WEE1 Compound for Treating Uterine Serous Carcinoma

Abstract

Disclosed herein are methods for identifying or selecting subjects having sensitivity to a WEE1 inhibitor, e.g., ZN-c3, and methods for using a WEE1 inhibitor, e.g., ZN-c3, to treat or inhibit cancer, including non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC).

Description

[Technical Field]

[0001] (Incorporation by reference of priority application) Any and all applications in which a claim of foreign or domestic priority is identified, for example, in an Application Data Sheet or claim filed with this application, including U.S. Provisional Application No. 63 / 370,580, filed August 5, 2022, which is expressly incorporated herein by reference in its entirety, are incorporated herein by reference pursuant to 37 CFR 1.57 and Rules 4.18 and 20.6. This application is related to International Application No. US2022 / 024079, which claims priority to U.S. Provisional Application No. 63 / 174,004, filed April 12, 2021, and U.S. Provisional Application No. 63 / 174,005, filed April 12, 2021, all of which are expressly incorporated herein by reference in their entireties.

[0002] FIELD OF THE INVENTION The present application generally relates to methods for identifying or selecting subjects or individuals having sensitivity to a WEE1 inhibitor, e.g., ZN-c3, and methods for using a WEE1 inhibitor, e.g., ZN-c3, to treat or inhibit cancer, including non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC). [Background technology]

[0003] DNA is constantly damaged by the environment. Light, chemicals, stress, and cellular replication result in single- or double-strand breaks along the DNA backbone. Organisms typically defend against DNA damage with repair proteins that reconnect or resynthesize damaged DNA. The correct functioning of these proteins is essential for life. Incorrect substitution of nucleotides into DNA can cause mutations (and other genetic alterations, including, but not limited to, insertions, deletions, and frameshifts), genetic disorders, and loss of protein function. A complete loss of DNA repair can lead to cell death, tumor progression, and cancer.

[0004] Cell cycle checkpoints are important for proper DNA repair and ensure that cells do not proceed with cell replication until their genome integrity is restored. WEE1 is a nuclear kinase involved in arresting the G2-M cell cycle checkpoint for DNA repair before the onset of mitosis. Normal cells repair damaged DNA during G1 arrest. Cancer cells often lack the G1-S checkpoint and rely on a functional G2-M checkpoint for DNA repair. WEE1 is overexpressed in various cancer types, and several inhibitors and / or decomposers of WEE1 are known to those skilled in the art. See, for example, International Publication Nos. 2019 / 173082 and 2020 / 069105.

[0005] Uterine serous carcinoma (USC) is a highly aggressive type II endometrial cancer. See, for example, Ferriss JS et al, Uterine serous carcinoma: key advances and novel treatment approaches. International Journal of Gynecologic Cancer 2021; 31:1165-1174. A phase II trial of adavosertib in recurrent uterine serous carcinoma has been reported. See Liu JF et al. Phase II study of the WEE1 inhibitor adavosertib in recurrent uterine serous carcinoma. J Clin Oncol 2021; 39. However, there are no existing therapies for treating USC and other cancers sensitive to WEE1 inhibitors. Existing therapeutic approaches remain limited, and therefore there is an urgent need for additional therapies and approaches to identify or select individuals who would benefit from such therapies. Summary of the Invention

[0006] Various embodiments described herein relate to methods of providing treatment, e.g., administration of the drug ZN-c3, to subjects or individuals with cancer, e.g., cancers that are sensitive to WEE1 inhibitors, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), and methods of identifying or selecting individuals or subjects that would benefit from receiving such treatment. The methods described herein may comprise administering a therapeutically effective amount of a WEE1 inhibitor, such as the drug ZN-c3, or a pharmaceutically acceptable salt thereof, to a subject or individual in need thereof, and may further comprise screening the subject or individual for modulation in a protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein, e.g., a polymorphism that confers sensitivity to a WEE1 inhibitor, e.g., the drug ZN-c3, e.g., a polymorphism selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W, in a biological sample obtained from the subject or individual. Also contemplated are methods for identifying or selecting a subject or individual having sensitivity to a WEE1 inhibitor, e.g., the drug ZN-c3, by analyzing a biological sample from such subject or individual to determine the presence or absence of a regulation in a protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein, e.g., a polymorphism that confers sensitivity to a WEE1 inhibitor, e.g., the drug ZN-c3, e.g., a polymorphism selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W. These and other embodiments are described in more detail below.

[0007] 1. A method for identifying or selecting a subject having sensitivity to the drug ZN-c3, comprising: identifying, in a biological sample obtained from such subject or individual, a regulation in protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such PPP2R1A protein, for example, one or more polymorphisms that confer sensitivity to the drug ZN-c3, preferably a polymorphism selected from P179R, S256F or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F or R183W, and selecting or identifying such subject as a subject having sensitivity to the drug ZN-c3 when such regulation, for example, any one or more polymorphisms in such PPP2R1A gene or protein, is identified.

[0008] 2. The method of alternative 1, further comprising administering ZN-c3 to said subject upon identification of such regulation or polymorphism in said biological sample.

[0009] 3. The method of any one of alternatives 1 or 2, wherein said subject has cancer.

[0010] 4. The method of alternative 3, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

[0011] 5. The regression or inhibition of such cancer is greater than 7%, e.g., 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more. The method according to any one of alternatives 2 to 4,

[0012] 6. The method of any one of alternatives 2-5, further comprising administering to said subject an agent or combination of agents that inhibits the PPP2R1A gene or protein.

[0013] 7. The method of alternative 6, wherein the agent or combination of agents comprises the PP2A catalytic inhibitor LB-100.

[0014] 8. The method of any one of alternatives 1-7, wherein the modulation of PPP2R1A comprises one or more gain-of-function mutations.

[0015] 9. The method of any one of alternatives 1-7, wherein the modulation of PPP2R1A comprises one or more loss-of-function mutations.

[0016] 10. The method of any one of alternatives 1-7, wherein modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

[0017] 11. The method of any one of alternatives 1-7, wherein the modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

[0018] 12. A method for inhibiting, ameliorating, or treating cancer or its sequelae in a subject, comprising identifying, in a biological sample obtained from such subject, a modulation in protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein, wherein such polymorphism confers sensitivity to the drug ZN-c3, for example a polymorphism selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W, and administering ZN-c3 to such subject when such polymorphism is identified in such biological sample.

[0019] 13. The method of alternative 12, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

[0020] 14. The method of alternative 12 or 13, wherein the cancer regression or inhibition is greater than 7%, e.g., 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

[0021] 15. The method of any one of alternatives 12-14, wherein said subject is further provided with an agent or combination of agents that inhibits the PPP2R1A gene or protein.

[0022] 16. The method of alternative 15, wherein the agent or combination of agents comprises the PP2A catalytic inhibitor LB-100.

[0023] 17. The method of any one of alternatives 12-16, wherein the modulation of PPP2R1A comprises one or more gain-of-function mutations.

[0024] 18. The method of any one of alternatives 12-16, wherein the modulation of PPP2R1A comprises one or more loss-of-function mutations.

[0025] 19. The method of any one of alternatives 12-16, wherein modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

[0026] 20. The method of any one of alternatives 12-16, wherein the modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

[0027] 21. The compound ZN-c3 for use in inhibiting, ameliorating, or treating cancer or its sequelae in a subject identified in a biological sample obtained from the subject as having a modulation in the protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or in a gene encoding such a PPP2R1A protein, for example a polymorphism that confers sensitivity to the drug ZN-c3, for example a polymorphism selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W.

[0028] 22. Compound ZN-c3 for use according to alternative 21, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

[0029] 23. Compound ZN-c3 for use according to alternative 21, wherein the regression or inhibition of cancer is more than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

[0030] 24. Compound ZN-c3 for use according to any one of alternatives 21 to 23, wherein said subject is further provided with a drug or combination of drugs that inhibits the PPP2R1A gene or protein.

[0031] 25. Compound ZN-c3 for use according to alternative 24, wherein such agent or combination of agents comprises the PP2A catalytic inhibitor LB-100.

[0032] 26. Compound ZN-c3 for use according to any one of alternatives 21 to 25, wherein the modulation of PPP2R1A comprises one or more gain-of-function mutations.

[0033] 27. Compound ZN-c3 for use according to any one of alternatives 21 to 25, wherein the modulation of PPP2R1A comprises one or more loss-of-function mutations.

[0034] 28. Compound ZN-c3 for use according to any one of alternatives 21 to 25, wherein the modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

[0035] 29. Compound ZN-c3 for use according to any one of alternatives 21 to 25, wherein the modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

[0036] 30. A method for inhibiting, ameliorating, or treating cancer or its sequelae in a subject, comprising administering to such subject an agent or combination of agents that inhibits the PPP2R1A gene or protein, and administering ZN-c3 to such subject.

[0037] 31. Any such drug or drug combination that inhibits the PPP2R1A gene or protein The method of alternative 30, wherein the method is provided to the subject separately.

[0038] 32. The method of alternative 30 or 31, wherein such agent or combination of agents that inhibits the PPP2R1A gene or protein is provided to such subject prior to administration of ZN-C3.

[0039] 33. The method of alternative 30, wherein such agent or combination of agents that inhibit the PPP2R1A gene or protein is provided to such subject at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 minute intervals.

[0040] 34. The method of any one of alternatives 30-33, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

[0041] 35. The method of any one of alternatives 30-34, wherein the cancer regression or inhibition is greater than 7%, e.g., 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

[0042] 36. The method of any one of alternatives 30-35, wherein such agent or combination of agents comprises the PP2A catalytic inhibitor LB-100.

[0043] 37. A product combination comprising the compound ZN-c3 and an agent or combination of agents that inhibit the PPP2R1A gene or protein for use in inhibiting, ameliorating, or treating cancer or its sequelae in a subject.

[0044] 38. The product combination for use according to alternative 37, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

[0045] 39. The product combination for use according to any one of alternatives 37 or 38, wherein the regression or inhibition of such cancer is greater than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

[0046] 40. A product combination for use according to any one of alternatives 37 to 39, wherein such agent or combination of agents comprises the PP2A catalytic inhibitor LB-100.

[0047] 41. Compound ZN-c3 for use according to any one of alternatives 1 to 20 or any one of alternatives 21 to 29, wherein the presence of such polymorphism is determined using next generation sequencing (NGS), sequencing, polymerase chain reaction (PCR), loop-mediated isothermal amplification, recombinase polymerase amplification, or antibody detection. [Brief explanation of the drawings]

[0048] [Figure 1] Figure 1 shows the cell density ratio of A427 (a primary non-small cell lung cancer cell line) cells after 72 hours of treatment with a 10-point dose response of ZN-c3, compared to cells transfected with a scrambled control siRNA. Cell density ratios were calculated using cell titer glow assays. PPP2R1A siRNA-transfected cells have an approximately 3.5-fold increase in sensitivity to ZN-c3 (Table 1). [Figure 2] 1 shows PPP2R1A mRNA expression in A427 cells treated with either scrambled control siRNA or siRNA against PPP2R1A. mRNA is expressed as a percentage of the scrambled control. [Figure 3] Figure 2 shows the cell density ratio of H1975 (a second non-small cell lung cancer cell line) cells after 72 hours of treatment with a 10-point dose response of ZN-c3, compared to cells transfected with a scrambled control siRNA. Cell density ratios were calculated using cell titer glow assays. PPP2R1A siRNA-transfected cells have an approximately 3.7-fold increase in sensitivity to ZN-c3 (Table 2). [Figure 4] 1 shows PPP2R1A mRNA expression in H1975 cells treated with either scrambled control siRNA or siRNA against PPP2R1A. mRNA is expressed as a percentage of the scrambled control. [Figure 5]Figure 3 shows the cell density ratio of Hs578T (breast cancer cell line) cells after 72 hours of treatment with a 10-point dose response of ZN-c3, compared to cells transfected with a scrambled control siRNA. Cell density ratios were calculated using cell titer glow assays. PPP2R1A siRNA-transfected cells have an approximately 2.4-fold increase in sensitivity to ZN-c3 (Table 3). [Figure 6] 1 shows the mRNA expression of PPP2R1A in Hs578T cells treated with either scrambled control siRNA or siRNA against PPP2R1A. mRNA is expressed as a percentage of the scrambled control. [Figure 7] Figure 4 shows the cell density ratio of SW620 (colorectal cancer cell line) cells after 72 hours of treatment with a 10-point dose response of ZN-c3, compared to cells transfected with a scrambled control siRNA. Cell density ratios were calculated using cell titer glow assays. PPP2R1A siRNA-transfected cells have an approximately 1.9-fold increase in sensitivity to ZN-c3 (Table 4). [Figure 8] 1 shows PPP2R1A mRNA expression in SW620 cells treated with either scrambled control siRNA or siRNA against PPP2R1A. mRNA is expressed as a percentage of the scrambled control. [Figure 9] Figure 5 shows the cell density percentage of SKOV3 (ovarian cancer cell line) cells after 72 hours of treatment with a 10-point dose response of ZN-c3, compared to cells transfected with a scrambled control siRNA. Cell density percentages were calculated using cell titer glow assays. PPP2R1A siRNA-transfected cells have an approximately 2.7-fold increase in sensitivity to ZN-c3 (Table 5). [Figure 10]1 shows PPP2R1A mRNA expression in SKOV3 cells treated with either scrambled control siRNA or siRNA against PPP2R1A. mRNA is expressed as a percentage of the scrambled control. [Figure 11] The cell density ratios of HEC1B and HEC-59 (endometrial cancer cell lines) cells after 72-hour treatment with a 10-point dose response of ZN-c3 are shown for cells transfected with siRNA against PPP2R1A compared with cells transfected with scrambled control siRNA. HEC1B has a missense mutation (W257L, L429F) in PPP2R1A, which reduces the effectiveness of siRNA against PPP2R1A in inducing sensitivity to ZN-c3. HEC-59 has wild-type PPP2R1A. Cell density ratios were calculated using cell titer glow assays. HEC1B PPP2R1A siRNA-transfected cells had an approximately 1.4-fold increase in sensitivity to ZN-c3, while HEC-59 PPP2R1A siRNA-transfected cells had an approximately 2.5-fold increase in sensitivity to ZN-c3 (Table 6). [Figure 12] Figure 1 shows Western blot analysis of endometrial cancer cell lines HEC1B and HEC-59 with and without siRNA-mediated knockdown of PPP2R1A, with vinculin used as a loading control. [Figure 13] Figure 7 shows the percent cell density of SKOV3 ovarian cancer cells after 72 hours of treatment with a 10-point dose response of ZN-c3, comparing cells transduced with a non-targeting guide RNA to cells with a gene knockout of PPP2R1A after transduction with a CRISPR guide RNA targeting PPP2R1A. Percent cell density is calculated using a cell titer glow assay. PPP2R1A CRISPR knockout cells have an approximately 1.7-fold increase in sensitivity to ZN-c3 (Table 7). [Figure 14] Figure 1 shows a Western blot analysis of the ovarian cancer cell line SKOV3 with and without CRISPR-mediated gene knockout of PPP2R1A, with β-actin used as a loading control. [Figure 15]Figure 8 shows the percent cell density of H1975 lung cancer cells after 72 hours of treatment with a 10-point dose response of ZN-c3, comparing cells transduced with a non-targeting guide RNA to cells with a gene knockout of PPP2R1A after transduction with a CRISPR guide RNA targeting PPP2R1A. Percent cell density is calculated using a cell titer glow assay. PPP2R1A CRISPR knockout cells have an approximately 1.9-fold increase in sensitivity to ZN-c3 (Table 8). [Figure 16] Figure 1 shows a Western blot analysis of H1975 lung cancer cells with and without CRISPR-mediated gene knockout of PPP2R1A, with β-actin used as a loading control. [Figure 17] Figure 9 shows the cell density percentage of OV17R ovarian cancer cells after 72 hours of treatment with a 10-point dose response of ZN-c3. OV17R harbors a S256F hotspot missense mutation in PPP2R1A. This cell line is highly sensitive to ZN-c3 treatment, likely due in part to the PPP2R1A mutation (Table 9). Cell density percentages are calculated using cell titer glow assays. [Figure 18] Figure 1 shows the percent inhibition of cell proliferation in HEC-59 endometrial cancer cells after treatment with ZN-c3 alone, the PP2A catalytic inhibitor LB-100 alone, or the combination of ZN-c3 and LB-100. The HEC-59 cell line is PPP2R1A wild-type. Single-agent treatment with either compound is not as effective as the combination therapy. Percent inhibition is calculated using a cell titer glow assay. The doses of each compound are shown in Table 10. [Figure 19] Table 11 shows the inhibition rate of cell proliferation in OV17R ovarian cancer cells after treatment with ZN-c3 alone, the PP2A catalytic inhibitor LB-100 alone, or the combination of ZN-c3 and LB-100. OV17R harbors a PPP2R1A missense mutation, S256F. The inhibition rate was calculated using a cell titer glow assay. The dose of each compound is shown in Table 11. [Figure 20]Figure 1 shows the percent inhibition of cell proliferation in HEC-59 endometrial cancer cells after treatment with ZN-c3 alone, the PP2A catalytic inhibitor LB-100 alone, or the combination of ZN-c3 and LB-100 following siRNA-mediated knockdown of PPP2R1A. Knockdown of PPP2R1A leads to increased sensitivity to ZN-c3 both as a single agent and in combination therapy. The percent inhibition is calculated using a cell titer glow assay. The doses of each compound are shown in Table 12. [Figure 21] Figure 1 shows the percent inhibition of cell proliferation in HEC1B endometrial cancer cells after treatment with ZN-c3 alone, the PP2A catalytic inhibitor LB-100 alone, or the combination of ZN-c3 and LB-100 following siRNA-mediated knockdown of PPP2R1A. Knockdown of PPP2R1A results in a slight increase in sensitivity, as previously observed. Percent inhibition is calculated using a cell titer glow assay. The doses of each compound are shown in Table 13. DETAILED DESCRIPTION OF THE INVENTION

[0049] WEE1 is a tyrosine kinase that is a key component of the ATR-mediated G2 cell cycle checkpoint control, which blocks the entry of mitosis in response to cellular DNA damage. ATR phosphorylates and activates CHK1, which in turn activates WEE1, leading to the selective phosphorylation of cyclin-dependent kinase 1 (CDK1) at Tyr15. WEE1 inhibits the G2 checkpoint, thereby stabilizing the CDK1-cyclin B complex and halting cell cycle progression. This process confers a survival advantage by allowing tumor cells time to repair damaged DNA before the onset of mitosis. Suppressing WEE1 inhibits the G2 checkpoint, causing cancer cells with DNA damage to enter unscheduled mitosis and undergo cell death by mitotic apoptosis. Therefore, inhibiting and / or degrading WEE1 has the potential to sensitize tumors to DNA-damaging agents such as cisplatin and induce tumor cell death.

[0050] definition Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. All patents, applications, published applications, and other publications referenced herein are incorporated by reference in their entirety unless otherwise stated. In the event that there are multiple definitions for a term herein, those in this section prevail unless stated otherwise.

[0051] As used herein, the terms "WEE1 inhibition," "WEE1 inhibitor," and similar terms refer to inhibiting the activity or function of WEE1 tyrosine kinase, for example, by degrading WEE1 tyrosine kinase and / or by reducing the activity of WEE1 tyrosine kinase in mediating the phosphorylation of CDK1. WEE1 inhibitors that function by degrading WEE1 tyrosine kinase may be referred to herein as WEE1 degraders.

[0052] As used herein, "subject" refers to an animal that is the object of treatment, observation, or experiment. The subject animal may be a mammal, such as, without limitation, a mouse, rat, rabbit, guinea pig, dog, cat, sheep, goat, cow, horse, primate (such as monkey, chimpanzee, and ape), and, in particular, a human. In some embodiments, the subject may be a human. In some embodiments, the subject may be a child and / or infant. In other embodiments, the subject may be an adult.

[0053] As used herein, the terms "treat," "treating," "treatment," "therapeutic," and "therapy" do not necessarily mean a complete cure or elimination of a disease or condition. Any alleviation of any undesirable signs or symptoms of a disease or condition, to any degree, can be considered treatment and / or therapy. Furthermore, treatment can include actions that may worsen a subject's overall feeling of health or appearance.

[0054] The terms "therapeutically effective amount" and "effective amount" are used to refer to the amount of an active compound (e.g., ZN-c3 or a pharmaceutically acceptable salt thereof) that elicits the indicated biological or medical response. For example, a therapeutically effective amount of such a ZN-c3 compound, salt, or composition may be the amount necessary to prevent, alleviate, or ameliorate the symptoms of a disease or condition, or to prolong the survival of the subject being treated. This response may occur in a tissue, system, animal, or human, and may include alleviation of signs or symptoms of the disease or condition being treated. Determining an effective amount is well within the capabilities of one of ordinary skill in the art in light of the disclosure provided herein. The therapeutically effective amount of a ZN-c3 compound, salt, or composition required as a dose will depend on the route of administration, the type of animal, including humans, being treated, and the physical characteristics of the particular animal under consideration. Dosages can be adjusted to achieve the desired effect and will depend on factors such as body weight, diet, concomitant medications, and other factors that would be recognized by those skilled in the medical field.

[0055] For example, an effective amount of a ZN-c3 compound, salt, or composition can be an amount that results in (a) reduction, alleviation, or elimination of one or more symptoms caused by USC, (b) reduction in tumor size, (c) tumor elimination, and / or (d) long-term disease stabilization (growth arrest) of the tumor. For example, an effective amount of a ZN-c3 compound, salt, or composition can be an amount that results in reduced WEE1 activity and / or phosphorylation (e.g., CDC2 phosphorylation). Reductions in WEE1 activity are known to those skilled in the art and can be determined by assaying WEE1 endogenous kinase activity and phosphorylation of downstream substrates.

[0056] As used herein, the term "dosing regimen" refers to the manner in which a ZN-c3 compound is administered to a subject, including the route of administration, the dose, and the dosing interval. A dosing regimen may include a "cyclic" dosing phase in which a specific dosage (e.g., 300 mg) is administered at regular intervals (e.g., once a day) for a specific period (e.g., 3 days). A dosing regimen may also include an "intermittent" dosing phase in which one or more dosing parameters, such as the dosage and / or the dosing interval, are varied or altered. For example, an intermittent dosing phase may include a "rest" phase in which the ZN-c3 compound is not administered or is administered at a reduced dosage and / or less frequently.

[0057] It is understood that the compounds described herein can be isotopically labeled. Substitution with isotopes such as deuterium can result in certain therapeutic advantages due to greater metabolic stability, such as increased in vivo half-life or reduced dosage requirements. Each chemical element represented in a compound structure may include any isotope of that element. For example, in a compound structure, a hydrogen atom may be explicitly disclosed or understood as being present in the compound. At any position in a compound where a hydrogen atom can be present, the hydrogen atom may be any isotope of hydrogen, including, but not limited to, hydrogen-1 (protium) and hydrogen-2 (deuterium). Thus, reference to a compound herein encompasses all possible isotopic forms unless the context clearly indicates otherwise.

[0058] When a range of values is provided, it is understood that the upper and lower limits, and every intervening value between the upper and lower limits of that range, are encompassed within an embodiment.

[0059] Terms and phrases used in this application, and variations thereof, particularly in the appended claims, should be construed as open-ended rather than limiting, unless expressly stated. For the foregoing examples, the term "including" should be construed to mean "including without limitation," "including but not limited to," etc.; as used herein, the term "comprising" is synonymous with "including," "containing," or "featuring," is inclusive or open-ended, and does not exclude additional, unrecited elements or method steps; the term "having" should be construed as "having at least," the term "including" should be construed as "including, but not limited to," and the term "example" is used to provide illustrative examples rather than an exhaustive or exclusive list of items under discussion. The use of terms such as "preferably," "preferred," "desired," or "desirable," and words of similar import, should not be understood as implying that a particular feature is critical, essential, or even important to its structure or function, but rather is intended merely to highlight alternative or additional features that may or may not be utilized in a particular embodiment. Additionally, the term "comprising" is intended to be synonymous with the phrases "having at least" or "including at least." When used in the context of a compound, composition, or device, the term "comprising" means that the compound, composition, or device includes at least the recited features or components, but may include additional features or components.

[0060] With respect to the use of virtually any plural and / or singular term herein, those skilled in the art can convert from plural to singular and / or from singular to plural as appropriate depending on the context and / or application. Various singular / plural permutations may be expressly stated herein for clarity. The indefinite article "a" or "an" does not exclude a plurality. The mere fact that certain measures are recited in mutually different dependent claims does not indicate that a combination of these measures cannot be used to advantage. Any reference signs in the claims should not be construed as limiting the scope thereof.

[0061] In some embodiments, methods for selecting or identifying subjects with sensitivity to the drug ZN-c3 are provided. In some embodiments, the methods include identifying or detecting modulation in protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein. For example, such modulation may be overexpression or underexpression of the PPP2R1A protein or a transcript encoding the PPP2R1A protein, or a polymorphism in the PPP2R1A protein or a gene or transcript encoding the PPP2R1A protein. In some embodiments, a polymorphism that confers sensitivity to the drug ZN-c3 is identified or detected in such methods. In some such embodiments, the polymorphism is selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W. The aforementioned regulation of the PPP2R1A protein or the gene or transcript encoding the PPP2R1A protein is determined by analyzing a biological sample obtained from such a subject with cancer, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC). In some embodiments, the method includes selecting such a subject as being sensitive to the drug ZN-c3 when any one or more of the aforementioned polymorphisms in the PPP2R1A gene or protein are identified. In some embodiments, once a subject is identified or selected as being sensitive to ZN-c3, the identified or selected subject is provided with ZN-c3, e.g., administered a ZN-c3 dosing regimen.

[0062] In some embodiments, methods are provided for inhibiting, ameliorating, or treating cancer, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), or its sequelae, in a subject. In some embodiments, the methods include identifying or detecting modulation in protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein; for example, such modulation may be overexpression or underexpression of the PPP2R1A protein or a transcript encoding the PPP2R1A protein, or a polymorphism in the PPP2R1A protein or a gene or transcript encoding the PPP2R1A protein. In some embodiments, a polymorphism that confers sensitivity to the drug ZN-c3 is identified or detected in such methods. In some embodiments, the polymorphism is selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W, in a biological sample obtained from such a subject. In some embodiments, ZN-c3 is administered to such a subject when such a polymorphism is identified in such a biological sample.

[0063] In some embodiments, the compound ZN-c3 is provided for use in inhibiting, ameliorating, or treating cancer or its sequelae in a subject identified as having a modulation in a protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding such a PPP2R1A protein, and detecting a polymorphism in a biological sample obtained from such subject, e.g., a polymorphism selected from any one or more of P179R, S256F, or R183W, or a polymorphism in a biological sample obtained from such subject. Nucleic acids encoding polymorphisms selected from any one or more of W confer sensitivity to the drug ZN-c3. In some such uses, the cancer is, for example, non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC).

[0064] In some embodiments, methods are provided for inhibiting, ameliorating, or treating cancer, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), or sequelae thereof, in a subject. In some embodiments, the methods include administering to the subject an agent or combination of agents that inhibits the PPP2R1A gene or protein, and administering ZN-c3 to the subject.

[0065] In some embodiments, a product combination is provided that includes the compound ZN-c3 and an agent or combination of agents that inhibit the PPP2R1A gene or protein for use in inhibiting, ameliorating, or treating cancer in a subject, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), or sequelae thereof.

[0066] In some embodiments, the method comprises administering ZN-c3 to the subject when a regulation in the PPP2R1A protein or a gene or transcript encoding the PPP2R1A protein, e.g., a polymorphism, preferably a polymorphism selected from any one or more of P179R, S256F, or R183W, or a polymorphism in a gene encoding any one or more of the polymorphisms selected from P179R, S256F, or R183W, is identified in the biological sample.

[0067] In some embodiments, the subject has cancer.

[0068] In some embodiments, the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, or endometrial cancer.

[0069] In some embodiments, such cancer regression or inhibition is greater than 7%, e.g., 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

[0070] In some embodiments, the methods further comprise administering to such a subject an agent or combination of agents that inhibits the PPP2R1A gene or protein.

[0071] In some embodiments, the agent or combination of agents provided to such a subject comprises the PP2A catalytic inhibitor LB-100.

[0072] In some embodiments, such agents or combinations of agents that inhibit the PPP2R1A gene or protein are provided separately to such a subject.

[0073] In some embodiments, such an agent or combination of agents that inhibits the PPP2R1A gene or protein is provided to such a subject prior to administration of ZN-C3.

[0074] In some embodiments, such agents or combinations of agents that inhibit the PPP2R1A gene or protein include 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 1 provided to such subject at 3, 14, 15, 16, 17, 18, 19, or 20 minute intervals.

[0075] In some embodiments, the presence of any one or more such polymorphisms, e.g., polymorphisms in genes encoding P179R, S256F, or R183W, is determined using next generation sequencing (NGS), sequencing, polymerase chain reaction (PCR), loop-mediated isothermal amplification, recombinase polymerase amplification, or antibody detection.

[0076] In some embodiments, the modulation of PPP2R1A comprises one or more gain-of-function mutations.

[0077] In some embodiments, the modulation of PPP2R1A comprises one or more loss-of-function mutations.

[0078] In some embodiments, modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

[0079] In some embodiments, the modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

[0080] WEE1 compound ZN-c3 is a selective, orally bioavailable small molecule inhibitor of WEE1, a key component of the G2 / M cell cycle checkpoint that prevents cells from entering mitosis and allows DNA damage to be repaired. ZN-c3 has demonstrated significant in vitro antitumor activity in multiple cell lines and xenograft models.

[0081] [Table 1]

[0082] ZN-c3 can be prepared as described in WO 2019 / 173082 (see, e.g., Example 9B therein), which is expressly incorporated herein by reference, particularly for the purpose of describing methods for making ZN-c3, as well as methods for making salts and pharmaceutical compositions thereof. If there is any discrepancy between the chemical name and structure of ZN-c3, the structure should be more significant.

[0083] Pharmaceutical Composition Some embodiments described herein relate to pharmaceutical compositions that may include an effective amount of a ZN-c3 compound or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier, diluent, excipient, or a combination thereof.

[0084] The term "pharmaceutical composition" refers to a mixture of ZN-c3 and / or its pharmaceutically acceptable salts with other chemical components, such as diluents or carriers. Pharmaceutical compositions facilitate the administration of a compound to an organism. Pharmaceutical compositions can also be obtained by reacting a compound with an inorganic or organic acid, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. Pharmaceutical compositions are generally and tailored to the specific intended route of administration.

[0085] The term "physiologically acceptable" defines a carrier, diluent, or excipient that does not neutralize the biological activity and properties of the ZN-c3 compound or cause substantial damage or injury to the animal to which the composition is intended to be delivered.

[0086] As used herein, "carrier" refers to a compound that facilitates the incorporation of a compound into cells or tissues. For example, and without limitation, dimethyl sulfoxide (DMSO) is a commonly used carrier that facilitates the uptake of many organic compounds into cells or tissues of a subject.

[0087] As used herein, "diluent" refers to an ingredient in a pharmaceutical composition that has no apparent pharmacological activity but may be pharmaceutically necessary or desirable. For example, a diluent may be used to bulk a potent drug whose mass is too small for manufacture and / or administration. A diluent may also be a liquid for dissolving a drug to be administered by injection, ingestion, or inhalation. A common form of diluent in the art is a buffered aqueous solution, such as, without limitation, phosphate buffered saline, which mimics the pH and isotonicity of human blood.

[0088] As used herein, "excipient" refers to an essentially inert substance added to a pharmaceutical composition to provide the composition with, but not limited to, bulk, consistency, stability, binding ability, lubrication, disintegration ability, etc. For example, stabilizers such as antioxidants and metal chelators are excipients. In one embodiment, the pharmaceutical composition includes an antioxidant and / or a metal chelator. A "diluent" is a type of excipient.

[0089] The pharmaceutical compositions described herein can be administered to human patients either by themselves or in pharmaceutical compositions in which they are mixed with other active ingredients, such as in combination therapy, or with carriers, diluents, excipients, or combinations thereof. The appropriate formulation depends on the selected route of administration. Techniques for formulating and administering the compounds described herein are known to those skilled in the art.

[0090] The pharmaceutical compositions disclosed herein can be prepared in a manner known per se, for example, by conventional mixing, dissolving, granulating, dragee-making, elutriating, emulsifying, encapsulating, entrapping, or tableting processes. Additionally, the ZN-c3 active ingredient is typically contained in an amount effective to achieve its intended purpose, and can be provided as a salt with a pharmaceutically compatible counterion.

[0091] Multiple art-recognized techniques for administering ZN-c3 compounds, salts, and / or compositions can be used, including, but not limited to, oral, rectal, pulmonary, topical, aerosol, injection, infusion, and parenteral delivery, including intramuscular, subcutaneous, intravenous, intramedullary injection, intrathecal, direct intraventricular, intraperitoneal, intranasal, and intraocular injection. In some embodiments, ZN-c3 or a pharmaceutically acceptable salt thereof can be administered orally.

[0092] The compositions may, if desired, be presented in a pack or dispenser device, which may contain one or more unit dosage forms containing the active ingredient. The pack may, for example, comprise metal or plastic foil, such as a blister pack. The pack or dispenser device may be accompanied by instructions for administration. The pack or dispenser may also be accompanied by a notice associated with the container in a format prescribed by a government agency regulating the manufacture, use, or sale of pharmaceuticals, which notice reflects the agency's approval of the drug form for human or animal administration. Such notice may, for example, be the label approved by the U.S. Food and Drug Administration for prescription drugs, or an approved product insert. The compatible compositions described herein may be Compositions comprising ZN-c3 and / or salts formulated in a suitable pharmaceutical carrier may also be prepared, placed in an appropriate container, and labeled for treatment of an indicated condition.

[0093] Uses and Methods of Treatment or Inhibition Some embodiments described herein relate to methods of treating or inhibiting cancer in a subject, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), comprising administering to the subject a therapeutically effective amount of ZN-c3 or a pharmaceutically acceptable salt thereof, preferably according to a dosing regimen.

[0094] Some embodiments described herein relate to the use of a therapeutically effective amount of ZN-c3 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating a subject with cancer, e.g., non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), preferably according to a dosing regimen. Other embodiments relate to ZN-c3 for use in treating, inhibiting, or ameliorating cancer in a subject, e.g., non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), comprising administering a therapeutically effective amount of ZN-c3 or a pharmaceutically acceptable salt thereof to the subject, preferably according to a dosing regimen. In various embodiments, the dosing regimen comprises oral administration of ZN-c3 to the subject. In many of these embodiments, the subject is identified or selected as having susceptibility to ZN-c3, and such susceptibility can be determined by analyzing a biological sample from the subject for regulation of the PPP2R1A protein or gene encoding PPP2R1A, e.g., overexpression of the PPP2R1A protein or transcript encoding the PPP2R1A protein, or underexpression of the PPP2R1A protein or transcript encoding the PPP2R1A protein. In some embodiments, the regulation of the PPP2R1A protein or gene or transcript encoding the PPP2R1A protein relates to a polymorphism in the PPP2R1A protein or gene or transcript encoding the PPP2R1A protein, e.g., any one or more of P179R, S256F, or R183W polymorphisms, or a gene or transcript encoding the P179R, S256F, or R183W polymorphism.

[0095] In various embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is 300 mg or more per day. For example, in some embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is about 300 mg per day. In other embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is about 350 mg per day. In other embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is about 200 mg per day. In some embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is 300 mg or more once per day (QD). In one embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 300 mg once per day (QD). In another embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 350 mg once per day (QD). In yet another embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 200 mg once per day (QD). In some embodiments, the therapeutically effective amount of ZN-c3 administered to a subject is 150 mg or more twice daily (BID). In one embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 150 mg twice daily (BID). In another embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 175 mg twice daily (BID). In yet another embodiment, the therapeutically effective amount of ZN-c3 administered to a subject is about 100 mg twice daily (BID).

[0096] In various embodiments, the dosing regimen includes a cyclical dosing phase in which the dose and frequency of ZN-c3 administration to the subject are fixed for a period of time, e.g., three or more consecutive days. For example, in one embodiment, the cyclical dosing phase includes a daily dose in the range of about 200 mg per day to about 350 mg per day, which is fixed for a period of at least three consecutive days. In one embodiment, the daily dose is administered once daily (QD). In another embodiment, the daily dose is administered twice daily. In one embodiment, the cyclical dosing phase comprises once-daily dosing for a period of at least three consecutive days.

[0097] In various embodiments, the dosing regimen includes an intermittent dosing phase in which the dose and frequency of ZN-c3 administration to the subject are altered. In one embodiment, the intermittent dosing phase includes at least one change in the amount of ZN-c3 administered to the subject daily. In one embodiment, the intermittent dosing phase includes at least one drug-free day, e.g., 1, 2, 3, 4, 5, 6, or 7 drug-free days. For example, in one embodiment, the dosing regimen includes a cyclical phase in which ZN-c3 is orally administered to the subject once daily for 5 days, followed by an intermittent or drug-free phase in which ZN-c3 is not administered for 2 days. Such a dosing regimen, which may be referred to as 5 days on / 2 days off, can be repeated or cycled as frequently as needed, depending on the particular case.

[0098] In various embodiments, the subject is a human. In some embodiments, the subject has cancer, e.g., non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC). In some embodiments, the USC is advanced USC. In some embodiments, the subject has USC that is recurrent USC.

[0099] In various embodiments, the subject has been previously treated with a previous treatment regimen for cancer, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC). Various treatment regimens are known to those skilled in the art or are under development. In one embodiment, the previous treatment regimen may include administration of a drug, antibody, CAR T cells, surgery, and / or radiation prior to administration of ZN-c3. For example, in various embodiments, the previous treatment may include at least one selected from carboplatin, paclitaxel, bevacizumab, trastuzumab, pembrolizumab, lenvatinib, or doxorubicin.

[0100] The amount of ZN-c3 compound or its pharmaceutically acceptable salt that is effective in treating cancer in a particular subject, such as non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, endometrial cancer, or uterine serous carcinoma (USC), will vary depending not only on the specific compound or salt selected, but also on the route of administration, the nature and / or symptoms of the disease or condition being treated, and the age and condition of the patient, and is ultimately at the discretion of the attending physician or clinician. When administering a pharmaceutically acceptable salt, the dosage may be calculated as the free base. As will be understood by those skilled in the art, in certain circumstances, it may be necessary to administer ZN-c3 compound, salt, or composition in amounts that exceed or even far exceed the dosage ranges described herein to effectively and aggressively treat particularly aggressive cases of USC.

[0101] The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example, as two, three, four or more sub-doses per day The sub-dose itself may be further divided, for example, into a number of discrete loosely spaced administrations.

[0102] As will be readily apparent to those skilled in the art, the useful in vivo dosage and specific mode of administration will vary depending on the age, weight, severity of the affliction, the mammalian species being treated, the specific ZN-c3 compound, salt, or composition used, and the specific application for which the therapy is being used. Determining effective dosage levels, i.e., the dosage levels necessary to achieve the desired results, can be accomplished by those skilled in the art using routine methods, such as human clinical trials, in vivo studies, and in vitro studies. For example, useful dosages of ZN-c3 or its pharmaceutically acceptable salts can be determined by comparing their activity and in vivo activity in animal models. Such comparisons can be made with established drugs such as carboplatin and / or paclitaxel.

[0103] Dosage amount and interval are adjusted individually to provide plasma levels of the active moiety sufficient to maintain a modulating effect or minimal effective concentration (MEC). The MEC may be determined by the MEC. The MEC varies for each compound but can be estimated from in vivo and / or in vitro data. The dosage required to achieve the MEC will depend on individual characteristics and the route of administration. However, HPLC assays or bioassays can be used to determine plasma concentrations. Dosage intervals can also be determined using the MEC value. Compositions should be administered using a regimen that maintains plasma levels above the MEC for 10-90% of the time, preferably 30-90% of the time, and most preferably 50-90% of the time. In cases of local administration or selective uptake, the effective local concentration of a drug may not be related to plasma concentration.

[0104] It should be noted that the attending physician would know how and when to terminate, interrupt, or adjust administration due to toxicity or organ dysfunction. Conversely, the attending physician would also know to adjust treatment to higher levels if the clinical response is not adequate (precluding toxicity). The magnitude of the dose administered in the management of the disease of interest will vary with the severity of the disease or condition to be treated and the route of administration. The severity of the disease or condition can, for example, be assessed, in part, by standard prognostic evaluation methods. Furthermore, the dose, and perhaps dose frequency, will also vary with the age, weight, and response of the individual patient. Programs equivalent to those discussed above can be used in veterinary medicine.

[0105] The ZN-c3 compounds, salts, and compositions disclosed herein can be evaluated for efficacy and toxicity using known methods. For example, toxicology for ZN-c3 can be established by determining in vitro toxicity against cell lines, such as mammalian cell lines, and preferably human cell lines. The results of such studies often predict toxicity in animals, such as mammals, or particularly humans. Alternatively, the toxicity of a particular compound in an animal model, such as a mouse, rat, rabbit, dog, or monkey, can be determined using known methods. The efficacy of a particular compound can be established using several recognized methods, such as in vitro methods, animal models, or human clinical trials. When selecting a model for determining efficacy, those skilled in the art can rely on the state of the art to guide them in selecting the appropriate model, dose, route of administration, and / or regimen.

[0106] In some embodiments, a subject's susceptibility to ZN-c3 is determined by analyzing the presence of one or more PPP2R1A polymorphisms, such as one or more of P179R, S256F, or R183W, or a gene or transcript encoding the P179R, S256F, or R183W polymorphism. Such polymorphisms can be identified, for example, by using whole genome sequencing, including next-generation sequencing (NGS). In some embodiments, PPP2R1A polymorphisms are identified using polymerase chain reaction (PCR)-based methods. In some embodiments, PCR-based methods include allele-specific PCR (ASPCR) using TaqMan probes, high-resolution melting analysis (HRM), digital PCR, and cold-PCR (cold-PCR). In some embodiments, PPP2R1A polymorphisms are identified using mass spectrometry. In some embodiments, the mass spectrometry used may be matrix-assisted laser desorption / ionization time-of-flight (MALDI-TOF-MS). In some embodiments, PPP2R1A polymorphisms are identified using small nucleotide polymorphism (SNP) arrays. In some embodiments, PPP2R1A polymorphisms are identified using commercially available SnAPshot multiplex kits. In some embodiments, PPP2R1A polymorphisms are identified using denaturing high performance liquid chromatography (DHPLC).

[0107] In some embodiments, the primers used in the NGS or PCR reaction are It is read through the locus or its complement.

[0108] In some embodiments, the PPP2R1A mutation is a hotspot mutation. In some embodiments, the genomic locus for each codon where the hotspot is located is S256:hg19:chr19:52,716,321-52,716,323, R183:hg19:chr19:52,715,981-52,715,983, and / or P179:hg19:chr19:52,715,969-52,715,971. In some embodiments, these hotspot mutations are found in uterine cancer, ovarian cancer, breast cancer, lung cancer, colorectal cancer, prostate cancer, salivary gland cancer, or other cancers.

[0109] In some embodiments, modulation in PPP2R1A, such as the polymorphisms described above, is detected. In some embodiments, the modulation comprises one or more gain-of-function mutations in PPP2R1A. In some embodiments, the modulation comprises one or more loss-of-function mutations in PPP2R1A. In some embodiments, the modulation comprises one or more mutations that result in overexpression of PPP2R1A. In some embodiments, the modulation comprises one or more mutations that result in underexpression of PPP2R1A. [Example]

[0110] Example 1 Dose response of cancer cell lines sensitized with PPP2R1A siRNA to ZN-c3 The effect of PPP2R1A knockdown on cancer cell susceptibility to ZN-c3 was evaluated. Various cancer cell lines were transfected with PPP2R1A siRNA or a control scrambled siRNA, and the cells were treated with a 10-point dose-response of ZN-c3 for 72 hours. Cell confluency was calculated using cell titer glow assays. PPP2R1A siRNA targeted the following bases: Chr.19:52189802-52226425. siRNA transfection was initiated 48 hours before treatment with ZN-c3. ZN-c3 dose-response assessment included ZN-c3 doses of 10 μM, 3.33 μM, 1.11 μM, 370.3 nM, 123.4 nM, 41.1 nM, 13.7 nM, 4.5 nM, and 1.5 nM.

[0111] The first non-small cell lung cancer cell line, A427, was treated with PPP2R1A siRNA or control siRNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the decrease in cell density (Figure 1). PPP2R1A siRNA-mediated knockdown of PPP2R1A was evaluated by comparing PPP2R1A expression in siCTRL and siPPPR12A-treated conditions (Figure 2). Knockdown of PPP2R1A by siRNA was successful, and A427 cells transduced with siPPP2R1A showed a 3.5-fold increase in sensitivity to ZN-c3 (Table 1).

[0112] [Table 2]

[0113] A second non-small cell lung cancer cell line, H1975, was treated with PPP2R1A siRNA or control siRNA, and transfected cells were evaluated for sensitivity to ZN-c3 by measuring the reduction in cell density (Figure 3). PPP2R1A siRNA-mediated knockdown of PPP2R1A was assessed by comparing PPP2R1A expression in siCTRL- and siPPPR12A-treated conditions (Figure 4). Knockdown of PPP2R1A by siRNA was successful, and cells transfected with siPPP2R1A were significantly less sensitive to ZN-c3. H1975 cells showed a 3.7-fold increase in sensitivity to ZN-c3 (Table 2).

[0114] [Table 3]

[0115] The breast cancer cell line Hs578T was treated with PPP2R1A siRNA or control siRNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the decrease in cell density (Figure 5). PPP2R1A siRNA-mediated knockdown of PPP2R1A was evaluated by comparing PPP2R1A expression in the siCTRL and siPPPR12A-treated conditions (Figure 6). Knockdown of PPP2R1A by siRNA was successful, and Hs578T cells transduced with siPPP2R1A showed a 2.4-fold increase in sensitivity to ZN-c3 (Table 3).

[0116] [Table 4]

[0117] The colorectal cancer cell line SW620 was treated with PPP2R1A siRNA or control siRNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the decrease in cell density (Figure 7). PPP2R1A siRNA-mediated knockdown of PPP2R1A was evaluated by comparing PPP2R1A expression in siCTRL and siPPPR12A-treated conditions (Figure 8). Knockdown of PPP2R1A by siRNA was successful, and SW620 cells transduced with siPPP2R1A showed a 1.9-fold increase in sensitivity to ZN-c3 (Table 4).

[0118] [Table 5]

[0119] The ovarian cell line SKOV3 was treated with PPP2R1A siRNA or control siRNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the decrease in cell density (Figure 9). PPP2R1A siRNA-mediated knockdown of PPP2R1A was evaluated by comparing PPP2R1A expression in siCTRL and siPPPR12A-treated conditions (Figure 10). Knockdown of PPP2R1A by siRNA was successful, and SKOV3 cells transduced with siPPP2R1A showed a 2.7-fold increase in sensitivity to ZN-c3 (Table 5).

[0120] [Table 6]

[0121] Endometrial cancer cell lines HEC1B and HEC-59 were treated with PPP2R1A siRNA or control siRNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the reduction in cell density (Figure 11). PPP2R1A siRNA-mediated knockdown of PPP2R1A was evaluated by Western blot analysis of PPP2R1A protein expression in siPPPR12A-treated cells, compared with vinculin, which was used as a control (Figure 12). siRNA-mediated knockdown of PPP2R1A was limited in HEC1B cells due to missense mutations W257L and L429F within PPP2R1A in this cell line. However, HEC-59 cells have wild-type PPP2R1A, and siRNA-mediated knockdown of PPP2R1A was successful in this cell line. HEC1B cells transfected with PPP2R1A siRNA showed a 1.4-fold increase in sensitivity to ZN-c3 (Table 6). HEC-59 cells transfected with PPP2R1A showed a 2.5-fold increase in sensitivity to ZN-c3 (Table 6). Thus, these results demonstrate the important role of PPP2R1A knockdown in sensitizing cancer cells to ZN-c3-mediated growth inhibition.

[0122] [Table 7]

[0123] Example 2 Dose Response of Cancer Cell Lines Sensitized with CRISPR-Mediated Knockout of PPP2R1A to ZN-c3 Because we observed that siRNA-mediated knockdown of PPP2R1A sensitized cancer cell lines to ZN-c3 treatment, we were also interested in assessing whether CRISPR-mediated PPP2R1A knockout would have a similar effect. Various cancer cell lines were transfected with CRISPR guide RNAs targeting PPP2R1A or non-targeting guide RNAs, and the cells were treated with a 10-point dose-response of ZN-c3 for 72 hours. The 10-point dose-response to ZN-c3 included the same doses used in Example 1. As in Example 1, no medium changes were performed during the 72-hour ZN-c3 dose-response evaluation period. Fewer than 20 cell passages were completed between CRISPR-mediated knockout and ZN-c3 treatment. CRISPR was performed by targeting PPP2R1A with a PPP2R1A sgRNA sequence. Cell density rates were calculated using cell titer glow analysis.

[0124] The ovarian cancer cell line SKOV3 was treated with CRISPR guide RNA targeting PPP2R1A or a non-targeting guide RNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the reduction in cell density (Figure 13). CRISPR-mediated knockout of PPP2R1A was evaluated by Western blot analysis of PPP2R1A protein expression in conditions treated with non-targeting guide RNA and PPPR12A-targeting CRISPR guide RNA, compared with beta-actin as a control (Figure 14). CRISPR-mediated knockout of PPP2R1A was successful, and SKOV3 cells transduced with PPP2R1A-targeting guide RNA showed a 1.7-fold increase in sensitivity to ZN-c3 (Table 7).

[0125] [Table 8]

[0126] The lung cancer cell line H1975 was treated with CRISPR guide RNA targeting PPP2R1A or a non-targeting guide RNA, and the transfected cells were evaluated for sensitivity to ZN-c3 by measuring the reduction in cell density (Figure 15). CRISPR-mediated knockout of PPP2R1A was evaluated by comparing PPP2R1A expression in conditions treated with guide RNA and PPPR12A-targeting CRISPR guide RNA (Figure 16). CRISPR-mediated knockout of PPP2R1A was successful, and H1975 cells transduced with PPP2R1A-targeting guide RNA showed a 1.9-fold increase in sensitivity to ZN-c3 (Table 8).

[0127] [Table 9]

[0128] The ovarian cancer cell line OV17R was evaluated for sensitivity to ZN-c3 by measuring the decrease in cell density (Figure 17). OV17R cells harbor a S256F hotspot missense mutation in PPP2R1A, thus rendering these cells highly sensitive to ZN-c3 without the need for additional mechanisms of PPP2R1A knockdown or knockout (Table 9).

[0129] [Table 10]

[0130] Example 3 Response of Cancer Cell Lines to Treatment with ZN-c3, LB-100, or ZN-c3+LB-100 Following the observation that both knockdown and knockout of PPP2R1A can sensitize cancer cells to Zn-c3, we evaluated the effects of pharmacological inhibition of PPP2R1A with the catalytic inhibitor LB-100. Various cancer cell lines were treated with Zn-c3 alone, the PP2A catalytic inhibitor LB-100 alone, or the combination of Zn-c3 and LB-100. As in Examples 1 and 2, the treatment period was 72 hours, and the inhibition rate was calculated using a cell titer glow assay.

[0131] The endometrial cancer cell line HEC-59 was evaluated for sensitivity to ZN-c3 alone, LB-100 alone, or the combination of ZN-c3 and LB-100 (Figure 18). The doses of ZN-c3 and LB-100 are shown in Table 10. ZN-c3 treatment alone was able to achieve approximately 25% inhibition, while LB-100 treatment achieved approximately half that value. However, combination therapy with both ZN-c3 and LB-100 was significantly more effective, reaching nearly 75% inhibition.

[0132] [Table 11]

[0133] The ovarian cancer cell line OV17R was evaluated for sensitivity to ZN-c3 alone, LB-100 alone, or the combination of ZN-c3 and LB-100 (Figure 19). The doses of ZN-c3 and LB-100 are shown in Table 11. ZN-c3 treatment alone was able to achieve approximately 30% inhibition, while LB-100 treatment achieved approximately 12% inhibition. Combination therapy with both ZN-c3 and LB-100 demonstrated a modest increase to approximately 40% inhibition over ZN-c3 alone. The slight increase in inhibition seen with combination therapy was due to the presence of wild-type PPP2R1A in the test sample. This is likely the result of the PPP2R1A missense mutation S256F present in OV17R, which is absent in the other cancer lines tested.

[0134] [Table 12]

[0135] The endometrial cancer cell line HEC-59 was evaluated for sensitivity to ZN-c3 alone, LB-100 alone, or the combination of ZN-c3 and LB-100 after treatment with siCTRL or siPPP2R1A (Figure 20). The doses of ZN-c3 and LB-100 are shown in Table 12. ZN-c3 treatment alone achieved approximately 12% inhibition, while LB-100 treatment achieved approximately half the value of siCTRL-treated cells. However, combination therapy with both ZN-c3 and LB-100 was significantly more effective, exceeding 25% inhibition in siCTRL-treated cells. Importantly, the inhibition achieved by ZN-c3 treatment alone and by ZN-c3 in combination with LB-100 was approximately twice as high in siPPP2R1A-treated cells as in siCTRL-treated cells. The inhibition achieved by LB-100 alone was only slightly enhanced in cells with PPP2R1A knockdown. These data demonstrate that combined pharmacological inhibition (LB-100) with Zn-C3 treatment against a background with reduced PPP2R1A expression provides a powerful combination approach to cancer therapy.

[0136] [Table 13]

[0137] The endometrial cancer cell line HEC1B was also evaluated for sensitivity to ZN-c3 alone, LB-100 alone, or the combination of ZN-c3 and LB-100 after treatment with siCTRL or siPPP2R1A (Figure 21). The doses of ZN-c3 and LB-100 are shown in Table 13. ZN-c3 treatment alone was able to achieve approximately 30% inhibition, while LB-100 treatment achieved 25% inhibition in cells treated with siCTRL. Combination therapy with both ZN-c3 and LB-100 was more effective than either individual treatment, achieving close to 50% inhibition in siCTRL-treated cells. The percent inhibition achieved by ZN-c3 treatment alone and by ZN-c3 in combination with LB-100 was nearly the same regardless of whether the cells were treated with siPPP2R1A or siCTRL. This result is similar to the evaluation of HEC1B in the previous example, where missense mutations W257L and L429F within PPP2R1A in this cell line limited the induction of sensitivity to ZN-c3.

[0138] [Table 14]

[0139] Example 4 Clinical trial of ZN-c3 in patients with PPP2R1A gene mutations. Genetic ablation and pharmacological inhibition of PPP2R1A sensitize cancer cell lines to ZN-c3 Because of the observed efficacy of ZN-c3, the next objective was to investigate the clinical response to ZN-c3 in human patients with hotspot mutations in PPP2R1A. Five USC patients with previously known genomic profiling information for PPP2R1A were evaluated as part of a clinical trial. Three patients had the P179R mutation and two patients had the S256F mutation of PPP2R1A (Table 14). All patients were evaluable for clinical response by at least one tumor assessment with imaging using computed tomography (CT) and / or magnetic resonance imaging (MRI). Four of five patients (80%) demonstrated clinical benefit with ZN-c3. The objective response rate for patients with PPP2R1A mutations was 20%.

[0140] For subject #1, a starting dose of 300 mg QD ZN-c3 was administered, and on day 44, the dose was reduced to 5 days on and 2 days off, and on day 51, the dose was further reduced to 200 mg QD, with the final dose given on day 230.

[0141] For subject number 2, a starting dose of 300 mg QD ZN-c3 was administered, and on day 36, the dose was reduced to 200 mg QD, and on day 50, the dose was further reduced to 200 mg for 5 days on and 2 days off, with the final dose given on day 84.

[0142] For subject #3, a starting dose of 300 mg QD ZN-c3 was administered, and on day 71, the dose was reduced to 200 mg QD, with the final dose given on day 172.

[0143] Subject #4 received a starting dose of 300 mg QD ZN-c3 with no dose reductions during the study, with the final dose provided on Day 64.

[0144] For subject number 5, a starting dose of 300 mg QD ZN-c3 was administered, and on day 36, the dose was reduced to 150 mg BID, with the final dose given on day 62.

[0145] [Table 15] cPR = confirmed partial response. SD = stable disease. PD = progressive disease.

[0146] Moreover, although the foregoing has been described in some detail by way of illustration and example for purposes of clarity and understanding, it will be understood by those skilled in the art that numerous and various modifications may be made thereto without departing from the spirit of the present disclosure. It should therefore be clearly understood that the forms disclosed herein are illustrative only and are not intended to limit the scope of the present disclosure, but on the contrary, encompass all modifications and alternative forms consistent with the true scope and spirit of the present invention.

Claims

1. 1. A method for identifying or selecting a subject having sensitivity to the drug ZN-c3, comprising: Identifying, in a biological sample obtained from said subject or individual, a regulation in the protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding said PPP2R1A protein, for example, one or more polymorphisms, preferably polymorphisms selected from P179R, S256F or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F or R183W, that confer sensitivity to said drug ZN-c3; and selecting or identifying the subject as having sensitivity to the drug ZN-c3 when said modulation, such as any one or more polymorphisms, in said PPP2R1A gene or protein is identified.

2. 10. The method of claim 1, further comprising administering ZN-c3 to the subject upon identification of the regulation or polymorphism in the biological sample.

3. 3. The method of claim 1, wherein the subject has cancer.

4. 4. The method of claim 3, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

5. 5. The method of any one of claims 2 to 4, wherein the cancer regression or inhibition is more than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

6. The method of any one of claims 2 to 5, further comprising administering to the subject an agent or combination of agents that inhibits the PPP2R1A gene or protein.

7. 7. The method of claim 6, wherein the drug or combination of drugs comprises the PP2A catalytic inhibitor LB-100.

8. The method of any one of claims 1 to 7, wherein said modulation of PPP2R1A comprises one or more gain-of-function mutations.

9. The method of any one of claims 1 to 7, wherein said modulation of PPP2R1A comprises one or more loss-of-function mutations.

10. The method of any one of claims 1 to 7, wherein said modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

11. The method of any one of claims 1 to 7, wherein said modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

12. 1. A method of inhibiting, ameliorating, or treating cancer or its sequelae in a subject, comprising: a regulation in a protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding said PPP2R1A protein in a biological sample obtained from said subject, said polymorphisms, e.g., P179R, S256F or identifying a polymorphism selected from P179R, S256F or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F or R183W, that confers sensitivity to the drug ZN-c3; and when said polymorphism is identified in said biological sample, administering ZN-c3 to said subject.

13. 13. The method of claim 12, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

14. 14. The method of claim 12 or 13, wherein the cancer regression or inhibition is more than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

15. The method of any one of claims 12 to 14, wherein the subject is further provided with a drug or combination of drugs that inhibits the PPP2R1A gene or protein.

16. 16. The method of claim 15, wherein the drug or combination of drugs comprises the PP2A catalytic inhibitor LB-100.

17. The method of any one of claims 12 to 16, wherein said modulation of PPP2R1A comprises one or more gain-of-function mutations.

18. The method of any one of claims 12 to 16, wherein said modulation of PPP2R1A comprises one or more loss-of-function mutations.

19. The method of any one of claims 12 to 16, wherein said modulation of PPP2R1A comprises one or more mutations that result in overexpression of PPP2R1A.

20. The method of any one of claims 12 to 16, wherein said modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

21. 1. The compound ZN-c3 for use in inhibiting, ameliorating, or treating cancer or its sequelae in a subject, wherein the subject has been identified as having a regulation in a protein phosphatase 2 scaffold subunit alpha (PPP2R1A) protein or a gene encoding said PPP2R1A protein, e.g., a polymorphism that confers sensitivity to said drug ZN-c3, e.g., a polymorphism selected from P179R, S256F, or R183W, or a nucleic acid encoding a polymorphism selected from P179R, S256F, or R183W, in a biological sample obtained from the subject.

22. 22. The compound ZN-c3 for use according to claim 21, wherein said cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

23. and wherein the cancer regression or inhibition is greater than 7%, e.g., 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

22. The compound ZN-c3 for use according to claim 21.

24. The compound ZN-c3 for use according to any one of claims 21 to 23, wherein the subject is additionally provided with a drug or combination of drugs that inhibits the PPP2R1A gene or protein.

25. The compound ZN-c3 for use according to claim 24, wherein said drug or combination of drugs comprises the PP2A catalytic inhibitor LB-100.

26. The compound ZN-c3 for use according to any one of claims 21 to 25, wherein said modulation of PPP2R1A comprises one or more gain-of-function mutations.

27. The compound ZN-c3 for use according to any one of claims 21 to 25, wherein said modulation of PPP2R1A comprises one or more loss-of-function mutations.

28. The compound ZN-c3 for use according to any one of claims 21 to 25, wherein said modulation of PPP2R1A comprises one or more mutations that lead to overexpression of PPP2R1A.

29. The compound ZN-c3 for use according to any one of claims 21 to 25, wherein said modulation of PPP2R1A comprises one or more mutations that result in underexpression of PPP2R1A.

30. 1. A method of inhibiting, ameliorating, or treating cancer or its sequelae in a subject, comprising: administering to the subject an agent or combination of agents that inhibits the PPP2R1A gene or protein; and administering ZN-c3 to the subject.

31. 31. The method of claim 30, wherein the agent or combination of agents that inhibit the PPP2R1A gene or protein are provided separately to the subject.

32. 32. The method of claim 30 or 31, wherein the agent or combination of agents that inhibits the PPP2R1A gene or protein is provided to the subject prior to administration of ZN-C3.

33. 31. The method of claim 30, wherein the agent or combination of agents that inhibit the PPP2R1A gene or protein is provided to the subject at 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 minute intervals.

34. 34. The method of any one of claims 30 to 33, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

35. 35. The method of any one of claims 30 to 34, wherein the cancer regression or inhibition is more than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

36. 36. The method of any one of claims 30 to 35, wherein the drug or combination of drugs comprises the PP2A catalytic inhibitor LB-100.

37. Compound ZN-c3 and a drug or drug that inhibits the PPP2R1A gene or protein and a combination thereof, wherein the product combination comprises a compound selected from the group consisting of hydroxybenzoates, ...

38. 38. The product combination for use according to claim 37, wherein the cancer comprises non-small cell lung cancer, breast cancer, colorectal cancer, ovarian cancer, uterine serous carcinoma (USC), or endometrial cancer.

39. 39. The product combination for use according to any one of claims 37 or 38, wherein the cancer regression or inhibition is more than 7%, such as 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50% or more.

40. 40. A product combination for use according to any one of claims 37 to 39, wherein said drug or drug combination comprises the PP2A catalytic inhibitor LB-100.

41. 30. The method of any one of claims 1 to 20 or the compound ZN-c3 for use according to any one of claims 21 to 29, wherein the presence of the polymorphism is determined using next generation sequencing (NGS), sequencing, polymerase chain reaction (PCR), loop-mediated isothermal amplification, recombinase polymerase amplification, or antibody detection.

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