VHL Mutation Drives Human Clear Cell Renal Cell Carcinoma Progression Through PI3K/AKT-Dependent Cholesteryl Ester Accumulation

Abstract

SummaryBackgroundCholesteryl ester (CE) accumulation in intracellular lipid droplets (LDs) is an essential signature of clear cell renal cell carcinoma (ccRCC), but its molecular mechanism and pathological significance remain elusive.MethodsEnabled by the label-free Raman spectromicroscopy, which integrated stimulated Raman scattering microscopy with confocal Raman spectroscopy on the same platform, we quantitatively analyzed LD distribution and composition at the single cell level in intact ccRCC cell and tissue specimensin situwithout any processing or exogenous labeling. Since we found that commonly used ccRCC cell lines actually did not show the CE-rich signature, primary cancer cells were isolated from human tissues to retain the lipid signature of ccRCC with CE level as high as the original tissue, which offers a preferable cell model for the study of cholesterol metabolism in ccRCC.FindingsSurprisingly, our results revealed that CE accumulation was induced by tumor suppressor VHL mutation, the most common mutation of ccRCC. Moreover, VHL mutation was found to promote CE accumulation by upregulating HIFα and subsequent PI3K/AKT/mTOR/SREBPs pathway. Inspiringly, inhibition of cholesterol esterification remarkably suppressed ccRCC aggressiveness through the reduced membrane cholesterol-mediated downregulations of the integrin and MAPK signaling pathways.InterpretationCollectively, our study improves current understanding of the role of CE accumulation in ccRCC and opens up new opportunities for treatment.FundingThis work was supported by National Natural Science Foundation of China (No. 62027824 and No. 91959120 to S. Yue), Fundamental Research Funds for the Central Universities (No. YWF-22-L-547 to S. Yue), PKU-Baidu Fund (No. 2020BD033 to L. Yao), Peking University First Hospital Scientific and Technological Achievement Transformation Incubation Guidance Fund (No.2022CX02 to L. Yao), Beijing Municipal Health Commission (No. 2020-2Z-40713 to L. Yao).Research in contextEvidence before this studyAs the most common and aggressive subtype of renal carcinoma, clear cell renal cell carcinoma (ccRCC) is known to be one of the most LD-rich cancers. Although a couple of studies have been conducted to investigate the mechanism underlying LD accumulation in ccRCC from the perspective of altered fatty acid metabolism, they overlook the fact that the major form of lipid accumulated in LDs of ccRCC is esterified cholesterol but not fatty acid. Since 1987, accumulation of esterified cholesterol, that is cholesteryl ester (CE), has been becoming an essential signature of ccRCC, but its biological mechanism and pathological significance remain elusive.Added value of this studyFirst, our study demonstrates the need for primary cancer cell, rather than cell line, as the suitable cell model to study cholesterol metabolism in ccRCC. In contrast with the high CE level in human ccRCC tissues, the commonly used ccRCC cell lines in literatures did not contain detectable CEs in LDs. This is very likely the culprit for current limited understanding of CE accumulation in ccRCC. Second, we extend the current understanding of cholesterol metabolism in ccRCC. VHL mutation was found to promote CE accumulation by upregulating HIFα and subsequent PI3K/AKT/mTOR/SREBPs pathway. Third, our study discovers cholesterol esterification as a vulnerable metabolic target for ccRCC treatment.Implications of all the available evidenceCollectively, our study improves current understanding of the role of CE accumulation in ccRCC and opens up new opportunities for treatment.