Targeting Lactate Transport in Small-Cell Lung Cancer

Abstract

Small cell lung cancer (SCLC) is the most aggressive lung cancer and though many patients respond well to frontline chemotherapy and radiation (RT), most recur quickly and have poor outcomes. Robust glucose uptake and glycolytic flux provides energy and metabolic intermediates needed to sustain rapid tumor growth; consequently, the production of excess lactic acid must be used as fuel or shuttled from the cytoplasm to the extracellular milieu. Monocarboxylate transporters (MCTs) are the primary transporters of lactate and require the co-chaperone protein CD147 for cell surface expression. CD147 aids in organizing diverse multiprotein complexes (e.g. MCTs, heterodimeric amino acid (AA) transporter heavy chain (CD98) and light chain AA transporters). We hypothesize that exploiting SCLC reliance on this multiprotein metabolic complex, particularly lactate transport, will perturb other interconnected metabolic pathways and will provide novel approaches to target SCLC. Human SCLC cell lines were assessed for expression, localization and interactions of metabolic complex proteins (e.g. CD147, MCT1, MCT4, CD98, and xCT) by western blot, confocal microscopy, gradient/subcellular fractionation and proximity ligation assays (PLA). Cells treated with SR-13800, a MCT1-specific inhibitor, were profiled for metabolic alterations using the Seahorse XF analyzer, radioactive lactate and AA transport assays and mass spectrometry. RT-induced DNA damage was quantified by γH2AX staining. Protein expression of MCT1 and MCT4 was assessed in SCLC xenografts in mice and intratumoral levels of SR-13800 was quantified after intraperitoneal (I.P) injection. Finally, tests for synergy of SR-13800 with other metabolic inhibitors were performed ex vivo. SCLCs express varying levels of metabolic complex proteins, with some expressing MCT1 or MCT4 exclusively. CD147, MCT1, CD98 and xCT localized in similar membrane fractions and CD147-MCT1 and CD98-xCT colocalized at the cell surface and were found to interact by PLA. Interestingly, CD147 and MCT1 also localized in mitochondria. SR-13800 treatment of MCT1- vs. MCT4-expressing cells decreased lactate transport and incited a shift to OXPHOS; electron transport chain analysis demonstrated this is primarily due to complex I/II activity. Differences in select metabolites were noted after SR-13800 treatment at 1 and 24 hr. SR-13800 when delivered with 2 Gy RT increased DNA damage. In vivo, MCT1 was highly expressed in the tumor whereas MCT4 was limited to necrotic regions. SR-13800 (30 mg/kg I.P) resulted in an intratumoral concentration of ∼600 nM after 24 hr. Lastly, synergism of SR-13800 with other metabolic inhibitors was observed ex vivo, especially those that disable central carbon metabolism. Disrupting lactate transport incites compensatory metabolic shifts that provide novel therapeutic opportunities when combined with other metabolic pathway inhibitors or standard of care therapies.