Cutting cholesterol curbs clear cell RCC

Abstract

0123456789();: Nature reviews | Urology New research suggests that clear cell renal cell carcinoma (ccRCC) cells are dependent on exogenous cholesterol for growth and survival. Also, elevated levels of circulating high-density lipoprotein (HDL) cholesterol increase the risk of developing ccRCC and increased dietary cholesterol intake promotes tumour growth. These observations have implications for targeting cholesterol transporters and for managing circulating cholesterol in people with this disease. “For decades, ccRCC has been diagnosed based on its clear cell phenotype. The tumours exhibit this appearance on histological staining owing to substantial accumulation of cholesterol and cholesterol esters in all cases. In our opinion, the role of the exaggerated cholesterol and cholesterol ester abundance in disease progression has been unclear,” Celeste Simon, corresponding author, tells Nature Reviews Urology. Surprisingly, the researchers observed that gene sets associated with cholesterol metabolism and biosynthesis were significantly downregulated in ccRCC in The Cancer Genome Atlas data subjected to gene-set enrichment analysis, suggesting that intracellular cholesterol metabolism is deregulated in ccRCC tumour cells “making them cholesterol auxotrophs and dependent on a means of cholesterol import,” explains Simon. Two-sample Mendelian randomization of summary association statistics of a genomewide association study (GWAS) including 10,784 people with RCC and 20,406 without showed that the odds of developing RCC were significantly increased when genetic alleles predicted elevated serum levels of HDL or HDL cholesterol. In vivo, mice injected with A498 cells receiving a high-cholesterol diet had larger tumour volumes than those who ate a cholesterol-free diet. To test whether ccRCC cells are dependent on exogenous cholesterol, the investigators cultured A498 and 786-O cells in delipidated serum, resulting in reduced proliferation and cell death. Supplementing the media with cholesterol rescued cell proliferation and viability. In human ccRCC samples, metabolic gene-set analysis showed that the HDL transporter scavenger receptor B1 (SCARB1) is significantly overexpressed, suggesting that ccRCC cells use SCARB1 to import cholesterol. SCARB1 expression was increased at all tumour stages of ccRCC, suggesting that deregulation of cholesterol metabolism is an early event in ccRCC development. In vitro, SCARB1 knockdown in A498 cells using doxycyclineinducible SCARB1 shRNAs or CRISPR–Cas9 technology inhibited HDL import and cell proliferation by inducing apoptosis and cell cycle arrest. Treatment with the small molecule block lipid transporter 1 (BLT1), which is a potent inhibitor of SCARB1-mediated lipid transport, also resulted in reduced proliferation associated with apoptosis and cell cycle arrest. HDL supplementation in all of these treatment conditions failed to rescue these cells. Lipiddeprived cells also exhibited significantly increased reactive oxygen species levels, which returned to baseline levels after HDL supplementation. Depleting SCARB1 in ccRCC cells grown in replete medium elevated reactive oxygen species levels. In vivo, A498 cell xenografts expressing doxycycline-inducible SCARB1 shRNA in mice that were fed a diet containing doxycycline had reduced tumour growth and SCARB1 expression and increased cell death. Treating A498 tumourbearing mice with BLT1 for 21 days resulted in inhibited tumour growth and increased circulating HDL levels. “Intriguingly, GWAS from some time ago indicated that SNPs in SCARB1 actually predispose individuals to developing renal cancer,” explains Simon, continuing “SCARB1 inhibitors already exist and have been investigated in clinical trials, and could be repurposed for treating kidney cancer.” Mechanistically, in cells in which cholesterol uptake is reduced, AKT phosphorylation is also reduced. Inhibition of AKT and PI3K signalling also resulted in reduced ccRCC cell proliferation, which was not reversed by HDL supplementation. Treating ccRCC cells with edelfosine, an alkyl-lysophospholipid cholesterol analogue that accumulates in lipid rafts and alters cellular signalling, decreased AKT phosphorylation and cell growth and increased apoptosis. Taken together, these findings show the dependence of ccRCC on exogenous cholesterol and reveal a mechanism by which cholesterol is imported into ccRCC cells. Importantly, this research shows the influence of circulating HDL cholesterol on ccRCC growth and patient risk and provides a new potential drug target for treatment of this disease. “Future studies will involve investigating the effects of SCARB1 inhibitors in a variety of preclinical models of renal cancer and then hopefully deploying them in the clinic,” concludes Simon.