An Accessible Approach to Molecular and Biomarker Testing for Risk Stratification in Oral Cavity Squamous Cell Carcinoma

INTRODUCTION: Oral cavity squamous cell carcinoma (OCSCC) is a common epithelial malignancy for which site-specific biomarkers with proven prognostic utility remain limited. In this study, we present one of the largest single-center, site-specific molecular analyses of OCSCC to date. Using two separate versions of multigene next-generation sequencing (NGS), we sought to identify a focused set of clinically actionable and widely accessible biomarkers that could support diagnosis, prognostication, and therapeutic decision-making. Our findings highlight the potential of a limited molecular panel to capture key prognostic information, supporting a more accessible and scalable approach to precision oncology in OCSCC. PATIENTS AND METHODS: Seventy-three adult patients (ages 19-85 years) with site-specific OCSCC treated at City of Hope between 2019 and 2023 were included. Tissue specimens were collected at initial diagnosis or recurrence and analyzed using NGS panels for both DNA and RNA. Sequencing identified single nucleotide variants, copy number alterations, and gene fusions. Associations between genomic alterations and overall survival (OS) were assessed using Kaplan-Meier curves and log-rank tests with significance set at p < 0.05. RESULTS: TP53 and TERT-promoter oncogenic variants were the most frequent alterations, co-occurring in 64.4% of cases regardless of disease stage (primary vs. recurrence). Pathogenic variants in CDKN2A, HRAS, PIK3CA, BRCA1, and CASP8 were enriched among patients who developed recurrence. In this group, copy number amplification of CCND1, FGF3, FGF4, FGF19, RPS6KA4, RPS6KB2, and MYC, along with copy number loss of CDKN2A and CDKN2B, were observed. Lower PD-L1 expression, absence of TERT-promoter mutations (PMs), and focal amplification of 11q13 were associated with poor overall survival in univariate analysis (all p < 0.05). CONCLUSIONS: Multigene NGS profiling revealed recurrent genomic alterations and prognostic biomarkers in OCSCC. Based on these findings, we propose an initial biomarker panel comprising TP53 and PD-L1 (22C3 clone) immunostains, along with TP53 and TERT PM analysis. Collectively, these assays capture key prognostic biomarkers associated with adverse outcomes, including low PD-L1 expression (CPS < 20) and the absence of TERT PMs in relation to overall survival. In addition, copy-number assessment of 11q13, MYC (8q), and CDKN2A (9p) may warrant further investigation for future clinical application.