Treatment of cancer with tg02

Abstract

This application relates to the treatment of cancer with TG02. The present disclosure provides methods of treating a patient with cancer with TG02 and a second therapeutic agent, for example, TG02 and an immune checkpoint inhibitor, TG02 and a COX-2 inhibitor, or TG02 and an immune checkpoint inhibitor and a COX-2 inhibitor.

Description

[0001] This application is a continuation of China Patent Application No. 201780032318.X. BACKGROUND

[0002] TECHNICAL FIELD

[0003] The present disclosure provides methods of treating a patient with cancer with TG02 and a second therapeutic agent, for example, TG02 and an immune checkpoint inhibitor, TG02 and a COX-2 inhibitor, and TG02 and an immune checkpoint inhibitor and a COX2 inhibitor.

[0004] BACKGROUND

[0005] TG02 is a purine-based multikinase inhibitor that inhibits CDK 1, 2, 5, 7, and 9 along with JAK2 and FLT3. It dose-dependently inhibits signaling pathways downstream of CDK, JAK2, and FLT3 in cancer cells, with the primary target being CDK. TG02 is antiproliferative in a broad range of tumor cell lines, inducing G1 cell cycle arrest and apoptosis. Primary cultures of progenitor cells derived from acute myeloid leukemia (AML) and polycythemia vera patients are very sensitive to TG02. Comparison with reference inhibitors that block only one of the primary targets of TG02 demonstrated the benefit of combined CDK and JAK2 / FLT3 inhibition in cell lines as well as in primary cells. See Goh et al., Leukemia 26:236-43 (2012). TG02 is also known as SB1317, and has the chemical name: (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8(27),9,11,16,21,23-decaene. TG02 is disclosed as Compound 1 in US 8,143,255. US 9,120,815 discloses various salts, for example, TG02 citrate, and crystalline forms of TG02. The chemical structure of TG02 is:

[0006] SUMMARY

[0007] In one aspect, the present disclosure provides methods of treating a patient with cancer, the method comprising administering to the patient a therapeutically effective amount of TG02. In another aspect, the patient’s cancer is characterized by overexpression of MYC, MCL1, or both.

[0008] In another aspect, the present application provides a therapeutic method of treating a cancer patient, the method comprising administering to the patient a therapeutically effective amount of TG02 and an immune checkpoint inhibitor, for example, a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a LAG3 inhibitor, a TIM3 inhibitor, or a cd47 inhibitor.

[0009] In another aspect, the present application provides a therapeutic method of treating a cancer patient, the method comprising administering to the patient a therapeutically effective amount of TG02 and a COX inhibitor, for example, alcloxa or 6-bromo-8-(methyl-D3)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid.

[0010] In another aspect, the present application provides a therapeutic method of treating a cancer patient, the method comprising administering to the patient a therapeutically effective amount of TG02, an immune checkpoint inhibitor, and a COX-2 inhibitor.

[0011] In another aspect, the present application provides a therapeutic method of treating a cancer patient having a tumor that overexpresses MYC, MCL1, or both.

[0012] In another aspect, the present application provides a kit comprising TG02, TG02 and an immune checkpoint inhibitor, TG02 and a COX-2 inhibitor, and TG02 and an immune checkpoint inhibitor and a COX-2 inhibitor.

[0013] In another aspect, the present application provides a pharmaceutical composition comprising TG02, a COX-2 inhibitor, for example, alcloxa or 6-bromo-8-(methyl-D3)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid, and a pharmaceutically acceptable excipient. BRIEF DESCRIPTION OF DRAWINGS

[0014] Figure 1 is a bar graph showing the in vitro activity of TG02, TMZ (temozolomide), and TG02 + TMZ in GSC923 cells.

[0015] Figure 2 is a bar graph showing the in vitro activity of TG02, TMZ, and TG02 + TMZ in LN18 cells.

[0016] Figure 3 is a bar graph showing the in vitro activity of TG02, TMZ, and TG02 + TMZ in T98G cells.

[0017] Figure 4 is a bar graph showing the in vitro activity of TG02, TMZ, and TG02 + TMZ in U251 cells.

[0018] Figure 5is a bar graph showing in vitro activity of TG02, TMZ, and TG02 + TMZ in U87 cells.

[0019] Figure 6 is a bar graph showing in vitro activity of TG02, TMZ, and TG02 + TMZ in LN299 cells.

[0020] Figure 7 is a bar graph showing in vitro activity of TG02, TMZ, and TG02 + TMZ in GSC827 cells.

[0021] Figure 8 is a bar graph showing in vitro cytotoxicity of TG02, TMZ, and TG02 + TMZ (T+T) in GSC923 cells.

[0022] Figure 9 is a bar graph showing in vitro cytotoxicity of TG02, TMZ, and TG02 + TMZ (T+T) in U251 cells.

[0023] Figure 10 is a bar graph showing lack of in vitro activity of TG02, TMZ, and TG02 + TMZ (T+T) in human pulmonary artery endothelial cells.

[0024] Figure 11 is a bar graph showing lack of in vitro activity of TG02, TMZ, and TG02 + TMZ (T+T) in human astrocytes.

[0025] Figure 12 is a dose response curve showing in vitro activity of TG02 and TG02 + TMZ in GSC923 cells.

[0026] Figure 13 is a dose response curve showing in vitro activity of TMZ and TG02 + TMZ in GSC923 cells.

[0027] Figure 14 is a dose response curve showing in vitro activity of TG02 and TG02 + TMZ in U251 cells.

[0028] Figure 15 is a dose response curve showing in vitro activity of TMZ and TG02 + TMZ in U251 cells.

[0029] Figure 16 is a schematic of TG02 and TMZ administration in a mouse glioma GL261 cell allograft model.

[0030] Figure 17is a line graph showing the percent survival after TG02, TMZ, and TG02 + TMZ administration in a mouse glioma GL261 cell allograft model.

[0031] Figure 18 is a line graph showing tumor burden after TG02, TMZ, and TG02 + TMZ administration in a mouse glioma GL261 cell allograft model.

[0032] Figure 19 is a graphical representation showing the effect of TG02 on MYC protein levels in hepatocellular carcinoma (HCC) cells.

[0033] Figure 20 is a dose response curve showing the effect of TG02 on MYC protein levels in HCC cells.

[0034] Figure 21 is a graphical representation showing the effect of TG02 on MYC protein levels in HCC tumor cells.

[0035] Figure 22 is a line graph showing in vivo activity of TG02 and TG02 + sorafenib in an orthotopic model of HepG2 HCC xenografts.

[0036] Figure 23 is a bar graph showing PD-L1 expression after treatment with TG02 in a transgenic mouse model of MYC-induced T-cell acute lymphoblastic leukemia.

[0037] Figure 24 is a bar graph showing CD47 expression after treatment with TG02 in a transgenic mouse model of MYC-induced T-cell acute lymphoblastic leukemia.

[0038] Figure 25 is a bar graph showing BCL-xL expression after treatment with TG02 in a transgenic mouse model of MYC-induced T-cell acute lymphoblastic leukemia.

[0039] Figure 26 is a bar graph showing MYC expression after treatment with TG02 in a transgenic mouse model of MYC-induced T-cell acute lymphoblastic leukemia.

[0040] Figure 27 is a line graph showing efficacy of TG02 in combination with anti-PD1 in a mouse syngeneic GL261 orthotopic glioblastoma model.

[0041] Figure 28 is a graphical representation showing that BT245 tumor cells exposed to TG02 show inhibition of MYC and MCL-1 expression.

[0042] Figure 29 is a histogram showing the area under the curve (AUC) of TG02-induced inhibition in glioblastoma (GBM) cells.

[0043] Figure 30 is a scatter plot showing high MYC expression is associated with low AUC in GBM cells.

[0044] Figure 31 is a series of six line plots showing TG02 in combination with radiation in glioblastoma cell lines.

[0045] Figure 32 is a histogram showing TG02 activity in 26 patient-derived GBM stem cell lines.

[0046] Figure 33 is a graphical representation showing the expression levels of CDK9, MYC, and Mcl-1 in patient-derived GBM stem cell lines after treatment with TG02. DETAILED DESCRIPTION

[0047] In one embodiment, the present disclosure provides a method of treatment for treating a patient having cancer, the method comprising administering to the patient a therapeutically effective amount of TG02, wherein one or more genes listed in Table 1 (see below) are differentially present in a biological sample taken from the patient as compared to a biological sample taken from a subject in another phenotypic state. In another embodiment, MYC overexpression is differentially present in the sample taken from the patient. In another embodiment, MCL1 overexpression is differentially present in the sample taken from the patient.

[0048] In another embodiment, the present disclosure provides a method of treatment for treating a patient having cancer, the method comprising administering to the patient a therapeutically effective amount of TG02 and an immune checkpoint inhibitor, wherein one or more genes listed in Table 1 (see below) are differentially present in a biological sample taken from the patient as compared to a biological sample taken from a subject in another phenotypic state. In another embodiment, MYC overexpression is differentially present in the sample taken from the patient. In another embodiment, MCL1 overexpression is differentially present in the sample taken from the patient. In another embodiment, TG02 is administered to the patient prior to the immune checkpoint inhibitor. In another embodiment, TG02 is administered to the patient after the immune checkpoint inhibitor. In another embodiment, TG02 is administered to the patient concurrently with the immune checkpoint inhibitor.

[0049] In another embodiment, the present disclosure provides a method of treatment of a patient having cancer, comprising administering to the patient a therapeutically effective amount of TG02, an immune checkpoint inhibitor, and a COX-2 inhibitor, wherein one or more genes listed in Table 1 (see below) are differentially present in a biological sample taken from the patient as compared to a biological sample taken from a subject in another phenotypic state. In another embodiment, MYC overexpression is differentially present in the sample taken from the patient. In another embodiment, MCL1 overexpression is differentially present in the sample taken from the patient. In another embodiment, TG02 is administered to the patient prior to the COX-2 inhibitor. In another embodiment, TG02 is administered to the patient after the COX-2 inhibitor. In another embodiment, TG02 is administered to the patient concurrently with the COX-2 inhibitor.

[0050] In another embodiment, the present disclosure provides a method of treatment of a patient having cancer, comprising administering to the patient a therapeutically effective amount of TG02 and an immune checkpoint inhibitor. In another embodiment, TG02 is administered to the patient prior to the immune checkpoint inhibitor. In another embodiment, TG02 is administered to the patient after the immune checkpoint inhibitor. In another embodiment, TG02 is administered to the patient concurrently with the immune checkpoint inhibitor.

[0051] In another embodiment, the present disclosure provides a method of treatment of a patient having cancer, comprising administering to the patient a therapeutically effective amount of TG02, an immune checkpoint inhibitor and a COX-2 inhibitor. In another embodiment, TG02 is administered to the patient prior to the COX-2 inhibitor. In another embodiment, TG02 is administered to the patient after the COX-2 inhibitor. In another embodiment, TG02 is administered to the patient concurrently with the COX-2 inhibitor.

[0052] In another embodiment, the present disclosure provides a kit comprising TG02 and an immune checkpoint inhibitor, and instructions for administering TG02 and an immune checkpoint inhibitor to a patient having cancer. In another embodiment, the kit further comprises a COX-2 inhibitor.

[0053] In another embodiment, the present disclosure provides a kit comprising TG02 and a COX-2 inhibitor, and instructions for administering TG02 and a COX-2 inhibitor to a patient having cancer.

[0054] In another embodiment, the kit is packaged in a manner that facilitates its use in practicing the methods of the present disclosure.

[0055] In another embodiment, the kit comprises TG02 (or a composition comprising TG02) packaged in a container (e.g., a sealed bottle or vessel) with a label affixed to the container or included in the kit that describes using TG02 or the composition to practice the methods of the application. In one embodiment, TG02 is packaged in unit dosage form. The kit can also include a device suitable for administering the composition in a predetermined administration route.

[0056] The present disclosure provides various methods of treatment, kits, and compositions related to cancer treatment. In one embodiment, the cancer is a solid tumor. In another embodiment, the cancer is a hematological malignancy. In another embodiment, the cancer is selected from the group consisting of adrenal gland cancer, acinar cell carcinoma, acoustic neuroma, acral lentiginous melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryocytic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adamantinoma, adenosquamous carcinoma, adipose tissue neoplasms, adrenocortical carcinoma, adult T-cell leukemia / lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft-part sarcoma, ameloblastic fibroma, anaplastic large-cell lymphoma, anaplastic thyroid carcinoma, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytic tumor, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B-cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary duct cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, bronchioalveolar carcinoma, bronchogenic carcinoma, burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, degos' disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, enteropathy-associated T-cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, ganglioneuroblastoma, gastrointestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of bone, glioma, glioblastoma, glioneuronal tumor, glioma of the brain, glucagonoma, gonadoblastoma, granulosa cell tumor, hermaphroditoblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatocellular carcinoma, hepatosplenic T-cell lymphoma, hodgkin's lymphoma, non-hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphoblastic leukemia, acute myeloid leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell carcinoma, mesothelioma, metastatic urothelial carcinoma, mixed mullerian tumor,myeloma, muscle tissue tumor, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurilemmoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligodendroglioma, oligodendrocytoma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, parietal sulcus carcinoma, papillary thyroid carcinoma, paraganglioma, pineoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, primary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, peritoneal pseudomyxoma, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Rieger's syndrome, rectal cancer, sarcoma, schwannomatosis, seminoma, Sertoli cell tumor, sex cord-stromal tumor, signet ring cell carcinoma, skin cancer, small round blue cell tumor, small cell carcinoma, soft tissue sarcoma, somatostatinoma, seborrheic keratosis, spinal cord tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma, Sezary disease, small intestine cancer, squamous cell carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, thecoma, thyroid cancer, transitional cell carcinoma, throat cancer, urachal cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Wossing's tumor, and Wilms' tumor.

[0057] In another embodiment, the cancer is selected from the group consisting of head and neck squamous cell carcinoma, esophageal adenocarcinoma, gastric adenocarcinoma, colon adenocarcinoma, hepatocellular carcinoma, cholangiocarcinoma of the biliary system, gallbladder adenocarcinoma, pancreatic cancer, breast ductal carcinoma in situ, breast cancer, lung adenocarcinoma, lung squamous cell carcinoma, bladder transitional cell carcinoma, bladder squamous cell carcinoma, cervical squamous cell carcinoma, cervical adenocarcinoma, endometrial carcinoma, penile squamous cell carcinoma, and cutaneous squamous cell carcinoma.

[0058] In another embodiment, the precancerous tumor is selected from the group consisting of leukoplakia of the head and neck, Barrett's esophagus, gastric metaplasia, colonic adenoma, chronic hepatitis, biliary metaplasia, pancreatic intraepithelial neoplasia, lung atypical adenomatous hyperplasia, urothelial dysplasia, cervical intraepithelial neoplasia, penile intraepithelial neoplasia, and cutaneous actinic keratosis.

[0059] In another embodiment, the patient has a tumor that overexpresses MYC, MCL1, or both. The tumor can be determined to overexpress MYC, MCL1, or both by methods known in the art.

[0060] In another embodiment, the cancer is selected from the group consisting of hepatocellular carcinoma, glioblastoma, lung cancer, breast cancer, head and neck cancer, prostate cancer, melanoma, and colorectal cancer.

[0061] In another embodiment, the cancer is selected from the group consisting of glioblastoma, hepatocellular carcinoma, non-small cell and small cell lung cancer, head and neck cancer, colorectal cancer, and triple negative breast cancer.

[0062] In another embodiment, the cancer has become resistant to a conventional cancer therapy. As used herein, the term "conventional cancer therapy" means any cancer drug or biologic, or combination of cancer drugs and / or biologies, that has been tested and / or approved for human therapeutic use by the U.S. Food and Drug Administration, the European Medicines Agency, or similar regulatory agency.

[0063] In another embodiment, the patient has been previously treated with an immune checkpoint inhibitor that does not contain TG02. For example, the previous immune checkpoint therapy can be an anti-PD-1 therapy.

[0064] In another embodiment, the patient has been previously treated with a COX-2 inhibitor that does not contain TG02.

[0065] In another embodiment, the present disclosure provides a pharmaceutical composition comprising TG02, a COX-2 inhibitor, and a pharmaceutically acceptable excipient.

[0066] In another embodiment, the present disclosure provides a method of treatment for treating a patient having a cancer, the method comprising administering to the patient a therapeutically effective amount of TG02, wherein the patient's phenotypic state is overexpression of MYC, overexpression of MCL1, or overexpression of both MYC and MCL1. In another embodiment, the cancer is selected from the group consisting of hepatocellular carcinoma, glioblastoma, lung cancer, breast cancer, head and neck cancer, prostate cancer, melanoma, and colorectal cancer.

[0067] In another embodiment, the present disclosure provides a method of treatment for treating a patient having a cancer, the method comprising administering to the patient a therapeutically effective amount of TG02 and a second therapeutic agent, wherein the second therapeutic agent is neither an immune checkpoint inhibitor nor a COX-2 inhibitor.

[0068] In another embodiment, the present disclosure provides a method of treatment for treating a patient having a cancer, the method comprising administering to the patient a therapeutically effective amount of TG02, an immune checkpoint inhibitor, and a third therapeutic agent, wherein the third therapeutic agent is not a COX-2 inhibitor.

[0069] In another embodiment, the present disclosure provides a method of treatment for treating a patient having a cancer, the method comprising administering to the patient a therapeutically effective amount of TG02, a COX-2 inhibitor, and a third therapeutic agent, wherein the third therapeutic agent is not an immune checkpoint inhibitor.

[0070] In another embodiment, the present disclosure provides a method of treatment for treating a patient having cancer, the method comprising administering to the patient a therapeutically effective amount of TG02, an immune checkpoint inhibitor, a COX-2 inhibitor, and a fourth therapeutic agent, wherein the fourth therapeutic agent is neither an immune checkpoint inhibitor nor a COX-2 inhibitor.

[0071] In another embodiment, the present disclosure provides personalized medicine for a cancer patient and comprises selecting a treatment option that has the highest likelihood of a successful outcome for the individual cancer patient. In another aspect, the present disclosure relates to an assay for use in predicting the outcome of treatment (e.g., likelihood of a favorable response or treatment success) for a patient having cancer.

[0072] In another embodiment, the present disclosure provides a method of selecting a patient (e.g., a human subject) for treatment of cancer with TG02, and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor, comprising obtaining a biological sample from the patient, e.g., blood cells, testing the biological sample from the patient for the presence of a biomarker, e.g., overexpression of MYC, overexpression of MCL1, or both, and selecting the patient for treatment if the biological sample contains the biomarker. In another embodiment, the method further comprises administering to the patient a therapeutically effective amount of TG02, and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor, if the biological sample contains the biomarker. Examples of cancer biomarkers are provided in Table 1. In another embodiment, the cancer is a solid tumor. In another embodiment, the cancer is a hematological malignancy. In another embodiment, the cancer is selected from the group consisting of hepatocellular carcinoma, glioblastoma, lung cancer, breast cancer, head and neck cancer, prostate cancer, melanoma, and colorectal cancer.

[0073] In another embodiment, the present disclosure provides a method of predicting the outcome of treatment for a patient having cancer, comprising obtaining a biological sample from the patient, testing the biological sample from the patient for the presence of a biomarker, e.g., overexpression of MYC, overexpression of MCL1, or both, wherein detection of the biomarker indicates that the patient will respond favorably to administration of a therapeutically effective amount of TG02, and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor. Favorable responses include, but are not limited to, reduction in tumor size and increase in progression-free or overall survival.

[0074] In another embodiment, the present disclosure provides a method of treating cancer, comprising administering to a patient (e.g., a human subject) having cancer a therapeutically effective amount of TG02 and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor, the patient's cells containing a biomarker. In another embodiment, the patient is selected for treatment with TG02 and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor after determining that the patient's cells contain overexpression of MYC. In another embodiment, the patient is selected for treatment with TG02 and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor after determining that the patient's cells contain overexpression of MCL1. In another embodiment, the patient is selected for treatment with TG02 and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor after determining that the patient's cells contain overexpression of MYC and overexpression of MCL1.

[0075] In another embodiment, a method of treating a patient having cancer comprises obtaining a biological sample from the patient, determining whether the biological sample contains a biomarker, e.g., overexpression of MYC, overexpression of MCL1, or both, and administering to the patient a therapeutically effective amount of TG02 and optionally an immune checkpoint inhibitor and / or a COX-2 inhibitor if the biological sample contains the biomarker. In another embodiment, the methods provided herein comprise determining whether the patient's cells contain overexpression of MYC. In another embodiment, the methods provided herein comprise determining whether the patient's cells contain overexpression of MCL1. In another embodiment, the methods provided herein comprise determining whether the patient's cells contain overexpression of MYC and MCL1.

[0076] In another embodiment, the present disclosure provides a method of treating a subject having cancer, the method comprising obtaining a biological sample from the subject, determining the expression level of MYC, MCL1, or both in the biological sample; and administering to the subject a therapeutically effective amount of TG02 and a second therapeutic agent, e.g., temozolomide, carfilzomib, sorafenib, bortezomib, doxorubicin, cisplatin, lenalidomide, dexamethasone, or Ara-C, if the biological sample shows overexpression of MYC, MCL1, or both.

[0077] In another embodiment, the patient was previously treated with an immune checkpoint inhibitor only. For example, the previous immune checkpoint treatment can be anti-PD-1 treatment.

[0078] In another embodiment, the patient was previously treated with a COX-2 inhibitor only.

[0079] I. Immune Checkpoint Inhibitor

[0080] Immune checkpoint inhibitors are therapies that block an inhibitor checkpoint of the immune system. Immune checkpoints can be stimulatory or inhibitory. Blocking inhibitory immune checkpoints can activate immune system function and can be used in cancer immunotherapy. Pardoll, Nature Reviews. Cancer 12:252-64 (2012). When tumor cells attach to specific T-cell receptors, they shut down activated T-cells. Immune checkpoint inhibitors prevent tumor cells from attaching to T-cells, causing the T-cells to remain activated. In effect, the synergy of cellular and soluble components can combat pathogens and damage caused by cancer. Modulation of immune system pathways can involve altering the expression or functional activity of at least one component of the pathway, which then modulates the response of the immune system. U.S. 2015 / 0250853. Examples of immune checkpoint inhibitors include PD-1 inhibitors, PD-L1 inhibitors, CTLA-4 inhibitors, LAG3 inhibitors, TIM3 inhibitors, cd47 inhibitors, and B7-H1 inhibitors. Thus, in one embodiment, the immune checkpoint inhibitor is selected from the group consisting of a PD-1 inhibitor, a PD-L1 inhibitor, a CTLA-4 inhibitor, a LAG3 inhibitor, a TIM3 inhibitor, and a cd47 inhibitor.

[0081] In another embodiment, the immune checkpoint inhibitor is a programmed cell death (PD-1) inhibitor. PD-1 is a T-cell co-inhibitory receptor that plays a key role in the ability of tumor cells to evade the host immune system. Blocking the interaction between PD-1 and PD-L1, a ligand for PD-1, enhances immune function and mediates anti-tumor activity. Examples of PD-1 inhibitors include antibodies that specifically bind to PD-1. Specific anti-PD-1 antibodies include, but are not limited to, nivolumab, pembrolizumab, STI-1014, and durvalumab. For a general discussion of the availability, methods of production, mechanism of action, and clinical studies of anti-PD-1 antibodies, see U.S. 2013 / 0309250, U.S. 6,808,710, U.S. 7,595,048, U.S. 8,008,449, U.S. 8,728,474, U.S. 8,779,105, U.S. 8,952,136, U.S. 8,900,587, U.S. 9,073,994, U.S. 9,084,776, and Naido et al., British Journal of Cancer 111 :2214-19 (2014).

[0082] In another embodiment, the immune checkpoint inhibitor is a PD-L1 (also known as B7-H1 or CD274) inhibitor. Examples of PD-L1 inhibitors include antibodies that specifically bind to PD-L1. Particular anti-PD-L1 antibodies include, but are not limited to, avelumab, atezolizumab, durvalumab, and BMS-936559. For a general discussion of availability, methods of production, mechanism of action, and clinical studies, see U.S. 8,217,149, U.S. 2014 / 0341917, U.S. 2013 / 0071403, WO 2015036499, and Naido et al., British Journal of Cancer 111 :2214-19 (2014).

[0083] In another embodiment, the immune checkpoint inhibitor is a CTLA-4 inhibitor. CTLA-4, also known as cytotoxic T-lymphocyte antigen 4, is a protein receptor that downregulates the immune system. CTLA-4 is characterized by "braking" the co-stimulatory molecules on antigen-presenting cells, which prevents interaction with CD28 on T cells and also generates a dominant inhibitory signal that limits T cell activation. Examples of CTLA-4 inhibitors include antibodies that specifically bind to CTLA-4. Particular anti-CTLA-4 antibodies include, but are not limited to, ipilimumab and tremelimumab. For a general discussion of availability, methods of production, mechanism of action, and clinical studies, see U.S. 6,984,720, U.S. 6,207,156, and Naido et al., British Journal of Cancer 111 :2214-19 (2014).

[0084] In another embodiment, the immune checkpoint inhibitor is a LAG3 inhibitor. LAG3, lymphocyte-activation gene 3, is a negative co- stimulatory receptor that regulates T cell homeostasis, proliferation, and activation. In addition, LAG3 has been reported to be involved in regulatory T cell (Treg) suppressive function. The majority of LAG3 molecules are retained in cells close to the microtubule organization center and are induced only after antigen-specific T cell activation. U.S. 2014 / 0286935. Examples of LAG3 inhibitors include antibodies that specifically bind to LAG3. Particular anti-LAG3 antibodies include, but are not limited to, GSK2831781. For a general discussion of availability, methods of production, mechanism of action, and studies, see U.S. 2011 / 0150892, U.S. 2014 / 0093511, U.S. 20150259420, and Huang et al., Immunity 21 :503-13 (2004).

[0085] In another embodiment, the immune checkpoint inhibitor is a TIM3 inhibitor. TIM3, T-cell immunoglobulin and mucin domain 3, is an immune checkpoint receptor whose function is to limit T H 1 and T C 1 cell responses. The TIM3 pathway is considered a target for anti-cancer immunotherapy because it is expressed on dysfunctional CD8 + T cells and Tregs, both reported immune cell populations that contribute to immune suppression in tumor tissue. Anderson, Cancer Immunology Research 2:393-98 (2014). Examples of TIM3 inhibitors include antibodies that specifically bind TIM3. For a general discussion of the availability, methods of production, mechanism of action, and studies of TIM3 inhibitors, see U.S. 20150225457, U.S. 20130022623, U.S. 8,522,156, Ngiow et al., Cancer Res 71 :6567-71 (2011), Ngiow et al., Cancer Res 71 :3540-51 (2011), and Anderson, Cancer Immunology Res 2:393-98 (2014).

[0086] In another embodiment, the immune checkpoint inhibitor is a cd47 inhibitor. See Unanue, E.R., PNAS 110:10886-87 (2013).

[0087] The term "antibody" is intended to include intact monoclonal antibodies, polyclonal antibodies, multispecific antibodies formed from at least two intact antibodies, and antibody fragments so long as they exhibit the desired biological activity. In another embodiment, "antibody" is intended to include soluble receptors that do not have an antibody Fc portion. In one embodiment, the antibody is a humanized monoclonal antibody and fragments thereof that are produced by recombinant genetic engineering.

[0088] Another class of immune checkpoint inhibitors includes polypeptides that bind to and block the PD-1 receptor on T cells without triggering the inhibitor signal transduction. These peptides include B7-DC polypeptides, B7-H1 polypeptides, B7-1 polypeptides, and B7-2 polypeptides, and soluble fragments thereof, as disclosed in U.S. Patent No. 8,114,845.

[0089] Another class of immune checkpoint inhibitors includes compounds having a peptide moiety that inhibits PD-1 signaling. Examples of these compounds are disclosed in U.S. Patent No. 8,907,053 and have the following structure:

[0090]

[0091] or a pharmaceutically acceptable salt thereof, wherein the compound comprises at least 5 amino acids, which can be used as a therapeutic agent capable of inhibiting the PD-1 signaling pathway.

[0092] Another class of immune checkpoint inhibitors includes inhibitors of certain metabolic enzymes, such as indoleamine 2,3 dioxygenase (IDO), which is expressed by infiltrating myeloid cells and tumor cells. The IDO enzyme suppresses the immune response by eliminating amino acids necessary for anabolic function in T cells or by synthesizing specific natural ligands that can alter the function of lymphocytes. Pardoll, Nature Reviews. Cancer 12:252-64 (2012); Cancer Immunol Immunother 58:153-57 (2009). Particular IDO blocking agents include, but are not limited to, L-leucine-1 -methyl tryptophan (L-1 MT) and 1 -methyl-tryptophan (1 MT). Qian et al., Cancer Res 69:5498-504 (2009); and et al., Cancer Immunol Immunother 58:153-7 (2009).

[0093] In one embodiment, the immune checkpoint inhibitor is Nivolumab, Pembrolizumab, Durvalumab, STI-1110, Afutuzumab, Atezolizumab, Durvalumab, STI-1014, Ipilimumab, Tremelimumab, GSK2831781, BMS-936559, or MED14736.

[0094] II. COX-2 Inhibitors

[0095] Cyclooxygenase-2 (COX-2) is an enzyme that promotes inflammation and plays a role in tumor progression. COX-2 inhibitors include non-selective inhibitors such as aspirin, ibuprofen, sulindac sulfone, sulindac sulfide, diclofenac, nafamostat, naproxen, indomethacin, and piroxicam, selective inhibitors such as celecoxib, rofecoxib, valdecoxib, ANS-398, Cay 10404, SC-236, and DUP 697, and preferential inhibitors such as meloxicam and nimesulide. Other COX-2 inhibitors include alclofenac, tilmacoxib, and cimicoxib. Any COX-2 inhibitor is contemplated for use in the therapeutic methods of the present disclosure. See Sobolewski et al., “Role of Cyclooxygenase-2 in Cell Proliferation and Cell Death in Human Malignancies,” International Journal of Cell Biology, vol. 2010, Article ID 215158, 21 pages, 2010. doi:10.1155 / 2010 / 215158.

[0096] In another embodiment, the COX-2 inhibitor is alclofenac. See Kirane et al., Clin. Cancer Res. 18:5031-5042 (2012).

[0097] In another embodiment, the COX-2 inhibitor is selected from the group consisting of:

[0098] 8-(Ethyl-D5)-6-(trifluoromethoxy)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0099] 6-Chloro-8-(methyl-D3)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0100] 6-Bromo-8-(methyl-D3)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0101] 8-Chloro-6-(methyl-D3)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0102] 6,8-Dibromo-5,7-(dimethyl-D6)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0103] 8-(1-Methylhexyl-D15)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0104] 6-Chloro-8-(1-methylhexyl-D15)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0105] 8-(Hexyl-D13)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid;

[0106] 7,8-(Dimethyl-D6)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid; and

[0107] 6-Chloro-8-(hexyl-D13)-2-(trifluoromethyl)-2H-chromene-3-carboxylic acid.

[0108] See US 2015 / 0133538.

[0109] In another embodiment, the COX-2 inhibitor is 6-bromo-8-(methyl-D3)-2- (trifluoromethyl)-2H-chromene-3-carboxylic acid.

[0110] III. Optional Therapeutic Agents

[0111] In certain methods of treatment of the disclosure, a second therapeutic agent is administered to a cancer patient in combination with TG02, a third therapeutic agent is administered to a cancer patient in combination with TG02 and an immune checkpoint inhibitor or in combination with TG02 and a COX-2 inhibitor, or a fourth therapeutic agent is administered to a cancer patient in combination with TG02, an immune checkpoint inhibitor, and a COX-2 inhibitor. The second, third, and fourth therapeutic agents used in the methods of treatment of the disclosure are referred to as“optional therapeutic agents.” Such optional therapeutic agents for use in treating a cancer patient are known in the art. In one embodiment, the optional therapeutic agent in combination with TG02 is an anti-cancer agent that is neither an immune checkpoint inhibitor nor a COX-2 inhibitor.

[0112] The optional therapeutic agent is administered in an amount to provide the desired therapeutic effect thereof. Effective dosage ranges for various optional therapeutic agents are known in the art, and the optional therapeutic agent is administered to an individual in need thereof within such established ranges.

[0113] TG02, the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent can be administered together as a single unit dose or separately as multiple unit doses, and in any order, e.g., where TG02 is administered prior to the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent, or vice versa. One or more doses of TG02, the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent can be administered to the patient.

[0114] In one embodiment, the optional therapeutic agent is an epigenetic drug. Herein, the term“epigenetic drug” refers to a therapeutic agent that targets an epigenetic regulator. Examples of epigenetic regulators include histone lysine methyltransferases, histone arginine methyltransferases, histone demethylases, histone deacetylases, histone acetylases, and DNA methyltransferases. Histone deacetylase inhibitors include, but are not limited to, vorinostat.

[0115] In another embodiment, the optional therapeutic agent is a chemotherapeutic agent or other anti-proliferative agent that can be administered in combination with TG02 or a pharmaceutically acceptable salt thereof to treat cancer. Examples of therapeutic and anti-cancer agents that can be used in combination with TG02 or a pharmaceutically acceptable salt thereof include surgery, radiation therapy (e.g., gamma-radiation, neutron beam radiation, electron beam radiation, proton therapy, brachytherapy, and systemic radioisotopes), endocrine therapy, biological response modifiers (e.g., interferons, interleukins, tumor necrosis factor (TNF)), hyperthermia and cryotherapy, agents to attenuate any adverse effects (such as anti-emetics), and any other approved chemotherapeutic drugs.

[0116] Non-limiting examples of anti-proliferative compounds include: aromatase inhibitors; anti-estrogens; anti-androgens; gonadotropin-releasing hormone agonists; topoisomerase I inhibitors; topoisomerase II inhibitors; microtubule active agents; alkylating agents, such as temozolomide; retinoids, carotenoids, or tocopherols; cyclooxygenase inhibitors; MMP inhibitors; mTOR inhibitors; antimetabolites; platinum compounds; methionine aminopeptidase inhibitors; bisphosphonates; anti-proliferative antibodies; heparanase inhibitors; Ras oncogene isoform inhibitors; telomerase inhibitors; proteasome inhibitors; compounds used in the treatment of hematologic malignancies; Flt-3 inhibitors; Hsp90 inhibitors; kinesin spindle protein inhibitors; MEK inhibitors; antitumor antibiotics; nitrosoureas; compounds targeting / decreasing the activity of a protein or lipid kinase; compounds targeting / decreasing the activity of a protein or lipid phosphatase, or any other anti-angiogenic compound.

[0117] Non-limiting exemplary aromatase inhibitors include steroids (such as atamestane, exemestane, and formestane), and non-steroids (such as aminoglutethimide, Roglethimide, pyridoglutethimide, trilostane, testolactone, ketoconazole, vorozole, fadrozole, anastrozole, and letrozole).

[0118] Non-limiting anti-estrogens include tamoxifen, fulvestrant, raloxifene, and raloxifene hydrochloride. Anti-androgens include, but are not limited to, bicalutamide. Gonadotropin-releasing hormone agonists include, but are not limited to, abarelix, goserelin, and goserelin acetate.

[0119] Non-limiting exemplary topoisomerase I inhibitors include topotecan, gimatecan, irinotecan, camptothecin and its analogs, 9-nitrocamptothecin, and the macromolecular camptothecin conjugate PNU-166148. Topoisomerase II inhibitors include, but are not limited to, anthracyclines (such as doxorubicin, daunorubicin, epirubicin, idarubicin, and nemorubicin); anthraquinones (such as mitoxantrone and losoxantrone); and podophillotoxines (such as etoposide and teniposide).

[0120] Microtubule active agents include microtubule-stabilizing, microtubule-destabilizing compounds, and microtubule polymerization inhibitors, including, but not limited to, taxoids (such as paclitaxel and docetaxel); vinca alkaloids (such as vinblastine, vinblastine sulfate, vincristine, and vincristine sulfate, as well as vinorelbine); discodermolide; colchicine; and epothilones and derivatives thereof.

[0121] Non-limiting exemplary alkylating agents include cyclophosphamide, ifosfamide, melphalan, and nitrosoureas, such as carmustine and lomustine.

[0122] Non-limiting exemplary matrix metalloproteinase inhibitors ("MMP inhibitors") include collagen peptidomimetic and non-peptidomimetic inhibitors, tetracycline derivatives, batimastat, marimastat, prinomastat, metastat, BMS-279251, BAY 12-9566, TAA211, MMI270B, and AAJ996.

[0123] Non-limiting exemplary mTOR inhibitors include compounds that inhibit the mammalian target of rapamycin (mTOR) and have anti-proliferative activity, such as sirolimus, everolimus, CCI-779, and ABT578.

[0124] Non-limiting exemplary antimetabolites include 5-fluorouracil (5-FU), capecitabine, gemcitabine, DNA demethylating compounds such as 5-azacytidine and decitabine, methotrexate, and edatrexate, and folic acid antagonists such as pemetrexed.

[0125] Non-limiting exemplary platinum compounds include carboplatin, cis-platin, cisplatin, and oxaliplatin.

[0126] Non-limiting exemplary methionine aminopeptidase inhibitors include benzamides or derivatives thereof and PPI-2458.

[0127] Non-limiting exemplary bisphosphonates include etridonic acid, clodronic acid, tiludronic acid, pamidronic acid, alendronic acid, ibandronic acid, risedronic acid, and zoledronic acid.

[0128] Non-limiting exemplary heparanase inhibitors include compounds that target, decrease, or inhibit heparan sulfate degradation, such as PI-88 and OGT2115.

[0129] Non-limiting exemplary compounds that target, decrease, or inhibit the oncogenic activity of Ras include farnesyl transferase inhibitors, such as L-744832, DK8G557, tipifarnib, and lonafarnib.

[0130] Non-limiting exemplary telomerase inhibitors include compounds that target, decrease, or inhibit telomerase activity, such as compounds that inhibit the telomerase receptor, such as imetelstat.

[0131] Non-limiting exemplary proteasome inhibitors include compounds that target, decrease, or inhibit telomerase activity, including but not limited to bortezomib. In some embodiments, the proteasome inhibitor is carfilzomib.

[0132] Non-limiting exemplary FMS-like tyrosine kinase inhibitors, which are compounds that target, decrease or inhibit FMS-like tyrosine kinase receptor (Flt-3R), include interferons, I-beta-D-arabinofuranosylcytosine (ara-c) and bisulfan; and ALK inhibitors, which are compounds that target, decrease or inhibit anaplastic lymphoma kinase.

[0133] Non-limiting exemplary Flt-3 inhibitors include PKC412, midostaurin, a staurosporine derivative, SU11248 and MLN518.

[0134] Non-limiting exemplary HSP90 inhibitors include compounds that target, decrease or inhibit the endogenous ATPase activity of HSP90; or compounds that degrade, target, decrease or inhibit HSP90 client proteins through the ubiquitin proteasome pathway. Compounds that target, decrease or inhibit the endogenous ATPase activity of HSP90 are specifically compounds, proteins or antibodies that inhibit the ATPase activity of HSP90, such as 17-allylamino, 17-demethoxygeldanamycin (17AAG), a geldanamycin derivative; other geldanamycin-related compounds; radicicol and HDAC inhibitors.

[0135] Non-limiting exemplary protein tyrosine kinase and / or serine and / or threonine kinase inhibitors or lipid kinase inhibitors include a) compounds that target, decrease or inhibit the activity of platelet-derived growth factor receptor (PDGFR), such as N-phenyl-2-pyrimidine-amine derivatives, such as imatinib, SU101, SU6668 and GFB111; b) compounds that target, decrease or inhibit the activity of fibroblast growth factor receptor (FGFR); c) compounds that target, decrease or inhibit the activity of insulin-like growth factor receptor I (IGF-IR), such as compounds that target, decrease or inhibit the activity of IGF-IR; d) compounds that target, decrease or inhibit the activity of the Trk receptor tyrosine kinase family, or ephrin B4 inhibitors; e) compounds that target, decrease or inhibit the activity of the Axl receptor tyrosine kinase family; f) compounds that target, decrease or inhibit the activity of the Ret receptor tyrosine kinase family; g) compounds that target, decrease or inhibit the activity of the Kit / SCFR receptor tyrosine kinase family, such as imatinib; h) compounds that target, decrease or inhibit the activity of the c-Kit receptor tyrosine kinase family, such as imatinib; i) compounds that target, decrease or inhibit the activity of c-Abl family members, gene fusion products thereof (such as Bcr-Abl kinase) and mutants, such as N-phenyl-2-pyrimidine-amine derivatives, such as imatinib or nilotinib; PD180970; AG957; NSC 680410; PD173955; or dasatinib; j) compounds that target, decrease or inhibit the activity of the protein kinase C (PKC) and Raf family members, MEK, SRC, JAK, FAK, PDK1, PKB / Akt and Ras / MAPK family members, and / or members of the cyclin-dependent kinase family (CDK), such as the cross- domain derivatives disclosed in U.S. Patent No. 5,093,330, such as midostaurin; other examples of compounds include: UCN-01, safingol, BAY 43-9006, myriocin, perifosine; ilmofosine; RO 318220 and RO 320432; GO 6976; isis 3521; LY333531 / LY379196; isoquinoline compounds; farnesyl transferase inhibitors; PD184352 or QAN697, or AT7519; k) compounds that target, decrease or inhibit the activity of protein tyrosine kinases, such as imatinib mesylate or tyrosine phosphorylation inhibitors, such as tyrosine phosphorylation inhibitor A23 / RG-50810; AG 99; tyrosine phosphorylation inhibitor AG 213; tyrosine phosphorylation inhibitor AG 1748; tyrosine phosphorylation inhibitor AG 490; tyrosine phosphorylation inhibitor B44; tyrosine phosphorylation inhibitor B44 (+) enantiomeric isomer; tyrosine phosphorylation inhibitor AG 555; AG 494;Tyrosine phosphorylation inhibitors AG 556, AG 957 and adaphostin (4-{[(2,5- dihydroxyphenyl)methyl]amino}-adamantyl benzoate; NSC 680410, adaphostin); 1) compounds targeting, decreasing or inhibiting the activity of the epidermal growth factor family of receptor tyrosine kinases (EGFR, ErbB2, ErbB3, ErbB4 in homo- and heterodimeric form) and mutants thereof, such as CP 358774, ZD 1839, ZM 105180; trastuzumab, cetuximab, gefitinib, erlotinib, OSI-774, CI-1033, EKB-569, GW-2016, antibodies E1.1, E2.4, E2.5, E6.2, E6.4, E2.11, E6.3 and E7.6.3, and 7H-pyrrolo[2,3-d]pyrimidine derivatives; and m) compounds targeting, decreasing or inhibiting the activity of the c-Met receptor.

[0136] Non-limiting exemplary compounds targeting, decreasing or inhibiting the activity of a protein or lipid phosphatase include inhibitors of phosphatase 1, phosphatase 2A or CDC25, such as okadaic acid or derivatives thereof.

[0137] Other antiangiogenic compounds include compounds having other mechanisms of activity unrelated to inhibition of protein or lipid kinase, such as thalidomide and TNP-470.

[0138] In addition, one or more non-limiting exemplary chemotherapy compounds can be used in conjunction with TG02 or a pharmaceutically acceptable salt thereof, including bevacizumab, daunorubicin, doxorubicin, Ara-C, VP-16, teniposide, mitoxantrone, idarubicin, carboplatin, PKC412, 6-mercaptopurine (6-MP), fludarabine phosphate, octreotide, SOM230, FTY720, 6-thioguanine, cladribine, 6-mercaptopurine, pentostatin, hydroxyurea, 2-hydroxy-lH-isoindole-l,3-dione derivatives, l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine or a pharmaceutically acceptable salt thereof, l-(4-chloroanilino)-4-(4-pyridylmethyl)phthalazine succinate, angiostatin, endostatin, anthranilic acid amides, ZD4190, ZD6474, SU5416, SU6668, bevacizumab, rhuMAb, rhuFab, macugon; FLT-4 inhibitors, FLT-3 inhibitors, VEGFR-2 IgGI antibodies, RPI 4610, bevacizumab, porfimer sodium, anecortave, triamcinolone, hydrocortisone, 11-a-epihydrocotisol, cortex olone, 17a-hydroxyprogesterone, cortexone, deoxycortone, testosterone, estrone, dexamethasone, fluocinolone, plant alkaloids, hormonal compounds and / or antagonists, biological response modifiers such as lymphokines or interferons, antisense oligonucleotides or oligonucleotide derivatives, shRNA and siRNA.

[0139] Many suitable optional therapeutic agents, e.g., anti-cancer agents, are contemplated for use in the treatment methods provided herein. Indeed, the methods provided herein can include, but are not limited to, administration of a variety of optional therapeutic agents, e.g., agents that induce apoptosis; polynucleotides (e.g., antisense, ribozymes, siRNA); polypeptides (e.g., enzymes and antibodies); biological mimetics (e.g., gossypol or BH3 mimetics); agents that bind to (e.g., oligomerize or complex with) Bcl-2 family proteins such as Bax; alkaloids; alkylating agents; antitumor antibiotics; antimetabolites; hormones; platinum compounds; monoclonal or polyclonal antibodies (e.g., antibodies that bind to anti-cancer drugs, toxins, defensins), toxins; radionuclides; biological response modifiers (e.g., interferons (e.g., IFN-a) and interleukins (e.g., IL-2)); adoptive immunotherapy agents; hematopoietic growth factors; agents that induce tumor cell differentiation (e.g., all-trans retinoic acid); gene therapy agents (e.g., antisense therapy agents and nucleotides); tumor vaccines; angiogenesis inhibitors; proteasome inhibitors; NF-KB modulators; anti-CDK compounds; HDAC inhibitors; and the like. Many other examples of optional therapeutic agents (e.g., chemotherapy compounds and anti-cancer therapies) suitable for co-administration with the disclosed compounds are known to those of skill in the art.

[0140] In certain embodiments, the anti-cancer agent comprises an agent that induces or stimulates apoptosis. Agents that induce or stimulate apoptosis include, for example, agents that interact with or modify DNA, such as by intercalation, cross-linking, alkylation or otherwise damaging or chemically modifying DNA. Agents that induce apoptosis include, but are not limited to, radiation (e.g., X-rays, gamma rays, UV); tumor necrosis factor (TNF) related factors (e.g., TNF family receptor proteins, TNF family ligands, TRAIL, antibodies against TRAIL-R1 or TRAIL-R2); kinase inhibitors (such as epidermal growth factor receptor (EGFR) kinase inhibitors. Other anti-cancer drugs include: vascular growth factor receptor (VGFR) kinase inhibitors, fibroblast growth factor receptor (FGFR) kinase inhibitors, platelet-derived growth factor receptor (PDGFR) kinase inhibitors, and Bcr-Abl kinase inhibitors (such as GLEEVEC)); antisense molecules; antibodies (e.g., HERCEPTIN, RITUXAN, ZEVALIN, and AVASTIN); anti-estrogens (such as raloxifene and tamoxifen); anti-androgens (e.g., flutamide, bicalutamide, finasteride, aminoglutethimide, ketoconazole, and corticosteroids); cyclooxygenase 2 (COX-2) inhibitors (such as celecoxib, meloxicam, NS-398, and non-steroidal anti-inflammatory drugs (NSAIDs)); anti-inflammatory drugs (e.g., diazolidinyl urea, DECADRON, DELTASONE, dexamethasone, dexamethasone concentration, DEXONE, HEXADROL, hydroxychloroquine, METICORTEN, ORADEXON, ORASONE, oxyphenbutazone, PEDIAPRED, phenylbutazone, PLAQUENIL, prednisolone, prednisone, PRELONE, and TANDEARIL); and cancer chemotherapeutics (such as irinotecan (CAMPTOSAR), CPT-11, fludarabine (FLUDARA), dacarbazine (DTIC), dexamethasone, mitoxantrone, MYLOTARG, VP-16, cisplatin, carboplatin, oxaliplatin, 5-FU, doxorubicin, gemcitabine, bortezomib, gefitinib, bevacizumab, TAXOTERE, or TAXOL); cell signaling molecules; ceramides and cytokines; streptonigrin, and the like.

[0141] In other embodiments, the methods of treatment provided herein comprise administering to a cancer patient a therapeutically effective amount of TG02 and at least one other anti-hyperproliferative or anti-neoplastic agent selected from the group consisting of alkylating agents, antimetabolites, and natural products (e.g., herbs and other plant and / or animal derived compounds).

[0142] Alkylating agents suitable for use in the methods of the present application include, but are not limited to: 1) nitrogen mustards (e.g., mechlorethamine, cyclophosphamide, ifosfamide, melphalan (L-sarcolysin); and chlorambucil); 2) ethylenimines and methyl triazenes (e.g., hexamethylmelamine and thioepa); 3) alkyl sulfonates (e.g., busulfan); 4) nitrosoureas (e.g., carmustine (BCNU); lomustine (CCNU); semustine (methyl-CCNU); and streptozotocin (streptozotocin)); and 5) triazines (e.g., dacarbazine (DTIC; dimethyltriazenylimidazole carboxamide)).

[0143] In some embodiments, antimetabolites suitable for use in the methods of the present application include, but are not limited to: 1) folic acid analogs (e.g., methotrexate (amethopterin)); 2) pyrimidine analogs (e.g., fluorouracil (5-fluorouracil; 5-FU), floxuridine (fluoro- deoxyuridine; FudR), and cytarabine (cytosine arabinoside)); 3) purine analogs (e.g., mercaptopurine (6-mercaptopurine; 6-MP), thioguanine (6-thioguanine; TG), and pentostatin (2'-deoxycoformycin)).

[0144] In still further embodiments, chemotherapeutic agents suitable for use in the methods of the present disclosure include, but are not limited to: 1) vinca alkaloids (e.g., vinblastine (VLB), vincristine); 2) epipodophyllotoxins (e.g., etoposide and teniposide); 3) antibiotics (e.g., dactinomycin (actinomycin D), daunorubicin (daunomycin; rubidomycin), doxorubicin, bleomycin, plicamycin (mithramycin), and mitomycin (mitomycin C)); 4) enzymes (e.g., L-asparaginase); 5) biological response modifiers (e.g., interferon-α); 6) platinum coordination complexes (e.g., cisplatin (cis-DDP) and carboplatin); 7) anthracenediones (e.g., mitoxantrone); 8) substituted ureas (e.g., hydroxyurea); 9) methyl hydrazine derivatives (e.g., procarbazine (N-methylhydrazine; MIH)); 10) adrenocortical suppressants (e.g., mitotane (o,p'-DDD) and aminoglutethimide); 11) adrenocorticosteroids (e.g., prednisone); 12) progestins (e.g., hydroxyprogesterone caproate, medroxyprogesterone acetate, and megestrol acetate); 13) estrogens (e.g., stilbestrol and ethylnyl estradiol); 14) antiestrogens (e.g., tamoxifen); 15) androgens (e.g., testosterone propionate and fluoxymesterone); 16) antiandrogens (e.g., flutamide); and 17) gonadotropin-releasing hormone analogs (e.g., leuprolide).

[0145] Any oncolytic agent conventionally used in the context of cancer therapy can be used in the treatment methods of the present disclosure. For example, the U.S. Food and Drug Administration (FDA) maintains a formulary of oncolytic agents approved for use in the United States. International counterpart agencies of the FDA maintain similar formularies. Those skilled in the art will appreciate that the “product label” required for all U.S. approved chemotherapeutic agents describes the approved indications, dosage information, toxicity data, etc. for the exemplified agent.

[0146] Anti-cancer agents also include compounds that have been identified as having anti-cancer activity. Examples include, but are not limited to, 3-AP, 12-O-tetradecanoylphorbol-13-acetate, 17AAG, 852A, ABI-007, ABR-217620, ABT-751, ADI-PEG 20, AE-941, AG-013736, AGRO100, alamthine, AMG706, antibody G250, antineoplaston, AP23573, apaziquone, APC8015, aptigene, ATN-161, atrasenten, azacitidine, BB-10901, BCX-1777, bevacizumab, BG00001, bicalutamide, BMS 247550, bortezomib, bryostatin-1, buserelin, calcitriol, CCI-779, CDB-2914, cefixime, cetuximab, CG0070, cilengitide, clofarabine, combretastin A4 phosphate, CP-675,206, CP-724,714, CpG 7909, curcumin, decitabine, DENSPM, doxercalciferol, E7070, E7389, ecteinascidin 743, efaproxiral, eflomithine, EKB-569, enzalutamide, erlotinib, etoricoxib, fenretinide, flavopiridol, fludarabine, flutamide, fotemustine, FR901228, G17DT, galiximab, gefitinib, genistein, glufosfamide, GTI-2040, histrelin, HKI-272, homoharringtonine, HSPPC-96, hu14.18 - interleukin-2 fusion protein, HuMax-CD4, iloprost, imiquimod, infliximab, interleukin-12, IPI-504, ilofungine, ixabepilone, lapatinib, lenalidomide, letrozole, leuprolide, LMB-9 immunotoxin, lonafarnib, lumiliximab, mafosfamide, MB07133, MDX-010, MLN2704, monoclonal antibody 3F8, monoclonal antibody J591, motexafin, MS-275, MVA-MUC1-IL2, nilotinib, nitrocomphostin, nolatrexed, Novadigm, NS-9, O6-benzylguanine, oblimersen sodium, ONYX-015, omalizumab, OSI-774, panitumumab, paraplatin, PD-0325901, pemetrexed, PHY906, pioglitazone, pirfenidone, plitidepsin, PS-341, PSC 833, PXD101, pyrazolo pyridine, R115777, RAD001, ranpimase, rapamycin analogs, recombinant human endostatin protein, rhuMab 2C4, rosiglitazone, rubitecan, S-1, S-8184, sandoplatin, SB-, 15992, SGN-0010, SGN-40, sorafenib, SR31747A, ST1571, SU011248, suberoylanilide hydroxamic acid, suramin, talabostat, talaporfin, tariquidar, temsirolimus, TGFa-PE38 immunotoxin, thalidomide, thymalfasin, tipifarnib, tirapazamine, TLK286, trabectedin, trimetrexate glucuronate, TroVax, UCN-1, valproic acid, vinflunine, VNP40101M, vorinostat, VX-680, ZD1839, ZD6474, zileuton, and zolazepam trihydrochloride.

[0147] For more detailed descriptions of anticancer agents and other optional therapeutic agents, one skilled in the art can refer to any number of instructive manuals, including but not limited to the Physician's Desk Reference and Goodman and Gilman's The Pharmaceutical Basis of Therapeutics, Tenth Edition, Hardman et al., eds., 2002.

[0148] In some embodiments, the methods provided herein comprise administering TG02 to a cancer patient in combination with radiation therapy. The methods provided herein are not limited by the type, amount, or delivery and dosing system used to deliver a therapeutic dose of radiation to the patient. For example, the patient can receive photon radiation therapy, particle beam radiation therapy, other types of radiation therapy, and combinations thereof. In some embodiments, the radiation is delivered to the patient using a linear accelerator. In other embodiments, the radiation is delivered using a gamma knife.

[0149] The source of radiation can be external or internal to the patient. External radiation therapy is the most common and involves directing a beam of high-energy radiation through the skin to the tumor site using, for example, a linear accelerator. While the beam of radiation is targeted at the tumor site, it is almost impossible to avoid exposure of normal healthy tissue. However, patients generally tolerate external radiation well. Internal radiation therapy involves implanting a source of emitted radiation, such as a bead, wire, pellet, capsule, particle, etc., including using a delivery system that specifically targets cancer cells (e.g., using particles attached to cancer cell binding ligands), into the body at or near the tumor site. Such implants can be removed after treatment, or left in the body inactive. Types of internal radiation therapy include, but are not limited to, brachytherapy, interstitial irradiation, intracavitary irradiation, radioimmunotherapy, and the like.

[0150] The patient can optionally receive a radiosensitizer (e.g., metronidazole, imidazole, intra-arterial Budr, intravenous iododeoxyuridine (IudR), nitroimidazole, 5-substituted-4-nitroimidazole, 2H-isoindolequinone, [[ (2-bromoethyl) -amino]methyl] -nitro-1H-imidazole-1-ethanol, nitroaniline derivative, DNA-affmityl hypoxia-selective cytotoxin, halogenated DNA ligand, 1,2,4-benzotriazine oxide, 2-nitroimidazole derivative, fluorine-containing nitroazole derivative, benzamide, nicotinamide, acridine-intercalator, 5-thiotetrazole derivative, 3-nitro-1,2,4-triazole, 4,5-dinitroimidazole derivative, hydroxylated tetrac, cisplatin, mitomycin, tirapazamine, nitrosoureas, mercaptopurine, methotrexate, fluorouracil, bleomycin, vincristine, carboplatin, epirubicin, doxorubicin, cyclophosphamide, vindesine, etoposide, paclitaxel, heat (hyperthermia), etc.), radioprotector (e.g., cysteamine, aminoalkyl dihydrogen phosphorothioate, amifostine (WR 2721), IL-1, IL-6, etc.). Radiosensitizers enhance the killing of tumor cells by radiation. Radioprotectors protect healthy tissue from the deleterious effects of radiation.

[0151] Any type of radiation can be administered to the patient, so long as the patient can tolerate the radiation dose without unacceptable negative side effects. Suitable types of radiation therapy include, for example, ionizing (electromagnetic) radiation therapy (e.g., X-rays or gamma rays) or particle beam radiation therapy (e.g., high linear energy transfer radiation). Ionizing radiation is defined as radiation that includes particles or photons that have sufficient energy to produce ionization, i.e., the gain or loss of electrons (e.g., as described in U.S. 5,770,581, which is incorporated herein by reference in its entirety). The effects of the radiation can be controlled, at least in part, by the clinician. In one embodiment, the radiation dose is fractionated to achieve maximum target cell exposure and reduced toxicity.

[0152] In one embodiment, the total dose of radiation administered to the patient is from about 0.01 Gray (Gy) to about 100 Gy. In another embodiment, from about 10 Gy to about 65 Gy (e.g., about 15 Gy, 20 Gy, 25 Gy, 30 Gy, 35 Gy, 40 Gy, 45 Gy, 50 Gy, 55 Gy, or 60 Gy) is administered over the course of the treatment. While in some embodiments, the entire dose of radiation can be administered over the course of a day, desirably the total dose is fractionated and administered over several days. Desirably, the radiation therapy is administered over the course of at least about 3 days, e.g., at least 5, 7, 10, 14, 17, 21, 25, 28, 32, 35, 38, 42, 46, 52, or 56 days (about 1-8 weeks). Thus, the daily radiation dose will include from about 1-5 Gy (e.g., about 1 Gy, 1.5 Gy, 1.8 Gy, 2 Gy, 2.5 Gy, 2.8 Gy, 3 Gy, 3.2 Gy, 3.5 Gy, 3.8 Gy, 4 Gy, 4.2 Gy, or 4.5 Gy), or 1-2 Gy (e.g., 1.5-2 Gy). The daily radiation dose should be sufficient to induce destruction of the targeted cells. In one embodiment, the radiation is not administered every day if extended over a period of time, allowing the animal to rest and effect the treatment. For example, for a weekly treatment, it is desirable to administer the radiation for 5 consecutive days, rather than 2 days, allowing for 2 days of rest per week. However, the radiation can be administered 1 day / week, 2 days / week, 3 days / week, 4 days / week, 5 days / week, 6 days / week, or all 7 days / week, depending on the responsiveness of the animal and any potential side effects. The radiation treatment can begin at any time during the treatment period. In one embodiment, the radiation begins in the first week or the second week and is administered for the remainder of the duration of the treatment period. For example, the radiation is administered for 6 weeks, including weeks 1-6 or weeks 2-6 of the treatment period, for the treatment of, e.g., a solid tumor. Alternatively, the radiation is administered for 5 weeks, including weeks 1-5 or weeks 2-5 of the treatment period. However, these exemplary radiation treatment schedules are not intended to limit the methods provided herein.

[0153] IV. Methods of Treatment

[0154] In the treatment methods provided herein, TG02, the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent (e.g., anti-cancer agent) can be administered to the cancer patient under one or more of the following conditions: under different cycles, under different durations, under different concentrations, by different routes of administration, etc.

[0155] In some embodiments, TG02 is administered prior to the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent, e.g., 0.5, 1, 2, 3, 4, 5, 10, 12, or 18 hours, 1, 2, 3, 4, 5, or 6 days, or 1, 2, 3, or 4 weeks prior to administration of the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent.

[0156] In some embodiments, TG02 is administered after the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent, e.g., 0.5, 1, 2, 3, 4, 5, 10, 12, or 18 hours, 1, 2, 3, 4, 5, or 6 days, or 1, 2, 3, or 4 weeks after administration of the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent.

[0157] In some embodiments, TG02, the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent are administered concurrently, but on different schedules, e.g., TG02 is administered daily, while the immune checkpoint inhibitor is administered once a week, once every two weeks, once every three weeks, or once every four weeks. In other embodiments, TG02 is administered once daily, while the immune checkpoint inhibitor, the COX-2 inhibitor, and / or the optional therapeutic agent are administered once a week, once every two weeks, once every three weeks, or once every four weeks.

[0158] The methods of treatment provided herein include administering TG02 to a cancer patient in an amount effective to achieve its intended purpose. Although individual needs vary, a determination of an optimal range of effective amount of each component is within the capability of those skilled in the art. Generally, TG02 can be administered in an amount from about 1 mg / kg to about 500 mg / kg, from about 1 mg / kg to about 100 mg / kg, or from about 1 mg / kg to about 50 mg / kg. The dose of the composition can be any dose, including but not limited to 30-600 mg / day. Specific doses include 50, 100, 200, 250, 300, 400, 500, and 600 mg / day. In one embodiment, TG02 is administered once daily for consecutive days 3-7 days prior to administration of the immune checkpoint inhibitor. In another embodiment, 250 mg / day of TG02 is administered. In another embodiment, 250 mg / day of TG02 is administered twice a week. In another embodiment, TG02 administration continues on the day of immune checkpoint inhibitor administration and for additional days until disease progression or until TG02 administration is no longer beneficial. These doses are examples of average cases, but there are individual cases where higher or lower doses should be used, and such doses are within the scope of the application. In practice, the physician will determine the dosage regimen that is most appropriate for an individual patient, which can vary with the age, weight, and response of the particular patient.

[0159] A unit oral dose of TG02 can comprise from about 0.01 to about 1000 mg, for example, from about 10 to about 500 mg, of TG02. In one embodiment, a unit oral dose of TG02 is 10 mg, 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 110 mg, 120 mg, 130 mg, 140 mg, 150 mg, 160 mg, 170 mg, 180 mg, 190 mg, 200 mg, 210 mg, 220 mg, 230 mg, 240 mg, 250 mg, 260 mg, 270 mg, 280 mg, 290 mg, or 300 mg. The unit dose can be administered once or multiple times per day, for example, in one or more tablets or capsules.

[0160] In addition to administering TG02 as a raw chemical, it can also be part of a pharmaceutical formulation or composition. In some embodiments, the pharmaceutical formulation or composition can include one or more pharmaceutically acceptable carriers, excipients, and / or auxiliaries. In some embodiments, the one or more carriers, excipients and auxiliaries aid in processing TG02 into a formulation or composition that can be used pharmaceutically. Formulations, particularly those suitable for oral or topical administration, can be presented as discrete units suitable for such modes of administration as, for example, tablets, pills, caplets, sachets, cachets, lozenges, aqueous or oily suspensions, aqueous or oily solutions, emulsions, jellies, gels, powders, sustained release formulations, elixirs, sprinkles, sprays, ointments, and the like; and those suitable for rectal administration, such as suppositories; and those suitable for parenteral administration, such as sterile solutions containing amounts of the active compound that are from about 0.01 to 99%, in one embodiment, from about 0.25 to 75%, in combination with the one or more carriers, excipients and / or auxiliaries.

[0161] The pharmaceutical compositions provided herein can be administered to any patient that can experience a beneficial effect from TG02. Foremost among these patients are mammals, such as humans, although the methods and compositions provided herein are not so limited. Other patients include veterinary animals (cows, sheep, pigs, horses, dogs, cats, etc.).

[0162] The pharmaceutical formulations provided herein are prepared by conventional mixing, granulation, sugar coating, dissolving or lyophilization processes. Thus, pharmaceutical formulations for oral use can be obtained by combining the active compounds with solid excipient, if desired or necessary, with the aid of suitable auxiliaries, then, optionally, grinding the resulting mixture, and processing the mixture, if desired or necessary, through sieves and the like.

[0163] Suitable excipients include: fillers such as sugars, such as lactose or sucrose, mannitol or sorbitol; cellulose preparations and / or calcium phosphates, such as tricalcium phosphate or dicalcium phosphate; and binders such as starch pastes using corn starch, wheat starch, rice starch, potato starch, gelatin, astragalus gum, methylcellulose, hydroxypropyl methylcellulose, sodium carboxymethyl cellulose, and / or polyvinylpyrrolidone. Disintegrants, such as those found in the starches mentioned above, may be added if desired; they may also be carboxymethyl starch, croscarmellose, agar or alginate or their salts, such as sodium alginate. Additives may be suitable flow conditioners and lubricants. Suitable additives include, for example, silica, talc, stearic acid or its salts, such as magnesium stearate or calcium stearate, and / or polyethylene glycol. The sugar core may be coated with a suitable coating, if desired, that is resistant to gastric juices. For this purpose, a concentrated sugar solution may be used, optionally containing gum arabic, talc, polyvinylpyrrolidone, polyethylene glycol and / or titanium dioxide, varnish solution, and a suitable organic solvent or solvent mixture. To produce a coating resistant to gastric juices, a suitable cellulose formulation, such as a solution of acetylcellulose phthalate or hydroxypropyl methylcellulose phthalate, may be used. Dyes or pigments may be added to the coating of tablets or lozenges to identify or characterize combinations of active compound dosages.

[0164] Other pharmaceutical formulations suitable for oral administration include push-fit capsules made of gelatin, and sealed soft capsules made of gelatin and plasticizers such as glycerin or sorbitol. Push-fit capsules may contain an active compound in particulate form, which may be mixed with fillers such as lactose, binders such as starch, and / or lubricants such as talc or magnesium stearate, and optionally, a stabilizer. In one embodiment, in soft capsules, the active compound is dissolved or suspended in a suitable liquid, such as fatty oil or liquid paraffin. A stabilizer may also be added.

[0165] Possible pharmaceutical formulations that can be used rectally include, for example, suppositories composed of one or more active compounds combined with a suppository base. Suitable suppositories bases are, for example, natural or synthetic triglycerides or paraffin hydrocarbons. Alternatively, gelatin rectal capsules, which are composed of an active compound and a base, may also be used. Possible bases include, for example, liquid triglycerides, polyethylene glycol, or paraffin hydrocarbons.

[0166] Suitable parenteral formulations include aqueous solutions and alkaline solutions of the active compound in its water-soluble form (e.g., water-soluble salts). Alternatively, a suitable oily injectable suspension of the active compound may be administered. Suitable lipophilic solvents or carriers include fatty oils such as sesame oil, or synthetic fatty acid esters such as ethyl oleate or triglycerides, or polyethylene glycol-400. Aqueous injectable suspensions may also contain substances that increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, and / or dextran. Optionally, the suspension may also contain a stabilizer.

[0167] The present disclosure encompasses the use of solvates of TG02. Solvates generally do not significantly alter the physical properties of a compound, and thus can be considered physio logically equivalent. The term "solvate" as used herein is a combination of TG02 with solvent molecules, physically associated and / or solvated, e.g., disolvate, monosolvate or hemisolvate, wherein the ratio of solvent molecules to TG02 is about 2: 1, about 1: 1 or about 1:2, respectively. Such physical associations involve varying degrees of ionization and covalency, including hydrogen bonding. In some cases, the solvate will be capable of isolation, such as when one or more solvent molecules are incorporated into the crystal lattice of the solid state form. "Solvate" includes solution phases and isolatable solvates. TG02 can exist in solvated as well as unsolvated forms with pharmaceutically acceptable solvents such as water, methanol, ethanol and the like, and the present disclosure is intended to include both solvated and unsolvated forms of TG02. One solvate is a hydrate. "Hydrate" refers to a specific subgroup of solvates wherein the solvent molecule is water. Solvates generally can be considered physiologically equivalent. The preparation of solvates is known in the art. See, e.g., M. Caira et al., J. Pharmaceut. Sci., 93(3):601-611 (2004), which describes the preparation of solvates of fluconazole with ethyl acetate and water. E.C. van Tonder et al., AAPS Pharm. Sci. Tech., 5(1):Article 12 (2004), and A.L. Bingham et al., Chem. Commun. 603-604 (2001) describe similar preparations of solvates, hemisolvates, hydrates and the like. Typical non-limiting methods of preparing solvates include dissolving TG02 in the desired solvent (organic, water or mixtures thereof) at temperatures above 20 °C to about 25 °C, then cooling the solution at a rate sufficient to form crystals, and isolating the crystals by known methods, e.g., filtration. Analytical techniques such as infrared spectroscopy can be used to confirm the presence of a solvent in the solvate crystals.

[0168] A therapeutically effective amount of TG02 and / or an immune checkpoint inhibitor and / or a COX-2 inhibitor and / or an optional therapeutic agent, formulated in accordance with standard pharmaceutical practice, is administered to a human patient in need thereof. Whether such treatment is indicated depends on the individual case and requires medical evaluation (diagnosis) that considers the signs, symptoms and / or functional disturbances present, the risk of developing particular signs, symptoms and / or functional disturbances, and other factors.

[0169] TG02, immune checkpoint inhibitor, COX-2 inhibitor and / or optional therapeutic agent can be administered by any suitable route, for example, by oral, buccal, inhalation, sublingual, rectal, vaginal, intracisternal or intrathecal (via lumbar puncture), transurethral, transnasal, transdermal (i.e., transcutaneous) or parenteral (including intravenous, intramuscular, subcutaneous, intracoronary, intradermal, intramammary, intraperitoneal, intraarticular, intrathecal, retrobulbar, intrapulmonary injection and / or surgical implantation into a specific site). Parenteral administration can be achieved using a needle or syringe or using high pressure technology.

[0170] Pharmaceutical compositions include those wherein TG02, immune checkpoint inhibitor, COX-2 inhibitor and / or optional therapeutic agent is administered in an effective amount to accomplish its intended purpose. The exact formulation, route of administration and dosage is chosen by the individual physician in view of the disease or condition being treated, and the dosage can be adjusted according to the patient's age, body weight and response to the therapy. Optimal dosages can vary depending on the type of disease or disorder being treated. In general, satisfactory results are indicated to be achieved at a total daily dose of TG02, immune checkpoint inhibitor, COX-2 inhibitor and / or optional therapeutic agent in the range of 0.01 to 100 mg / kg or 0.5 to 1000 mg / m2. The dosage should not exceed 1000 mg / kg or 50 mg / kg, respectively. The pharmaceutical compositions can be administered on a regimen of 1 to 4 times per day, week, month or year.

[0171] Toxicity and therapeutic efficacy of TG02, immune checkpoint inhibitor, COX-2 inhibitor and / or optional therapeutic agent can be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., for determining the maximum tolerated dose (MTD) of a certain compound, which is defined as the highest dose not causing toxicity in patients. The dose ratio between the maximum tolerated dose and the therapeutic effect, e.g., inhibition of tumor growth, is the therapeutic index. The dosage can vary within this range depending upon the dosage form employed and the route of administration utilized. Determination of a therapeutically effective amount is within the skill of the art, given the teachings of the detailed disclosure herein.

[0172] The therapeutically effective amount of TG02, immune checkpoint inhibitor, COX-2 inhibitor and / or optional therapeutic agent required to treat a given disease or condition will vary according to the nature of the condition, the length of time required for the activity, and the age and state of the patient, and will ultimately be at the discretion of the attendant physician. For example, the dosage and interval between doses can be adjusted to provide TG02 and immune checkpoint inhibitor plasma levels sufficient to maintain therapeutic effect. The desired dose can be given in a single dose, or as multiple doses, at appropriate intervals, for example, 1, 2, 3, 4, or more sub-doses per day. Multiple doses are usually required. For example, TG02 and immune checkpoint inhibitor of the application can be administered at a frequency of: once per day; four doses delivered at a once-daily dose with four days between doses (q4d x 4); four doses delivered at a once-daily dose with three days between doses (q3d x 4); one dose delivered per day with five days between doses (qd x 5); a once-daily dose for three weeks (qwk3); five daily doses, two days apart, followed by another five daily doses (5 / 2 / 5); or any dosage regimen determined to be appropriate in the circumstances.

[0173] The immune checkpoint inhibitor is administered in a therapeutically effective amount. When the immune checkpoint inhibitor is a monoclonal antibody, 1-20 mg / kg is administered intravenously every 2-4 weeks. For example, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 200 mg, 300 mg, 400 mg, 500 mg, 600 mg, 700 mg, 800 mg, 900 mg, 1000 mg, 1100 mg, 1200 mg, 1300 mg, 1400 mg, 1500 mg, 1600 mg, 1700 mg, 1800 mg, 1900 mg, and 2000 mg of the antibody can be administered.

[0174] For example, when the immune checkpoint inhibitor is the anti-PD-1 antibody nivolumab, 3 mg / kg can be administered by intravenous infusion over 60 minutes every two weeks. When the immune checkpoint inhibitor is the anti-PD-1 antibody pembrolizumab, 2 mg / kg can be administered by intravenous infusion over 30 minutes every two or three weeks. When the immune checkpoint inhibitor is the anti-PD-Ll antibody atezolizumab, 10 mg / kg can be administered by intravenous infusion at a frequency of every 2 weeks. Disis et al., J. Clin Oncol. 33 (2015) (Suppl; abstr 5509). When the immune checkpoint inhibitor is the anti-PD-Ll antibody MPDL3280A, 20 mg / kg can be administered by intravenous infusion every 3 weeks. Herbst et al., Nature 515:563-80 (2014). When the immune checkpoint inhibitor is the anti-CTLA-4 antibody ipilimumab, 3 mg / kg can be administered by intravenous infusion over 90 minutes every three weeks. When the immune checkpoint inhibitor is the anti-CTLA-4 antibody tremelimumab, 15 mg / kg can be administered by intravenous infusion every 12 weeks. Naido et al., British Journal of Cancer 111 :2214-19 (2014); Drugs R D, 10:123-32 (2010). When the immune checkpoint inhibitor is the anti-LAG3 antibody GSK2831781, 1.5-5 mg / kg can be administered by intravenous infusion over 120 minutes or more every 2-4 weeks. When the immune checkpoint inhibitor is an anti-TIM3 antibody, 1-5 mg / kg can be administered by intravenous infusion over 30-90 minutes or more every 2-4 weeks. When the inhibitor of the indoleamine 2,3-dioxygenase (IDO) pathway is the inhibitor indoximod in combination with temozolomide, indoximod is escalated from 18.5 mg / kg / dose BID to 27.7 mg / kg / dose BID, 200 mg / m 2 Temozolomide.

[0175] The COX-2 inhibitor is also administered in a therapeutically effective amount, such as about 1 mg / kg to about 500 mg / kg, about 1 mg / kg to about 100 mg / kg, or about 1 mg / kg to about 50 mg / kg. A unit oral dose of the COX-2 inhibitor can comprise about 0.01 to about 1000 mg, such as about 1 to about 250 mg, of the COX-2 inhibitor. In one embodiment, a unit oral dose of the COX-2 inhibitor is 5 mg, 10 mg, 20 mg, 30 mg, 40 mg, 50 mg, 60 mg, 70 mg, 80 mg, 90 mg, 100 mg, 110 mg, 120 mg, 130 mg, 140 mg, 150 mg, 160 mg, 170 mg, 180 mg, 190 mg, 200 mg, 210 mg, 220 mg, 230 mg, 240 mg, or 250 mg. The unit dose can be given once or multiple times per day, for example, in one or more tablets or capsules.

[0176] In one embodiment, the immune checkpoint inhibitor is an antibody, and 1-20 mg / kg is administered every 2-4 weeks by intravenous infusion. In another embodiment, 50-2000 mg of the antibody is administered every 2-4 weeks by intravenous infusion. In another embodiment, TG02 is administered prior to administration of the antibody. In another embodiment, TG02 is administered 3-7 days prior to the day of administration of the antibody. In another embodiment, TG02 is also administered on the day of administration of the antibody, and for consecutive days thereafter until disease progression or until TG02 administration is no longer beneficial.

[0177] In one embodiment, the cancer patient has a tumor with a biomarker, such as overexpression of MYC and / or MCL1, and 2 mg / kg of pembrolizumab is administered every three weeks by intravenous infusion and 30-600 mg of TG02 is administered for 3-7 days prior to pembrolizumab administration, on the day of pembrolizumab administration, and thereafter until disease progression or until there is no therapeutic benefit.

[0178] In another embodiment, the cancer patient has a tumor with a biomarker, such as overexpression of MYC and / or MCL1, and 3 mg / kg of nivolumab is administered every 2 weeks by intravenous infusion and 30-600 mg of TG02 is administered orally for 3-7 days prior to nivolumab administration, on the day of nivolumab administration, and thereafter until disease progression or until there is no therapeutic benefit.

[0179] In another embodiment, the cancer patient has a tumor with a biomarker, such as overexpression of MYC and / or MCL1, and is administered 3 mg / kg of nimotuzumab via intravenous infusion every 2 weeks and 30-600 mg of TG02 orally twice a week prior to, on the day of, and after nimotuzumab administration until disease progression or until there is no therapeutic benefit.

[0180] In another embodiment, treating a cancer patient with an immune checkpoint inhibitor and TG02 induces an anti-proliferative response more quickly than when the immune checkpoint inhibitor is administered alone.

[0181] In another embodiment, treating a cancer patient with a COX-2 inhibitor and TG02 induces an anti-proliferative response more quickly than when the COX-2 inhibitor is administered alone.

[0182] V. Biomarkers

[0183] The term "biomarker" as used herein refers to any biological compound, such as a gene, protein, protein fragment, peptide, polypeptide, nucleic acid, etc., that can be detected and / or quantified in a cancer patient or in a biological sample obtained from a cancer patient. The biomarker can be an intact molecule, or it can be a portion or fragment thereof. In one embodiment, the expression level of the biomarker is measured. The expression level of the biomarker can be measured, for example, by detecting the protein or RNA, such as mRNA, level of the biomarker. In some embodiments, a portion or fragment of the biomarker can be detected or measured, for example, by an antibody or other specific binding agent. In some embodiments, a measurable aspect of the biomarker is correlated with a given state of the patient, such as a particular stage of cancer. For biomarkers detected at the protein or RNA level, such measurable aspects can include, for example, the presence, absence, or concentration (i.e., expression level) of the biomarker in a cancer patient, or in a biological sample obtained from a cancer patient. For biomarkers detected at the nucleic acid level, such measurable aspects can include, for example, the allelic form of the biomarker or the type, rate, and / or degree of mutation of the biomarker, also referred to herein as the mutation status.

[0184] For biomarkers based on protein or RNA expression level measurements, it can be considered that the expression levels measured between different phenotypic states are different, e.g., if the average or median expression level of a biomarker in different groups is statistically significant. Common tests for statistical significance include t-tests, ANOVA, Kruskal-Wallis, Wilcoxon, Mann-Whitney microarray importance analysis, resubstitution ratio, etc. Biomarkers, alone or in combination, provide a measure of the relative likelihood that a subject belongs to one or another phenotypic state. They are thus useful, inter alia, as markers of disease, and as indicators of the likely beneficial patient outcome from a particular treatment regimen.

[0185] Biomarkers include, but are not limited to, the genes listed in Table 1. In one embodiment, the measurable aspect of a biomarker is its expression state. In one embodiment, the measurable aspect of a biomarker is its mutation state.

[0186] Table 1

[0187]

[0188]

[0189]

[0190]

[0191]

[0192]

[0193]

[0194]

[0195]

[0196]

[0197]

[0198]

[0199]

[0200]

[0201]

[0202]

[0203]

[0204]

[0205]

[0206]

[0207]

[0208]

[0209]

[0210]

[0211]

[0212]

[0213]

[0214]

[0215]

[0216]

[0217]

[0218]

[0219]

[0220]

[0221]

[0222]

[0223]

[0224]

[0225]

[0226]

[0227]

[0228]

[0229]

[0230]

[0231]

[0232]

[0233]

[0234]

[0235]

[0236]

[0237]

[0238]

[0239]

[0240]

[0241]

[0242]

[0243]

[0244]

[0245]

[0246]

[0247]

[0248]

[0249]

[0250] In one embodiment, the biomarker is MYC. In one embodiment, the measurable aspect of MYC is its expression status. In one embodiment, the biomarker is overexpression of MYC.

[0251] Accordingly, in certain aspects of the disclosure, the biomarker is MYC, which is differentially present in a subject in one phenotypic state, e.g., a patient having cancer, e.g., hepatocellular carcinoma (HCC), glioblastoma (GBM), lung cancer, breast cancer, head and neck cancer, prostate cancer, melanoma, or colorectal cancer, as compared to another phenotypic state, e.g., a normal, non-diseased subject or a patient having cancer but not overexpressing MYC.

[0252] Biomarker criteria can be predetermined, determined simultaneously, or determined after a biological sample is obtained from a subject. Biomarker criteria used with the methods described herein can, for example, include data from samples of subjects without cancer; data from samples of subjects with cancer, e.g., GBM, that is not progressive, recurrent, and / or metastatic cancer; data from samples of subjects with cancer, e.g., GBM, that is progressive, recurrent, and / or metastatic cancer. Comparisons can be made to establish predetermined threshold biomarker criteria for different categories of subjects, e.g., diseased versus non-diseased subjects. Criteria can be performed in the same assay, or can be known standards from a previous assay.

[0253] In one embodiment, the biomarker is MCL1. In one embodiment, the measurable aspect of MCL1 is its expression status. In one embodiment, the biomarker is overexpression of MCL1.

[0254] A biomarker is differentially present between groups of different phenotypic states if the average or median expression or mutation level of the biomarker is different (i.e., higher / lower) between the groups. Thus, a biomarker provides an indication of whether a subject, e.g., a cancer patient, belongs to one phenotypic state or another phenotypic state.

[0255] Accordingly, in certain aspects of the disclosure, the biomarker is MCL1, which is differentially present in a subject in one phenotypic state, e.g., a patient having cancer, e.g., hepatocellular carcinoma (HCC), glioblastoma (GBM), lung cancer, breast cancer, head and neck cancer, prostate cancer, melanoma, colorectal cancer, medulloblastoma, or generally brain tumor, as compared to another phenotypic state, e.g., a normal, non-diseased subject or a cancer patient not overexpressing MCL1.

[0256] In addition to a single biological compound, e.g., MYC or MCL1, the term "biomarker" as used herein is intended to encompass groups, panels, or arrays of multiple biological compounds. For example, a combination of MYC and MCL1 can comprise a biomarker. The term "biomarker" can include one, two, three, four, five, six, seven, eight, nine, ten, fifteen, twenty, twenty-five, thirty, or more biological compounds.

[0257] Any of a number of methods known in the art can be used to determine the expression level or mutation status of a biomarker in a patient. Any method known in the art for quantifying a particular protein and / or detecting MYC and / or MCL1 expression, or the expression or mutation level of any other biomarker in a patient or biological sample can be used in the methods of the present disclosure. Examples include, but are not limited to, PCR (polymerase chain reaction), or RT-PCR, Northern blot, Western blot, ELISA (enzyme-linked immunosorbent assay), RIA (radioimmunoassay), gene chip analysis of RNA expression, immunohistochemistry, or immunofluorescence. See, e.g., Slagle et al., Cancer 83:1401 (1998). Certain embodiments of the present disclosure include methods of assaying biomarker RNA expression (transcription). Other embodiments of the present disclosure include methods of determining protein expression in a biological sample. See, e.g., Harlow et al., Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y., (1988) and Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, New York, 3rd ed., (1995). For northern blot or RT-PCR analysis, RNA is isolated from tumor tissue samples using RNase-free techniques. Such techniques are known in the art.

[0258] In one embodiment of the present disclosure, a biological sample is obtained from a patient and cells in the biopsy are assayed to determine biomarker expression or mutation status.

[0259] In one embodiment of the present disclosure, PET imaging is used to determine biomarker expression.

[0260] In another embodiment of the disclosure, Northern blot analysis of the transcription of the biomarker in a tumor cell sample is performed. Northern analysis is a standard method for detecting and / or quantifying the level of mRNA in a sample. Initially, RNA is isolated from the sample to be tested using Northern blot analysis. In the analysis, the RNA sample is first separated by size by electrophoresis in an agarose gel under denaturing conditions. The RNA is then transferred to a membrane, cross-linked and hybridized with a labeled probe. Typically, Northern hybridization involves in vitro polymerization of a radiolabeled or non-isotopically labeled DNA, or the production of an oligonucleotide as a hybridization probe. Typically, the membrane holding the RNA sample is pre-hybridized or blocked prior to probe hybridization to prevent the probe from coating the membrane, thereby reducing non-specific background signal. Following hybridization, unhybridized probe is typically removed by washing in several changes of buffer. The stringency of the washing and hybridization conditions can be designed, selected and implemented by one of ordinary skill in the art. Detection is accomplished using a detectably labeled probe and appropriate detection methods. Radiolabeled and non-radiolabeled probes and their use are well known in the art. The presence and / or relative expression level of the biomarker determined can be quantified using, for example, densitometry.

[0261] In another embodiment of the disclosure, RT-PCR is used to determine biomarker expression and / or mutation status. RT-PCR allows for the real-time detection of the progress of PCR amplification of a target gene. The design of primers and probes required to detect the expression and / or mutation status of the biomarkers of the disclosure is within the skill of one of ordinary skill in the art. RT-PCR can be used to determine the level of RNA encoding the biomarkers of the disclosure in a tumor tissue sample. In one embodiment of the disclosure, RNA from a biological sample is isolated under RNase-free conditions and converted to DNA by treatment with reverse transcriptase. Methods for reverse transcriptase conversion of RNA to DNA are well known in the art. A description of PCR is provided in the following references: Mullis et al., Cold Spring Harbor Symp. Quant. Biol. 51 :263 (1986); EP 50,424; EP 84,796; EP 258,017; EP 237,362; EP 201,184; U.S. Patent Nos. 4,683,202; 4,582,788; 4,683,194.

[0262] RT-PCR probes depend on the 5'-3' nuclease activity of the DNA polymerase used for PCR to hydrolyze an oligonucleotide probe that hybridizes to the target amplicon (the biomarker gene). RT-PCR probes are oligonucleotides that have a fluorescent reporter dye attached to the 5' end and a quencher moiety coupled to the 3' end (or vice versa). These probes are designed to hybridize to an internal region of the PCR product. In the unhybridized state, the proximity of the fluor and quencher molecules prevents detection of the fluorescent signal of the probe. During PCR amplification, as the polymerase copies the template to which the RT-PCR probe is bound, the 5'-3' nuclease activity of the polymerase cleaves the probe. This separates the fluorescent and quencher dyes, and FRET no longer occurs. Thus, fluorescence increases in each cycle in a manner proportional to the amount of probe cleaved. The fluorescent signal emitted by the reaction can be measured or tracked over time using conventional and universal techniques using commercially available equipment.

[0263] In another embodiment of the disclosure, expression of the protein encoded by the biomarker is detected by western blot analysis. Western blotting (also known as immunoblotting) is a method for detecting proteins in a given tissue homogenate or extract sample. It separates denatured proteins by mass using gel electrophoresis. The proteins are then transferred from the gel to a membrane (such as nitrocellulose or polyvinylidene difluoride (PVDF)), where they are detected using a primary antibody that specifically binds the protein. The bound antibody can then be detected by a secondary antibody conjugated to a detectable label (e.g., biotin, horseradish peroxidase, or alkaline phosphatase). Detection of the secondary label signal indicates the presence of the protein.

[0264] In another embodiment of the disclosure, expression of the protein encoded by the biomarker is detected by enzyme-linked immunosorbent assay (ELISA). In one embodiment of the disclosure, a "sandwich ELISA" involves coating a plate with a capture antibody; adding a sample, where any antigen present binds to the capture antibody; adding a detection antibody that also binds the antigen; adding an enzyme-linked secondary antibody that binds to the detection antibody; and adding a substrate that is converted by the enzyme on the secondary antibody to a detectable form. Detection of a signal from the secondary antibody indicates the presence of the biomarker antigen protein.

[0265] In another embodiment of the disclosure, expression of the biomarker is assessed by using a gene chip or microarray. These techniques are within the ability of one of ordinary skill in the art.

[0266] VI. DEFINITIONS

[0267] The present disclosure provides various methods of treatment, kits, and pharmaceutical compositions comprising TG02. The term "TG02" as used herein refers to (16E)-14-methyl-20-oxa 5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8(27),9,11,16,21,23-decaene in any crystalline or amorphous form, or free base or pharmaceutically acceptable salt or solvate. In one embodiment, TG02 refers to the free base of (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8(27),9,11,16,21,23-decaene. In another embodiment, TG02 refers to a pharmaceutically acceptable salt of (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-1(25),2(26),3,5,8(27),9,11,16,21,23-decaene. The pharmaceutically acceptable salts of TG02 can be prepared either during the final isolation and purification of TG02, or separately by reacting TG02 with the appropriate acid. Examples of acids that can be employed to provide pharmaceutically acceptable salts include inorganic acids such as nitric, boric, hydrochloric, hydrobromic, sulfuric, and phosphoric acids, and organic acids such as oxalic, maleic, succinic, and citric acids. Non-limiting examples of salts of TG02 include, but are not limited to: hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate, 2-hydroxyethansulfonate, phosphate, hydrogen phosphate, acetate, adipate, alginate, aspartate, benzoate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, formate, succinate, fumarate, maleate, ascorbate, isethionate, salicylate, methanesulfonate, mesitylenesulfonate, naphthalenesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, picrate, pectinate, persulfate, 3-phenylpropionate, picrate, pivalate, propionate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, p-toluenesulfonate, undecanoate, lactate, citrate, tartrate, gluconate, methanesulfonate, ethanedisulfonate, benzenesulfonate, and p-toluenesulfonate.

[0268] In another embodiment, TG02 refers to the citrate salt of (16E)-14-methyl-20-oxa- 5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-l(25),2(26),3,5,8(27),9,11,16,21,23-decaene. This is referred to as TG02 citrate or (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa-l(25),2(26),3,5,8(27),9,11,16,21,23-decaene-citric acid.

[0269] The term "biological sample" as used herein refers to any tissue or fluid from a patient that is suitable for detecting biomarker, e.g., MYC and / or MCL1 expression status. Examples of useful biological samples include, but are not limited to, biopsy tissue and / or cells, e.g., solid tumors, lymph nodes, inflamed tissue, tissue and / or cells involved in a condition or disease, blood, plasma, serum, cerebrospinal fluid, saliva, urine, lymphatic fluid, cerebrospinal fluid, and the like. Other suitable biological samples will be familiar to those of ordinary skill in the relevant art. Biomarker expression and / or mutations of a biological sample can be analyzed using any technique known in the art, and the biological sample can be obtained using techniques well within the ordinary skill of a clinician. In one embodiment of the disclosure, the biological sample comprises blood cells.

[0270] The terms "a," "an," and "the" and similar referents (especially in the context of the claims) are to be construed to cover both the singular and the plural, unless otherwise indicated. The use of the term "or" in the context of this specification is to be construed as inclusive or open-ended, for example, as "and / or" unless otherwise indicated. The use of the term "at least one" will be understood to include one as well as one or more of a number of items, unless otherwise indicated. Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated in the specification as if it were individually recited herein. The use of any and all examples, or exemplary language (e.g., "such as") provided herein, is intended merely to better illuminate the application and does not pose a limitation on the scope of the application unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the application.

[0271] In this text the term "about" includes the recited number ± 10%. Thus, about "10" means 9-11.

[0272] As used herein, the terms "treat," "treatment," and the like, refer to eliminating, reducing, or ameliorating a disease or condition and / or its associated symptoms. While not excluding, treatment of a disease or condition does not require that the disease or condition or its associated symptoms be completely eliminated. In one embodiment, however, administration of TG02 and / or an immune checkpoint inhibitor and / or a COX-2 inhibitor results in complete remission of the cancer.

[0273] As used herein, the term "therapeutically effective amount" refers to an amount of a therapeutic agent sufficient to result in an improvement in one or more symptoms of a disorder, or prevent progression of a disorder, or cause regression of a disorder. For example, with respect to treatment of cancer, in one embodiment, a therapeutically effective amount refers to an amount of a therapeutic agent that results in a therapeutic response, such as normalization of blood cell counts, reduction in tumor growth rate, reduction in tumor mass, reduction in number of metastases, increase in time to tumor progression, and / or increase in patient survival time by at least about 2%, at least about 5%, at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, or at least about 100% or more.

[0274] The term "pharmaceutically acceptable carrier" or "pharmaceutically acceptable vehicle" includes any and all solvents, dispersion media, diluents, or other liquid vehicles into which the pharmaceutical composition is formulated. Suitable pharmaceutically acceptable carriers include aqueous vehicles and non-aqueous vehicles. Standard pharmaceutical carriers and their formulation are described in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 19th Edition, 1995.

[0275] The term "container" refers to any receptacle and closure suitable for storing, shipping, dispensing, and / or handling a pharmaceutical product.

[0276] The term "insert" refers to information accompanying a pharmaceutical product that describes how to administer the product, as well as the required safety and efficacy data to allow physicians, pharmacists, and patients to make informed decisions regarding use of the product. The package insert is often referred to as the "label" of the pharmaceutical product.

[0277] "Co-administration," "administration in combination," "administration concurrently," and similar phrases refer to the concurrent administration of two or more agents to a patient being treated. "Concurrent" means that each agent is administered at the same point in time or sequentially at different points in time in any order. However, if not administered at the same time, it means that each agent is administered to the patient sequentially and sufficiently close in time to provide the desired therapeutic effect and to be able to act in concert. For example, TG02 can be administered concurrently or sequentially in any order at different points in time as an immune checkpoint inhibitor and / or COX-2 inhibitor and / or optional therapeutic agent. TG02 and the immune checkpoint inhibitor and / or COX-2 inhibitor and / or optional therapeutic agent can be administered separately in any appropriate form and by any suitable route. When TG02 and the immune checkpoint inhibitor and / or COX-2 inhibitor and / or optional therapeutic agent are not administered at the same time, it is understood that they can be administered to a patient in need in any order. For example, TG02 can be administered prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks prior), concurrently with, or after (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) the administration of the immune checkpoint inhibitor and / or COX-2 inhibitor to an individual in need thereof. In embodiments, the administration of TG02 and the immune checkpoint inhibitor and / or COX-2 inhibitor are separated by 1 minute, 10 minutes, 30 minutes, less than 1 hour, 1 hour, 1-2 hours, 2-3 hours, 3-4 hours, 4-5 hours, 5-6 hours, 6-7 hours, 7-8 hours, 8-9 hours, 9-10 hours, 10-11 hours, 11-12 hours, no more than 24 hours, or no more than 48 hours. In one embodiment, the administration of the components of the combination therapy are separated by about 1 minute to about 24 hours. In one embodiment, TG02 is administered 3-7 days prior to the day of the immune checkpoint inhibitor. In another embodiment, TG02 is also administered on the day of the immune checkpoint inhibitor administration and continues to be administered until disease progression or TG02 treatment is no longer beneficial. Examples

[0278] Example 1

[0279] This study is being conducted to compare the progression-free or overall survival rates of cancer participants using pembrolizumab (p) or nivolumab (n) versus p or n in combination with TG02, who are treatment naive or have progressed after prior therapy and have been selected for overexpression of MYC and / or MCL1 status. Participants will be randomized to receive standard anti-PD-1 therapy plus placebo or standard anti-PD-1 therapy plus TG02.

[0280] Primary outcome measures: progression-free survival (PFS) and / or overall survival (OS)

[0281] Secondary outcome measures: overall response rate (ORR) and / or duration of response

[0282] Eligibility

[0283] Age eligible for study: 18 years of age and older

[0284] For medulloblastoma patients - 6 months or older

[0285] Gender eligible for study: Both

[0286] Inclusion criteria:

[0287] Histologically or cytologically confirmed diagnosis of cancer that is not amenable to local treatment

[0288] Must have signed informed consent for the related study; must provide a newly obtained tissue / biopsy specimen (or specimen obtained within 60 days of consent)

[0289] Radiographically measurable disease

[0290] Eastern Cooperative Oncology Group performance status of 0 or 1

[0291] Patient has disease that overexpresses MYC and / or MCL1

[0292] Exclusion criteria:

[0293] Chemotherapy, radiation therapy, or biologic therapy within four weeks prior to first dose of study drug, or has not recovered from AEs due to cancer therapy administered more than four weeks prior

[0294] Participation or has participated in a study of an investigational agent or use of an investigational device within 30 days prior to first dose of study drug

[0295] During the study, any other form of systemic or local anti-neoplastic therapy is anticipated to be required

[0296] Chronic systemic steroid therapy within two weeks prior to the planned date of first dose of randomization therapy or any other form of immunosuppressive medication

[0297] Known history of malignancy other than adequately treated basal or squamous cell skin cancer, superficial bladder cancer, carcinoma in situ of the cervix, breast cancer, or other cancer in situ

[0298] Known active central nervous system (CNS) metastases and / or carcinomatous meningitis

[0299] Known history of active autoimmune disease or history of autoimmune disease or syndrome requiring systemic steroids or immunosuppressive agents

[0300] Prior treatment with any other anti-programmed cell death (PD) agent

[0301] Active infection requiring systemic therapy

[0302] Known history of human immunodeficiency virus (HIV)

[0303] Active hepatitis B or hepatitis C

[0304] Current use of illicit drugs (including use for recreational purposes) or recent (within the last year) substance abuse (including alcohol)

[0305] Pregnant or breastfeeding, or expecting to become pregnant or conceive a child during the projected duration of the study.

[0306] Protocol:

[0307] The first group of patients is given 2-10 mg / kg pembrolizumab (or flat dose equivalent) by intravenous infusion every three weeks and 100, 200, or 300 mg of TG02 orally once daily until disease progression or until TG02 is no longer beneficial. TG02 administration is initiated 3-7 days prior to the start of pembrolizumab treatment, continued on the day of pembrolizumab administration, and continued until disease progression or until TG02 is no longer beneficial. The control patients are given 2-10 mg / kg pembrolizumab (or flat dose equivalent) by intravenous infusion every three weeks.

[0308] The second group of patients receives 3 mg / kg nivolumab by intravenous infusion over 60 minutes every 2 weeks and 100, 200, or 300 mg of TG02 orally once daily. TG02 administration is initiated 3-7 days prior to the start of nivolumab treatment, continued on the day of nivolumab administration, and continued until disease progression or until TG02 is no longer beneficial. The control patients receive 3 mg / kg nivolumab by intravenous infusion over 60 minutes every 2 weeks.

[0309] Results:

[0310] In patients with tumors overexpressing MYC and / or MCL1, TG02 in combination with pemirozilab or nimozilab produced superior antitumor clinical activity compared to immune checkpoint inhibitors alone. Unexpected objective responses associated with lack of tumor progression and prolonged long-term survival were obtained compared to historical controls using only (antibody). In one implementation, patients receiving TG02 and an immune checkpoint inhibitor achieved a progression-free survival (or time to progression) extension of at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months. In another implementation, at least some patients receiving TG02 and an immune checkpoint inhibitor achieved a response duration extension of at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months.

[0311] Example 2

[0312] An open-label phase 2 study evaluated the combination of checkpoint blockade immunotherapy and TG02 in patients who had relapsed or were refractory to standard anti-PD-1 therapy (p) or (n).

[0313] Level 1 endpoint: ORR

[0314] Secondary endpoints: PFS, OS, response time, security

[0315] Inclusion criteria:

[0316] Histologically confirmed cancer diagnosis is not suitable for local treatment.

[0317] The Eastern Cooperative Oncology Group (ECOG) performance status is 0 or 1.

[0318] At least one measurable lesion

[0319] Sufficient organ function

[0320] Previous treatment with anti-PD-1 or anti-PD-L1 antibodies

[0321] The patient has a disease that overexpresses MYC and / or MCL1.

[0322] Exclusion criteria:

[0323] Chemotherapy, targeted small molecule therapy, radiation therapy, or biological cancer therapy (including monoclonal antibodies), within 4 weeks prior to the first dose of trial treatment, or failure to recover from an adverse event caused by a previously administered drug (<= Grade 1 or baseline).

[0324] During the study, any other form of systemic or local anti-tumor therapy is expected to be needed.

[0325] Known active central nervous system (CNS) metastases and / or carcinomatous meningitis.

[0326] History of clinically significant autoimmune disease, or syndromes requiring systemic steroids or immunosuppressive agents.

[0327] Received systemic steroid treatment or any other form of immunosuppressive treatment within 1 week prior to first dose of study treatment.

[0328] Vaccinated with live vaccines within 4 weeks prior to first dose of trial treatment.

[0329] History or evidence of active pneumonia.

[0330] Human immunodeficiency virus (HIV) positive.

[0331] Active hepatitis B or hepatitis C.

[0332] Pregnant, breastfeeding, or expecting to conceive or become pregnant within the projected duration of the trial treatment up to 120 days after the last dose of study drug.

[0333] Dosing regimen:

[0334] Table 1 TG02 + checkpoint inhibitor combination dosing and regimen

[0335]

[0336]

[0337] TG02 dosing (50-400 mg) weekly, starting at least 5 days prior to initiation of checkpoint inhibitor treatment, continuing until disease progression or investigator decision outcome

[0338] Combining TG02 with at least one checkpoint inhibitor in patients with tumors that overexpress MYC and / or MCL1 can reverse immune evasion and induce clinically relevant responses in patients who were previously non-responders or non- treated by checkpoint inhibitor therapy or in patients with de novo cancers. Unexpected objective responses associated with lack of tumor progression and prolonged long-term survival extension are obtained compared to historical controls using (antibodies) alone. In one embodiment, patients receiving TG02 and at least one immune checkpoint inhibitor achieve a response duration (or progression-free survival) extension of at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months. In another embodiment, at least some patients receiving TG02 and at least one immune checkpoint inhibitor achieve a response duration extension of at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months.

[0339] Example 3

[0340] Randomized phase 2 placebo-controlled study of pembrolizumab + TG02 vs. pembrolizumab + placebo in previously treated participants with locally advanced unresectable or metastatic colorectal cancer exhibiting MYC and / or MCL1 overexpression status.

[0341] Primary endpoint: PFS

[0342] Secondary endpoints: ORR, duration of response

[0343] Inclusion criteria:

[0344] Histologically confirmed locally advanced unresectable or metastatic high-grade colorectal cancer

[0345] Previously treated with at least two lines of approved standard therapy, must include fluoropyrimidine, oxaliplatin, irinotecan, bevacizumab and cetuximab or panitumumab

[0346] Eastern Cooperative Oncology Group performance status of 0 or 1

[0347] Patient has disease overexpressing MYC and / or MCL1

[0348] Expected life span of more than 3 months

[0349] At least one measurable lesion

[0350] Fertile female participants should be willing to use 2 methods of birth control or surgical sterilization or avoid heterosexual sexual activity from the time of informed consent through 120 days after the last dose of study drug

[0351] Male participants should agree to use an appropriate method of contraception from the first dose of study treatment until 120 days after the last dose of study drug

[0352] Sufficient organ function

[0353] Exclusion criteria:

[0354] Currently participating in another study and receiving investigational treatment, participating in a study of a study drug, and receiving investigational treatment within 4 weeks of the first dose of study drug in this study, or using study equipment within 4 weeks of the first dose of study drug in this study

[0355] Active autoimmune disease requiring systemic treatment within the past 2 years

[0356] Diagnosis of immunodeficiency or receiving systemic steroid therapy or any other form of immunosuppressive therapy within 7 days prior to the first dose of study drug treatment

[0357] Known active central nervous system (CNS) metastasis and / or carcinomatous meningitis

[0358] Prior monoclonal antibody (mAb), chemotherapy, targeted small molecule therapy or radiation therapy within 2 weeks prior to study Day 1 or not recovered (i.e., < Grade 1 or baseline) from adverse events due to prior administered drugs

[0359] Prior use of an anti-Programmed Death (PD)-l, anti-PD-Ll or anti-PD-L2 drug therapy or the participant has previously participated in a Merck pembrolizumab (MK-3475) clinical trial

[0360] Known other malignancy that is progressing or requires active treatment, with the exception of a skin basal cell carcinoma or a skin squamous cell carcinoma that has undergone potentially curative therapy or a cervical carcinoma in situ

[0361] Vaccination with live vaccines within 30 days prior to the planned start of study drug

[0362] Known history of human immunodeficiency virus (HIV)

[0363] Known active hepatitis B or hepatitis C

[0364] Known history or evidence of any interstitial lung disease or active non-infectious pneumonitis

[0365] Active infection requiring systemic treatment

[0366] Known psychiatric disorder or substance abuse disorder that would interfere with cooperation with trial requirements

[0367] Pregnancy or breastfeeding, or the desire to become pregnant or to breastfeed, from the start of the screening visit until 120 days after the last dose of study drug

[0368] Dosing regimen:

[0369] Patients receive 2-10 mg / kg pembrolizumab intravenously every 3 weeks and 1, 2 or 3 mg / kg of TG02 orally 3-7 days prior to pembrolizumab administration and on the day of pembrolizumab administration, continuing thereafter until disease progression or until it is no longer beneficial. Control patients receive 2 mg / kg pembrolizumab by intravenous infusion every three weeks.

[0370] Results:

[0371] TG02 in combination with pembroziizumab provides better clinical activity in the same patients than pembroziizumab alone when used in patients with tumors that overexpress MYC and / or MCL1. Unexpected objective responses associated with lack of tumor progression and long-term survival prolongation were obtained in patients compared to historical controls using (antibodies) alone. In one embodiment, patients receiving TG02 and pembroziizumab achieve at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months prolongation of time to progression (or progression free survival). In another embodiment, at least some patients receiving TG02 and pembroziizumab achieve at least 2 months, at least 4 months, at least 6 months, at least 8 months, at least 10 months, or at least 12 months prolongation of duration of response.

[0372] Example 4

[0373] TG02 in combination with carfilzomib for carfilzomib (CFZ) refractory multiple myeloma (MM) patients

[0374] Methods

[0375] An open-label phase lb study enrolled MM patients who had previously received >2 lines of therapy. The primary objective was to determine the maximum tolerated dose (MTD) of TG02 in combination with carfilzomib (TG02 / CFZ). Secondary objectives included anti-tumor activity and safety. TG02 was administered once daily on days 1, 4, 8, 11, 15, 18 of a 28-day schedule (BIW). The starting dose of TG02 was 150 mg. TG02 was escalated in 50 mg increments to 300 mg. CFZ was administered according to the prescribing information. Response was assessed using standard criteria.

[0376] Results

[0377] Fourteen patients were enrolled for dose escalation and ten patients were enrolled for expansion at the MTD. Patients received a high number of prior treatments: median of 6 prior treatments [min 3; max 15] and 92% of patients received CFZ in a prior treatment regimen. The best response to prior treatments was progressive disease in 46% of patients. The MTD was 250 mg TG02 in combination with CFZ. Two dose-limiting toxicities were observed in the 300 mg cohort (including Grade 4 (Gr) sepsis and Gr 4 neutropenia). The most common drug-related adverse events (AEs) were diarrhea (Gr 1-2: 71%, Gr 3: 17%), nausea (Gr 1-2: 79%), vomiting (Gr 1-2: 50%), fatigue (Gr 1-2: 38%, Gr 3: 4%), anorexia (Gr 1: 21%), anemia (Gr 1-2: 4%, Gr 3: 17%), and thrombocytopenia (Gr 3: 8%, Gr 4: 13%). Six patients (25%) discontinued treatment due to AEs. Serious AEs occurred in 50% of patients; only acute renal failure and febrile neutropenia occurred in more than one patient (8% each). The severity of AEs was similar to TG02 as a single agent. The incidence of diarrhea was increased with TG02 / CFZ dosing (88% vs 67%), but the incidence of other AEs was similar to TG02 as a single agent. Responses were evaluable in 14 patients who received TG02 at the MTD. The overall response rate (≥PR) was 27%; the clinical benefit rate (≥MR) was 45% (1 very good partial response, 2 partial responses, and 2 minimal responses). All responders (MR or better) were CFZ-refractory in a prior treatment regimen. Durable stable disease was observed in 27% of patients.

[0378] CONCLUSIONS

[0379] The safety profile of TG02 BIW / CFZ was similar to TG02 alone. The most common drug-related AEs were diarrhea, nausea, and vomiting; grade 4 AEs were uncommon. Objective responses were observed in CFZ-refractory patients.

[0380] Example 5

[0381] TG02 activity in glioma cell and allograft models

[0382] Several standard glioblastoma multiforme (GBM) cell lines and one O6-methylguanine DNA methyltransferase (MGMT)-expressing stem cell line were treated with TG02, temozolomide (TMZ), or a combination of TG02 and TMZ in a 72-hour cell proliferation assay. 6 - a stem cell line expressing the methylguanine DNA methyltransferase (MGMT). See Figures 1-3 TG02, TMZ, and TG02 + TMZ combinations were also tested in a cell line that does not express MGMT. SeeFigures 4-7 Cells were seeded onto 12-well plates and treated with 50 nM TG02, 100 μΜ TMZ, or TG02 + TMZ for 72 hours. Cell viability was determined by cell counting.

[0383] The cytotoxic effects of TG02, TMZ, and TG02 in combination with TMZ were examined by colony formation assays in GSC923 and U251 cells. See Figure 8 and 9 .

[0384] Pulmonary artery endothelial cells and human astrocytes were tested with 50 nM TG02, 100 μΜ TMZ, or TG02 + TMZ for 72 hours. Cells were then changed to normal culture media and cultured for an additional 7 days. Cell viability was determined by cell counting. See Figure 10 and 11 .

[0385] GSC923 (Figures Figure 12 and 13 ) and U251 (Figures Figure 14 and 15 ) cells were exposed to various concentrations of TG02, TMZ, and TG02 + TMZ for 72 hours and cell viability was examined by cell counting. The synergistic effect of TG02 + TMZ was determined by the combination index (CI). CI values were calculated by COMPUSYN software and are shown in Table 2 for GSC923 cells and Table 3 for U251 cells. CI < 1 is synergistic, CI = 1 is additive, and CI > 1 is antagonism of the two compounds in combination.

[0386] Table 2

[0387] Figure 16 Figure 17 0.25 0.077 0.5 0.136 0.75 0.242 0.90 0.430

[0388] Table 3

[0389] Figure 18 Figure 19 0.25 0.029 0.5 0.104 0.75 0.376 0.90 1.333

[0390] Figure 20 is a schematic of drug administration in a mouse glioma GL261 cell allograft model. Mouse glioma GL261 cells were stereotactically injected into the striatum of female C57BL / 6 albino mice (n = 5-7 per group) and then treated with vehicle, TG02, TMZ, and TG02 + TMZ combination. Median overall survival was observed to be 24, 24.5, 27.5, and 32 days, respectively. See Figure 21 Results were analyzed using the log-rank test with a trend in GraphPad Prism software (Chi-square = 9.063, df = 1, P value = 0.0026**). Survival was determined by using PerkinElmer Spectrum bioluminescence imaging (BLI) determined tumor burden. The intensity of BLI was calculated and normalized to the initial intensity at day 5. See Figure 22 .

[0391] Example 6

[0392] TG02 activity in hepatocellular carcinoma (HCC) cells and xenograft models

[0393] The effect of TG02 on MYC expression in HCC cell lines was investigated. Five HCC cell lines with varying MYC expression levels were treated with 0.5 μΜ TG02 for 24 hours and MYC expression was assessed by western blot. See Figures 23-26 MYC expression levels were reduced by TG02 treatment in HepG2, SNU398 and HUH-1 cell lines, but not in the JHH-5 cell line. Hep3B cells do not express MYC.

[0394] Eight HCC cell lines with high or low MYC expression were then treated with TG02 in vitro. TG02 treatment resulted in inhibition of cell proliferation in all HCC cell lines tested. TG02 was selectively more potent in cell lines with high levels of MYC expression compared to those with low levels of MYC expression, with mean IC 50 values of 524 nM and 84 nM, respectively. See Figure 23 .

[0395] Inhibition of MYC expression was also measured in vivo. HepG2 hepatocellular carcinoma xenografts were grown orthotopically in Balb / c nude mice. TG02 or vehicle was given orally at 50 mg / kg to 5 mice; tumors were collected 8 hours after treatment and MYC protein expression levels were measured by western blot. MYC expression was observed in each of the control tumors. In the TG02 treated group, MYC expression levels were reduced in 4 of 5 animals, MYC was essentially depleted in 2 animals and partially reduced in the other 2 animals. See Figure 25 .

[0396] TG02 was evaluated as a single agent or in combination with sorafenib for therapeutic efficacy in treating the orthotopic HepG2 human hepatocarcinoma xenograft model in BALB / c nude mice. Mice were randomized into treatment groups based on baseline serum AFP levels that track tumor volume in the liver on day 19 post inoculation. TG02 was given orally at 50 mg / kg twice weekly and was reduced to 40 mg / kg. Sorafenib was given orally at 15 mg / kg daily. TG02 as a single agent had a modest effect on tumor volume. TG02 in combination with sorafenib resulted in significant anti-tumor activity. See Figure 27 .

[0397] Example 7

[0398] TG02-mediated CDK9 inhibition

[0399] Tet-off transgenic mouse model of MYC-induced T-cell acute lymphoblastic leukemia (MYC T-ALL) that overexpresses and is dependent on MYC was treated with 100 or 500 mM of TG02.

[0400] MYC is a transcription factor that regulates the expression of multiple gene products involved in cell proliferation, growth, differentiation, and apoptosis. The MYC gene is genetically activated and overexpressed in many human cancers, and this overexpression is causally related to tumorigenesis, leading to malignant growth and immune evasion. See, e.g., Fernandez et al., Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res. 19: 2677-2687 (2013); Carter et al., Blood 105: 4043-4050 (2005); Casey et al., Science 352: 227-231 (2016); Hannah, A. L., Curr. Mol. Med. 5: 625-642 (2005); and Parcells et al., Stem Cells Dayt. Ohio 24: 1174-1184 (2006).

[0401] As shown in Figure 28 , PD-L1 expression was detected by RT-PCR when MYC was “on” (t = 0 hours). But when MYC was “turned off,” both CD47 and PD-L1 expression were significantly reduced in a time-dependent manner. TG02 caused a time- and dose-dependent downregulation of PD-L1 and CD47 mRNA expression in MYC T-ALL cells (by Figure 29 and 24 ), and both BCL-xL and MYC expression were downregulated (by Figure 30 and 26 ), respectively). Reduction of CD47 and PD-L1 expression on tumor cells can lead to reduced immune evasion and increased tumor cell death.

[0402] Example 8

[0403] TG02 in combination with anti-PD-1

[0404] TG02 and PD-1 mAb (anti-PD-1) were tested in combination in an orthotopic GL261 glioma model. GL261 allografts were established for 3 days, and then mice were randomized into 6 treatment groups (n = 8) based on bioluminescent tumor volume.

[0405] Mice were treated with vehicle, TG02 alone (20 or 40 mg / kg), PD-1 mAb alone (500 μg), and the combination of TG02 and anti-PD-1. Median survival times for the mice were 27.5, 26.5, 33, 32, 78, and over 95 days, respectively Figure 31

[0406] Mice treated with TG02 alone at 40 mg / kg (0.009), PD-1 mAb alone (0.003), TG02 20 mg / kg + PD-1 mAb (0.0001), and TG02 40 mg / kg + PD-1 mAb (0.0001) had a significant survival benefit compared to the vehicle group.

[0407] Example 9

[0408] TG02 induces cell death and synergizes with radiation in MYC-driven glioblastoma

[0409] The relationship between TG02 anti-tumor activity and MYC expression was tested in a panel of patient-derived GBM cell lines (PDCLs). IC 50 TG02 inhibited 6 of 12 PDCLs at less than 0.2 μΜ. See Table 4.

[0410] Table 4

[0411]

[0412] Downregulation of MYC and Mcl-1 was observed as early as 6 hours in the MYC-amplified BT245 cell line, and complete downregulation was observed at 24 hours, consistent with a significant increase in apoptosis Figure 32 The AUC of TG02-induced cell viability inhibition was calculated in this cell line panel to correlate with MYC expression levels Figure 33 TG02 was a more potent inhibitor of PDCLs, exhibiting high MYC expression ​ In vitro sensitivity (AUC) was negatively correlated with MYC expression in GBM cells (P value = 0.02)

[0413] Example 10

[0414] Radiation is an effective treatment for glioblastoma. However, tumor resistance and recurrence occurs in all patients.

[0415] A panel of GBM PDCLs, see Example 9, was selected for evaluation of the combination of TG02 and radiotherapy for the treatment of glioblastoma ​ ​). Cells were first treated with increasing concentrations of TG02. Cells were treated with increasing doses of radiation over 30 minutes and cell proliferation was measured 72 hours after treatment. TG02 alone had anti-proliferative activity in these cell lines. The addition of TG02 enhanced the effect of radiation in a synergistic manner. The combination of TG02 and radiation exceeded the Bliss predicted model (more than 10% change compared to Bliss predicted model), demonstrating synergy between TG02 and radiation in multiple PDCLs.

[0416] Example 11

[0417] TG02 activity correlates with MYC expression in glioblastoma cell lines

[0418] TG02 activity was evaluated in a panel of 26 patient-derived GBM stem cell lines for activity against GBM stem cell proliferation ​ ). TG02 was effective in this panel, with 16 cell lines achieving IC 50 values of less than 250 nM.

[0419] Expression levels of CDK9 and downstream markers, including MYC and Mcl-1, were measured to explore whether there was a correlation between protein expression and IC 50 values in this GBM panel. High MYC expression was found to correlate with higher sensitivity to TG02 treatment ​

[0420] Having now fully described the methods, compounds and compositions of the present application, it will be apparent to one of ordinary skill in the art that numerous modifications, substitutions and alterations to the methodologies, compounds and compositions of the present application can be made without departing from the scope of the methods, compounds and compositions of the present application or any embodiments thereof. All patents, patent applications and publications cited herein are fully incorporated by reference herein in their entirety.​

Claims

1. Use of TG02 in the manufacture of a medicament for treating a patient with cancer, wherein TG02 is (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa- 1(25),2(26),3,5,8(27),9,11,16,21,23-decaene or a pharmaceutically acceptable salt thereof, wherein TG02 is administered to the patient with a therapeutically effective amount of radiation therapy, and wherein the cancer is a glioma.

2. The use of claim 1, wherein the TG02 is a citrate salt of (16E)-14-methyl-20-oxa-5,7,14,26-tetraazatetracyclo[19.3.1.1(2,6).1(8,12)]heptacosa- 1(25),2(26),3,5,8(27),9,11,16,21,23-decaene.

Images