Sunitinib is one of the most widely used targeted therapeutics for renal cell carcinoma (RCC), but acquired resistance against
targeted therapies remains a major clinical challenge. To dissect mechanisms of acquired resistance and unravel reliable
predictive biomarkers for sunitinib in RCC, we sequenced the exons of 409 tumor-suppressor genes and oncogenes in paired
tumor samples from an RCC patient, obtained at baseline and after development of acquired resistance to sunitinib. From
newly arising mutations, we selected, using in silico prediction models, six predicted to be deleterious, located in G6PD,
LRP1B, SETD2, TET2, SYNE1, and DCC. Consistently, immunoblotting analysis of lysates derived from sunitinib-desensitized
RCC cells and their parental counterparts showed marked differences in the levels and expression pattern of the proteins
encoded by these genes. Our further analysis demonstrates essential roles for these proteins in mediating sunitinib
cytotoxicity and shows that their loss of function renders tumor cells resistant to sunitinib in vitro and in vivo. Finally, sunitinib
resistance induced by continuous exposure or by inhibition of the six proteins was overcome by treatment with cabozantinib or
a low-dose combination of lenvatinib and everolimus. Collectively, our results unravel novel markers of acquired resistance to
sunitinib and clinically relevant approaches for overcoming this resistance in RCC