Abstract 1611: Statins inhibit the collaborative metastasis mediated by VHL heterogeneity in clear cell renal cell carcinoma (ccRCC) via a non-cholesterol pathway
Authors: Ishihara Moe , Hu Junhui , Cano-Ruiz Celine , Jackson Maia , Wu Aimee , Conway Stuart , Damoiseaux Robert D. , Wu Lily 
Published: April 2023

Abstract
BACKGROUND: Despite the advancement of targeted therapies and immunotherapies, the outcome of clear cell renal cell carcinoma (ccRCC) remains poor at the metastatic stage, with a median survival of mere 7 months. This devastating outcome is partially due to an inadequate understanding of the metastatic mechanism of ccRCC. Previously, we found that two distinct cell populations co-exist in the malignancy: one with the functional von-Hippel Lindau (VHL) gene (VHL(+)) and the other without (VHL(-)). This heterogeneity in VHL plays an essential role, where the VHL(-) cells serve as the metastatic driver and promote VHL(+) cells to disseminate to distant organs such as the lungs. In this study, we sought to find small molecules that can selectively inhibit the VHL(-) cells' growth and survival. We hypothesize that these molecules will effectively inhibit ccRCC metastases by targeting the metastatic driver population.
METHODS: CRISPR knockout was conducted to generate VHL(-) cells from a murine RCC cell line, RENCA. Both VHL(-) and (+) cells were then fluorescently labeled and went through high-throughput screening (HTS) with 2,530 FDA-approved drugs. We followed a ratiometric black-box approach to identify selective modulators of cell growth and survival of VHL(-) cells without affecting VHL(+) counterparts. CRISPR knockin was conducted on human ccRCC cells, as well, generating VHL(+) and (-) pairs of 786O, RCC4, and a patient-derived primary cell line.
RESULTS: HTS yielded 9 hits that selectively inhibited the survival of the metastatic drivers VHL(-) cells. 4 out of 9 hits belonged to the same drug family called statins. Statins bind to HMG-CoA reductase (HMGCR) and help lower cholesterol, thus are widely prescribed to high-risk patients for cardiovascular diseases. Interestingly, not all statins showed the VHL(-) suppressing effect, resulting in IC50 varying from 0.6uM to &gt;50mM. Atorvastatin and rosuvastatin, the two most potent and prescribed statins for cholesterol-lowering with the highest HMGCR affinity, did not show as much efficacy as other statins. This observation also held true in human ccRCC cell lines, suggesting an involvement of a non-cholesterol/HMGCR pathway in statins’ inhibitory effect on VHL(-) cells. Animal studies were conducted using fluvastatin, which had the lowest IC50 in our model. Intraperitoneal injection of 15mg/kg fluvastatin Q.O.D for 2 weeks successfully suppressed the growth of primary tumors, especially by eliminating VHL(-) cells, and most importantly, prevented lung metastases in the treatment group, whereas all of the control group developed metastases.
CONCLUSIONS: Translational potential of this project is enormous. By investigating the mechanism of action further, we could potentially reposition the FDA-approved, cheap, and widely available small molecules as a novel anti-metastatic therapeutic in ccRCC.
Citation Format: Moe Ishihara, Junhui Hu, Celine Cano-Ruiz, Maia Jackson, Aimee Wu, Stuart Conway, Robert D. Damoiseaux, Lily Wu. Statins inhibit the collaborative metastasis mediated by VHL heterogeneity in clear cell renal cell carcinoma (ccRCC) via a non-cholesterol pathway [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1611.