Disruption of nucleotide homeostasis confers cancer cell susceptibility to oxidative phosphorylation inhibition independently of energy depletion

Cancer metabolism is highly heterogenous and flexible with the Warburg effect or oxidative phosphorylation (OXPHOS) prevailing in a cancer type- and context-dependent manner. Past studies have demonstrated that targeting OXPHOS robustly inhibits glycolysis-deficient cancer cell viability and tumourigenicity. However, the therapeutic potential of OXPHOS inhibition in metabolically flexible glycolysis-competent cancers is unclear. Furthermore, whether the depletion of OXPHOS-derived ATP or the abolition of OXPHOS-supported biosynthesis is the major determinant of cancer cell susceptibility remains obscure. To address these questions, I exposed a panel of metabolically flexible glycolysis-competent cancer cell lines to OXPHOS inhibitors and tested cell survival and proliferation. Patient-derived xenograft models of colorectal cancer in NSG mice were used for in vivo validation. Here I provide evidence that OXPHOS inhibition potently diminishes metabolically flexible glycolysis-competent cancer cell proliferation and tumourigenicity without causing devastating energy stress. The inhibition of cell proliferation by OXPHOS inhibitors is associated with S-phase cell cycle arrest and the enrichment of the G2/M DNA-damage checkpoint regulation pathway, suggestive of replication stress. Indeed, IACS-010759 (IACS) treatment significantly reduces the purine/pyrimidine nucleotide pools, which is primarily caused by aspartate deficiency resulting from a shortage in the electron acceptor NAD+. The supplementation of exogenous nucleosides, aspartate, or pyruvate that can accept electron generating NAD+, into the culture medium rescues cells from IACS-induced cell cycle arrest. Instructively, inhibition of cytosolic aspartate transaminase (GOT1), which catalyzes cytosolic aspartate biosynthesis when mitochondrial aspartate production is dampened, renders cancer cells more susceptible to OXPHOS inhibition in vitro and in vivo. Collectively, these results indicate that 1) disruption of nucleotide homeostasis is a major determinant of cancer cell susceptibility to OXPHOS inhibition; 2) OXPHOS inhibition is a promising avenue for the treatment of cancers that are metabolically flexible and glycolysis competent; and 3) GOT1 targeting is potentially a useful approach to improve the therapeutic efficacy of OXPHOS inhibition for cancer treatment.