Introduction: Renal oncocytoma (RO) is a benign tumor found in up to 25% of resected small renal masses. Identification prior to surgery remains a diagnostic challenge and there are limited methods for pre-operative identification. Although recurring mutations have been identified in genes associated with complex I (NADH Dehydrogenase) of the electron transport chain in RO, the functional significance is unknown due to the lack of available models. Here we aim to use a patient-derived cell line model to investigate how complex I mutations affect mitochondrial function and respiration, hoping that improved characterization could lead to methods of detection of this tumor. Methods: RO and normal kidney (NK) specimens from patients undergoing surgical resection were obtained. YUOC1 cell line was derived from a 38-year-old female with a 4cm renal mass during a partial nephrectomy which was pathologically confirmed to be RO. Long-range PCR was performed to sequence tumor and blood mitochondrial DNA (mtDNA). MtDNA content was measured by real-time PCR. Protein levels of Complex I to IV were analyzed using Western Blot. Mitochondrial morphology was examined using electron microscopy. Cellular oxygen consumption rate and extracellular acidification rate were measured by Seahorse XF96 analyzer. Normal kidney cells, YUNK1, were treated with a complex I inhibitor to model Complex I loss. Results: We successfully established a patient-derived RO cell line (YUOC1), which harbored a somatic ND5 mutation (c89_90insA, N30X) with near homoplasmy (>80% mutation frequency). This led to a loss of ND5 expression, loss of complex I activity, and an increased mtDNA content. Cellular respiration of YUOC1 demonstrated near loss of oxidative phosphorylation and reliance on aerobic glycolysis. Supplementation of succinate for complex II was able to increase oxidative phosphorylation, indicating the rest of the cellular respiration chain was relatively intact. Complex I inhibition with rotenone in a normal kidney cell line (YUNK1) recapitulated an upregulation of mitochondrial biogenesis, indicating complex I inhibition is sufficient to cause changes in mitochondrial phenotype in ROs. Conclusions: This study helps to characterize some of the metabolic changes associated with RO complex I loss which leads to mitochondrial dysfunction and altered cellular respiration. Further development of models will allow future strategies aimed at identification of nutrient dependencies and metabolic imaging to limit surgical resection of small asymptomatic ROs. SOURCE OF Funding: AUA Research Scholar Award
