The rising incidence of human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma (OPSCC) necessitates advancements in risk stratification to optimize treatment outcomes and improve the quality of life for patients. Despite its favorable prognosis compared to HPV-negative OPSCC, current clinical staging and biomarkers, such as p16 status, are limited in their ability to distinguish between high- and low-risk patients within HPV-associated OPSCC. This limitation results in the overtreatment of low-risk patients, exposing them to unnecessary toxicity, and the undertreatment of high-risk patients who require more aggressive interventions. This review critically evaluates current stratification methods, including clinical assessments, de-escalation trials, and candidate molecular biomarkers for risk stratification. Emerging approaches such as immune markers, viral genomic integration patterns, and other molecular markers offer promising avenues for enhanced prognostic accuracy. By integrating advanced risk stratification methods, tailored treatment approaches may one day be developed to balance oncologic efficacy with reduced treatment-related morbidity. This review underscores the need for continued research into predictive biomarkers and adaptive treatment strategies to better address the diverse risk profiles of HPV-associated OPSCC patients.

Purpose: Human papillomavirus (HPV) integration (HPVint) is associated with carcinogenesis and tumor progression in HPV-associated cancers, including head and neck squamous cell carcinomas (HNSCC). Although its impact on human DNA has been well characterized, its relationship with clinical outcomes remains unconfirmed. Experimental Design: We analyzed HPVint events from 261 HPV-associated HNSCC bulk and single-cell RNA sequencing (scRNA-seq) samples from five cohorts, including 62 from a new University of Michigan cohort, and DNA HPVint events from 102 HPV(+) HNSCC participants in two of the cohorts. We investigated the consequences of HPVint both with respect to human and HPV gene expression and clinical outcomes (recurrence and overall survival). Results: By leveraging this large meta-cohort of HNSCC, we first reveal an oncogenic gene network based on the recurrent HPVint locations in the human genome and gene expression alterations, highlighting key recurrent and overexpressed genes, including NR4A2, CD274, and CCER1, and genes from the CAMK and KLF families. We then stratify HPVint-positive participants by risk using HPV RNA features, specifically spliced HPV–human fusion transcripts (E1* integration) and HPV gene expression ratios, showing that subsets of participants have worse clinical outcomes based on these two candidate biomarkers. Conclusions: By focusing on RNA instead of DNA, we expand our understanding of the carcinogenic mechanisms of HPVint, in part addressing the conflicting findings of whether HPVint is associated with aggressive phenotypes and worse clinical consequences, and provide potential biomarkers to advance precision oncology in HPV-associated HNSCC. Newly identified genes with recurrent integration events may serve as candidates for targeted therapy.

Supplementary Figures and Table Titles

Purpose:Human papillomavirus (HPV) integration (HPVint) is associated with carcinogenesis and tumor progression in HPV-associated cancers, including head and neck squamous cell carcinomas (HNSCC). Although its impact on human DNA has been well characterized, its relationship with clinical outcomes remains unconfirmed.Experimental Design:We analyzed HPVint events from 261 HPV-associated HNSCC bulk and single-cell RNA sequencing (scRNA-seq) samples from five cohorts, including 62 from a new University of Michigan cohort, and DNA HPVint events from 102 HPV(+) HNSCC participants in two of the cohorts. We investigated the consequences of HPVint both with respect to human and HPV gene expression and clinical outcomes (recurrence and overall survival).Results:By leveraging this large meta-cohort of HNSCC, we first reveal an oncogenic gene network based on the recurrent HPVint locations in the human genome and gene expression alterations, highlighting key recurrent and overexpressed genes, including NR4A2, CD274, and CCER1, and genes from the CAMK and KLF families. We then stratify HPVint-positive participants by risk using HPV RNA features, specifically spliced HPV–human fusion transcripts (E1* integration) and HPV gene expression ratios, showing that subsets of participants have worse clinical outcomes based on these two candidate biomarkers.Conclusions:By focusing on RNA instead of DNA, we expand our understanding of the carcinogenic mechanisms of HPVint, in part addressing the conflicting findings of whether HPVint is associated with aggressive phenotypes and worse clinical consequences, and provide potential biomarkers to advance precision oncology in HPV-associated HNSCC. Newly identified genes with recurrent integration events may serve as candidates for targeted therapy.