Metabolic effects of cooper in renal cancer

Clear cell renal cell carcinoma (ccRCC) is a frequent and malignant renal cancer with a glycolytic phenotype due to the loss of VHL tumor suppressor and activation of HIF transcription factors. Up to 50% of patients relapse within five years after surgical resection. Thus, there is an urgent clinical need to understand the molecular mechanisms leading to ccRCC relapse and advancement. Pathogenic mechanisms underlying ccRCC progression represent a key knowledge gap. Our recent discoveries demonstrate a copper accumulation in advanced ccRCC. In this proposal, we interrogate impact for copper-dependent metabolic reprogramming in driving ccRCC progression. Copper (Cu) is a metal cofactor of enzymes, including cytochrome c oxidase complex (CuCOX) essential for mitochondrial respiration, and a metalloallosteric regulator of cell proliferation and survival. Using patients’ primary ccRCCs, we found accumulation of Cu and increased CuCOX strongly correlated with advanced ccRCC and relapse. In cell line xenograft models, functional evidence shows that dietary Cu drives growth of tumors and stimulates formation of CuCOX in cancer cells. We discovered that Cu enhances electron transfer chain (ETC) activity with important functional consequences. High Cu induces (i) assembly of the respiratory supercomplex (RSC) associated with regulatory subunit, COX7A2L, and (ii) accumulation and remodeling of cardiolipins (CLs), phospholipids of the inner mitochondrial membrane necessary for ETC activity. Surprisingly, despite glycolytic phenotype, Cu-reprogrammed cells recover mitochondrial respiration, become hyperdependent on glucose and on the activity of CuCOX, and therefore are highly sensitive to CuCOX inhibitors, including hydrogen sulfide (H2S). Importantly, we discovered that the endocytic process of macropinocytosis is responsible for major proportion of Cu uptake by renal cancer cells. Small molecules that function as donors of H2S are in preclinical trials, while inhibitors of MP are developed for pharmacotherapies, prompting investigation of these pathways as targets for therapeutic interventions. We propose that chronic accumulation of Cu in RCC cells promotes glucose oxidation via TCA cycle and ETC activity causing glucose auxotrophy and stimulating bioenergy and biosynthesis required for tumor growth. To determine the source of Cu, we will investigate macropinocytosis as little understood but major mechanism of Cu uptake and its role in allocation of Cu to CuCOX (Aim 1). To understand Cu-dependent regulation of ETC activity, we will investigate mechanisms of RSC formation, cardiolipin synthesis and the flux of glucose carbon (Aim 2). Finally, because Cu-reprogramming creates new functional vulnerability to CuCOX inhibitors, we will investigate effects of H2S in cell death and tumor formation (Aim 3). We will use several model systems, including RCC cells, ex-vivo tumor fragments and patient derived tumor grafts. The study will identify basic mechanisms by which high level of Cu in cancer cells regulate their metabolism supporting tumor growth. The research will lead to the development of new biomarkers and therapies for cancer