Multiplexed in vivo small molecule screening for immediate drug repositioning reveals fluvastatin and pimozide as potential therapeutics in metastatic pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is frequently diagnosed at an advanced stage. Metastasis of PDAC prior to detection as well as its high intrinsic resistance to all currently available therapeutic approaches renders it one of the most-difficult-to-treat cancers. Thus far, conventional chemotherapy is the only treatment option that improves survival in advanced-stage patients. New therapeutic options are therefore urgently needed. However, the development and application of new compounds for treatment can last several years to decades. Hence, testing compounds and drugs that are already approved for treatment in other diseases in drug repositioning approaches provides a promising new avenue to identify novel treatment options without the need to perform expensive and time-consuming drug development steps. To identify new compounds that target metastatic PDAC, a previously developed multiplexed small molecule screening platform that allows investigation of the effect of hundreds of compounds on metastatic seeding of cancer cells in a rapid, unbiased, and quantitative high-throughput manner in vivo was applied. With this platform, both the ICCB library of Known Bioactive Molecules and the FDA library of approved inhibitors were screened in a direct drug repositioning approach. Subsequently, five top candidates that significantly inhibited metastatic seeding of PDAC cells in vivo were identified. Utilization of an in vivo mouse model that interrogates the whole metastatic cascade validated that both fluvastatin and pimozide, two FDA-approved compounds, specifically inhibit metastasis formation of pancreatic cancer cells. These two compounds were therefore chosen for further investigation. Functional in vitro assays revealed that pimozide particularly inhibits the migratory ability of pancreatic cancer cells. Microarray and proteome analyses depicted an upregulation of ubiquitin (UbI) conjugation and mRNA splicing pathways. Target-identification experiments will be necessary to further decipher pimozide’s mechanism of action during the metastatic process in pancreatic cancer. To investigate how statins impact the metastatic process in pancreatic cancer, STRING analysis of microarray, proteome and phospho-proteome was performed. It was shown that statin treatment induces a partial Epithelial-to-mesenchymal transition (EMT)-like cell state via cytoskeletal morpho-dynamic changes and upregulation of Integrin, Rho-GTPases, SRC and TGF-β signaling. Further functional and mechanistic in vitro and in vivo analyses revealed that cells that are unable to undergo this phenotypic shift become apoptotic, but cells that upregulate ERK signaling escapes the induction of apoptosis. The new partial EMT-like cell state induced by statin treatment initially increases their ability to migrate and resist anoikis. However, cancer cells are trapped in this EMT-like cell state that blocks their cellular plasticity and prohibits their return to an epithelial cell state, which results in reduced metastasis formation in vivo. Taken together, this study emphasizes the application of drug repositioning approaches as a promising avenue to identify novel compounds for treating advanced-stage PDAC and the potential of continuous statin treatment as anti-metastatic therapy in cancer patients