Recharacterizing the FDA approved drug pyrvinium pamoate as a clinically relevant HuR inhibitor in pancreatic ductal adenocarcinoma

HuR is an RNA binding protein involved in a coordinated cellular survival response to stressors. Upon insults such as chemotherapy, HuR translocates from the nucleus to the cytoplasm where it binds the 3’UTR of target mRNAs. HuR’s interaction with target mRNAs leads to the upregulation of target genes and ultimately treatment resistance. This is particularly relevant in the case of pancreatic ductal adenocarcinoma (PDA). PDA is highly resistant to radiotherapy and standard chemotherapy such as FOLFIRINOX or gemcitabine/nab-paclitaxel. Using a tumor microarray (TMA), we found 79% of patient tumor samples (n=70) were positive for active cytoplasmic HuR, while little to no cytoplasmic localization was detected in normal tissue. In addition, HuR CRISPR knockout cell lines have a xenograft lethal phenotype. The aim of our current study is to target HuR by re-purposing the anti-helminthic, FDA approved small molecule pyrvinium pamoate (PP) to inhibit HuR’s translocation and sensitize PDA cells to concurrent therapies. PP has been shown in bladder cancer to inhibit the translocation of HuR in vitro and in vivo. We have reproduced this in multiple PDA cell lines and have shown impressive drug potency with IC50s as low as 38nM in 2D cultures of PDA cell lines and PDX lines and 16nM in a 3D mouse PDA organoid model. We have demonstrated that inhibition of HuR translocation is likely to occur through secondary effectors AMPK and CDK1. We have also demonstrated that PP’s inhibition of HuR function may be through direct inhibition of target binding. In comparison to other published HuR inhibitors PP inhibits the binding of HuR to targets more potentlt with nanomolar IC50’s. We confirmed this work through HuR RNA Immunoprecipitation experiments and determined that PP inhibited the ability of HuR to bind target mRNA. We generated HuR deficient CRISPR lines to and demonstrated that lack of HuR sensitizes PDA cells to various therapeutics, an effect which is exacerbated in physiologically relevant low glucose settings. We next demonstrated that PP can synergize with several therapeutics including the CDK4/6 inhibitors Abemaciclib and Palbociclib in PDA cells and that this synergy is increased in low glucose setting. This synergistic effect is ameliorated in HuR deficient CRISPR cell lines, indicating that PP achieves this synergistic potential through the inhibition of HuR. We performed targeted phosphoproteomics and found that PP robustly inhibited critical mTOR pathway members as well as validating previous reports that it can inhibit the WNT pathway. Finally, we have demonstrated that PP has a dose dependent effect on PDA tumor growth in vivo with IP and PO dosing regimens. This work supports the recharacterization of PP as a potentially effective therapeutic agent for the treatment of PDA. Early phase clinical trials of PP in human subjects are being planned for 2019. Citation Format: Christopher W. Schultz, Teena Dhir, Samantha Z. Brown, Saswati Chand, Wei Jiang, Grace A. McCarthy, Alex O. Haber, Charles J. Yeo, Austin Goetz, Avinoam Nevler, Oloruntoba Bolaji, Jonathan R. Brody. Recharacterizing the FDA approved drug pyrvinium pamoate as a clinically relevant HuR inhibitor in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3058.