A Compound that Inhibits Glycolysis in Prostate Cancer Controls Growth of Advanced Prostate Cancer

Purpose Metastatic castration-resistant prostate cancer remains incurable regardless of recent therapeutic advances. Prostate cancer tumors display highly glycolytic phenotypes as the cancer progresses. Non-specific inhibitors of glycolysis have not been utilized successfully for chemotherapy, because of their penchant to cause systemic toxicity. This study reports the preclinical activity, safety, and pharmacokinetics of a novel small molecule preclinical candidate, BKIDC-1553, with antiglycolytic activity. Experimental design We tested a large battery of prostate cancer cell lines for inhibition of cell proliferation, in vitro. Cell cycle, metabolic and enzymatic assays were used to demonstrate their mechanism of action. A human PDX model implanted in mice and a human organoid were studied for sensitivity to our BKIDC preclinical candidate. A battery of pharmacokinetic experiments, absorption, distribution, metabolism, and excretion experiments, and in vitro and in vivo toxicology experiments were carried out to assess readiness for clinical trials. Results We demonstrate a new class of small molecule inhibitors where antiglycolytic activity in prostate cancer cell lines is mediated through inhibition of hexokinase 2. These compounds display selective growth inhibition across multiple prostate cancer models. We describe a lead BKIDC-1553 that demonstrates promising activity in a preclinical xenograft model of advanced prostate cancer, equivalent to that of enzalutamide. BKIDC-1553 demonstrates safety and pharmacologic properties consistent with a compound that can be taken into human studies with expectations of a good safety margin and predicted dosing for efficacy. Conclusion This work supports testing BKIDC-1553 and its derivatives in clinical trials for patients with advanced prostate cancer. ### Competing Interest Statement SRP is the President, WCVV is the Chief Executive Officer, DJM is the Chief Scientific Officer, and KKO is the Secretary of ProsTech Inc. ProsTech Inc. was created to commercialize therapies for prostate cancer, but as yet, has no commercial claims on BKIDCs or any other potential commercial products. Other authors declare that they have no competing interests. The patent numbers related to this article are as follows. Use Patent in Cancer Field: US 10350211 issued, Composition of Matter Patents: US 10632122 and US 10307425 both issued; and US patent application 17/758,577 pending. None of these four are sublicensed for cancer or producing royalties. * AA : antimycin A ACC : acetyl-CoA Carboxylase ACN : acetonitrile adeno-PCa : adenocarcinoma prostate cancer ADME : adsorption-distribution-metabolism-excretion AMPK : AMP-activated protein kinase AR : androgen receptor ARSi : androgen receptor signaling inhibitors AUC : area under the curve BKI : bumped kinase inhibitor BKIDCs : bumped kinase inhibitor (BKI) derived compounds BSA : bovine serum albumin CAR : cell-associated radioactivity Cl : clearance CMC : Chemistry Manufacturing and Controls CRPC : castration-resistant prostate cancer CYP : cytochrome P450 CytB : cytochalasin B DAPI : 4, 6-diamidino-2-phenylindole DAB : 3,3’-diaminobenzidine 2-DG : 2-deoxyglucose 2-DG-6-P : 2-DG-6-phosphate DHAP : dihydroxyacetone phosphate DPBS : Dulbecco’s phosphate-buffered saline ECAR : extracellular acidification rate FBS : fetal bovine serum FCCP : carbonyl cyanide 4-(trifluoromethoxy)phenylhydrazone FDA : Food and Drug Administration FDG : fluorodoxyglucose GLP : Good Laboratory Practice G6P : glucose-6-phosphate hERG : human ether-a-go-go related gene HK : hexokinase hr : human recombinant IC50 : 50% inhibition values IND : Investigative New Drug IRS-1 : insulin receptor substrate-1 ITS : Insulin-Transferrin-Selenium KO : knock-out KRB : Krebs-Ringer bicarbonate solution LC-MS : liquid chromatography coupled with mass spectrometry MTS : 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt NE : neuroendocrine OCR : oxygen consumption rate Oligo : oligomycin 3-OMG : 3-O-methyl D-glucose OXPHOS : oxidative phosphorylation OXPHOSi : OXPHOS inhibitors Papp : apparent permeability coefficient PARP : poly ADP ribose polymerase PCa : prostate cancer PDL : poly-D-lysine PDX : patient-derived xenograft PEP : phosphoenolpyruvate PG : phosphoglycerate PK : pharmacokinetics PKM2 : pyruvate kinase M2 PP : pyrazolopyrimidine PrP : pyrrolopyrimidine RLU : relative luminescence units RPPA : reverse-phase protein arrays RPS6 : ribosomal S6 protein SCID : severe combined immunodeficient SD : standard deviation SEM : standard error of mean SO : sodium oxamate UPLC-HRMS : ultra-high-performance liquid chromatography-high resolution mass spectrometry V : volume of distribution