additional therapeutic agents are palbociclib and ZN-c5.

[0242] In some embodiments, additional therapeutic agents include palbociclib and taragastrant.

[0243] In some embodiments, additional therapeutic agents are palbociclib and AZD9496.

[0244] In some embodiments, additional therapeutic agents are palbociclib and clotrimazole.

[0245] In some embodiments, additional therapeutic agents are palbociclib and fenticonazole.

[0246] In some embodiments, additional therapeutic agents include palbociclib and SHR9549.

[0247] In some embodiments, additional therapeutic agents are palbociclib and parazestrant.

[0248] In some embodiments, additional therapeutic agents are ribociclib and clomiphene.

[0249] In some embodiments, additional therapeutic agents are ribociclib and cyclofenil.

[0250] In some embodiments, additional therapeutic agents are ribociclib and anodrine.

[0251] In some embodiments, additional therapeutic agents are ribociclib and propalestrol.

[0252] In some embodiments, additional therapeutic agents are ribociclib and napoxidine.

[0253] In some embodiments, additional therapeutic agents are ribociclib and olmeloxifen.

[0254] In some embodiments, additional therapeutic agents are ribociclib and raloxifene.

[0255] In some embodiments, additional therapeutic agents include ribociclib and toremifene.

[0256] In some embodiments, additional therapeutic agents are ribociclib and rasofoxifen.

[0257] In some embodiments, additional therapeutic agents are ribociclib and bazedoxifene.

[0258] In some embodiments, additional therapeutic agents are ribociclib and ospemifene.

[0259] In some embodiments, additional therapeutic agents are ribociclib and afimoxifen.

[0260] In some embodiments, additional therapeutic agents are ribociclib and enclomifene.

[0261] In some embodiments, additional therapeutic agents are ribociclib and cerophene.

[0262] In some embodiments, additional therapeutic agents include ribociclib and arzoxifen.

[0263] In some embodiments, additional therapeutic agents are ribociclib and tamoxifen.

[0264] In some embodiments, additional therapeutic agents are ribociclib and etactyl.

[0265] In some embodiments, additional therapeutic agents are ribociclib and fulvestrant.

[0266] In some embodiments, additional therapeutic agents are ribociclib and brillanestrane.

[0267] In some embodiments, additional therapeutic agents are ribociclib and elastorant.

[0268] In some embodiments, additional therapeutic agents include ribociclib and ziledetrant.

[0269] In some embodiments, additional therapeutic agents include ribociclib and amsenestrant.

[0270] In some embodiments, additional therapeutic agents are ribociclib and camizestrant.

[0271] In some embodiments, additional therapeutic agents are ribociclib and lintodestrant.

[0272] In some embodiments, additional therapeutic agents are ribociclib and imurunestrant.

[0273] In some embodiments, additional therapeutic agents include ribociclib and LSZ102.

[0274] In some embodiments, additional therapeutic agents include ribociclib and LY3484356.

[0275] In some embodiments, additional therapeutic agents are ribociclib and ZN-c5.

[0276] In some embodiments, additional therapeutic agents include ribociclib and taragastrant.

[0277] In some embodiments, additional therapeutic agents include ribociclib and AZD9496.

[0278] In some embodiments, additional therapeutic agents are ribociclib and clotrimazole.

[0279] In some embodiments, additional therapeutic agents are ribociclib and fenticonazole.

[0280] In some embodiments, additional therapeutic agents include ribociclib and SHR9549.

[0281] In some embodiments, additional therapeutic agents are ribociclib and parazestrant.

[0282] In some embodiments, additional therapeutic agents are abemaciclib and clomiphene.

[0283] In some embodiments, additional therapeutic agents are abemaciclib and cyclofenil.

[0284] In some embodiments, additional therapeutic agents are abemaciclib and anoldrin.

[0285] In some embodiments, additional therapeutic agents are abemaciclib and proparestrol.

[0286] In some embodiments, additional therapeutic agents are abemaciclib and napoxidine.

[0287] In some embodiments, additional therapeutic agents are abemaciclib and olmeroxifen.

[0288] In some embodiments, additional therapeutic agents are abemaciclib and raloxifene.

[0289] In some embodiments, additional therapeutic agents are abemaciclib and toremifene.

[0290] In some embodiments, additional therapeutic agents are abemaciclib and rasofoxifene.

[0291] In some embodiments, additional therapeutic agents are abemaciclib and bazedoxifene.

[0292] In some embodiments, additional therapeutic agents are abemaciclib and ospemifene.

[0293] In some embodiments, additional therapeutic agents are abemaciclib and afimoxifen.

[0294] In some embodiments, additional therapeutic agents are abemaciclib and enclomifene.

[0295] In some embodiments, additional therapeutic agents are abemaciclib and cerophene.

[0296] In some embodiments, additional therapeutic agents include abemaciclib and alzoxifen.

[0297] In some embodiments, additional therapeutic agents include abemaciclib and tamoxifen.

[0298] In some embodiments, additional therapeutic agents are abemaciclib and etactyl.

[0299] In some embodiments, additional therapeutic agents are abemaciclib and fulvestrant.

[0300] In some embodiments, additional therapeutic agents are abemaciclib and brillanestrant.

[0301] In some embodiments, additional therapeutic agents are abemaciclib and elastorant.

[0302] In some embodiments, additional therapeutic agents include abemacicli and ziledetrant.

[0303] In some embodiments, additional therapeutic agents include abemaciclib and amsenestrant.

[0304] In some embodiments, additional therapeutic agents are abemaciclib and camizestrant.

[0305] In some embodiments, additional therapeutic agents are abemaciclib and lintodestrant.

[0306] In some embodiments, additional therapeutic agents are abemaciclib and imurunestrant.

[0307] In some embodiments, additional therapeutic agents include abemaciclib and LSZ102.

[0308] In some embodiments, additional therapeutic agents include abemaciclib and LY3484356.

[0309] In some embodiments, additional therapeutic agents are abemaciclib and ZN-c5.

[0310] In some embodiments, additional therapeutic agents include abemaciclib and taragastrant.

[0311] In some embodiments, additional therapeutic agents are abemaciclib and AZD9496.

[0312] In some embodiments, additional therapeutic agents are abemaciclib and clotrimazole.

[0313] In some embodiments, additional therapeutic agents are abemaciclib and fenticonazole.

[0314] In some embodiments, additional therapeutic agents include abemaciclib and SHR9549.

[0315] In some embodiments, additional therapeutic agents are abemaciclib and parazestrant.

[0316] The method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with two additional independently selected therapeutic agents, such as a HER2 inhibitor, SERM / SERD, CDK4 / 6 inhibitor, MEK inhibitor, checkpoint inhibitor (e.g., anti-PD-1 or PD-L1 antibody, anti-CLTA 4 antibody), multikinase inhibitor, and PI3K inhibitor.

[0317] In some embodiments, the method involves compound 1 or a pharmaceutically acceptable salt thereof, and two additional independently selected therapeutic agents, such as trastuzumab, pertuzumab, trastuzumab emtansine, famtrastuzumab deruxtecan, lapatinib, neratinib, dacomitinib, afatinib, tucatinib, erlotinib, pirotinib, tanespimycin, dacomitinib, peritinib, margetuximab, clomiphene, cyclophenyl, propalestrol, olmeroxifene, raloxifene, toremifene, rasofoxifene, bazedoxifene, ospemifene, enclo This includes administering miphene, cerophene, tamoxifen, fulvestrant, esestrant, camizestrant, lintodestrant, clotrimazole, fenticonazole nivolumab, pembrolizumab, trastuzumab emtansine, atezolizumab, durvalumab, avelumab, ipilimumab, palbociclib, ribociclib, abemaciclib, trilaciclib, darpiciclib, trametinib, cobimetinib, binimetinib, selumetinib, mildametinib, pimacertib, alpelisib, idelalisib, duvelisib, copanlisib, and umbralisib.

[0318] In some embodiments, additional therapeutic agents are fulvestrant and lapatinib.

[0319] In some embodiments, additional therapeutic agents are fulvestrant and trametinib.

[0320] In some embodiments, additional therapeutic agents are fulvestrant and binimetinib.

[0321] In some embodiments, additional therapeutic agents are fulvestrant and alpelicib.

[0322] In some embodiments, additional therapeutic agents include fulvestrant and trilasiclib.

[0323] In some embodiments, additional therapeutic agents are fulvestrant and darpiciclib.

[0324] In some embodiments, additional therapeutic agents are fulvestrant and palbociclib.

[0325] In some embodiments, additional therapeutic agents include fulvestrant and roniclib.

[0326] In some embodiments, additional therapeutic agents include fulvestrant and dinacyclib.

[0327] In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with an ERα inhibitor or degrading agent (e.g., SERM or SERD).

[0328] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with a CDK4 / 6 inhibitor.

[0329] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with a HER2 inhibitor.

[0330] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with alpelisib.

[0331] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof with an ERα inhibitor or a degrading agent.

[0332] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof with a CDK4 / 6 inhibitor.

[0333] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof and a HER2 inhibitor.

[0334] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof with an alpelisib.

[0335] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof, an ERα inhibitor or degrading agent, and a CDK4 / 6 inhibitor.

[0336] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof, fulvestrant, and a CDK4 / 6 inhibitor.

[0337] In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof, fulvestrant, and palbociclib. In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof, fulvestrant, and ribociclib. In some embodiments, the method essentially consists of administering compound 1 or a pharmaceutically acceptable salt thereof, fulvestrant, and abemaciclib.

[0338] In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with fulvestrant. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with fulvestrant, where fulvestrant is administered in doses ranging from about 250 mg to about 500 mg. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with fulvestrant, where fulvestrant is administered in doses ranging from about 250 mg to about 500 mg. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with fulvestrant, where fulvestrant is administered in two 5 mL injections on days 1, 15, 29 and once monthly thereafter, with each 5 mL injection containing 250 mg of fulvestrant.

[0339] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with lapatinib. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with lapatinib, where lapatinib is administered in doses ranging from about 1250 mg to about 1500 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with lapatinib, where lapatinib is administered in doses ranging from about 1250 mg to about 1500 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with lapatinib, where lapatinib is administered in the form of five tablets once daily, each tablet containing 250 mg of lapatinib. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with lapatinib, where lapatinib is administered once daily in the form of six tablets, each tablet containing 250 mg of lapatinib.

[0340] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered in a dose ranging from about 150 mg to about 400 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered in a dose of about 150 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered in a dose of about 200 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered in a dose of about 300 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered in a dose of approximately 400 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where abemaciclib is administered as a tablet of 50 mg, 100 mg, 150 mg, or 200 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where 150 mg of abemaciclib is administered as a tablet twice daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with abemaciclib, where 200 mg of abemaciclib is administered as a tablet twice daily.

[0341] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with a MEK inhibitor.

[0342] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered in a dose ranging from about 1 mg to about 2 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered in a dose of about 1 mg if the subject is a pediatric patient weighing about 26 kg to about 37 kg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered in a dose of about 1.5 mg if the subject is a pediatric patient weighing about 38 kg to about 50 kg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered in a dose of about 2 mg if the subject is a pediatric patient weighing about 51 kg or more. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered in a dose of about 2 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where trametinib is administered as a 0.5 mg or 2 mg tablet. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where 2 mg of trametinib is administered once daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, where 1.5 mg of trametinib is administered once daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with trametinib, wherein 1 mg of trametinib is administered once daily.

[0343] In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered in a dose ranging from about 30 mg to about 90 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered in a dose of about 30 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered in a dose of about 45 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered in a dose of about 60 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered in a dose of about 90 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where binimetinib is administered as a 15 mg tablet. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where 45 mg of binimetinib is administered as three 15 mg tablets twice daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with binimetinib, where 30 mg of binimetinib is administered as two 15 mg tablets twice daily.

[0344] In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where palbociclib is administered in a dose ranging from about 75 mg to about 125 mg. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where palbociclib is administered in a dose of about 75 mg. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where palbociclib is administered in a dose of about 100 mg. In some embodiments, the method involves administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where palbociclib is administered in a dose of about 125 mg. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where palbociclib is administered as a 75 mg, 100 mg, or 125 mg tablet. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where 75 mg of palbociclib is administered once daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where 100 mg of palbociclib is administered once daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where 125 mg of palbociclib is administered once daily. In some embodiments, the method comprises administering compound 1 or a pharmaceutically acceptable salt thereof with palbociclib, where 125 mg of palbociclib is administered once daily as a 125 mg tablet.

[0345] Pharmaceutical composition Some embodiments include a compound of formula (I) or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable excipients.

[0346] Some embodiments provide a pharmaceutical composition comprising compound 1 or a pharmaceutically acceptable salt thereof and one or more pharmaceutically acceptable excipients.

[0347] In some embodiments, the pharmaceutical composition includes one or two additional therapeutic agents, i.e., as a combination of fixed doses. In some embodiments, the pharmaceutical composition includes one additional therapeutic agent. In some embodiments, the pharmaceutical composition includes two additional therapeutic agents.

[0348] Administration In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered in doses ranging from about 1 mg to about 500 mg, for example, about 1 mg to about 450 mg, about 1 mg to about 400 mg, about 1 mg to about 350 mg, about 1 mg to about 300 mg, about 1 mg to about 250 mg, about 1 mg to about 200 mg, about 1 mg to about 150 mg, about 1 mg to about 100 mg, about 1 mg to about 50 mg, about 1 mg to about 40 mg, about 1 mg to about 30 mg, about 1 mg to about 25 mg, about 1 mg to about 20 mg, about 1 mg to about 15 mg, about 1 mg to about 10 mg, or about 1 mg to about 5 mg.

[0349] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered in doses ranging from about 1 mg to about 100 mg, for example, about 1 mg to about 80 mg, about 1 mg to about 75 mg, about 1 mg to about 60 mg, about 1 mg to about 50 mg, about 1 mg to about 40 mg, about 1 mg to about 30 mg, or about 1 mg to about 20 mg.

[0350] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered in doses ranging from about 20 mg to about 100 mg, for example, about 20 mg to about 80 mg, about 30 mg to about 80 mg, about 30 mg to about 70 mg, about 30 mg to about 60 mg, about 20 mg to about 50 mg, about 40 mg to about 70 mg, about 40 mg to about 60 mg, or about 45 mg to about 55 mg.

[0351] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered in doses ranging from about 1 mg to about 50 mg, for example, about 1 mg to about 45 mg, about 1 mg to about 40 mg, about 1 mg to about 35 mg, about 1 mg to about 30 mg, about 1 mg to about 25 mg, about 1 mg to about 20 mg, about 1 mg to about 15 mg, about 1 mg to about 10 mg, or about 1 mg to about 5 mg.

[0352] In some embodiments, the total daily dose of compound 1 or a pharmaceutically acceptable salt thereof is as described herein. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered once daily. In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered twice daily.

[0353] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered as monotherapy in the doses described herein.

[0354] In some embodiments, compound 1 or a pharmaceutically acceptable salt thereof is administered in combination with one or more additional therapeutic agents described herein, in the dosage of compound 1 described herein. In some embodiments, the dosage of one or more additional therapeutic agents is the standard dosage of one or more additional therapeutic agents, for example, as indicated on a regulatory authority-approved label (e.g., USFDA or EMA). [Examples]

[0355] Preparation of compounds Compound 1: (R)-1-(2-aminopyrimidine-5-yl)-3-(1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethyl)urea

[0356] [ka]

[0357] Step 1 Phenylenol-2,5-diamine (3.0 g, 27.2 mmol) and NaHCO3 (11.4 g, 135.9 mmol) were added dropwise to a mixture in THF (300 mL) at 0°C with phenylcarbonochloride (4.5 g, 28.5 mmol). The mixture was stirred at 25°C for 4 hours. After the reaction, the reaction mixture was concentrated to obtain the residue, which was purified by silica gel chromatography column (0-10% DCM / MeOH) to obtain phenyl(2-aminopyrimidine-5-yl)carbamate (2.2 g, 34%) as a brown solid. MS(ESI):C 11 H 10 The calculated mass of N4O2 is 230.1, and the measured m / z value is 231.1 [M+H]. + .

[0358] Step 2 To a solution of 1-(3,5-difluoro-2-hydroxyphenyl)ethane-1-one (20 g, 116.2 mmol) in DMF (200 mL), methyl 2-bromoacetate (19.4 g, 127.9 mmol) and K2CO3 (24.1 g, 174.4 mmol) were added at room temperature. The reaction mixture was stirred at room temperature for 2 hours. After the reaction, the insoluble material was filtered off, and DBU (17.7 g, 116.2 mmol) was added to the filtrate. This mixture was stirred again at 80°C for 2 hours. After the reaction, the reaction mixture was concentrated to obtain a residue, which was purified by silica gel chromatography column (0-35% PE / DCM) to obtain methyl 5,7-difluoro-3-methylbenzofuran-2-carboxylate (8.4 g, 32%) as a grayish-white solid. MS(ESI):C 11 The calculated mass of H8F2O3 is 226.0, and the measured m / z value is 227.0 [M+H]. + .

[0359] Step 3 To a solution of 5,7-difluoro-3-methylbenzofuran-2-carboxylate (8.1 g, 35.8 mmol) in THF (160 mL), LiAlH4 (21.5 mL, 21.5 mmol, 1 M in THF) was added at 0°C. The reaction mixture was stirred at 0°C for 2 hours, then slowly quenched with saturated potassium carbonate aqueous solution (150 mL), and extracted with EA (200 mL x 3). The combined organic layers were washed with brine, dried over Na2SO4, and concentrated to obtain a residue. This residue was purified by silica gel chromatography column (0-30% PE / EA) to obtain (5,7-difluoro-3-methylbenzofuran-2-yl)methanol (6.5 g, 91%) as a grayish-white solid. MS(ESI):C 10 The calculated mass of H8F2O2 is 198.0, and the measured m / z value is 181.0 [M-H2O+H]. + .

[0360] Step 4 To a solution of (5,7-difluoro-3-methylbenzofuran-2-yl)methanol (6.5 g, 32.6 mmol) in ACN (65 mL), IBX (13.7 g, 49.0 mmol) was added at room temperature. The reaction mixture was stirred at 80 °C for 2 hours. The insoluble material was then filtered off, and the filtrate was concentrated to obtain the residue. This residue was purified by silica gel chromatography column (0-15% PE / EA) to obtain 5,7-difluoro-3-methylbenzofuran-2-carbaldehyde (6.1 g, 95%) as a grayish-white solid. MS(ESI):C 10 The calculated mass of H6F2O2 is 196.0, and the measured m / z value is 197.1 [M+H]. + .

[0361] Step 5 To a solution of 5,7-difluoro-3-methylbenzofuran-2-carbaldehyde (6.1 g, 31.1 mmol) in DMF (92 mL), trimethyl(trifluoromethyl)silane (8.89 g, 62.2 mmol) and K2CO3 (2.1 g, 15.5 mmol) were added at 0°C. The reaction mixture was stirred at room temperature for 0.5 hours, and then K2CO3 (4.3 g, 31.0 mmol) from another batch was added to the reaction mixture. The mixture was stirred at room temperature for 16 hours, and then H2O (2.8 g, 115.5 mmol) was added, and the reaction mixture was further stirred at 0°C for 1 hour. The mixture was quenched with ice water and extracted with EA (200 mL x 3). The combined organic layers were washed with brine, dried over Na2SO4, and concentrated to obtain the residue, which was purified by silica gel chromatography column (0-20% PE / EA) to obtain 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-ol (6.5 g, 78%) as a bright yellow oil. MS(ESI):C 11 Calculated mass of H7F5O2: 266.0, measured m / z value: 249.1 [M-H2O+H] + .

[0362] Step 6 IBX (584 mg, 2.08 mmol) was added to a solution of 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-ol (370 mg, 1.39 mmol) in ACN (10 mL). The reaction mixture was refluxed for 16 hours. After the reaction, the mixture was filtered and washed with EA. The filtrate was collected and concentrated to obtain the residue, which was purified by silica gel chromatography column (0-20% PE / EA) to obtain 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-one (320 mg, 87%) as a yellow oil. 1 H NMR (400MHz, DMSO) δ 7.82-7.74 (m, 2H), 2.65 (s, 3H).

[0363] Step 7 A mixture of 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-one (320 mg, 1.21 mmol), hydroxylamine hydrochloride (585 mg, 8.48 mmol), and NaOAc (992 mg, 12.10 mmol) in EtOH (10 mL) was refluxed for 16 hours. After the reaction, the mixture was concentrated and redissolved in MeOH (10 mL), to which Raney Ni (50 mg) and 1 drop of ammonia were added. The mixture was stirred under H2 at room temperature for 6 hours. After the reaction, the mixture was filtered, and the filtrate was concentrated to obtain the residue, which was purified by silica gel chromatography column (0-50% PE / EA) to obtain 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-amine (150 mg, 47%) as a pale yellow oil. MS(ESI):C 11 The calculated mass of H8F5NO is 265.0, and the measured m / z value is 249.1 [M-NH3+H]. + .

[0364] Step 8 A mixture of 1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethane-1-amine (150 mg, 0.57 mmol) and DIEA (219 mg, 1.70 mmol) in DMF (10 mL) was mixed with phenyl(2-aminopyrimidine-5-yl)carbamate (143 mg, 0.62 mmol) at 0°C. The reaction mixture was then stirred at room temperature for 16 hours, diluted with water, and extracted with EA (50 mL x 3). The combined organic layer was concentrated to obtain the residue, which was purified by silica gel chromatography column (0-50% PE / EA) to obtain (rac)-1-(2-aminopyrimidine-5-yl)-3-(1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethyl)urea (150 mg, 66%) as a pale yellow oil. MS(ESI):C 16 H 12 The calculated mass of F5N5O2 is 401.1, and the measured mass in m / z is 402.1 [M+H]. + .

[0365] Step 9 (rac) 1-(2-Aminopyrimidin-5-yl)-3-(1-(5,7-difluoro-3-methylbenzofuran-2-yl)-2,2,2-trifluoroethyl)urea (150 mg) was separated by chiral HPLC to obtain the (S)-enantiomer of Compound 1 (Peak 1, 61 mg, 41%) and Compound 1 (Peak 2, 58 mg, 39%). MS (ESI): C 16 H 12 Calculated mass of C15H11F5N5O2 401.1, measured m / z 402.1 [M+H] + .

[0366] (S)-Enantiomer of Compound 1 (Peak 1): 1 H NMR (400 MHz, DMSO-d6) δ 8.21 (s, 2H), 8.20 (s, 1H), 7.81 (d, J = 8.0 Hz, 1H), 7.46 - 7.39 (m, 2H), 6.40 (s, 2H), 6.09 - 5.98 (m, 1H), 2.29 (s, 3H).

[0367] Compound 1 (Peak 2): 1 H NMR (400 MHz, DMSO-d6) δ 8.19 (s, 2H), 8.18 (s, 1H), 7.79 (d, J = 8.0 Hz, 1H), 7.46 - 7.34 (m, 2H), 6.37 (s, 2H), 6.03 - 5.97 (m, 1H), 2.27 (s, 3H).

[0368] Assay Homogenous Time-Resolved Fluorescence (HTRF)-pAKT-T47D Compound 1 was assayed using Homogenous Time-Resolved Fluorescence (HTRF).

[0369] Materials, Reagents, and Equipment Gibco RPMI 1640 medium, without phenol red; Gibco RPMI 1640 medium; Gibco trypsin-EDTA (0.5%), without phenol red; Gibco DPBS; Trypan blue solution 0.4% (Corning); Avantor Seradigm Premium Grade fetal bovine serum (FBS); Greiner 784080-384 well TC treated white plate; pAKT(Ser473)HTRF; Gibco Insulin, human recombinant, zinc soluble; Gibco Recovery Cell Culture Freezing Medium; Countess II FL automated cell counter (ThermoFisher); Countess II Slides (ThermoFisher); Microscope; and PHERAstar FSX microplate reader (BMG LABTECH, Inc.).

[0370] procedure The scinamic cell line ID was T47D.1, HTRF detection was performed using pAKT(S473), and the PI3Kα H1047R mutation was present. The seeding density was 5000 cells, the time point was 1 hour, and the culture medium used was RPMI + 10% FBS (without phenol red) + 0.2 units / ml bovine insulin.

[0371] Cell culture maintenance: ●Cell density was not allowed to reach 100% confluence. When the cells reached approximately 80% confluence, they were divided into a 1:5 ratio. ○The cells were divided twice a week (Monday and Friday). ○Cells exceeding passage 18 were not used (maintenance for approximately 2 months). Antibiotics were not used in tissue culture maintenance or assays.

[0372] To freeze cells: 1. Trypsin-treated cells were collected and counted. The cells were pelletized at 1000 rpm for 5 minutes, and the supernatant was aspirated. 2. The pelleted cells were gently resuspended in 3e6 cells / 1mL of Gibco Freezing Medium. For example, if there were 9e6 total cells, the cell pellet was resuspended in 3mL of Gibco Freezing Medium. 3. Aliquots of 1 mL of resuspended cells / cryovials were measured. Cells were frozen at -80°C in a suitable cell freezing vessel (i.e., Mr.Frosty or Corning CoolCell freezing system). 4. The cells were transferred to a Liquid Nitrogen Cryotank for long-term storage.

[0373] To thaw the cells: 1. The cells were removed from the liquid nitrogen tank. The cryovials were thawed in a 37°C water bath until small "ice pellets" remained. They were then sprayed with 70% ethanol and transferred to a TC / BSC hood. 2. 9 mL of fresh medium was added to a 15 mL conical tube. 10 mL of fresh medium was added to a T75 TC-treated flask. 3. Gently transfer 1 mL of cells from the cryovial to a 15 mL conical tube containing the culture medium. 4. The cells were pelleted by centrifuging at 1000 rpm for 5 minutes. 5. Aspirated culture medium / frozen culture medium. 6. Gently resuspend the cell pellet in 5 mL of fresh medium and transfer it to a T75 flask containing 10 mL of fresh medium. Place the flask in a 37°C incubator (5% CO2).

[0374] protocol Day 1 The procedure is as follows: 1. Prepared ARP: a. Using Echo, 12.5 nL of 10 mM source solution was stamped onto a destination plate. The plates were immediately sealed and frozen at -20°C if not used on the same day. b. When using a frozen ARP, the plate was thawed and rotated at 1000 rpm for 1 minute.

[0375] 2. Preparation of cells (adhesion): a. The culture medium was aspirated from the cells. The cells were washed with sterile 1XPBS. The PBS was aspirated and an appropriate amount of trypsin was added. b. After completely trypsinizing the cells, the cells were resuspended in appropriate culture medium. The cells were then transferred to 15 mL or 50 mL conical tubes. c. Cells were counted using a Countess II cell counter.

[0376] 3. Cell Plating: a. Cells were prepared at an appropriate plating density. Using Multidrop Combi, 12 μL of diluted cells per well were dispensed into columns 1-23 of a Greiner 7840 80-384 well TC-treated white plate. 12 μL of appropriate phenol-free medium was added to column 24 only. b. The plates were placed in a tissue culture incubator at 37°C for the appropriate processing time (see the “Assay” table).

[0377] 4. Prepared HTRF lysis buffer a. Calculate the amount of HTRF lysis buffer master mix required to perform the desired experiment + any extra dead volume required for dispensing (4 μL per well is needed). Dilute the blocking reagent in 4 × lysis buffer in a 1:25 ratio (i.e., 0.1 mL of blocking reagent solution + 2.4 mL of 4 × lysis buffer). b. Add 4 μL of lysis buffer master mix to all wells containing the sample or DMSO. Centrifuge the plate at 1000 rpm for 1 minute. c. Incubate at room temperature for 30 minutes.

[0378] 5. Prepared HTRF antibody a. The amount of HTRF antibody master mix required to perform the desired experiment + any extra dead volume required for dispensing (4 mL per well) was calculated. Eu Cryptate antibody and d2 antibody were added to the detection buffer in a ratio of 1:40 each (i.e., 100 μL Eu Cryptate + 100 μL d2 Cryptate + 3800 μL detection buffer). b. 4 μL of antibody master mix was added to each of the 24 wells of the column containing only culture medium. c. The plate was centrifuged at 1000 rpm for 1 minute. A "humidity chamber" was created by covering the plate and placing it in a Ziploc bag lined with a damp paper towel or similar material, and the plate was incubated overnight at room temperature, away from light.

[0379] Day 2 6. Measurements were taken using PHERAstar / Envision with the HTRF protocol. All wells were read when the plate was read.

[0380] IC1 of compound 1 in the T47D pAKT assay 50 The (nM) was 31–42 nM.

[0381] Surface plasmon resonance (SPR) PI3K binding can also be determined by SPR. SPR experiments are performed using a Biacore 8K instrument. Biotinylated recombinant PI3Kα H1047R protein containing a full-length p110-α subunit with the H1047R mutation and an N-terminal AviTag, complexed with a cleaved p85-α subunit (amino acid residues 322-694), is used. The protein is first incubated with 1 μM wartmannin at room temperature for 30 minutes to covalently block the ATP binding site, and then immobilized on a streptavidin sensor chip by flowing the protein through the sensor chip at a concentration of typically 20 μg / mL and a flow rate of 2 μL / min for 1200 seconds. Compound binding affinity is measured in multi-cycle dynamic mode at a flow rate of 90 μL / min, an association time of 90 seconds, and a dissociation time of 240 seconds. The running buffer contained 50 mM Tris, pH 7.5, 150 mM NaCl, 0.01% Brij35, 1 mM DTT, 1 mM MgCl2, 0.05% Tween-20, and 2% DMSO. The temperature was maintained at 25°C during the experiment, and the data were fitted to a 1:1 coupling model.

[0382] crystallization Crystals of the PI3Kα(H1047R) / p85α heterodimer, compounded with GDC-0077 and compound 1, were obtained at a concentration of 10 mg / ml (20 mM Tris / HCl, 150 mM NaCl, 1 mM TCEP, pH 8.0). These were pre-incubated for 1 hour with 1.5 excess GDC-0077 and compound 1 (150 mM in dimethyl sulfoxide (DMSO)). Then, 0.8 μl of the protein solution was mixed with 0.8 μl of reservoir solution (0.1 M MES pH 6.8, 0.5 M NaCl, 5% (w / v) polyethylene glycol 3350) and equilibrated at 293 K over 60 μl of reservoir solution. Crystals that diffracted well were selected for data collection after 5 days. Crystals were cryoprotected in a reservoir solution supplemented with 30% ethylene glycol and rapidly frozen in liquid nitrogen before data acquisition. A complete 2.9 Å dataset of PI3Kα(H1047R) / p85α / GDC-0077 / compound 1 crystals was acquired at the European Synchotron Radiation Facility (ESRF, Grenoble, FR, beamline ID30a1). The data were integrated, analyzed, and scaled by program XDS, Pointless, and STARANISO from within the autoPROC pipeline, respectively.

[0383] Structural solutions and refinement. An isomorphic reference model of PI3Kα compounded with p85α and GDC-0077 was used as a starting model for suppression refinement of the H1047R dataset using REFMAC5. The final model was obtained through several refinements using the REFMAC5 program, followed by BUSTER. Atomic displacement factors were modeled using a single isotropic B factor per atom and a single TLS group per chain. Non-crystallographic symmetry restrictions were used. Restrictions for compound GDC-0077 and compound 1 were generated using GRADE from Global Phasing with the big planes option. The final model consisted of two heterodimers in the asymmetric unit, both of which bound to compound 1 and GDC-0077. Chain A and B heterodimers dissolved better than the other heterodimers and were therefore used in the analysis of binding to compound 1. Crystal structure statistics are reported in Table 13. Images were generated using PyMOL (www.pymol.org).

[0384] Table 13. X-ray crystallographic statistics of Compound 1 [Table 4] * The value in parentheses represents the highest resolution shell. RMS stands for Root Mean Square.

[0385] PI3Kα ATPase assay Full-length WT, M1043X, H1047X, or G1049R enzymes (1–10 nM) were incubated with a vehicle or compound at room temperature for 1 hour, followed by the addition of ATP (final 90 μM) to initiate the enzymatic reaction. The assay buffer contained 50 mM Tris, 150 mM NaCl, 0.01% Brij 35, 15 mM MgCl2, 0.05% Tween-20, and 1 mM DTT. ADP production was measured at room temperature after 100 minutes of incubation using the ADP-Glo ​​kit (Promega#V9102).

[0386] Kinome-selective profiling In vitro kinase profiling assays of compound 1 were evaluated across 373 kinases (KinaseProfiler®, IC). 50 Profiler(TM); Eurofins Cerep, Le Bois l'Eveque, France).

[0387] Cell assay For details on the cell lines, please refer to Table 14.

[0388] Table 14. Cell lines and plating densities used for HTRF (pAKT) and viability (CTGLO). [Table 5]

[0389] Uniform time-resolved fluorescence (HTRF) A 384-well phospho-AKT(S473)HTRF(PerkinElmer#64AKSPEH) assay was used for target association. Each well was seeded with 12.5 μL of phenol red-free medium at the cell counts shown in Supplementary Table S3, followed by incubation at 37°C for 1 hour, and then read using a PHERAstar plate reader.

[0390] Cell proliferation Cell viability was measured using CellTiterGlo (Promega#G9243). The cell lines shown were seeded in 50 μL of medium in a 384-well plate according to Supplementary Table S3 and treated at 37°C with 5% CO2 for 72 hours.

[0391] Compound 1 was submitted to the Broad Institute PRISM high throughput cell viability screen (www.theprismlab.org / ).

[0392] Human adipose cells 3 H-2-deoxyglucose uptake Primary human subcutaneous adipocytes were treated with the test compound or vehicle for 1 hour, followed by 10 nM insulin and 3 H-2-deoxyglucose (Zen Bio Durham, NC USA, assay #CA-25, lot #SL0071) was added. Cytochalasin B treatment controlled nonspecific glucose uptake. Corrected counts / min were determined using a scintillation counter.

[0393] Animal testing All animal handling and treatment procedures were carried out in accordance with the Association for Assessment and Accreditation of Laboratory Animal Care guidance and approved Institutional Animal Care and Use Committee guidelines. Cell line xenografts were performed in BALB / c nude mice, with the exception of the T47D model using NSG mice transplanted with 17-β-estradiol tablets (0.5 mg, 90-day release). PK / PD using xenograft tumors was established using standard protocols. PDX models were performed using XenoSTART (San Antonio, TX). Compound 1 and alpelisib were formulated in 30% 2-hydroxypropyl-β-cyclodextrin pH 8. Tissue Western blotting was performed using standard protocols with flash-frozen tissue in radioimmunoprecipitation assay buffer containing a protease inhibitor. Primary antibodies from Cell Signaling Technologies were used: pAKT (S473) (AB_2315049), AKT (AB_1147620), and Vinculin (AB_2728768), and secondary antibodies: IRDye 680CW goat anti-mouse IgG (AB_10956588) and IRDye 800CW goat anti-rabbit IgG (AB_621843) from LiCOR. IHC samples were fixed in 10% NBF for 24 hours, transferred to 70% ethanol, and tumor pAKT (Cell Signaling #4060) was embedded, sectioned, stained, and quantified. Plasma and tissue bioanalysis of compound 1 or alperisib was measured after protein precipitation using liquid chromatography with tandem mass spectrometry. All methods and limitations of quantification were appropriate in terms of specificity and sensitivity to support PK analysis.

[0394] The OGTT and ITT tests in BALB / c nude mice were performed 5 days after treatment. Food was removed for 5 hours, the drug was administered, and then 2 g / kg of oral glucose (OGTT) or 0.75 U / kg of intraperitoneal insulin (ITT) (Lilly Inc, France, #H1079) was administered 1 hour later. After tail vein sampling or terminal blood sampling, blood glucose levels were measured at the indicated time points (One touch Glucose Meter, Roche, ACCU-CHEK Performa #06454038). Insulin was measured by ELISA (Crystal Chem, #90082). Metabolic profiling in CAL33 tumor-bearing mice was performed after a 4-hour fast, and then the drug was administered 1 hour before oral administration of 300 mg of 13 13C-labeled glucose (Cambridge Isotope Labs #CLM-1396-0). Tissues were collected immediately before (0 hour) or 30 minutes after labeled glucose administration, snap-frozen, and analyzed (NYU Metabolomics Core using the Hybrid Metabolomics protocol, RRID:SCR_017935).

[0395] Example 1: Isobologram as a measure of combination benefit Determination of a constant enzyme activity determined by varying substrate and inhibitor concentrations.

[0396] The following are three models for measuring synergy: 1. Loewe additivity can define additivity as a drug combined with itself where no interaction occurs. The Loewe additivity method is widely used, but it assumes that the compounds have the same Ymax and Hill slope. 2. The Bliss independence model is based on the concept of pharmacological independence. 3. The HSA model, which defines additivity as the maximum effect of the most potent compound, is a combination. See Di Veroli, et al., Bioinformatics. 2016. Sep 15;32(18):2866-8.

[0397] The quantification and graphing of synergistic effects were as follows. Both the combination index and isobologram were derived from Loewe's arithmetic addition. IC 50 The synergistic effect was quantified by calculating the combination index (CI) using the following formula. See Altenburger, et al., Handbook of Hazardous Materials, 1993, pp. 15-27. A CI of less than 0.75 indicates a synergistic effect, and a CI greater than 1.25 indicates an antagonistic effect. A CI between 0.75 and 1.25 is additive. The linear concentrations of the two drugs lie on different axes, and the IC of the combination... 50 This is commonly plotted. Linear isoballs representing Loewe additivity lie between equal concentration values. See Figures 1A to 1E.

[0398] Example 2: Compound 1 in combination with fulvestrant supports clinical benefits. Strong inhibition and quiescence of tumor growth were observed with compound 1 at a dose of 100 mg / kg. Fulvestrant alone showed little to no inhibition of growth. Consistent tumor regression was observed only with the combination of compound 1 and fulvestrant. See Figures 2A-2C and Table 3.

[0399] Table 3 [Table 6]

[0400] Example 3: Compound 1 in combination with palbociclib supports clinical benefits. Strong inhibition and quiescence of tumor growth were observed with compound 1 at 100 mg / kg and palbociclib alone. Tumor regression was observed only with the combination of compound 1 and palbociclib. See Figures 3A-3BC and Table 4.

[0401] Table 4 [Table 7]

[0402] Example 4: CDX model study using GP2D colon adenocarcinoma cells and Detroit 562 head and neck squamous cell carcinoma (HNSCC) cells GP2D and Detroit562 tumor cell lines were cultured and transplanted into immunodeficient BALB / c nude mice. The mice were subjected to the treatments shown in Tables 5 and 6.

[0403] Table 5 [Table 8]

[0404] Table 6 [Table 9]

[0405] Example 5: PDX model study using ST1056 breast cancer cells and ST433 head and neck squamous cell carcinoma (HNSCC) cells ST1056 tumor cell lines were biopsied and transplanted into athymic immunodeficient mice. Table 7 shows the details of the study. The mice were treated as shown in Table 8.

[0406] Table 7 [Table 10]

[0407] Table 8 [Table 11] * Administer in a fixed volume dose.

[0408] ST433 tumor cell lines were biopsied and transplanted onto Fox Chase SCID (CB17 / Icr-Prkdcscid / IcrIcoCrl). Table 9 shows the details of the experiment, and the mice were treated as shown in Table 10.

[0409] Table 9 [Table 12]

[0410] Table 10 [Table 13]

[0411] Example 6: Evaluation of the antitumor effect of Compound 1 in combination with fulvestrant in a T-47D human breast cancer xenograft model in immunodeficient mice. The cell line T-47D was used in this ER + HER2 - Used in the CDX breast cancer model, T-47D (PI3Kα H1047R The ) was transplanted into immunodeficient mice.

[0412] Compound 1 (100 mg / kg) monotherapy induced robust tumor regression (141% TGI). Compound 1 in combination with fulvestrant (5 mg QW) was well-tolerated and resulted in more consistent and deeper tumor regression. Table 11 shows that fulvestrant monotherapy (G2) or the submaximal dose of Compound 1 (50 mg / kg, G3) resulted in tumor growth inhibition, respectively (52% and 92%, respectively), but when combined (G5), the combination demonstrated a benefit exceeding either monotherapy alone, with 16% tumor regression.

[0413] Table 11 shows the antitumor activity of T-47D xenografts. Twenty days after treatment, the mean tumor volume (TV) in the vehicle control group was 413 mm². 3 The results showed that, compared to vehicle controls, fulvestrant at 50 mg / kg QD and 5 mg / mouse sc demonstrated antitumor activity when analyzed by two-way RM ANOVA followed by a post-hoc comparison of mean Tukey, respectively, with a TGI of 52% (P ≤ 0.01).

[0414] Compound 1 at 50 mg / kg QD and 100 mg / kg QD showed dose-dependent antitumor activity with TGI values ​​of 92% (P ≤ 0.0001) and 141% (P ≤ 0.0001), respectively. Compound 1 at 50 mg / kg QD, combined with fulvestrant 5 mg / mouse sc, showed strong antitumor activity with a TGI value of 116% (P ≤ 0.0001). Compound 1 at 100 mg / kg QD, combined with fulvestrant 5 mg / mouse sc, also showed strong antitumor activity with a TGI value of 155% (P ≤ 0.0001). Table 11 shows that G2 or G3 versus G5 demonstrated therapeutic synergies with tumor regression, which were observed only in combination.

[0415] Table 11 [Table 14]

[0416] A: Mean ± SEM; B: TGI = (1 - T / C) × 100%, T / C = 100% × (Final TV treatment - Initial TV treatment) / (Final TV vehicle - Initial TV vehicle); C: Vehicle control; NS = No significance. * P ≤ 0.05; ** P ≤ 0.01; *** P ≤ 0.001; **** P ≤ 0.0001, followed by a two-way RM ANOVA, and then Tukey's posterior comparison of the mean.

[0417] Example 7: Evaluation of Compound 1 as monotherapy and in combination with other antitumor agents in participants with advanced solid tumors. This clinical trial is a multipart, open-label Phase 1 / 2 study evaluating the safety, tolerability, pharmacokinetics (PK), and preliminary antitumor activity of compound 1 in participants with advanced solid tumors containing specific mutations.

[0418] Part 1 evaluates Compound 1 as monotherapy in participants with breast cancer and other solid tumor types. Part 2 evaluates Compound 1 therapy in combination with fulvestrant in participants with breast cancer.

[0419] Each study part includes a 28-day screening period followed by treatment with compound 1 as monotherapy or in combination therapy. Participants remain in the study part to which they were initially enrolled throughout their participation in the study (i.e., they do not move to other study parts).

[0420] Table 12. Arms and interventions [Table 15]

[0421] Key evaluation criteria Part 1.1 (Dose escalation): 1. MTD: The number and percentage of participants who experienced at least one DLT during the first 28 days of treatment. 2. OBD: PK, pharmacodynamics, ORR, TEAE / SAE ≥ Grade 2. Type, frequency, and severity of TEAE according to CTCAE v5.0 criteria.

[0422] Part 1.2 (Dosage Selection) and Part 2.1: 1. PK, pharmacodynamics, ORR, and safety parameters

[0423] Part 1.2 (Dose Expansion), Part 1.3, and Part 2.2: 1. ORR is defined as the percentage of participants who have a PR or CR based on RECIST 1.1.

[0424] Selected inclusion criteria 1. Having a metastatic or locally advanced and unresectable, progressive or refractory solid malignant tumor. 2. Having a new or recent tumor biopsy (collected at the time of screening, if feasible) or stored tumor specimen within 12 months prior to screening. 3. Having a tumor that possesses a recorded PI3Kα mutation, as determined by a PCR or NGS-based assay as an FDA-approved test in the United States, obtained from either a tumor or a plasma sample, or as part of routine clinical care in a CLIA-accredited or similarly accredited laboratory. 4. Having at least one measurable neoplastic lesion according to RECIST 1.1 5. The person was 18 years of age or older at the time of signing the ICF. 6. Having an ECOG performance status score of 0 or 1 at the time of screening.

[0425] Selected exclusion criteria 1. Having a history of a solid tumor or hematological malignancy histologically different from the cancer being tested (within 2 years prior to screening). 2. Symptomatic brain or spinal cord metastases 3. Having a tumor with a PTEN mutation / deletion and an activating mutation in AKT or mTOR, as confirmed by a CLIA-accredited or similarly accredited laboratory, or having a tumor with a PTEN mutation / deletion and an activating mutation in AKT (e.g., E17K), as confirmed by a CLIA-accredited or similarly accredited laboratory. 4. Having an established diagnosis of type 1 diabetes mellitus, or having uncontrolled type 2 diabetes mellitus requiring antidiabetic medication. 5. Cohorts A0, A1, A2, A3, A4, and B: Except in specific circumstances, have previously received treatment with PI3K / AKT / mTOR inhibitors. 6. Treatment with any topical or systemic antineoplasm therapy or investigational anticancer drug within 14 days or 4 half-lives, whichever is longer, prior to the start of the study treatment up to the maximum washout period of 6.28 days. 7. Toxicity from previous anticancer therapy that has not resolved to baseline levels or CTCAE grade ≤ 1, excluding alopecia and peripheral neuropathy. 8. The patient received radiotherapy within 14 days prior to the start of the experimental procedure.

[0426] Example 8. Compound 1 as a muta-selective allosteric PI3Kα inhibitor Compound 1 was confirmed to be a potent binder for H1047R PI3Kα by surface plasmon resonance (SPR; Figure 5B). Its low nanomolar binding affinity (equilibrium dissociation constant, K) for the H1047R mutation was confirmed. D In addition to approximately 2.9 nM, compound 1 had a binding affinity of 1 / 20th to WT PI3Kα (K D (approximately 56 nM). In contrast, duvericib (a non-selective orthosteric PI3Kα inhibitor) showed nearly equal binding affinity to mutant and WT PI3Kα. Although not bound by theory, the H1047R mutant selectivity of compound 1 is mainly due to its faster association constant (k) for H1047R versus WT PI3Kα. on ) and a slightly slower dissociation constant (k off This suggests that the allosteric site occupied by compound 1 may be more accessible in the H1047R mutant form of the enzyme, as it can be driven by (Figure 5B).

[0427] The biochemical potency and mutaselectivity of compound 1 were compared with alperisib in a panel of common oncogenic mutation PI3Kα morphologies (Figure 5A). Compound 1 was found to be the most common variant H1047R(IC1). 50 It is a potent and selective inhibitor of all kinase domain mutations in PI3Kα found in cancer, including those of approximately 9.4 nM, and is effective against WT PI3Kα (IC1). 50 It was found to have 14-fold selectivity for approximately 131 nM. Under these same assay conditions, compound 1 showed limited selectivity for the hotspot helical domain mutation (E542K / E545K) PI3Kα, while alpericib did not show mutation selectivity, as previously reported (Fritsch C, et al. Mol Cancer Ther 2014;13(5):1117-29).

[0428] Compound 1 also demonstrated exquisite kinome-wide selectivity (Figure 5G). Biochemical screening using 373 kinases, representing approximately 70% of the human kinome, including PI3Kβ, PI3Kδ, and PI3Kγ isoforms, showed that only AurB kinase was inhibited by >50% at 10 μM (IC). 50 (=1.6 μM). Subsequent follow-up studies confirmed that compound 1 exhibited limited AurB inhibition in cells at concentrations up to 10 μM.

[0429] Compared with the published WT and H1047R-PI3Kα X-ray crystal structures, the co-crystal structure of compound 1 revealed that compound 1 occupies a novel allosteric site formed due to a major conformational shift at residues 936-940, along with other smaller local rearrangements (Figure 5D) (Figure 5C, Table 13). Specifically, residues F937 and L938 occupy positions where they can directly collide with compound 1. Rearrangements of the side chain and main chain atoms of these residues resulted in the rearrangement of F937 and L938, creating space for the allosteric site. Furthermore, the activation loop is well dissolved by the compound 1 co-structure compared to the existing H1047R structure (3hhm and 3hiz; Mandelker D, et al. Proc Natl Acad Sci USA 2009;106(40):16996-7001). Compound 1 makes several specific protein contacts within its allosteric site (Figure 5E).

[0430] Example 9. Compound 1 selectively targets PI3Kα activity and cell viability in PI3Kα mutant cells. Compound 1 was tested and compared to alpericib in a panel of 10 human tumor cell lines carrying the PI3Kα kinase domain mutation (Table 15). In addition to H1047R, the panel included three cell lines carrying the second most common kinase domain mutation, H1047L (EFM19, GP2D, and OAW42 cell lines), as well as three double mutant forms (BT20, CAL148, and NCIH1048 cell lines). SKBR3 breast cancer cells were included as a comparator for WT selectivity, depending on the amplified WT PI3Kα activity for proliferation (Cerami E, et al. Cancer Discov 2012;2(5):401-4, Gao J, et al. Sci Signal 2013;6(269):pl1).

[0431] Target association was evaluated using phosphorylation (serine 473) as a marker of PI3Kα / AKT pathway activity. Figure 6B shows the dose-response curves of compound 1 and alpelisib in PI3Kα kinase domain mutant T47D and Cal33 cells, as well as WT SKBR3 cells. Compound 1 effectively inhibited mutant PI3Kα activity across the panel of cell lines and IC 50 The values ​​range from approximately 15 to 319 nM, which corresponds to approximately 28 to 268 nM for an alpha-spherical IC. 50 The values ​​were similar (Table 15). Compound 1 was more potent than alpelisib in 9 out of 11 cell lines, with the only exception being WT-PI3Kα-SKBR3 cells, which was expected based on the mutant selectivity of compound 1 (correlation plot Figure 6C, Table 15). ER + HER2 - In the breast cancer benchmark T47D (H1047R PI3Kα) cell line, compound 1 was 9-fold more selective than SKBR3 (WT PI3Kα) cells. In contrast, alpelisib did not show any difference in selectivity between mutant and WT-driven cell lines. When cell viability was tested in the same cell line, there was a strong correlation between target association (pAKT) and cell viability (Pearson correlation coefficient = 0.8 [logarithmic scale]) (Figure 6A). These results confirm that pAKT is a translational biomarker associated with mutation-selective inhibitors such as compound 1.

[0432] Table 15. Target association and cell survival activity of Compound 1 in a panel of human tumor cell lines. [Table 16]

[0433] GI 50 , the concentration of a compound that reduces total cell proliferation by 50%; GMean, geometric mean; RRID, research resource identifier.

[0434] Next, the activity of compound 1 was evaluated in a high-throughput cell viability panel of approximately 900 tumor cell lines to identify sensitivity markers. Consistent with the selectivity profile described above, cell lines with PI3Kα kinase domain mutations were significantly more sensitive than WT cell lines (Figure 6D). Unexpectedly, tumor cells with PI3Kα helical domain mutations exhibited nearly the same level of proliferation sensitivity to compound 1, despite initial results using recombinant proteins (Figure 5E). The increased sensitivity of helical domain mutant cell lines compared to WT PI3Kα cell lines may be explained by the PI3Kα dependence of the mutant cell lines for proliferation, which is not the case for most WT cell lines. PI3Kα mutations located in the hotspot helical and kinase domains appeared to be highly sensitive to compound 1 (Figure 6D). PTEN inactivating mutations conferred resistance to PI3Kα inhibition in this study (P<0.0081), consistent with previous reports (Razavi P, et al. Nat Cancer 2020;1(4):382-93).

[0435] The effect of compound 1 on WT PI3Kα was evaluated by assessing insulin-mediated glucose uptake in human primary adipocytes (Hauner H, Int J Obes Relat Metab Disord 1998;22(5):448-53). Differentiated primary human subcutaneous adipocytes were pretreated with alpelisib or compound 1, and then [ 3The patient was supplemented with [H]-2-deoxyglucose and 10 nM insulin. Alpelisib inhibited glucose uptake at a low concentration of 100 nM and nearly completely inhibited it at 10 μM, but the concentration of compound 1 (EC) required to achieve 50% inhibition was... 50 The concentration was ≥10 μM (Figure 6E). The maximum effect of compound 1 on glucose uptake (E max ) was 38%, but alperisib was 88% E max This resulted in deeper suppression. The overlay of the cell viability dose-response curve obtained in T47D cells (Figure 6A) shows the potentially improved therapeutic index of compound 1 compared to alpelisib in the relevant human cell line.

[0436] Example 10. Treatment with Compound 1 yielded robust antitumor efficacy in PI3Kα mutant tumors in mice without metabolic dysregulation. The in vivo pharmacological characterization of compound 1 was designed to establish the metabolic safety and antitumor efficacy profile of compound 1 compared to alperisib. Aperisib was selected at a once-daily (QD) dose of 50 mg / kg because it was effective in a published mouse xenograft model despite known glucose dysregulation (Fritsch C, et al. (2014) Mol Cancer Ther. 13(5):1117-29). At this dose, alperisib plasma exposure (AUC) in mice (approximately 75,000 ng) * hr / mL) is approximately twice the exposure to the maximum approved human dose (approximately 33,000 ng). * (hr / mL) exceeded (Juric D, et al. J Clin Oncol 2018;36(13):1291-9). Low doses (20 mg / kg) were also included. Based on the pharmacokinetic (PK) profile of compound 1, the QD doses of 30 and 100 mg / kg were found to be the inhibitory concentrations (IC) of 80% of the associated cancer cell lines. 80 Although it was expected that the levels would be bracketed, the QD dose of 300 mg / kg exceeded these levels (Figure 12).

[0437] Regarding metabolic regulation, the main consequence of WT PI3Kα inhibition is the blockade of insulin action (e.g., insulin resistance), impaired glucose processing, and hyperglycemia (Fruman DA, Chiu H, Hopkins BD, Bagrodia S, Cantley LC, Abraham RT. The PI3K pathway in human disease. Cell 2017;170(4):605-35; James DE, Stockli J, Birnbaum MJ. The aetiology and molecular landscape of insulin resistance. Nat Rev Mol Cell Biol 2021;22(11):751-7).

[0438] The effects of alpelisib and compound 1 doses on insulin sensitivity were analyzed using an insulin tolerance test (ITT) and oral glucose tolerance test (OGTT) after 5 days of repeated administration in non-tumor-carrying female BALB / c nude mice, a sex / strain frequently used in xenograft studies. Alpelisib treatment resulted in a dose-dependent decrease in glucose treatment in both ITT and OGTT, consistent with insulin resistance (Figures 7A / 7B and 7C / 7D, respectively). In contrast, compound 1 treatment was not associated with a significant change in glucose AUC, but there was a non-statistically significant increase at a 300 mg / kg dose in ITT. Notably, compound 1 had no effect on body weight or fasting blood glucose levels 5 days after treatment. These studies established that repeated doses of compound 1 at 100 mg / kg QD were well tolerable without metabolic dysregulation, while alpelisib induced evident insulin resistance at a 50 mg / kg dose level.

[0439] Example 11. Benchmarking of Compound 1 in a Cal33(H1047R PI3Kα) human HNSCC xenograft model. The Cal33(H1047R PI3Kα) head and neck squamous cell carcinoma (HNSCC) cell model was selected to benchmark in vivo therapeutic activity and pharmacodynamic biomarkers because this cell line showed moderate sensitivity to PI3Kα inhibition in culture (Figure 6A). The study consisted of three arms: an efficacy group in which animals received the test product for 28 days to evaluate the effects of alperisib and compound 1 on glucose uptake and glucose oxidation in target tissues; a PK / pharmacodynamic (PK / PD) group in which animals received compound 1 or alperisib treatment for 3 days; and an animal with tumors [U- 13 This group consisted of those who received an oral bolus of [1C]-glucose.

[0440] In the efficacy arm, both compounds showed a dose-dependent reduction in tumor volume, with compound 1 at 30 mg / kg exhibiting similar efficacy to alpelisib at 20 mg / kg, and compound 1 at 100 mg / kg exhibiting similar efficacy to alpelisib at 50 mg / kg (Figure 8A). Both compounds were well-tolerated at all doses, and there was no change in body weight (Figure 8B). Alpelisib treatment elevated serum insulin 1 hour after administration on day 28 (P=0.0585, Figures 8C and 9), and showed a similar trend in glucose (P<0.083, Figure 8D). In all subsequent cell-line derived xenograft (CDX) studies, alpelisib 50 mg / kg caused a significant increase in serum insulin 1 hour after administration, while compound 1 at 100 mg / kg did not (Table 16).

[0441] Table 16. Summary of the CDX trial: Tumor growth inhibition and insulin levels (1 hour after administration). [Table 17] Inhibition of tumor growth or a Regression (negative TGI). b One hour after administration. P ≤ 0.05 * , 0.01** , 0.001 *** , 0.0001 **** .

[0442] Negative TGI values ​​indicate % regression. NS: not significant; NSG: NOD (no significant) scid gamma. Target-associated PD biomarkers (pAKT / AKT ratio) and tumor drug levels of compound 1 were measured 1, 4, and 12 hours after administration on day 3 (PK / PD group), and 1 and 6 hours after the final administration (day 28). The curve-fit relationship between tumor-drug concentrations and pAKT / AKT levels in Cal33 tumor xenografts and in vitro is shown in Figure 8E. The IC calculated from this curve fit was also calculated. 50 When corrected for matrix binding, the concentration in tumors was approximately 45 nM compared to approximately 18 nM from cell cultures (Pearson r correlation coefficient = -0.613; P < 0.0001), thereby demonstrating a conclusive in vitro target association:in vivo correlation. In this xenograft study, 82% tumor growth inhibition (TGI) in the compound 1 100 mg / kg QD dose group was observed, along with a 57% mean pAKT suppression (treated / vehicle AUC). 1-12h ) was associated with the alpelisib 50 mg / kg QD group, and in the alpelisib 50 mg / kg QD group, 79% of TGI was associated with a mean pAKT suppression of 66% (Figure 8F). Unlike alpelisib, compound 1 did not reduce skeletal muscle pAKT / AKT (Figure 8G).

[0443] Inhibition of PI3Kα is known to suppress glucose metabolism in tumors and host tissues (Hopkins BD, et al. Nature 2018;560(7719):499-503, Juric D, et al. (2018) J Clin Oncol 36(13):1291-9, Dockx Y, et al. Mol Imaging 2021;2021:5594514, Sarker D, et al. Clin Cancer Res 2015;21(1):77-86).

[0444] To evaluate the effects of alpelisib and compound 1 on this process, Cal33 tumor-carrying mice were pre-treated with the drug or vehicle, and then U- was administered by forced oral administration. 13 C-glucose was administered. After 30 minutes, tumor and skeletal muscle metabolites were extracted and quantified using liquid chromatography-mass spectrometry (Lopes M, et al. Cell Rep 2021;37(2):109833). Oral bolus administration resulted in robust labeling of over 70% of circulating glucose carbon in all groups (Figure 8J). In tumors, both alpelisib and compound 1 mediated the formation of the TCA intermediate, a marker of glucose oxidation. 13 It significantly reduced C uptake (Figure 4H). However, only alpelisib-treated mice demonstrated a reduction in glucose oxidation in skeletal muscle (Figure 8H). This reduction occurred despite higher levels of circulating insulin in this group (30-minute alpelisib vs. vehicle), suggesting insulin resistance in this tissue (Figure 8I). These data support that compound 1 selectively inhibits mutant PI3Kα but does not inhibit the WT enzyme found in host tissues.

[0445] Example 12. Compound 1 is effective across a panel of PI3Kα-mutated CDX and PDX tumors without evidence of insulin resistance. This detailed metabolic characterization of compound 1 and its activity in the Cal33 xenograft model established that the optimal compound 1 dose is 100 mg / kg QD and demonstrated that compound 1 is more effective than alpelisib (20 mg / kg) at a clinically matched dose. Therefore, compound 1 at 100 mg / kg was carried over to a panel of PI3Kα-mutated CDX and patient-derived xenograft (PDX) models representing several cancers. The models included colorectal cancer (GP2D), lung cancer (NCIH1048), HNSCC (Detroit 562), and HR. - HER2 +This study included breast cancer (HCC1954). All studies were conducted in BALB / c nude mice, with high-dose alpelisib (50 mg / kg QD) included as a benchmark.

[0446] The key efficacy endpoints were TGI or regression, and tolerability measures included body weight and insulin levels one hour post-administration at the end of the study. Neither compound 1 nor alpelisib showed treatment-specific effects on body weight (Figures 11F-11I). TGI with compound 1 at 100 mg / kg QD treatment was similar to or better than that with 50 mg / kg QD alpelisib treatment (Figure 11E, Table 17). Compound 1 demonstrated robust efficacy in GP2D tumor xenografts expressing H1047L PI3Kα, the second most dominant kinase domain mutation. The waterfall plot showed tumor regression in half of the animals treated with compound 1, but no regression was observed with alpelisib treatment (Figure 11E). In the Detroit 562 HNSCC model, both compounds showed robust efficacy, including tumor regression, in 5 out of 9 test animals. Similarly, treatment with compound 1 and alpelisib was effective in treating NCIH1048 lung cancer containing the dual (H1047R / K411R)PI3Kα mutation, as well as HCC1954 HR + HER2 + It provided comparable control of tumor growth in a breast cancer model (Figure 11E). In all of the above studies, a significant increase in serum insulin was observed in animals administered alpelisib 50 mg / kg one hour after administration, but compound 1 at 100 mg / kg was not associated with an increase in insulin (Table 17). Both treatment with alpelisib and compound 1 caused significant TGI, but HCC1954 cells showed the lowest response among all cell lines tested in the CDX panel. HCC1954 cells were also weaker responders to both drugs in cell culture (Figure 6A) and HER2 +This may reflect the decreased efficacy of PI3Kα inhibitor monotherapy in cancer, which is thought to be related to compensatory HER3 activation (Serra V, et al. Oncogene 2011;30(22):2547-57, Chandarlapaty S, et al. Cancer Cell 2011;19(1):58-71).

[0447] Table 17. Summary of the CDX trial: Tumor growth inhibition and insulin levels (1 hour after administration). [Table 18] Inhibition of tumor growth or a Regression (negative TGI). b One hour after administration. P ≤ 0.05 * , 0.01 ** , 0.001 *** , 0.0001 **** .

[0448] Negative TGI values ​​indicate % regression. NS: not significant; NSG: NOD scid gamma. Monotherapy with compound 1 and alpelisib was evaluated in two breast cancer PDX models, ST1056 (kinase domain mutation [H1047R]) and ST1799 (double mutation, E542K / H1065L). A third HNSCC xenograft model (ST2652) carrying only the helical domain mutation (E542K) was also evaluated. Compound 1 was highly effective in all three models, including the (E542K) helical domain mutation, and was generally similar to alpelisib (Figures 11A-11C). A decrease in pAKT / AKT was observed 4 hours after administration in all models for both compound 1 and alpelisib (Figures 11A-11C).

[0449] Example 13. Safety and efficacy of clinically relevant combination therapies The monotherapy and combination therapy of compound 1 and fulvestran were evaluated in a T47D cell xenograft model. ER + HER2- In the PDX model, monotherapy with compound 1, fulvestrant, and palbociclib, as well as pairwise and tripartite combinations of these compounds, were evaluated.

[0450] The T47D cell line represents an important benchmark for the mutant PI3Kα mechanism and is a well-established estrogen-dependent ER cell previously characterized by alpericib treatment. + HER2 - This is a breast cancer xenograft model (Fritsch C, et al. (2014) Mol Cancer Ther 13(5):1117-29). Compound 1 monotherapy demonstrated dose-dependent TGI, with the 50 mg / kg QD dose achieving a similar TGI to high-dose alpelisib (50 mg / kg QD), and the 100 mg / kg QD dose of Compound 1 resulting in significant tumor regression in all animals (Figure 4A). Fulvestrant monotherapy provided only about 50% TGI, and the addition of either 50 or 100 mg / kg doses of Compound 1 resulted in regression in most xenografts (20% and 70% regression, respectively). The combination of fulvestrant monotherapy and Compound 1 was well tolerable based on body weight (Figures 4F-4G). Single-dose PK / PD measurements were performed in tumors collected 4 hours after administration of 100 mg / kg of compound 1 and 24 hours after fulvestrant. Phospho-AKT and pS6, biomarkers of PI3Kα pathway activity, were moderately reduced by fulvestrant, but compound 1 resulted in significant inhibition, as assessed by Western blotting and IHC (Figures 4B and 4C, respectively).

[0451] Combination studies including the use of compound 1, palbociclib, and fulvestrant in combination were conducted in invasive ER + HER2 -The study was extended to a breast cancer PDX model. ST1056 tumors grew rapidly in the vehicle, palbociclib monotherapy, fulvestrant monotherapy, and fulvestrant and palbociclib combination groups, requiring animals to be removed from the study on day 17 (Figure 4D). Compound 1 resulted in sustained tumor growth suppression in the majority of animals for more than 49 days. The addition of palbociclib to compound 1 did not provide further efficacy but was well tolerable until day 71 when the group was discontinued. The most significant responses occurred in the combination of compound 1 and fulvestrant, as well as in the triple combination of compound 1, fulvestrant, and palbociclib. Tumor suppression lasted from day 28 to day 94 in all animals in both groups. Treatment was stopped on day 94, and moderate regrowth was observed only one month after treatment when the study was discontinued (Figure 4D). Monotherapy with compound 1, and tripartite therapy including compound 1 and palbociclib in combination with fulvestrant, were all well tolerated based on changes in body weight (Figure 4E).

Claims

1. A method for treating cancer in a person requiring cancer treatment, wherein the person requires, (a) Compound 1 or a pharmaceutically acceptable salt thereof, (b) A method comprising administering one or more independently selected additional therapeutic agents selected from the group consisting of selective estrogen receptor modulators (SERMs), selective estrogen receptor degraders (SERDs), CDK4 / 6 inhibitors, HER2 inhibitors, EGFR inhibitors, immune checkpoint inhibitors, MEK inhibitors, RAS inhibitors, RAF inhibitors, PIM (e.g., PIM1, PIM2, and PIM3) inhibitors, and any combination thereof.

2. The method according to claim 1, wherein the one or more independently selected additional therapeutic agents is one additional therapeutic agent.

3. The method according to claim 1 or 2, wherein the additional therapeutic agent is SERM / SERD.

4. The method according to claim 1 or 2, wherein the additional therapeutic agent is a CDK4 / 6 inhibitor.

5. The method according to claim 1 or 2, wherein the additional therapeutic agent is a HER2 inhibitor.

6. The method according to claim 1 or 2, wherein the additional therapeutic agent is an EGFR inhibitor.

7. The method according to claim 1 or 2, wherein the additional therapeutic agent is an immune checkpoint inhibitor.

8. The method according to claim 1 or 2, wherein the additional therapeutic agent is a MEK inhibitor.

9. The method according to claim 1 or 2, wherein the additional therapeutic agent is a RAS inhibitor.

10. The method according to claim 1 or 2, wherein the additional therapeutic agent is an RAF inhibitor.

11. The method according to claim 1, wherein the one or more independently selected additional therapeutic agents are two independently selected additional therapeutic agents.

12. The method according to claim 1 or 11, wherein one of the additional therapeutic agents is SERM / SERD, and the other additional therapeutic agents are a HER2 inhibitor, a CDK4 / 6 inhibitor, or a MEK inhibitor.

13. The method according to claim 1 or 11 to 12, wherein one of the additional therapeutic agents is SERM / SERD, and the other additional therapeutic agent is a HER2 inhibitor.

14. The method according to claim 1 or 11 to 12, wherein one of the additional therapeutic agents is SERM / SERD, and the other additional therapeutic agent is a CDK4 / 6 inhibitor.

15. The method according to claim 1 or 11 to 12, wherein one of the additional therapeutic agents is SERM / SERD, and the other additional therapeutic agent is a MEK inhibitor.

16. The method according to any one of claims 1 to 3 or 11 to 15, wherein the SERM / SERD is clomiphene, cyclophenyl, propalestrol, olmeroxifene, raloxifene, toremifene, rasofoxifene, bazedoxifene, ospemifene, enclomifene, cerophene, tamoxifene, fulvestrant, elastrant, camizestrant, lintodestrant, clotrimazole, parazestrant, or fenticonazole.

17. The method according to any one of claims 1 to 3 or 11 to 16, wherein the SERM / SERD is full vestrant.

18. The method according to any one of claims 1, 2, 4, 11, 12, 14, or 17, wherein the CDK4 / 6 inhibitor is palbociclib, ribociclib, abemaciclib, or trilaciclib.

19. The method according to any one of claims 1, 2, 4, 11, 12, 14, 17, or 18, wherein the CDK4 / 6 inhibitor is palbociclib.

20. The method according to any one of claims 1, 2, 4, 11, 12, 14, 17, or 18, wherein the CDK4 / 6 inhibitor is abemaciclib.

21. The method according to any one of claims 1, 2, 5, 11, 12, or 14, wherein the HER2 inhibitor is trastuzumab, pertuzumab, trastuzumab emtansine, famtrastuzumab deruxtecan, lapatinib, neratinib, dacomitinib, afatinib, tucatinib, erlotinib, pirotinib, tanespimycin, dacomitinib, peritinib, or margetuximab.

22. The method according to any one of claims 1, 2, 5, 11, 12, 14, or 21, wherein the HER2 inhibitor is lapatinib.

23. The method according to any one of claims 1, 2, 6, or 11, wherein the EGFR inhibitor is gefitinib, erlotinib, afatinib, neratinib, osimertinib, vandetanib, cetuximab, necitumumab, razertinib, amivantanab, or panitumumab.

24. The method according to any one of claims 1, 2, 7, or 11, wherein the immune checkpoint inhibitor is nivolumab, pembrolizumab, semiprimab, atezolizumab, durvalumab, avelumab, or ipilimumab.

25. The method according to any one of claims 1, 2, 4, 11, 12, 14, or 15, wherein the MEK inhibitor is darpiciclib, trametinib, cobimetinib, binimetinib, selumetinib, mildametinib, or pimacertib.

26. The method according to any one of claims 1, 2, 4, 11, 12, 14, 15, or 25, wherein the MEK inhibitor is trametinib.

27. The method according to any one of claims 1, 2, 4, 11, 12, 14, 15, or 25, wherein the MEK inhibitor is binimetinib.

28. The method according to any one of claims 1, 2, 9, or 11, wherein the RAS inhibitor is sotracib.

29. The method according to any one of claims 1, 2, 10, or 11, wherein the RAF inhibitor is vemurafenib, dabrafenib, encorafenib, sorafenib, berbarafenib, or napolafenib.

30. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant.

31. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant and lapatinib.

32. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant and abemaciclib.

33. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant and palbociclib.

34. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant and trametinib.

35. The method according to claim 1, wherein the one or more additional therapeutic agents are fulvestrant and binimetinib.

36. The method according to any one of claims 1 to 35, wherein the cancer is selected from breast cancer, lung cancer, endometrial cancer, esophageal cancer, stomach cancer, ovarian cancer, colorectal cancer, bladder cancer, head and neck cancer, thyroid cancer, prostate cancer, glioma, and cervical cancer.

37. The method according to any one of claims 1 to 36, wherein the cancer is breast cancer.

38. The aforementioned breast cancer is HER2 ＋ The method according to any one of claims 1 to 37, wherein the patient has breast cancer.

39. The aforementioned breast cancer is HER2 － The method according to any one of claims 1 to 37, wherein the patient has breast cancer.

40. The aforementioned breast cancer, ER ＋ The method according to any one of claims 1 to 37, wherein the patient has breast cancer.

41. The method according to any one of claims 1 to 37, wherein the breast cancer is triple-negative breast cancer.

42. The method according to any one of claims 1 to 36, wherein the cancer is lung cancer.

43. The method according to any one of claims 1 to 36, wherein the cancer is endometrial cancer.

44. The method according to any one of claims 1 to 36, wherein the cancer is esophageal cancer.

45. The method according to any one of claims 1 to 36, wherein the cancer is gastric cancer.

46. The method according to any one of claims 1 to 36, wherein the cancer is ovarian cancer.

47. The method according to any one of claims 1 to 36, wherein the cancer is colorectal cancer.

48. The method according to any one of claims 1 to 36, wherein the cancer is bladder cancer.

49. The method according to any one of claims 1 to 36, wherein the cancer is head and neck cancer.

50. The method according to any one of claims 1 to 36, wherein the cancer is thyroid cancer.

51. The method according to any one of claims 1 to 36, wherein the cancer is prostate cancer.

52. The method according to any one of claims 1 to 36, wherein the cancer is a glioma.

53. The method according to any one of claims 1 to 36, wherein the cancer is cervical cancer.

Images