Targeting Cancer Metabolism to Enhance Tumor Radiation Response

Abstract

A key emerging hallmark of cancer includes a deregulated cellular metabolism that can be reprogrammed to preferentially exhibit dependence on glycolysis over oxidative phosphorylation (OXPHOS) even in the presence of oxygen (Warburg effect). Pre-clinical data suggests that targeting lactate dehydrogenase (LDH) enzyme and components of mitochondrial OXPHOS metabolism in tumor cells can have a potent anti-tumor effects, yet the interactions of these anti-tumor approaches with standard therapies, such as RT is less clear. clonogenic assay, immunoblotting, tumor regrowth delay, Seahorse studies, LC/MS Mass spectrometry We characterized novel and specific inhibitors targeting LDH enzyme A and B isoform (LDHi) and novel OXPHOS inhibitor (IACS-010759) targeting mitochondrial complex I in combination with ionizing radiation (IR) for their anti-cancer and radio-sensitization effects across various tumor types. Targeting LDH enzyme in vitro leads to significant (p<0.001) reduction in extracellular lactate levels and radiosensitization of variety of tumor cell lines with a dose modification factor (DMF) ranging between 1.5-2.2. Clonogenic survival studies of variety of pancreatic, prostate and lung cancer cells indicated that inhibiting LDH in conjunction with IR, can also enhance radiosensitivity under both hypoxic (1% oxygen) (DMF = 2.0) and normoxic (DMF = 1.65) conditions while not affecting non-glycolytic/normal cells (1522, skin fibroblasts, DMF = 1.0) in vitro. Further, we established that this enhanced radiosensitivity upon LDH inhibition could be partly attributed to reduced DNA repair as seen by enhanced expression of gamma-H2AX (4 fold), reduced ATP generation (80% reduction compared to control) as well as altered expression of glucose and lactate transporter proteins (Glut-1, MCT-1, MCT-4, PFKP and PKM2) as seen by immunoblotting. Tumor regrowth delay studies with MiaPaCa and H460 xenografts demonstrated differential LDH response with no significant tumor radiosensitization. We attributed this failure to tumor metabolic reprogramming confirmed by increased oxygen consumption rate of tumor cells upon LDH inhibition in vitro. Targeting OXPHOS pathway by IACS-010759 in conjunction with LDHi promoted metabolic synthetic lethality in vitro (survival reduced to 10-5). Similar synthetic lethality was also observed in the H460 xenograft; additive enhancement in radiation response. However, inhibition of OXPHOS by complex I inhibitor and fractionated IR led to additive tumor regrowth delay in both MiaPaCa and H460 xenograft models. Furthermore, steady state global metabolomics study of inhibiting LDH and complex I of mitochondria either alone or in combination lead to significant decrease in glycolytic and TCA cycle intermediates leading to severe energy stress. In summary, our results presently indicate that targeting mitochondrial metabolism may be a more effective means of enhancing tumor radiosensitivity.