Abstract 3366: Epithelial to mesenchymal transition mediates sensitivity to viral oncolysis in vitro and in a xenograft flank tumor model of non small cell lung cancer

Abstract

Abstract Background: Epithelial to mesenchymal transition (EMT) has been implicated in treatment resistance and can be characterized by the loss of epithelial cell markers such as E-cadherin and gain of mesenchymal markers such as vimentin. We hypothesized that EMT may play a role in resistance to viral oncolysis in non-small cell lung cancer (NSCLC). Methods: Multiple human NSCLC cell lines obtained from the American Tissue Culture Collection (H292, H1437, H258, H441, H1975, H1299, H1703, SW1573, H2228, A549) were evaluated for expression of E-cadherin, Vimentin and other markers via RT-PCR and Western blot. Infectability of all NSCLC cell lines was evaluated by fluorescence-activating cell sorting (FACS) following infection with a virus encoding GFP. Cell surface levels of Nectin-1 and Herpesvirus Entry Mediator (HVEM) were measured by FACS. Antibody neutralization assays were performed to elucidate viral entry mediators in NSCLC. A murine xenograft flank tumor model was created using candidate cell lines from each cellular phenotype. Tumors were analyzed for EMT protein and transcript markers. Following treatment with an oncolytic Herpes virus (hrR3), xenograft flank tumors were evaluated for tumor volume, viral distribution, and single step burst assay. Results: Western Blots on cell lysates revealed the presence of E-cadherin in H292, H1437, H358, H441, H1975. The cell lines SW1573, H1703, H1299, and H2228 lacked E-cadherin and expressed higher protein levels of Vimentin, and significantly higher levels of Zeb-1 mRNA suggesting a mesenchymal phenotype. Cell lines harboring epithelial markers were more were more infectable using the GFP-encoded virus (50% vs 32.9%, p=0.031) and more susceptible to viral oncolysis at low multiplicity of infection (MOI) compared to those with mesenchymal markers (55.5 vs 37.5% cell death, p=0.042). Nectin-1 cell surface expression was more abundant in epithelial phenotypes, whereas HVEM showed no differential expression across cell phenotypes. Forty-eight hours following treatment with hrR3, epithelial cell tumors (H292) exhibited significantly more viral staining (29.2% vs 2.6%, p=0.02) and a higher burst size (1.9×106 vs 5.4×104 pfu/mg of tissue, p=0.0013) compared to mesenchymal cell tumors (SW1573). When followed for tumor volume, the epithelial phenotype tumors responded earlier to viral oncolysis compared to mesenchymal tumors, but ultimately, tumor reduction was similar. Conclusion: Infectivity and viral oncolysis of NSCLC cell lines appears to be related to EMT. Nectin-1 appears to be the major receptor for viral infectivity in NSCLC. The mesenchymal phenotype shows less infectivity and reduced viral propagation in vivo, but flank tumor burden after treatment was similar to the epithelial phenotype. Future studies will examine mesenchymal to epithelial transition and its affect on viral oncolysis. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3366. doi:10.1158/1538-7445.AM2011-3366