Treatment of cancer with the pdk inhibitor 4-chloro-n-(2-(4-chlorobenzyl)-3-oxo-2,3-dihydro-1,2,4-thiadiazol-5-yl)benzamide

Abstract

Provided herein are methods of treating cancer by administering a treatment compound to a subject in need thereof. The present disclosure also relates generally to methods of inhibiting the activity of a pyruvate dehydrogenase kinase (PDK) using the treatment compound to a subject in need thereof. The treatment compound is a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or a prodrug thereof.

Description

METHODS OF TREATING CANCER WITH A PYRUVATE DEHYDROGENASE KINASE INHIBITOR CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Patent Application No. 63 / 517,525, filed August 3, 2023, which is incorporated herein by reference in its entirety. REFERENCE TO AN ELECTRONIC SEQUENCE LISTING

[0002] The contents of the electronic sequence listing (286502001040SEQLIST.xml; Size: 43,276 bytes; and Date of Creation: July 24, 2024) is herein incorporated by reference in its entirety. FIELD

[0003] The present disclosure relates generally to methods of treating, preventing, or delaying the onset of cancer by administering a compound provided herein. The present disclosure also provides combination therapies that can be used to treat cancer. BACKGROUND

[0004] A number of human cancers enact a metabolic shift to glycolysis and away from oxidative phosphorylation via increased PDK expression / activity. This phenomenon, often referred to as the Warburg Effect, has been shown to protect cancer cells from apoptosis and to increase cell proliferation. See Atas E., Oberhuber M., and Kenner L., The Implications of PDK1–4 on Tumor Energy Metabolism, Aggressiveness and Therapy Resistance. Front Oncol, 2020.10: p.583217.

[0005] Current ongoing research is designed to develop pharmacological intervention to counter the Warburg Effect and restore metabolic flexibility in cancer cells. Expression and increased activity of pyruvate dehydrogenase kinases (PDKs) results in increased cell proliferation, increased invasiveness, and increased resistance to chemotherapeutic drugs in certain types of cancer. See Wang J, Qian Y, Gao M. Overexpression of PDK4 is associated with cell proliferation, drug resistance and poor prognosis in ovarian cancer. Cancer Manag Res. 2019;11:251-262. Inhibition of PDK by various mechanisms can potentially reverse theseeffects, as has been shown in several studies. See Tiersma, J.F.; Evers, B.; Bakker, B.M.; Jalving, M.; de Jong, S. Pyruvate Dehydrogenase Kinase Inhibition by Dichloroacetate in Melanoma Cells Unveils Metabolic Vulnerabilities. Int. J. Mol. Sci.2022, 23, 3745 and Kim CJ, Terado T, Tambe Y, Mukaisho K, Kageyama S, Kawauchi A and Inoue H: Cryptotanshinone, a novel PDK 4 inhibitor, suppresses bladder cancer cell invasiveness via the mTOR / ȕ-catenin / N-cadherin axis. Int J Oncol 2021, 59: 40. Nonetheless, highly potent PDK inhibitors that show favorable toxicological profiles have not yet been developed.

[0006] Accordingly, there exists a strong need to develop new PDK inhibitors as anti-cancer agents. BRIEF SUMMARY

[0007] It has been surprisingly discovered that the compound of formula (I)or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is capable of restoring metabolic flexibility in muscle tissue and cells of a subject in need thereof that suffers from various diseases or complications, particularly cancer. The compound of formula (I) is also referred to as 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. In some variations, the subject is a human (e.g., a human patient). Restoration of metabolic flexibility is attributed, in part, to the discovery that the compound of formula (I) is an inhibitor of the expression of pyruvate dehydrogenase kinases (PDKs) and is capable of increasing tumor oxygenation and oxidative phosphorylation following administration to a subject in need thereof (e.g., a cancer patient), which can result in reduced proliferation and / or increased apoptosis in certain cancer cells.

[0008] In some aspects, provided herein are methods and compositions for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof, comprising administering a compound of formula (I) or apharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the PDK is pyruvate dehydrogenase kinase 4 (PDK4).

[0009] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat bladder cancer. In some embodiments, the bladder cancer is metastatic bladder cancer.

[0010] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat colon cancer. In some such embodiments, the colon cancer is associated with a mutation in KRAS. In some embodiments, the colon cancer is metastatic colon cancer.

[0011] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat lung cancer. In some such embodiments, the lung cancer is associated with a mutation in KRAS. In other such embodiments, the lung cancer is epidermal growth factor receptor (EGFR)-positive lung cancer. In some embodiments, the lung cancer is metastatic lung cancer. In some embodiments, the lung cancer is non-small cell lung cancer (NSCLC). In some embodiments, the NSCLC is associated with mutations in EGFR.

[0012] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat breast cancer. In some such embodiments, the breast cancer is tamoxifen-resistant breast cancer. In some embodiments, the breast cancer is metastatic breast cancer. In some embodiments, the breast cancer is triple negative breast cancer.

[0013] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat bladder cancer. In some embodiments, the bladder cancer is metastatic bladder cancer.

[0014] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat melanoma. In some embodiments, the melanoma is metastatic melanoma. In some embodiments, the metastatic melanoma in the subject (e.g., patient) had been treated previously with an immunotherapy (e.g., an anti-PD1 or anti-CTLA4antibody). In some such embodiments, the metastatic melanoma had not responded to or had limited response to the immunotherapy.

[0015] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat hepatocellular carcinoma (HCC).

[0016] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat cervical cancer. In some embodiments, the cervical cancer of the subject (e.g., patient) has become resistant to doxorubicin. In some embodiments, the cervical cancer of the subject (e.g., patient) has become resistant to etoposide.

[0017] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to target cancer stem cells. In some such embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to target cancer stem cells that are resistant to radiation therapy (radiotherapy).

[0018] In another aspect, also provided herein is a method of inducing apoptosis in apoptosis-inducible cancer cells comprising administering to said apoptosis-inducible cancer cells an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments the cancer cells are bladder cancer cells, gastric cancer cells, colon cancer cells, breast cancer cells, ovarian cancer cells, melanoma cells, or lung cancer cells.

[0019] In another aspect, compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is administered to a subject (e.g., patient) that has a cancer that is resistant to one or more chemotherapeutic agents and / or is resistant to radiation therapy (radiotherapy).

[0020] In another aspect, the disclosure provides methods of reducing drug resistance to a chemotherapeutic agent, said method comprising administering to a subject (e.g., cancer patient) a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the chemotherapeutic agent is cisplatin. In some embodiments, the chemotherapeutic agent is 5-flurouracil. In some embodiments, the chemotherapeutic agent iserlotinib. In some embodiments, the chemotherapeutic agent is paclitaxel. In some embodiments, the chemotherapeutic agent is doxorubicin.

[0021] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with cisplatin to treat a subject in need thereof, e.g., a patient with cancer. In some embodiments, the cancer is bladder cancer, gastric cancer, breast cancer, lung cancer, HCC, cervical cancer, ovarian cancer or colon cancer.

[0022] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with an EGFR inhibitor to treat lung cancer (e.g., NSCLC) in a subject in need thereof (e.g., a patient). In some embodiments the EGFR inhibitor is erlotinib, gefitinib, lapatinib, lazertinib or necitumumab.

[0023] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with paclitaxel to treat lung cancer (e.g., NSCLC) in a subject in need thereof (e.g., a patient).

[0024] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination doxorubicin to treat hepatocellular carcinoma (HCC) or cervical cancer in a subject in need thereof (e.g., a patient).

[0025] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with a recombinant arginase. In some such embodiments, the cancer is breast cancer (e.g., triple negative breast cancer). In other such embodiments, the cancer is HCC. In yet other embodiments, the cancer is cervical cancer.

[0026] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with a Cox2 inhibitor, propranolol, metformin, salinomycin, or phenformin.

[0027] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is a PD-1 inhibitor (e.g., cemipilimab, nivolumab or pembrolizumab). In some embodiments, the immune checkpointinhibitor is a PD-L1 inhibitor (e.g., atezolizumab, avelumab, or durvalumab). In some embodiments, the immune checkpoint inhibitor is a CTLA-4 inhibitor (e.g., ipilimumab or tremelimumab).

[0028] In another aspect, the disclosure provides methods of overcoming tumor radioresistance by administering to a subject suffering from cancer (e.g., cancer patient) a pharmaceutically effect amount of a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered prior to the subject receiving radiotherapy.

[0029] In any of the preceding embodiments, the compound of formula (I) can be administered as a salt. In some embodiments, the salt is an alkali metal salt. In some embodiments, the salt is a sodium salt. In some embodiments, the alkali metal salts of 4-chloro- N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide are of the formula:, wherein X+represents the alkali metal (e.g. lithium, rubidium, cesium, sodium, or potassium) cation.

[0030] In some embodiments, the compound administered in accordance with the disclosure is a prodrug of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. In some embodiments, the prodrug is has the following structure:, or a salt thereof, wherein R1is selected from the group consisting of -C(O)-C2H4-CO2H and - PO3H2, or a salt or solvate thereof.

[0031] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject (e.g., patient) in need of at a dose of from about 100 mg to about 1,000 mg. All references to dosage amounts discussed herein refer to the free acid (protonated) form. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 200 mg to about 1,000 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 400 mg to about 800 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 100 mg to about 300 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg or about 500 mg.

[0032] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need of (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 40 ^g / mL to about 200 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 40 ^g / mL to about 80 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein theadministration results in a steady state blood plasma concentration of the compound of formula (I) of from about 70 ^g / mL to about 120 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 90 ^g / mL to about 160 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need of (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 100 ^g / mL to about 150 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof s administered daily to the human subject and results in a steady state blood plasma concentration of the compound of formula (I) of from about 120 ^g / mL to about 140 ^g / mL.

[0033] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need of (e.g., patient), wherein the administration results in a steady state area under the time curve (AUC0-24) of the compound of formula (I) of from about 1,000 h*^g / mL to about 4,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject or a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24of the compound of formula (I) of from about 1,000 h*^g / mL to about 2,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 1,500 h*^g / mL to about 4,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24of the compound of formula (I) of from about 1,800 h*^g / mL to about 3,100 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results ina steady state AUC0-24of the compound of formula (I) of from about 1,500 h*^g / mL to about 2,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 2,500 h*^g / mL to about 4,000 h*^g / mL In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24of the compound of formula (I) of from about 3.000 h*^g / mL to about 3,500 h*^g / mL.

[0034] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 40 ^g / mL to about 200 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmax of the compound of formula (I) of from about 70 ^g / mL to about 200 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 80 ^g / mL to about 140 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmax of the compound of formula (I) of from about 70 ^g / mL to about 100 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 1200 ^g / mL to about 150 ^g / mL.

[0035] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in an oral dosage form. In some embodiments, the oral dosage form is a capsule. In other embodiments, the oral dosage form is a tablet.

[0036] In other aspects, also provided is the use of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in a therapy, including any of the methods described herein. For instance, in some embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof; for reducing drug resistance to a chemotherapeutic agent; for inducing apoptosis in apoptosis-inducible cancer cells.

[0037] In certain aspects, also provided is the use of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for the manufacture of a medicament. In some embodiments, the medicament is for any of the methods described herein. For instance, in some embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in the manufacture of a medicament for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof; for reducing drug resistance to a chemotherapeutic agent; for inducing apoptosis in apoptosis-inducible cancer DESCRIPTION OF THE FIGURES

[0038] The present application can be understood by reference to the following description taken in conjunction with the accompanying figures.

[0039] FIG.1 depicts the stimulation in skeletal muscle glucose uptake after administering the compound of formula (I) to STZ mice. FIG.1(a): Timeline in days for STZ-treatment and FDG-PET scanning. (FIG.1(b)) Representative PET / CT images of FDG uptake in an untreated mouse (upper panel), and a mouse treated with the compound of formula (I) (lower panel). (FIG. 1(c)) Estimated muscle and heart maximal glucose uptake (MRglu) at baseline (Scan 1), 9-10 days after the last STZ injection (Scan 2), and after 1 week of treatment (Scan 3) with 0.5mg / g of the compound of formula (I) (n=5) or no treatment (n=5). (FIG. 1(d)) Glycogen content in muscle and heart of control (n=6-18) and STZ mice untreated (n=14-36) or treated (n=13-27) with the compound of formula (I) from day 15. (FIG. 1(e)) Relative mRNA levels of Txnip, Slc2a1, Slc2a4, Hk2, Pkm, Ppargc1a, Pdk4, Pdha1, Shda, and Cox8b in muscle of control mice(n=5-7), untreated STZ mice (n=5-9) and STZ mice treated with the compound of formula (I) (n=6-8). (FIG.1(f)) Relative mRNA levels of Txnip, Slc2a1, Slc2a4, Pdk4, Ucp2 and Ucp3 in heart of control mice (n=5), untreated STZ mice (n=9) and STZ mice treated with the compound of formula (I) (n=6-7). (FIG.1(g)) Transmitral doppler in STZ treated mice at baseline (Scan 1), 15 days after STZ start (Scan 2,) and after 1 week treatment (Scan 3) with 0.5mg / g of the compound of formula (I) (n=9) or no treatment (n=9). Data are presented as mean ± SEM. Statistical significance between untreated STZ mice and STZ mice treated with the compound of formula (I) was determined by Student’s t-test (FIG.1(c): Muscle, FIG.1(g)) or by Wilcox-test (FIG. 1(c): Heart, FIGS. 1(d)-1(f)) (*p<0.05, **p<0.01, ***p<0.001), and between control mice and STZ mice by Wilcoxon test (FIGS.2(d)-2(f)) (#p<0.05,##p<0.01,###p<0.001), and between Scans in (FIG. 1(c), FIG. 1(g)) by 1-way repeated ANOVA followed by paired Student’s t-test (¤p<0.05).

[0040] FIG.2 depicts the quantification of protein levels by western blot analyses of p-T172 AMPK, p-S79 ACC, and TXNIP protein levels in extracts from vastus muscle in control (n= 5) and STZ mice untreated (n=9) or treated with 0.5 mg / g of the compound of formula (I) (n=9) from day 15.

[0041] FIG.3 depicts how the compound of formula (I) dose-dependently averts hyperglycemia in db / db mice. Fasted glucose (FIG.3(a)) and insulin (FIG.3(b)) levels with area under the curve (AUC) in BKS and db / db mice untreated or treated with 0.5 and 1.0 mg / g of the compound of formula (I), respectively (n=15-20 / group). HOMA-IR (FIG. 3(c)) and HOMA- ȕ (FIG.3(d)) (calculated from FIG.3(a) and FIG.3(b)) with area under the curve (AUC) in BKS and db / db mice untreated or treated with 0.5 and 1.0 mg / g of the compound of formula (I). (FIG. 3(e)) Glycogen content in muscle and heart of BKS (n=5-9) and db / db mice untreated (n=9-10) or treated with the compound of formula (I) (n=6-7). (FIG.3(f)) Relative mRNA levels of Txnip, Slc2a1, Slc2a4, Hk2, Pkm, Ppargc1a, Pdk4, Pdha1, Sdha, Cox8b, Ucp2, and Ucp3 in muscle of BKS (n=6-13) and db / db mice untreated (n=15-22) or treated with 1.0 g / kg of the compound of formula (I) (n=8-15). (FIG. 3(g)) Relative mRNA levels of Txnip, Slc2a1, Slc2a4, Pdk4, Ucp2, and Ucp3 in heart of BKS (n=4-5), untreated db / db mice (n=8-9) and db / db mice treated with a compound of formula (I) (n=6-7). Data are presented as mean ± SEM. Statistical significance between untreated db / db mice and db / db mice treated with the compound of formula(I) was determined by Welch’s ANOVA followed by Games-Howell post hoc test (FIGS.3(a)- 3(d)), Student’s t-test (FIG.3(e)) or Wilcoxon test (FIGS.3(f), 3(g)) (*p<0.05, **p<0.01, ***p<0.001), and between BKS and db / db mice was determined by Wilcoxon test (#p<0.05,##p<0.01,###p<0.001) (FIGS.3(a)-3(d), 3(f), 3(g)) or by Student’s t-test (FIG.3(e)).

[0042] FIG.4 shows that treatment with the compound of formula (I) does not increase serum levels of lactate in STZ and db / db mice. Fasted glucose (FIG.4(a)) and lactate (FIG. 4(b)) levels in STZ mice at day 15 (i.e. before treatment) and at day 22 after 1 week treatment with 0.5 mg / g of the compound of formula (I) (n=9 / group). Fasted glucose (FIG. 4(c)) and lactate (FIG. 4(d)) levels with area under the curve (AUC) in BKS and db / db mice untreated or treated with 0.5 and 1.0 mg / g of the compound of formula (I), respectively (n=7-9 / group). Data are presented as mean ± SEM. Statistical significance between timepoints in (FIG. 4(a)) was determined by Student’s t-test *P<0.05, and between untreated and the db / db mice treated with the compound of formula (I) was determined by Welch’s ANOVA followed by Games-Howell post hoc test (*P<0.05, **P<0.01, ***P<0.001), and between BKS and db / db mice was determined by Wilcoxon test in (FIG.4(c), FIG.4(d)) (##P<0.01).

[0043] FIG.5 depicts the induction of mitochondrial uncoupling by the compound of formula (I). Respirometry plots of intact differentiated C2C12 myotubes + / - 4 h treatment with 0.625, 1.25 and 2.5 ^M of the compound of formula (I) sequentially injected with oligomycin, FCCP, and a cocktail of rotenone and antimycin showing Oxygen consumption Rate (OCR) (FIG. 5(a)) and extracellular acidification rate (ECAR) (FIG.5(b)). FIG.5(c): OCR vs ECAR plot from last baseline measurement (measurement 3 in panel FIG. 5(a) and FIG. 5(b)). FIG. 5(d): Mitochondrial function parameters calculated from OCR data in panel FIG.5(a). Data are presented as mean ± SEM. n=5 / condition. Statistical significance in FIG.5(d) between untreated and cells treated with the compound of formula (I) was determined by one-way ANOVA followed by Tukey's post hoc test (¤p<0.05,¤¤p<0.01). DETAILED DESCRIPTION

[0044] The following description sets forth exemplary compositions, methods, parameters and the like. It should be recognized, however, that such description is not intended as alimitation on the scope of the present disclosure, but is instead provided as a description of exemplary embodiments. Methods of Treatment

[0045] Provided herein are methods and compositions for treating and / or preventing cancer by administering an effective amount compound of formula (I):or a pharmaceutically acceptable salt, solvate, or prodrug thereof. The term “effective amount” used herein refers to an amount of a compound or composition sufficient to treat a specified cancer or other cell proliferation such as ameliorate, palliate, lessen, and / or delay one or more of its symptoms. Generally, an effective amount comprises an amount sufficient to cause a tumor to shrink and / or to decrease the growth rate of the tumor (such as to suppress tumor growth) or to prevent or delay another cell proliferation. In some embodiments, an effective amount is an amount sufficient to delay development. In some embodiments, an effective amount is an amount sufficient to prevent or delay occurrence and / or recurrence. An effective amount can be administered in one or more administrations. The effective amount of the drug or composition may: (i) reduce the number of cancer cells; (ii) reduce tumor size; (iii) inhibit, retard, slow to some extent and preferably stop cancer cell infiltration into peripheral organs; (iv) inhibit (i.e., slow to some extent and preferably stop) tumor metastasis; (v) inhibit tumor growth; (vi) prevent or delay occurrence and / or recurrence of tumor; and / or (vii) relieve to some extent one or more of the symptoms associated with the cancer. In the disclosure below, reference to administration of a compound of formula (I) will include various salt forms, solvates and prodrugs, as described herein.

[0046] In some aspects, provided herein are methods and compositions for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof, comprising administering a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the PDK ispyruvate dehydrogenase kinase 1 (PDK1). In some embodiments, the PDK is pyruvate dehydrogenase kinase 2 (PDK2). In some embodiments, the PDK is pyruvate dehydrogenase kinase 3 (PDK3). In some embodiments, the PDK is pyruvate dehydrogenase kinase 4 (PDK4). In some embodiments, the PDK is two or more of PDK1, PDK2, PDK3 and PDK4.

[0047] The compound of formula (I) is capable of reducing metabolic inflexibility in cancer cells of a subject (e.g., patient), a phenomenon attributed, in part, to the discovery that the compound of formula (I) inhibits PDKs (e.g., PDK4) and increases glucose oxidation in the cancer cells. As a result, the compound of formula (I) can reduce anabolic pathways relied on by cancer cells that results in proliferation. By inhibiting PDKs, the compound of formula (I) is able to induce apoptosis of cancer cells, and hence can be a potent anti-cancer therapeutic. Moreover, the compound of formula (I) can synergize with other anti-cancer therapies. Accordingly, the compound of formula (I) can be used in combination with other anti-cancer agents, as disclosed herein, or can be used as an adjuvant with other anti-cancer therapies,

[0048] As set forth in the Examples, the compound of formula (I) stimulates glucose uptake and glucose utilization in cells. Moreover, as shown herein, the compound of formula (I) promotes gene expression profiles favoring glucose oxidation rather than glycogen storage. Therefore, the compound of formula (I) behaves differently than pan-AMPK activators known in the art, which promote glycogen storage rather than glucose oxidation.

[0049] The increase in glucose uptake in muscle tissue without an increase in glycogen and the observation of increased energy expenditure in animal models prompted in vitro experiments of mitochondrial function. As shown in the Examples, in isolated myotubes, the compound of formula (I) increased basal oxygen consumption and extracellular acidification rate, suggesting increased glucose utilization by increased flux though the tricarboxylic acid (TCA) cycle and oxidative phosphorylation by mitochondrial uncoupling. Thus, the current data indicate that the compound of formula (I) is a dual AMPK activator and mitochondrial uncoupler.

[0050] Additionally, the compound of formula (I) promotes gene expression profiles favoring glucose oxidation in the muscles and heart. Skeletal muscle thioredoxin-inhibiting protein (TXNIP) expression levels are negatively correlated with glucose uptake. As described herein, administration of the compound of formula (I) reduces cardiac TXN1P levels. Inparticular Txnip mRNA and protein levels were reduced in skeletal muscle of the compound of formula (I) treated compared with hyperglycemic mice (see FIG.1(e), and FIG. 2). Moreover, skeletal muscle expression of Slc2a4, encoding Glut4,was reduced in STZ mice compared with controls but normalized in the compound of formula (I) treated hyperglycemic mice and that of Slc2a1, encoding Glut1, tended to be increased. Similarly, the increased expressions of Pyruvate Dehydrogenase Kinase 4, Pdk4, a negative regulator of pyruvate dehydrogenase (PDH) and thus oxidative glucose metabolism, and Uncoupling protein 3 (Ucp3), which favor lipids as fuel substrate, in hyperglycemic mice were reduced in compound of formula (I) treated mice (FIG. 1(e)), indicating that the compound of formula (I) counteracts metabolic inflexibility in skeletal muscle of hyperglycemic diabetic mice. Txnip expression was increased also in hearts of hyperglycemic mice but comparatively reduced in compound of formula (I) treated hyperglycemic mice. Moreover, cardiac expressions of Slc2a1 and Slc2a4 were decreased in untreated hyperglycemic mice but tended to be increased in compound of formula (I) treated mice (FIG.1(f)). Thus, the compound of formula (I) promoted gene expression changes favoring glucose uptake, oxidative glucose metabolism, and ATP generation.

[0051] Hyperglycemia has been shown to rapidly increase cardiac expression of Pdk4 and Ucp3, and to provoke metabolic inflexibility and cardiac dysfunction in mice. Untreated STZ mice showed gradually decreased E peak velocity and increased isovolumetric relaxation time (IVRT), indicating impaired diastolic function, whereas 1 week treatment with the compound of formula (I) reduced IVRT and increased peak E velocity and thus E / A ratio (FIG.1(g), Table 4).

[0052] In one aspect, provided herein are methods and compositions for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof, comprising administering a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the PDK is pyruvate dehydrogenase kinase 4 (PDK4).

[0053] In some embodiments, the cancer is a solid tumor. In some such embodiments, the solid tumor is breast cancer, ovarian cancer, esophageal cancer, melanoma, brain cancer, lung cancer, head and neck cancer, oral cancer, colorectal cancer, breast cancer, prostate cancer, pancreatic cancer, liver cancer, testicular cancer, endometrial cancer, or uterine cancer. Othertypes of solid tumors are named for the particular cells that form them, for example, sarcomas formed from connective tissue cells (for example, bone cartilage, fat), carcinomas formed from epithelial tissue cells (for example, breast, colon, pancreas) and lymphomas formed from lymphatic tissue cells (for example, lymph nodes, spleen, thymus). In some of the foregoing embodiments, the cancer is metastatic cancer.

[0054] In some embodiments, the cancer is a blood cancer. In some such embodiments, the blood cancer is acute lymphocytic leukemia, chronic lymphocytic leukemia, or multiple myeloma.

[0055] In some embodiments, provided herein is a method of treating cancer, comprising administering a compound of formula (I), wherein the cancer is associated with a hypoxic tumor. In some embodiments, the cancer is associated with a hypoxic tumor s leukemia, breast cancer, cervical cancer, brain cancer, renal cancer, liver cancer, lung cancer, pancreatic cancer, colorectal cancer, head and neck cancer, prostate cancer, vulvar cancer, skin cancer, or sarcoma. %. In some embodiments, the hypoxic tumor has a median oxygen level below 1%. In some embodiments, the hypoxic tumor has a median oxygen level below 1% or below 0.5%.

[0056] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat bladder cancer. In some embodiments, the bladder cancer is metastatic bladder cancer.

[0057] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat colon cancer. In some such embodiments, the colon cancer is associated with a mutation in KRAS. In some embodiments, the colon cancer is metastatic colon cancer.

[0058] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat lung cancer. In some such embodiments, the lung cancer is associated with a mutation in KRAS. In other such embodiments, the lung cancer is epidermal growth factor receptor (EGFR)-positive lung cancer. In some embodiments, the lung cancer is metastatic lung cancer. In some embodiments, the lung cancer is non-small celllung cancer (NSCLC). In some embodiments, the NSCLC is associated with mutations in EGFR.

[0059] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat breast cancer. In some such embodiments, the breast cancer is tamoxifen-resistant breast cancer. In some embodiments, the breast cancer is metastatic breast cancer. In some embodiments, the breast cancer is triple negative breast cancer. In some embodiments, the breast cancer is hormone receptor-positive human epidermal growth factor receptor 2-negative breast cancer (HR+HER2-mBC).

[0060] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat melanoma. In some embodiments, the melanoma is metastatic melanoma. In some embodiments, the melanoma is associated with an activating mutation in the BRAF oncogene. In some embodiments, the metastatic melanoma in the subject (e.g., patient) had been treated previously with an immunotherapy (e.g., an anti-PD1 or anti- CTLA4 antibody). In some such embodiments, the metastatic melanoma had not responded to or had limited response to the immunotherapy. In some embodiments, the metastatic is or has become resistant to at least one BRAF inhibitor. In some such embodiments, the BRAF inhibitor is selected from vemurafenib, dabrafenib, encorafenib, trametinib, binimetinib and cobimetinib.

[0061] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat bladder cancer. In some embodiments, the bladder cancer is metastatic bladder cancer.

[0062] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to treat hepatocellular carcinoma (HCC). In some embodiments, the cervical cancer of the subject (e.g., patient) has become resistant to doxorubicin. In some embodiments, the cervical cancer of the subject (e.g., patient) has become resistant to etoposide.

[0063] In some embodiments, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to target cancer stem cells. In some such embodiments,the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is used to target cancer stem cells that are resistant to radiation therapy (radiotherapy).

[0064] Also provided herein is a method of inducing apoptosis in apoptosis-inducible cancer cells comprising administering to said apoptosis-inducible cancer cells an effective amount of a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments the cancer cells are bladder cancer cells, gastric cancer cells, colon cancer cells, breast cancer cells, ovarian cancer cells, melanoma cells, or lung cancer cells.

[0065] In another aspect, compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is administered to a subject (e.g., patient) that has a cancer that is resistant to one or more prior treatments (e.g., chemotherapeutic agents) and / or is resistant to radiation therapy (radiotherapy). In some embodiments, the prior treatment was treatment with a platinum-based agent, a taxane, a nucleoside analog, an immune-checkpoint inhibitor, a Cox-2 inhibitor, an anthracycline, a pyrimidine analog, a topoisomerase inhibitor, an mTOR inhibitor, a proteasome inhibitor, an angiogenesis inhibitor, a B-Raf inhibitor, or a tyrosine kinase inhibitor. In some embodiments, the cancer is resistant to treatment with docetaxel, paclitaxel, paclitaxel, cisplatin, or gemcitabine.

[0066] In another aspect, the disclosure provides methods of reducing drug resistance to a chemotherapeutic agent, said method comprising administering to a subject suffering from cancer (e.g., cancer patient) a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the chemotherapeutic agent is cisplatin. In some embodiments, the chemotherapeutic agent is 5-flurouracil. In some embodiments, the chemotherapeutic agent is erlotinib. In some embodiments, the chemotherapeutic agent is paclitaxel. In some embodiments, the chemotherapeutic agent is doxorubicin. In some embodiments, the chemotherapeutic agent is gemcitabine. In some embodiments, the chemotherapeutic agent is an immune checkpoint inhibitor.Treatment Compounds

[0067] In some embodiments of the foregoing, the treatment compound includes exemplary compounds as described in further detail below. The compound used in the methods provided herein may include salts, solvates, or prodrugs thereof.

[0068] In some aspects, the disclosure provides methods of treating cancer in a subject in need thereof (e.g., patient), comprising administering an effective amount of a compound of formula (I) (4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide), or a salt, solvate or a prodrug thereof.

[0069] In certain embodiments, the compound is an alkali metal salt of 4-chloro-N-[2-[(4- chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. “Alkali metals” are metals found, along with hydrogen, in group I of the periodic table. The alkali metals are lithium, sodium, potassium, rubidium, cesium, and francium. It will therefore be understood that an “alkali metal salt” is a chemical compound consisting of an assembly of cations of one or more alkali metals and associated anions. Accordingly, the term “an alkali metal salt of 4-chloro-N-[2-[(4- chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide” refers to a compound comprising alkali metal cations (e.g., lithium, rubidium, cesium, sodium, and potassium) and anions of 4- chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. For example, alkali metal salts of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5- yl]benzamide are as depicted below:. wherein X+represents the alkali metal (e.g. lithium, rubidium, cesium, sodium, or potassium) cation.

[0070] It will be understood that a “sodium salt” is a chemical compound consisting of an assembly of cations of sodium and associated anions. Accordingly, the term “a sodium salt of 4- chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide” refers to a compound comprising sodium cations and anions of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3- oxo-1,2,4-thiadiazol-5-yl]benzamide. For example:, wherein Na+represents the sodium cation.

[0071] The skilled person will recognize that, when dissolved in a suitable solvent (e.g., water), the alkali metal salt of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5- yl]benzamide may dissociate into its anionic and cationic components.

[0072] Throughout this specification, structures may or may not be presented with chemical names. Where any question arises as to nomenclature, the structure prevails. Where it is possible for the compound to exist as a tautomer (e.g., in an alternative resonance form), the depicted structure represents one of the possible tautomeric forms, wherein the actual tautomeric form(s) observed may vary depending on environmental factors such as solvent, temperature, or pH. All tautomeric (and resonance) forms and mixtures thereof are included within the scope of the Invention. For example, the following tautomers are included within the scope of the invention:

[0073] For the avoidance of doubt, alkali metal salts of 4-chloro-N-[2-[(4- chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide are solid under ambient conditions, and thus the scope of the invention includes all amorphous, crystalline, and part crystalline forms thereof.

[0074] Alkali metal salts of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol- 5-yl]benzamide may be prepared in accordance with techniques that are well known to those skilled in the art. For example, 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol- 5-yl]benzamide may be reacted with the appropriate alkali metal hydroxide, or an alternative alkali metal base compound. Salt switching techniques may also be used to convert one salt into another salt.

[0075] Sodium salts of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5- yl]benzamide may be prepared in accordance with techniques that are well known to those skilled in the art. For example, 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol- 5-yl]benzamide may be reacted with sodium hydroxide, or an alternative sodium base compound. Salt switching techniques may also be used to convert one salt into another salt.

[0076] Where the salt is prepared from 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4- thiadiazol-5-yl]benzamide, 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5- yl]benzamide may be prepared in accordance with techniques that are well known to those skilled in the art. For example, 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol- 5-yl]benzamide may be made in accordance with the techniques described in international patent application WO 2011 / 004162.

[0077] Unless indicated otherwise, all technical and scientific terms used herein will have their common meaning as understood by one of ordinary skill in the art to which this invention pertains.

[0078] In particular embodiments, the alkali metal salt of 4-chloro-N-[2-[(4- chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide is a sodium or potassium salt of 4- chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide. In oneembodiment, the salt is a sodium salt of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4- thiadiazol-5-yl]benzamide.

[0079] In some embodiments, the compound used in the methods provided herein is a compound of formula (II):or a salt thereof, wherein R1is selected from the group consisting of -C(O)-C2H4-CO2H and - PO3H2, or a salt or solvate thereof. In some embodiments, this compound is able to metabolise in vivo to form 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl] benzamide. In certain embodiments, this compound is a salt. For example, salts of this compound include:wherein X+represents an alkali metal, alkaline earth metal or quaternary ammonium (e.g. lithium, magnesium, calcium, ammonium, tetramethylammonium and, particularly, sodium and potassium) cation, with appropriate stoichiometric adjustments being made in view of charges of the ions. In certain embodiments, X+represents an alkali metal (e.g. lithium, rubidium, cesium or, particularly, sodium or potassium) cation. Pharmaceutical Dosage Forms

[0080] The treatment compound is administered to a human subject in need thereof in the form of a pharmaceutical formulation, which is also referred to herein as a pharmaceutical dosage form.

[0081] In one embodiment, the treatment compound is the sole active pharmaceutical ingredient present in the dosage form. In a further embodiment, treatment compound is present in the dosage form alongside one or more other active pharmaceutical ingredients, or may be administered as part of a combination therapy with one or more other active pharmaceutical ingredients.

[0082] In particular embodiments, the treatment compound is provided in the form of particles having a particle size distribution defined by a D90 of less than about 10 μm (e.g. as measured using laser diffraction). In one embodiment, the particles containing the treatment compound may have a particle size distribution defined by a D90 of less than about 10 μm (e.g. from about 5 μm to about 10 μm) (e.g. as measured using laser diffraction). The particle size distribution may alternatively be defined by a D90 of less than about 8 μm (e.g. from about 5 μm to about 8 μm). In a further embodiment, the particles consisting of the treatment compound may have a particle size distribution defined by a D50 of less than about 6 μm (e.g. from about 0.5 μm to about 6 μm). In a yet further embodiment, the particle size distribution of the particles consisting of the treatment compound may further be a defined by a D10 of less than about 2 μm (e.g. from about 0.2 μm to about 2 μm). The particle size distribution parameters mentioned above may be applicable, individually or in combination. For example, in particular embodiments, the dosage form comprises particles containing the treatment compound, said particles having a particle size distribution defined by a D90 of less than about 10 μm and a D50 of less than about 6 μm. Still further, said particles may have a particle size distribution defined by a D90 of less than 9 μm; a D50 of less than 6 μm or less than 5 μm; and a D10 of less than 2 μm or less than 1.5 μm. The particle size distribution of particles containing the treatment compound may be measured by laser diffraction, using, for example a commercially available particle size analyzer. Oral Dosage Forms

[0083] In some variations, the treatment compound may be provided in the form of a tablet or a capsule. For example, capsules such as soft gelatin capsules may be prepared containing the treatment compound alone, or together with a suitable vehicle, e.g. vegetable oil, fat, etc. Similarly, hard gelatin capsules may contain the treatment compound alone, or in combination with solid powdered ingredients such as a disaccharide (e.g. lactose or saccharose), an alcohol sugar (e.g. sorbitol or mannitol), a vegetable starch (e.g. potato starch or corn starch), a polysaccharide (e.g. amylopectin or cellulose derivatives), or gelling agent (e.g. gelatin).

[0084] The pharmaceutical dosage forms described herein may be prepared in accordance with standard and / or accepted pharmaceutical practice. The pharmaceutical dosage forms will generally be provided as a mixture comprising the treatment compound and one or more pharmaceutically acceptable excipients. The one or more pharmaceutically acceptable excipients may be selected with due regard to the intended route of administration in accordance with standard pharmaceutical practice. Such pharmaceutically acceptable excipients are preferably chemically inert to the active compound and are preferably have no detrimental side effects or toxicity under the conditions of use. Suitable pharmaceutical formulations may be found in, for example, Remington The Science and Practice of Pharmacy, 19th ed., Mack Printing Company, Easton, Pennsylvania (1995). A brief review of methods of drug delivery may also be found in, e.g., Langer, Science 249, 1527 (1990).

[0085] The skilled person will understand that the pharmaceutical dosage forms described herein may act systemically, and may therefore be administered accordingly using suitable techniques known to those skilled in the art. The pharmaceutical dosage form as described herein will normally be administered orally, e.g., as an oral pharmaceutical dosage form. Thus, in some variations, provided is an oral pharmaceutical dosage form comprising from about 100 to about 1000 mg of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5- yl]benzamide, or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In one variation, provided is an oral pharmaceutical dosage form comprising from about 200 to about 1000 mg of a sodium salt of 4-chloro-N-[2-[(4-chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0086] Dosage forms intended for oral administration may further comprise an enteric coating in order to prevent or minimize dissolution or disintegration in the gastric environment.As such, oral preparations (e.g., capsules or tablets) coated by an enteric coating may provide targeted release of the treatment compound in the small intestine. For example, the enteric coating may be present on surface of the formulation (e.g., on the surface of a tablet or a capsule), or each of the particles containing the treatment compound may be coated with the enteric coating. Thus, in particular embodiments, the pharmaceutical dosage form used in the method of the invention further comprises an enteric coating.

[0087] In certain embodiments, the enteric coating is present on the pharmaceutical dosage form, and in some variations, said coating may be provided as an outer layer on the pharmaceutical dosage form.

[0088] Alternatively, particles containing the treatment compound may be individually coated with the enteric coating, and said coated particles may be prepared into the pharmaceutical dosage form. Thus, in particular embodiments, the pharmaceutical dosage form contains particles comprising the treatment compound and each particle is coated with the enteric coating.

[0089] The term “enteric coating” refers to a substance (e.g., a polymer) that is incorporated into an oral medication (e.g., applied onto the surface of a tablet, a capsule, particles or pellets) and that inhibits dissolution or disintegration of the medication in the gastric environment. Enteric coatings are typically stable at the highly acidic pH found in the stomach, but break down rapidly in the relatively basic pH of the small intestine. Therefore, enteric coatings prevent release of the active ingredient in the medication until it reaches the small intestine.

[0090] Any enteric coating known to the skilled person may be used in the present invention. Particular enteric coating materials that may be mentioned include those which comprise beeswax, shellac, an alkylcellulose polymer resin (e.g. ethylcellulose polymers, carboxymethylethylcellulose, or hydroxypropyl methylcellulose phthalate) or an acrylic polymer resin (e.g. acrylic acid and methacrylic acid copolymers, methyl methacrylate copolymers, ethoxyethyl methacrylates, cyanoethyl methacrylate, methacrylate copolymers, methacrylic acid copolymer, aminoalkyl methacrylate copolymer, poly(acrylic acid), poly(methacrylic acid), methacrylic acid alkylamide copolymer, poly(methyl methacrylate), poly(methacrylic acid) (anhydride), polymethacrylate, methyl methacrylate copolymer, poly(methyl methacrylate)copolymer, polyacrylamide, poly(methacrylic acid anhydride), and glycidyl methacrylate copolymers), cellulose acetate phthalate and polyvinyl acetate phthalate.

[0091] In some variations, the pharmaceutical compositions comprise the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, and at least one pharmaceutically acceptable excipient. In particular, the at least one pharmaceutically acceptable excipient may be a lubricant, a binder, a filler, a surfactant, a diluent, an anti- adherent, a coating, a flavoring, a colorant, a glidant, a preservative, a sweetener, a disintegrant, an adsorbent, a buffering agent, an antioxidant, a chelating agent, a dissolution enhancer, a dissolution retardant, or a wetting agent.

[0092] Particular pharmaceutically acceptable excipients that may be mentioned include mannitol, PVP (polyvinylpyrrolidone) K30, lactose, saccharose, sorbitol, starch, amylopectin, cellulose derivatives, gelatin, or another suitable ingredients, as well as disintegrating agents and lubricating agents such as sodium lauryl sulfate, Na-docusate, magnesium stearate, calcium stearate, sodium stearyl fumarate and polyethylene glycol waxes. In the preparation of a pharmaceutical dosage form of the treatment compound for oral administration, particles containing the treatment compound (preferably milled) may be mixed, either together or separately, with mannitol, PVP (polyvinylpyrrolidone) K30 and sodium lauryl sulfate.

[0093] In the preparation of a pharmaceutical dosage form, the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, may be mixed, either together or separately, with one or more of the pharmaceutical excipients (including basic excipients) listed above.

[0094] Mixtures of the treatment compound and one or more pharmaceutically acceptable excipients may be processed into pellets or granules, or compressed into tablets. Thus, pharmaceutical dosage form of the method of the inventions may be a tablet, mini-tablets, blocks, pellets, particles, granules, or a powder for oral administration.

[0095] Pharmaceutical formulations that may be mentioned include those in which the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, is present in a total amount that is at least 1% (or at least 10%, at least 30% or at least 50%) byweight of the formulation. That is, the weight ratio of the treatment compound to the totality of the components (i.e., the treatment compound and all pharmaceutical excipients, e.g. adjuvants, diluents and carriers) of the pharmaceutical formulation is at least 1:99 (or at least 10:90, at least 30:70 or at least 50:50). Dosage Amounts on Oral Dosage Forms

[0096] A “therapeutically effective amount”, an “effective amount” or a “dosage” as used herein refers to an amount of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, that is sufficient to produce a desired effect, which can be a therapeutic and / or beneficial effect. The effective amount or dosage will vary with the age or general condition of the individual or subject (e.g., a human), the severity of the condition being treated, the particular agents administered, the duration of the treatment, the nature of any concurrent treatment, the pharmaceutically acceptable carrier used, and like factors within the knowledge and expertise of those skilled in the art.

[0097] The term “about,” as used herein when referring to a measurable value such as an amount of a compound, dose, time, temperature, and the like, refers to variations of 20%, 10%, 5%, 1%, 0.5%, or even 0.1% of the specified amount. It is contemplated that, at each instance, such terms may be replaced with the notation “±10%”, or the like (or by indicating a variance of a specific amount calculated based on the relevant value). It is also contemplated that, at each instance, such terms may be deleted.

[0098] It will be understood that the dosage amounts and pharmacokinetic parameters described below relate to treating patients with particular diseases, specifically heart diseases as set forth herein. The dosage amounts and pharmacokinetic parameters also can apply to human subjects taking the drugs for fitness enhancement or cardiac improvement. As set forth above, in some embodiments, lower amounts of the compound of formula (I) may be required to improve fitness or cardiac function than when the compound is administered to a subject (e.g., patient) with a heart disease.

[0099] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally oncedaily to a subject (e.g., patient) in need of at a dose of from about 100 mg to about 1,000 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 200 mg to about 1,000 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 400 mg to about 800 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of from about 100 mg to about 300 mg. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily at dose of about 100 mg, about 200 mg, about 300 mg, about 400 mg or about 500 mg.

[0100] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 40 ^g / mL to about 200 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 40 ^g / mL to about 80 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 70 ^g / mL to about 120 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 90 ^g / mL to about 160 ^g / mL. In some embodiments, compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state blood plasma concentration of the compound of formula (I) of from about 100 ^g / mL to about 150 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof s administered daily to the human subject and results in a steadystate blood plasma concentration of the compound of formula (I) of from about 120 ^g / mL to about 140 ^g / mL.

[0101] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 1,000 h*^g / mL to about 4,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 1,000 h*^g / mL to about 2,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 1,500 h*^g / mL to about 4,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 1,800 h*^g / mL to about 3,100 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24of the compound of formula (I) of from about 1,500 h*^g / mL to about 2,000 h*^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24 of the compound of formula (I) of from about 2,500 h*^g / mL to about 4,000 h*^g / mL In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a steady state AUC0-24of the compound of formula (I) of from about 3.000 h*^g / mL to about 3,500 h*^g / mL.

[0102] In any of the preceding embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily a subject in need thereof (e.g., patient), wherein the administration results in a Cmax of the compound of formula (I) of from about 40 ^g / mL to about 200 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 70 ^g / mL to about 200 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 80 ^g / mL to about 140 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmax of the compound of formula (I) of from about 70 ^g / mL to about 100 ^g / mL. In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof can be administered orally once daily to a subject in need thereof (e.g., patient), wherein the administration results in a Cmaxof the compound of formula (I) of from about 1200 ^g / mL to about 150 ^g / mL.

[0103] In some embodiments, the compound of formula (I) is provided in a tablet with the components shown in Table 1. Table 1: Exemplary composition of tablet comprising Na salt tablet of formula (I) Component Content per tablet Function Reference to standard Compound of formula (I) 424.24 mg*Drug substance Na saltAPIspecification Microcrystalline cellulose 206.59 mg Filler / Binder Ph. Eur. Lactose monohydrate 413.17 mg Filler Ph. Eur. Sodium starch glycolate 90.00 mg Disintegrant Ph. Eur. Colloidal silicon dioxide 30.00 mg Glidant Ph. Eur. Magnesium stearate 36.00 mg Lubricant Ph. Eur Total 1200.00 mg *Corresponds to 400 mg of free acid form of the compound of formula (I) Combination Therapies

[0104] The skilled person will understand that the method of the invention may comprise (e.g., be combined with) further treatment(s) for the same condition. Such combinations are selected based on the condition to be treated, cross-reactivities of ingredients and pharmaco- properties of the combination. In one embodiment, provided is a compound as described herein, or pharmaceutically acceptable salt thereof, used in combination with another anti-cancer therapy, such as a chemotherapeutic agent, an immunotherapeutic agent, a gene therapeutic agent or a combination thereof. For example, when treating cancer, the compounds and compositions provided herein can be combined with other anti-cancer therapeutic agents, surgical procedures, radiation procedures or a combination of any of the foregoing. The treatment methods described herein also contemplate combination therapy.

[0105] As used herein, by “combination therapy” is meant a therapy that includes two or more different anti-cancer compounds or therapeutic agents. Thus, in one aspect, a combination therapy comprising a compound detailed herein and another compound or therapeutic agent is provided. In some variations, the combination therapy optionally includes one or more pharmaceutically acceptable carriers or excipients, non-pharmaceutically active compounds and / or inert substances. In various embodiments, treatment with a combination therapy may result in an additive or even synergistic (e.g., greater than additive) result compared to administration of a single compound of the disclosure alone. In some embodiments, a lower amount of each compound is used as part of a combination therapy compared to the amount generally used for individual therapy. Preferably, the same or greater therapeutic benefit is achieved using a combination therapy than by using any of the individual compounds alone. In some embodiments, the same or greater therapeutic benefit is achieved using a smaller amount (e.g., a lower dose or a less frequent dosing schedule) of a compound in a combination therapy than the amount generally used for individual compound or therapy. Preferably, the use of a small amount of compound results in a reduction in the number, severity, frequency and / or duration of one or more side-effects associated with the compound.

[0106] It is also possible to combine a compound of the disclosure with one or more other active ingredients in a unitary dosage form for simultaneous or sequential administration to a subject (e.g., patient). The combination therapy may be administered as a simultaneous orsequential regimen. When administered sequentially, the combination may be administered in two or more administrations.

[0107] When treating a disease or disorder that is ameliorated by activating AMPK, the salt of the invention may be administered in conjunction with one or more other (i.e. different) therapeutic agents that are useful in treating that disease or disorder.

[0108] Such combination treatments may involve the administration of the salt of the invention to the subject in conjunction (e.g., sequentially or simultaneously) with the different therapeutic agent in the same formulation, or preferably in a separate formulation. By “administration in conjunction with” (and similarly “administered in conjunction with”) we include that the respective active ingredients are administered, sequentially or simultaneously, as part of a medical intervention directed towards treatment of the relevant condition. By simultaneously, we mean that the salt of the invention and the different therapeutic agent are administered alongside one another, either in a single pharmaceutical dosage form comprising both active ingredients or in separate dosage forms administered at the same time.

[0109] Thus, in relation to the present invention, the term “administration in conjunction with” (and similarly “administered in conjunction with”) includes that the salt of the invention and the different therapeutic agent are administered either together, or sufficiently closely in time, to enable a beneficial effect for the subject (e.g., patient) that is greater, over the course of the treatment of the relevant condition, than if either agent is administered alone in the absence of the other component over the same course of treatment. Determination of whether a combination provides a greater beneficial effect in respect of, and over the course of, treatment of a particular condition will depend upon the condition to be treated, but may be achieved routinely by the skilled person.

[0110] Further, in the context of the present invention, the term “in conjunction with” includes that one or other of the two active ingredients may be administered (optionally repeatedly) prior to, after, and / or at the same time as, administration of the other. When used in this context, the terms “administered simultaneously” and “administered at the same time as” include instances in which the individual doses of the salt of the invention and the differenttherapeutic agent are administered within 6 hours, 3 hours, 2 hours, 1 hour, 45 minutes, 30 minutes, 20 minutes or 10 minutes) of each other.

[0111] The methods of the invention (and oral dosage forms used in such methods) disclosed herein may also have the advantage that the dose-efficient methods using the salt of the invention may be more efficacious than, be less toxic than, be longer acting than, be more potent than, produce fewer side effects than, be more easily absorbed than, and / or have a better pharmacokinetic profile (e.g. higher oral bioavailability and / or lower clearance) than over other therapies known in the prior art, whether for use in the above-stated indications or otherwise. In particular, methods of the invention may have the advantage that they are more efficacious and / or exhibit advantageous properties in vivo such as fewer side effects as a result of the dose- efficient characteristics of the salt of the invention.

[0112] Particular examples of other anti-cancer agents that can be used in combination with a compound of formula (I) include, but are not limited to, a platinum-based chemotherapeutic agent (such as cisplatin, carboplatin, or oxaliplatin), a kinase inhibitor (such as bortezomib, erlotinib, gefitinib, imatinib, vemurafenib, vismodegib, ibrutinib), a topoisomerase inhibitor (such as a Topoisomerase I inhibitor (e.g., irinotecan or topotecan) or a Topoisomerase II inhibitor (e.g., etoposide or teniposide), an anthracycline (such as daunorubicin, doxorubicin, epirubicin, idarubicin, mitoxantrone, or valrubicin), a histone deacetylase inhibitor (such as vorinostat or romidepsin), a bromodomain inhibitor, other epigenetic inhibitors, a taxane (such as paclitaxel or docetaxel), an anti-angiogenic inhibitor, a nucleotide analog or precursor analog (such as azacitidine, azathioprine, capecitabine, cytarabine, doxifluridine, 5-fluorouracil, gemcitabine, hydroxyurea, mercaptopurine, methotrexate, or tioguanine), or pemetrexed.

[0113] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with cisplatin to treat a subject (e.g., patient) with cancer. In some embodiments, the cancer is bladder cancer, gastric cancer, breast cancer, lung cancer, HCC. ovarian cancer or colon cancer.

[0114] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with an EGFR inhibitor to treatlung cancer (e.g., NSCLC) in a subject in need thereof (e.g., patient). In some embodiments the EGFR inhibitor is erlotinib, gefitinib, lapatinib, lazertinib or necitumumab.

[0115] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with paclitaxel to treat lung cancer (e.g., NSCLC) in a subject in need thereof (e.g., patient).

[0116] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination doxorubicin to treat HCC in a subject in need thereof (e.g., patient). In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination doxorubicin to treat cervical cancer in a subject in need thereof (e.g., patient).

[0117] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with a recombinant arginase. In some such embodiments, the cancer is breast cancer (e.g., triple negative breast cancer). In other such embodiments, the cancer is HCC.

[0118] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with a COX2 inhibitor, propranolol, metformin, salinomycin, or phenformin.

[0119] In some embodiments, the compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof is administered in combination with an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is a PD-1 inhibitor (e.g., cemipilimab, nivolumab or pembrolizumab). In some embodiments, the immune checkpoint inhibitor is a PD-L1 inhibitor (e.g., atezolizumab, avelumab, or durvalumab). In some embodiments, the immune checkpoint inhibitor is a CTLA-4 inhibitor (e.g., ipilimumab or tremelimumab).

[0120] In another aspect, the disclosure provides methods of overcoming tumor radioresistance by administering to a subject suffering from cancer (e.g., cancer patient) a pharmaceutically effect amount of a compound of formula (I) or a pharmaceutically acceptable salt, solvate, or prodrug thereof. In some embodiments, the compound of formula (I) or apharmaceutically acceptable salt, solvate, or prodrug thereof is administered prior to the subject (e.g., patient) receiving radiotherapy. Uses of the Compound

[0121] In certain aspects, provided is the use of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in a therapy, including any of the methods described herein. For instance, in some embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof. In other embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for reducing drug resistance to a chemotherapeutic agent. In yet other embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for inducing apoptosis in apoptosis-inducible cancer cells.

[0122] In certain aspects, also provided is the use of a compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, for the manufacture of a medicament. In some embodiments, the medicament is for any of the methods described herein. For instance, in some embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in the manufacture of a medicament for treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a subject (e.g., human patient) in need thereof. In other embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in the manufacture of a medicament for reducing drug resistance to a chemotherapeutic agent. In yet other embodiments, provided is the use of the compound of formula (I), or a pharmaceutically acceptable salt, solvate, or prodrug thereof, in the manufacture of a medicament for inducing apoptosis in apoptosis-inducible cancer cells.EXAMPLES

[0123] The presently disclosed subject matter will be better understood by reference to the following Examples, which are provided as exemplary of the invention, and not by way of limitation.

[0124] Herein, and unless otherwise indicated, “Cmpd-(I)” or “Compound of formula (I)” referred to in the experiments detailed in the following Examples, and is 4-chloro-N-[2-[(4- chlorophenyl)methyl]-3-oxo-1,2,4-thiadiazol-5-yl]benzamide.

[0125] Mice. Mice were housed in the certified animal facility, with 12 h light / dark cycle and ad libitum access to the respective diets. Leptin receptor-deficient male BKS.Cg-Dock7m + / + Leprdb / J (db / db), and C57BLKS / J (BKS) mice were obtained. F1 mice were obtained from breeding male C57BL / 6J mice with female CBA / CaCrl. Nine weeks old male F1 mice were treated with multiple low dose streptozotocin (50 mg / kg*day for 5 consecutive days; freshly prepared in 0.1 mM sodium citrate, pH 4.5) to induce diabetes. Mice were ad libitum fed D10001 diet or D10001 formulated with the compound of formula (I) at 0.25 mg / g, 0.5 mg / g and 1 mg / g of the compound of formula (I) (CAS # 1261289-04-6). Cohorts of BKS, db / db and F1 mice were housed in groups of 4-5 mice / cage. Mice with apparent health problems such as >10% reduction in body weight or fighting were excluded with no differences between groups. In the different cohorts, mice were randomly allocated to the cages, based on weight and fasting blood glucose and allocated cage-wise, or if possible individually, to different treatments in order to minimize influence of starting weight and glucose homeostasis. For in vitro analyses such as western blot, qPCR, histology and immunohistology work, samples from 5-9 mice / diet were randomly selected. All in vivo analyses were performed between 9 am to 3 pm.

[0126] Echocardiography. For echocardiography, mice were sedated using 1.5-2% isoflurane, in 0.8Lāmin-1O2 (g), and placed on a temperature-controlled table. Chest hair was removed using hair removal cream. Respiration and ECG were monitored during the scan, and anaesthesia adjusted to avoid depression of respiration. Total scan time did not exceed 15 minutes. Stroke volume, cardiac output and wall thicknesses were measured in the parasternal long-axis view using Bmode and M-mode images. Diastolic left ventricle inflow used transmitral doppler in the apical four-chamber view. Off-line analysis was done in a blinded manner.

[0127] Mitochondrial respiration analyses. C2C12 myoblasts were in growth media, 10% fetal bovine serum and 20U / ml Penicillin-Streptomycin. To obtain myotubes, C2C12 myoblasts were seeded in poly-L-Lysin coated XF96 plates at 7500 cells / cm2, cultivated in growth media for 3 days to 80% confluence, thereafter switched to differentiation media (DM; growth media with FBS replaced by 2% horse serum for 6 days with media changes every 2 days. Respiration assays were performed using a Seahorse XFe96 Extracellular flux analyzer according to the manufacturer’s “Mito-stress” protocol and mitochondrial parameters. Briefly, differentiated myotubes were pre-treated by switching to growth media with 1% horse serum together with the compound of formula (I) (sodium salt formulation) for 4h. Myotubes were then equilibrated for 1h in Seahorse assay medium (1mM Na-Pyruvate, 10mM Glucose, 2mM L-Glutamine) adjusted to pH7.4 and supplemented with the compound of formula (I) as during pretreatment. Measurements of oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) was collected for baseline, followed by sequential addition of 1^M oligomycin, 2^M FCCP, and 0.5^M rotenone + 0.5^M antimycin A. Mitochondrial function parameters were calculated according to the manufacturer’s recommendations.

[0128] Western Blot analysis. Vastus muscles were isolated from non-fasted mice and crushed in a mortar using a pestle and liquid nitrogen. Vastus muscles and INS-1E cell samples were homogenized in ice cold protein lysis buffer (100mM Tris pH 6.8, 2% SDS), with protease inhibitor cocktail and phosphatase inhibitor cocktail and sonicated. The vastus supernatant was collected after 10 min at 14,000 rpm. Protein concentration was measured using the BCA kit after which samples were diluted in Laemmli buffer and denaturized. 20 ^g of vastus and 7 ^g of INS-1E cell protein samples were separated on 4–15 % CriterionTM TGX Stain-FreeTM Protein Gels and blotted to low fluorescent PVDF or Nitrocellulose membrane respectively. Primary and secondary antibodies are listed Table 2. Values were normalized to stain-free total protein signal or to a beta-actin.Table 2

[0129] qRT PCR. Total RNA from isolated islets was prepared using RNeasy Micro Kit. Total RNA from gastrocnemius muscles and left cardiac ventricles was prepared using RNeasy Fibrous Tissue Mini kit after first crushing the tissue in liquid nitrogen using a pestle. First strand cDNA synthesis was done using SuperScript III according to the manufacturer’s instructions. Primers used for qRT-PCR are listed in Table 3. Expression of Tbp was used for normalization of samples from islets, gastrocnemius muscle and left cardiac ventricle. Normalization by Tbp was validated by Rpl32 in left cardiac ventricle samples and gastrocnemius muscles. Table 3

[0130] Islet isolation and culture. Mouse islets were isolated essentially and cultured for 48h in media (RPMI 1640 medium, 1% fetal bovine serum, 10 mM HEPES, 1 mM sodiumpyruvate, 50^M 2-mercaptoethanol, 50U / ml Pen:Strep) supplemented with 11mM glucose or 22mM glucose + / - 5^M of the compound of formula (I). Human islets from nondiabetic donors were cultured for 48 h in media (CMRL medium, 10 % fetal bovine serum, 20 U / ml Pen:Strep and 1X GlutaMax) supplemented with 5.5mM glucose or 25mM glucose + / - 5^M of the compound of formula (I).

[0131] RNA-seq. RNA-seq was performed. RNA-seq libraries were prepared from 150ng total RNA using the Illumina TruSeq stranded mRNA Library Kit followed by 100 bp paired-end sequencing on a NovaSeq 6000 Illumina Sequencer. RNA-seq reads were processed. Fastq files with 100-nt paired-end sequenced reads were quality-checked, aligned to the mouse or human genome (GRCm39 or GRCh38. Fragment - gene hits were counted. Subsequent analysis was performed in R using packages DESeq2 and clusterProfiler. Normalization and differential expression analysis were performed using DESeq2 with batch correction (referring to islet preparation batch and human donor respectively), independent filtering (alpha = 0.05) and False Discovery Rate (FDR) < 0.05. Genes with baseMean expression > filterThreshold were considered to be expressed and to constitute the gene background for subsequent ORA and GSEA analysis using clusterProfiler with a pvalueCutoff = 0.05 and a qvalueCutoff =0.2 for overrepresented gene categories.

[0132] Cell lines. INS-1E and C2C12 were commercially available and therefore not authenticated following purchase. The cells were free of mycoplasma as determined by PCR.

[0133] Statistical analyses. The experimental design was considered to consist of a non- diabetic control group (BKS or Control (i.e. vehicle injected F1 mice), a diabetic control (db / db mice or STZ mice, i.e. STZ injected F1 mice) and compound-of-formula-(I)-treated group(s) (db / db mice or STZ mice kept on a diet formulated with the compound of formula (I)). First, the validity of the diabetic models was tested by comparing the control and diabetic groups. Second, the effect of the compound-of-formula-(I) diet was evaluated by comparing the compound-of- formula-(I)-treated group(s) to the diabetic group. The parametric nature and heteroscedasticity of the data were evaluated by Shapiro-Wilk test and Leven’s test and used to select the appropriate statistical test. For two-group comparisons, heteroscedastic t-test or Wilcoxon-test were used together with holm correction for multiple testing. For multiple groups, ANOVAanalysis, were used followed by the appropriate post hoc analysis (see figure legends). P<0.05 was considered as statistically significant. Data analysis, statistical analysis and visualization were performed in R using the tidyverse, rstatix, and ggpubr packages. Example 1: The compound of formula (I) promote gene expression profile favoring glucose oxidation.

[0134] Skeletal muscle TXNIP expression levels are negatively correlated with glucose uptake. Concurrent with enhanced skeletal muscle glucose uptake, Txnip mRNA and protein levels were reduced in skeletal muscle of STZ mice treated with the compound of formula (I) compared with untreated STZ mice (FIG.1(e), 2). Moreover, skeletal muscle expression of Slc2a4, encoding Glut4, was reduced in STZ mice compared with controls but normalized in STZ mice treated with the compound of formula (I) and that of Slc2a1, encoding Glut1, tended to be increased (p=0.059) in STZ mice treated with the compound of formula (I) (FIG.1(e)). Similarly, the increased expressions of Pyruvate Dehydrogenase Kinase 4, Pdk4, a negative regulator of pyruvate dehydrogenase (PDH) and thus oxidative glucose metabolism, and Uncoupling protein 3 (Ucp3), which favor lipids as fuel substrate, in STZ mice were reduced in mice treated with the compound of formula (I) (FIG. 1(e)), suggesting that the compound of formula (I) counteracts metabolic inflexibility in skeletal muscle of STZ diabetic mice. Txnip expression was increased also in hearts of STZ mice but comparatively reduced in STZ mice treated with the compound of formula (I) (FIG.1(f)). Moreover, cardiac expressions of Slc2a1 and Slc2a4 were decreased in untreated STZ mice but tended (p=0.088 for both) to be increased in mice treated with the compound of formula (I) (FIG.1(f)). The expressions of Pdk4, Ucp2, and Ucp3 were increased in heart of untreated STZ mice but attenuated (p= 0.055 for Ucp2) in STZ mice treated with the compound of formula (I) (FIG.1(f)). Thus, the compound of formula (I) promoted gene expression changes favoring glucose uptake, oxidative glucose metabolism, and ATP generation in skeletal muscle and heart of STZ mice (FIGS.1(e), 1(f)). Hyperglycemia has been shown to rapidly increase cardiac expression of Pdk4 and Ucp3, and to provoke metabolic inflexibility and cardiac dysfunction in mice

[0034] . Untreated STZ mice showed gradually decreased E peak velocity and increased isovolumetric relaxation time (IVRT), indicating impaired diastolic function, whereas 1 week treatment with the compound of formula (I) reduced IVRT and increased peak E velocity and thus E / A ratio (FIG.1(g), Table 4). Theimproved filling of the left ventricle in mice treated with the compound of formula (I) also resulted in increased stroke volume and cardiac output (Table 4). Altogether these findings show that in diabetic STZ mice, the compound of formula (I) ameliorated hyperglycemia by stimulating insulin independent glucose uptake and utilization, mitigated glycogen accumulation, and reverted diabetic cardiomyopathy. Table 4

[0135] Table 4 shows echocardiographic measurements of left ventricular dimensions in para-sternal long axis (PLAX) B-mode or M-mode. HR, heart rate; SV, stroke volume; CO, cardiac output; EDV, end-diastolic volume; ESV, end-systolic volume; AWd / s, anterior wall thickness in diastole / systole; LVIDd, / s left ventricular inner diameter in diastole / systole; PWd / s, posterior wall thickness in diastole / systole; EF, ejection fraction; FS, fractional shortening; E / A, ratio of E and A peak wave velocity; Decel time, deceleration time of E wave from peak toprojected baseline; IVRT, isovolumetric relaxation time. Statistics used One-way repeated ANOVA with Tukey’s post-hoc test. *P<0.05 between RD and the compound of formula (I) at same timepoint.#P<0.05 compared to baseline. ¤P<0.05 compared to scan 2. Data are mean±SD. Example 2: The compound of formula (I) ameliorates hyperglycemia and promotes gene expression profiles favoring glucose oxidation in muscle of db / db mice.

[0136] To explore the potential of the compound of formula (I) to ameliorate hyperglycemia in the context of severe insulin resistance, db / db mice were treated with a diet formulated with the compound of formula (I) at a concentration of 0.5 or 1.0 mg / g of the compound of formula (I), denoted Cmpd-(I)-(0.5) and Cmpd-(I)-(1.0), for 9w. BKS mice were used as controls. At 6w of age, compensatory hyperinsulinemia was evident in db / db mice (FIGS.3(a), 3(b)). Untreated db / db mice failed to compensate for the ensuing insulin resistance and blood glucose levels rapidly increased as a result of declining insulin levels (FIGS.3(a), 3(b)). The compound of formula (I) dose-dependently increased insulin levels compared both with starting values and with untreated db / db mice and thus attenuated an increase in blood glucose levels (FIGS.3(a), 3(b)). These findings provide evidence that the compound of formula (I) preserved a compensatory beta-cell insulin secretory response and homeostasis model assessment of beta-cell function (HOMA-beta) calculations showed that the decline in beta-cell function was attenuated in db / db mice treated with the compound of formula (I) (FIG.3(c)). The compound-of-formula- (I)-mediated attenuation of hyperglycemia in db / db mice was paralleled by reduced glycogen accumulation in skeletal muscle and heart (FIGS.3(d)), suggesting that the compound of formula (I) stimulated glucose utilization also in diabetic db / db mice. The decreased expression of Txnip, Pdk4 and Ucp3 together with the increased expression of Slc2a4, Peroxisome proliferator-activated receptor-gamma coactivator (PGC)-1alpha (Ppargc1a), a positive regulator of mitochondrial biogenesis and respiration, and Cox8b, a driver of oxidative phosphorylation, in skeletal muscle of db / db mice treated with the compound of formula (I) compared with untreated db / db mice supports this notion (FIG.3(f)). The increased cardiac expression of Slc2a1 and the attenuated expressions of Txnip, Pdk4, and Ucp3 in treated compared with untreated db / db mice (FIG. 3(g)) is also consistent with that observed in STZ mice treated with the compound of formula (I). Notably, the compound of formula (I) did not increase serum lactate levels in STZ mice treated with the compound of formula (I) and db / dbmice (FIG.4). Taken together, these findings provide evidence that, in both STZ and db / db diabetic mice, the compound of formula (I) averts gene signatures associated with metabolic inflexibility that in turn is associated with diabetes and diabetic cardiomyopathy. Example 3: The compound of formula (I) stimulates mitochondrial uncoupling in myotubes.

[0137] The increase in glucose utilization and reduced glycogen content observed in skeletal muscle and heart of STZ and db / db mice treated with the compound of formula (I) suggests that the compound of formula (I) increases energy expenditure by generating a metabolic demand either via futile cycling and / or mitochondrial uncoupling. Studies of mitochondrial and glycolytic function in intact, differentiated C2C12 myotubes were performed to elucidate a potential uncoupling potential for the compound of formula (I). The compound of formula (I) dose-dependently increased oxygen consumption rate (OCR), a measure of oxidative phosphorylation, in C2C12 myotubes, and the ratio of basal OCR to basal ECAR (extracellular acidification rate) (FIGS.5(a)-5(c)), indicating that the compound of formula (I) increased the cellular preference for oxidative metabolism. The compound of formula (I) also showed a diminished reduction in OCR in response to oligomycin, which blocks the ATP synthase, and thus increased proton leak (FIGS. 5(a), 5(d)). However, consistent with our previously published findings, the compound of formula (I) did not significantly reduce cellular ATP levels (FIG. 5(d)). Altogether these results show that the compound of formula (I) increased cellular respiration by functioning as mitochondrial uncoupler, providing evidence that the compound of formula (I), by stimulating TCA flux and oxidative metabolism, induces a metabolic demand that enhances energy expenditure.

Claims

CLAIMS What is claimed is:

1. A method of treating cancer associated with pyruvate dehydrogenase kinase (PDK) overexpression or activation in a human in need thereof, comprising administering an effective amount of a compound of formula (I):or a pharmaceutically acceptable salt thereof.

2. The method of claim 1, wherein said administering restores metabolic flexibility in the cancer cells of the human.

3. The method of claim 1 or 2, wherein the PDK is PDK4.

4. The method of any one of claims 1-3, wherein the cancer in the human is resistant to one or more anti-cancer agents.

5. The method of claim 4, wherein the one or more anti-cancer agents is a platinum-based agent, a taxane, a nucleoside analog, an immune-checkpoint inhibitor, a Cox-2 inhibitor, an anthracycline, a pyrimidine analog, a topoisomerase inhibitor, an mTOR inhibitor, a proteasome inhibitor, an angiogenesis inhibitor, a B-Raf inhibitor, or a tyrosine kinase inhibitor, or any combination thereof.

6. The method of any one of claims 1 to 5, wherein the cancer in the human is resistant to radiation therapy.

7. The method of any one of claims 1 to 6, wherein said administration increases tumor oxygenation.

8. The method of claim 7, wherein the tumor is a hypoxic tumor.

9. The method of any one of claims 1 to 8, wherein the cancer has metastasized.

10. The method of any one of claims 1 to 9, wherein the cancer is bladder cancer, colon cancer with mutation in KRAS, lung cancer with mutation in KRAS, gastric cancer, breast cancer, or ovarian cancer, or any combination thereof.

11. The method of claim 10, wherein the cancer is colon cancer with mutation in KRAS.

12. The method of claim 11, further comprising administering 5-fluorouracil to the human.

13. The method of claim 10, wherein the cancer is tamoxifen-resistant breast cancer.

14. The method of claim 13, further comprising administering tamoxifen to the human.

15. The method of claim 10, wherein the cancer is bladder cancer.

16. A method of reducing drug resistance to a chemotherapeutic agent, said method comprising administering to a human suffering from cancer a compound of formula (I):or a pharmaceutically acceptable salt thereof.

17. The method of claim 16, wherein the chemotherapeutic agent is cisplatin.

18. The method of clam 16 or 17, wherein the cancer is bladder cancer, gastric cancer, breast cancer, lung cancer, HCC, ovarian cancer, or colon cancer, or any combination thereof.

19. The method of claim 16, wherein the chemotherapeutic agent is an EGFR inhibitor.

20. The method of claim 19, wherein the EGFR inhibitor is erlotinib, gefitinib, lapatinib, lazertinib or necitumumab, or any combination thereof.

21. The method of claim 19 or 20, wherein the cancer is non-small cell lung cancer (NSCLC).

22. The method of claim 16, wherein the chemotherapeutic agent is paclitaxel.

23. The method of claim 22, wherein the cancer is non-small cell lung cancer (NSCLC).

24. The method of claim 16, wherein the chemotherapeutic agent is doxorubicin.

25. The method of claim 24, wherein the cancer is hepatocellular carcinoma (HCC) or cervical cancer.

26. The method of claim 16, wherein the chemotherapeutic agent is a Cox2 inhibitor, propranolol, metformin, salinomycin, or phenformin, or any combination thereof.

27. The method of claim 16, wherein the chemotherapeutic agent is an immune checkpoint inhibitor.

28. The method of claim 27, wherein the immune checkpoint inhibitor is an anti-PD1 antibody.

29. The method of claim 27, wherein the immune checkpoint inhibitor is an anti-PDL1 antibody.

30. The method of claim 27, wherein the immune checkpoint inhibitor is an anti-CTLA4 antibody.

31. A method of inducing apoptosis in apoptosis-inducible cancer cells, comprising administering to said apoptosis-inducible cancer cells an effective amount of a compound of formula (I):or a pharmaceutically acceptable salt thereof.

32. The method of any one of claims 1 to 31, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered orally once daily to the human at a dose of from about 100 mg to about 1,000 mg.

33. The method of any one of claims 1 to 31, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered orally once daily to the human at a dose of from about 100 mg to about 500 mg.

34. The method of any one of claims 1 to 31, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered orally once daily to the human at a dose of from about 200 mg to about 400 mg.

35. The method of any one of claims 1 to 31, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered daily to the human and results in a steady state blood plasma concentration of the compound of formula (I) of from about 40 ^g / mL to about 120 ^g / mL.

36. The method of any one of claims 1 to 35, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered daily to the human subject and results in a steady state AUC0-24 of the compound of formula (I) is from about 1,000 h*^g / mL to about 4,000 h*^g / mL.

37. The method of any one of claims 1 to 36, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is an alkali metal salt.

38. The method of claim 37, wherein the alkali metal salt is a sodium salt.

39. The method of any one of claims 1 to 38, wherein the compound of formula (I) or a pharmaceutically acceptable salt thereof is administered in a pharmaceutical composition.

40. The method of claim 39, wherein the pharmaceutical composition is a tablet or capsule.

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