Genomic Enrichment and Functional Impact of TP53 and CYLD Alterations in Recurrent and Metastatic HPV-Associated Head and Neck Cancer

PURPOSE: Recurrent and metastatic human papillomavirus (HPV)-associated head and neck squamous cell carcinoma (R/M HPV+ HNSCC) remains largely incurable, with genetic drivers incompletely defined. We profiled the genetic landscape of R/M HPV+ HNSCC and functionally characterized genetic alterations strongly enriched in this cancer type. EXPERIMENTAL DESIGN: We identified genetic alterations uniquely enriched in 159 R/M HPV+ tumors. High-priority alterations were functionally modeled, examining proliferation, clonogenicity, migration/invasion, apoptosis, therapy response, in vivo growth, metastasis, and immune contexture. RESULTS: Compared with HPV+ primary tumors, R/M HPV+ tumors were enriched for TP53 mutations (prespecified FDR threshold met; OR, 6.23; P = 0.02) and were associated with poorer survival. Within R/M disease, cylindromatosis lysine 63 deubiquitinase (CYLD) alterations were specific to HPV+ tumors (21% vs. 0% in HPV-). TP53 mutations were predominantly clonal and associated with whole-genome duplication. Expression of TP53 gain-of-function (GOF) mutants (R175H, G245C, R273C) in HPV+ HNSCC cells increased clonogenic survival, migration/invasion, lung metastatic burden in vivo, and cisplatin IC50, without altering radiation sensitivity. CYLD knockdown accelerated cellular growth yet increased radiosensitivity. Transcriptomic analyses linked CYLD loss to NF-κB/TNFα pathway activation, a T cell-inflamed microenvironment, and checkpoint upregulation. CONCLUSIONS: R/M HPV+ HNSCC is genomically and functionally shaped by two axes with therapeutic implications: TP53 GOF mutations promote metastatic phenotypes and cisplatin resistance, whereas CYLD loss defines an HPV-specific subset with enhanced radiation sensitivity and immune activation. These data support using TP53 and CYLD as predictive biomarkers to guide investigation into precision strategies for systemic therapy choices, p53-targeted/Wee1 strategies, and radiotherapy-immunotherapy combinations in high-risk or R/M HPV+ HNSCC.