Abstract 4882: Multi-omics analysis of mechanisms underlying sensitivity of Merkel cell carcinoma to pyrvinium pamoate

Abstract

Abstract Background: Merkel Cell Carcinoma (MCC) is a highly aggressive neuroendocrine cutaneous malignancy arising from either Ultraviolet-induced mutagenesis or Merkel cell polyomavirus (MCPyV) integration. Pyrvinium Pamoate (PP), an FDA-approved anthelminthic drug, has been reported to exhibit anti-tumor abilities in multiple cancers. By integrating transcriptomic, network, and molecular analyses, we discovered that PP is broadly effective against Merkel cell carcinoma cell lines, and that PP targets multiple proposed MCC tumorigenesis pathways. Methods: We performed time-series RNA-seq profiling on a MCPyV inducible cell model and on PP-treated MCC cell lines, and inferred gene regulatory networks associated with PP response. We then integrated the network models with publicly available MCC tumor profiles and the LINCS L1000 database to explore the targets of PP in MCC. In addition, we performed protein, RNA, growth rate, metabolic, and apoptosis assays on multiple MCC cell lines exposed to PP. Results: Analyzing the effect of PP in the L1000 cancer cell line panel suggested that PP could inhibit key drivers of MCC tumorigenesis. We therefore tested the anti-proliferation and pro-apoptosis effects of PP in MCC cell lines. PP effectively inhibits proliferation of MCC at concentrations as low as 100 nM and in a dose and time-dependent manner. RNA-seq analysis of PP treated MCC cells suggested that PP triggers the p53-mediated intrinsic apoptotic signaling pathway, impairs the mitochondria’s oxidative phosphorylation function and induces endoplasmic reticulum (ER) stress. Immunoblotting intrinsic apoptotic pathway protein markers and measuring the IC50 of PP and the MDM2 inhibitor Nutlin-3a in p53WT and p53mut or p53−/− MCC cell lines, we found that PP does not induce cell apoptosis only through p53 activation, but also through other mechanisms such as increasing Unfolded Protein Response (UPR) stress and decreasing expression of the anti-apoptotic protein survivin (BIRC5). Extracellular flux analysis revealed decreased oxidative phosphorylation rate in PP treated MCC cells. Western blot analysis suggests that PP directly inhibits mitochondria complexes genes ATP5A, MTCO1 and NDUF58 and induces UPR-ER stress. PP has also been reported to inhibit the canonical Wnt pathway. We found that WNT5B, the ligand for the non-canonical Wnt pathway, was significantly decreased in MCC tumors and in our MCPyV inducible cell model, and that PP can upregulate WNT5B significantly. MCC cells treated with WNT5B recombinant protein showed a retarded MCC phenotype. Conclusion: Our results suggest that PP exerts anti-tumor activity through both non-specific growth inhibition - via the modulation of mitochondrial function, ER stress, and apoptotic signaling - and more specifically by perturbing the non-canonical Wnt pathway. Citation Format: Jiawen Yang, James T. Lim, Paul Victor, Rick G. Schnellmann, James A. DeCaprio, Megha Padi. Multi-omics analysis of mechanisms underlying sensitivity of Merkel cell carcinoma to pyrvinium pamoate. [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 4882.