Abstract 3020: Periodic fasting and cholesterol biosynthesis inhibitors achieve a synergistic antitumor activity in gastrointestinal cancers

Abstract Periodic fasting enhances the activity of several chemotherapeutics, molecularly-targeted drugs, endocrine agents and immune checkpoint inhibitors. Identifying drugs that are approved for non-oncological conditions, but acquire antitumor properties through fasting could pinpoint new cancer cell liabilities and define new treatment options. By screening over 800 approved drugs in PK9 pancreatic ductal adenocarcinoma (PDAC) cells, we identified several azoles (antifungal agents), including clotrimazole (CTZ), as agents whose cytotoxic activity against cancer cells is synergistically enhanced by starvation conditions. Since azoles inhibit 14alpha-demethylase, which is a key enzyme for cholesterol (CE) biosynthesis, we hypothesized that starvation and azoles would cooperate by blunting CE production in PDAC cells. Consistent with this notion, we found that simvastatin (an HMG-CoA reductase inhibitor) and terbinafine (TRB), which obstruct squalene epoxidase (SQLE), another key enzyme from the CE biosynthetic pathway, also had their antitumor effects strongly enhanced by starvation. Combined starvation and CTZ or starvation and TRB reduced intracellular CE in Capan-1 cells both in vitro and in vivo. Methyl-beta-cyclodextrin, which depletes intracellular CE, and starvation also showed a synergistic interaction in Capan-1 and in MiaPaCa2 cells (another PDAC cell line). Finally, culture media supplementation with a water-soluble CE formulation prevented the synergistic interaction between starvation and CE biosynthesis inhibitors. TRB potentiation through starvation was countered through supplementation with insulin, IGF1 and leptin, which are downregulated during fasting. Consistent with CE being an essential constituent of membrane lipid rafts, that harbor the growth- and survival-promoting PI3K/AKT signaling cascade, we found combined CTZ or TRB and starvation to markedly downregulate phosphorylated AKT in Capan-1 xenografts. Similar results were obtained in MiaPaCa2 cells. Capan-1 cell transduction with constitutively active, myristoylated AKT protected them from combined CTZ or TRB and starvation, indicating that AKT inhibition mediates the synergistic interaction between CE production inhibitors and fasting. Weekly 48h fasting enhanced CTZ and TRB antitumor effects in vivo in Capan-1 xenograft-bearing mice and, when combined with TRB, lowered circulating LDL (while increasing HDL) cholesterol as compared to TRB alone. Overall, these findings support the potential of fasting-based diets in combination with CE biosynthesis inhibitors against PDAC. Citation Format: Amr Khalifa, Ana Guijarro, Asmaa Namatalla, Moustafa Ghanem, Matteo Lambertini, Alessio Nencioni, Irene Caffa. Periodic fasting and cholesterol biosynthesis inhibitors achieve a synergistic antitumor activity in gastrointestinal cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3020.