Abstract 935: HPV16 E6 splicing induces partial epithelial-mesenchymal transition in oropharyngeal squamous cell carcinoma

Abstract The incidence of HPV16-positive oropharyngeal squamous cell carcinoma (HPV+ OPSCC) is rising at an alarming rate and has overtaken cervical cancer to become the most prevalent HPV+ cancer in the United States. Although most patients with HPV+ OPSCC respond favorably to standard chemoradiation, a substantial proportion of patients have highly aggressive tumors that recur. Biomarkers are needed to reliably stratify patients into risk subgroups. Using multiple complementary models that recapitulate HPV+ OPSCC, our group demonstrated that enhanced splicing of HPV16 E6 oncogene increased migration and invasion. In vivo, HPV+OPSCC with higher spliced E6 (E6*) generated more isolated clusters of tumor cells that spread further away from the main tumor relative to the original full-length isoform (E6FL). This is consistent with an adverse phenotype seen in human HPV+ OPSCC. However, the mechanisms of how spliced E6 enhanced invasion remain unclear.Epithelial-mesenchymal transition (EMT) drives invasion in OPSCC, and is commonly characterized by a loss of epithelial genes and gain in mesenchymal genes. However, recent studies also identified a partial EMT (p-EMT) phenotype, where cancer cells co-express both epithelial and mesenchymal genes. In this study, we explored whether spliced E6 enhances invasion via EMT in OPSCC. Immunofluorescence staining of E-cadherin (epithelial phenotype) and vimentin (mesenchymal phenotype) was performed on formalin-fixed paraffin-embedded tissue of 46 human HPV+ OPSCC. Analysis of the tumor regions on whole-slide images showed that patients with increased E6* relative to E6FL had reduced membrane localization of E-cadherin. In contrast, total expression of E-cadherin and vimentin were unchanged. This finding corroborates with our in vivo chick chorioallantoic membrane model and in vitro studies in OPSCC cell lines, where E-cadherin was more localized at cell-cell junctions in cancer cells with increased E6 splicing. Therefore, we propose that splicing of HPV16 E6 oncogene promotes p-EMT by inhibiting translocation of epithelial E-cadherin from the cell membrane to cytoplasm to reduce cell-cell adhesion. Overall, our findings may shed valuable insights into the mechanisms driving aggressive HPV+ OPSCC and substantiate E6* as a prognostic biomarker to distinguish HPV+ OPSCC risk subgroups for personalized treatment. Citation Format: Yvonne Xinyi Lim, Min Liu, Allison Furgal, Jake McGue, Shiting Li, Bailey Garb, Timothy Frankel, Veera Baladandayuthapani, Laura Rozek, Maureen Sartor, Nisha D'Silva. HPV16 E6 splicing induces partial epithelial-mesenchymal transition in oropharyngeal squamous cell carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 935.