ABSTRACT
Due to increased lactate production during glucose metabolism, tumor cells heavily rely on efficient lactate transport to avoid intracellular lactate accumulation and acidification. Monocarboxylate transporter 4 (MCT4/SLC16A3) is a lactate transporter that plays a central role in tumor pH modulation. The discovery and optimization of a novel class of MCT4 inhibitors (hit 9a), identified by a cellular screening in MDA-MB-231, is described. Direct target interaction of the optimized compound 18n with the cytosolic domain of MCT4 was shown after solubilization of the GFP-tagged transporter by fluorescence cross-correlation spectroscopy and microscopic studies. In vitro treatment with 18n resulted in lactate efflux inhibition and reduction of cellular viability in MCT4 high expressing cells. Moreover, pharmacokinetic properties of 18n allowed assessment of lactate modulation and antitumor activity in a mouse tumor model. Thus, 18n represents a valuable tool for investigating selective MCT4 inhibition and its effect on tumor biology.
Subject(s)
Antineoplastic Agents/therapeutic use , Monocarboxylic Acid Transporters/antagonists & inhibitors , Muscle Proteins/antagonists & inhibitors , Neoplasms/drug therapy , Picolinic Acids/therapeutic use , Sulfonamides/therapeutic use , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Drug Screening Assays, Antitumor , Female , HEK293 Cells , Humans , Lactic Acid/metabolism , Mice, Inbred C57BL , Mice, Nude , Mice, SCID , Molecular Structure , Picolinic Acids/chemical synthesis , Picolinic Acids/pharmacology , Structure-Activity Relationship , Sulfonamides/chemical synthesis , Sulfonamides/pharmacology , Xenograft Model Antitumor AssaysABSTRACT
In oncology, the "Warburg effect" describes the elevated production of energy by glycolysis in cancer cells. The ubiquitous and hypoxia-induced 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) plays a noteworthy role in the regulation of glycolysis by producing fructose-2,6-biphosphate (F-2,6-BP), a potent activator of the glycolysis rate-limiting phosphofructokinase PFK-1. Series of amides and sulfonamides derivatives based on a N-aryl 6-aminoquinoxaline scaffold were synthesized and tested for their inhibition of PFKFB3 in vitro in a biochemical assay as well as in HCT116 cells. The carboxamide series displayed satisfactory kinetic solubility and metabolic stability, and within this class, potent lead compounds with low nanomolar activity have been identified with a suitable profile for further in vivo evaluation.
Subject(s)
Amides/chemistry , Phosphofructokinase-2/antagonists & inhibitors , Quinoxalines/chemistry , Quinoxalines/pharmacology , Sulfonamides/chemistry , HCT116 Cells , Humans , Kinetics , SolubilityABSTRACT
Energy and biomass production in cancer cells are largely supported by aerobic glycolysis in what is called the Warburg effect. The process is regulated by key enzymes, among which phosphofructokinase PFK-2 plays a significant role by producing fructose-2,6-biphosphate; the most potent activator of the glycolysis rate-limiting step performed by phosphofructokinase PFK-1. Herein, the synthesis, biological evaluation and structure-activity relationship of novel inhibitors of 6-phosphofructo-2-kinase/fructose-2,6-biphosphataseâ 3 (PFKFB3), which is the ubiquitous and hypoxia-induced isoform of PFK-2, are reported. X-ray crystallography and docking were instrumental in the design and optimisation of a series of N-aryl 6-aminoquinoxalines. The most potent representative, N-(4-methanesulfonylpyridin-3-yl)-8-(3-methyl-1-benzothiophen-5-yl)quinoxalin-6-amine, displayed an IC50 of 14â nm for the target and an IC50 of 0.49â µm for fructose-2,6-biphosphate production in human colon carcinoma HCT116 cells. This work provides a new entry in the field of PFKFB3 inhibitors with potential for development in oncology.
Subject(s)
Drug Discovery , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacology , Phosphofructokinase-2/antagonists & inhibitors , Quinoxalines/chemistry , Quinoxalines/pharmacology , Cell Survival/drug effects , Crystallography, X-Ray , Dose-Response Relationship, Drug , Enzyme Inhibitors/chemical synthesis , HCT116 Cells , Humans , Lactic Acid/antagonists & inhibitors , Lactic Acid/biosynthesis , Models, Molecular , Molecular Structure , Phosphofructokinase-2/metabolism , Quinoxalines/chemical synthesis , Structure-Activity RelationshipABSTRACT
USP2a is a deubiquitinase responsible for stabilization of cyclin D1, a crucial regulator of cell-cycle progression and a proto-oncoprotein overexpressed in numerous cancer types. Here we report that lithocholic acid (LCA) derivatives are inhibitors of USP proteins, including USP2a. The most potent LCA derivative, LCA hydroxyamide (LCAHA), inhibits USP2a, leading to a significant Akt/GSK3ß-independent destabilization of cyclin D1, but does not change the expression of p27. This leads to the defects in cell-cycle progression. As a result, LCAHA inhibits the growth of cyclin D1-expressing, but not cyclin D1-negative cells, independently of the p53 status. We show that LCA derivatives may be considered as future therapeutics for the treatment of cyclin D1-addicted p53-expressing and p53-defective cancer types.
