Diagnostic Accuracy of p16 Immunohistochemistry as a Marker of High-Risk HPV in Invasive Laryngeal Squamous Cell Carcinoma: A Systematic Review.

Background and

Objectives

p16 immunohistochemistry (IHC) is widely used as a surrogate marker for high-risk human papillomavirus (HPV)-driven carcinogenesis in oropharyngeal squamous cell carcinoma. Diagnostic reliability in laryngeal squamous cell carcinoma (LSCC) is still up for debate, particularly because HPV DNA detection, p16 overexpression, and transcriptionally active HPV infection may be discordant at this anatomical site. This systematic review aimed to assess the diagnostic performance of p16 IHC as a surrogate marker for high-risk HPV status in primary invasive LSCC. Materials and

Methods

A systematic review was conducted in accordance with PRISMA principles and prospectively registered in PROSPERO. PubMed, Scopus, and Web of Science, and citation searches were used to identify studies reporting paired p16 IHC and tumour-based molecular HPV testing in invasive LSCC. Eligible HPV reference standards included HPV DNA PCR/genotyping, DNA in situ hybridization, RNA in situ hybridization, and E6/E7 mRNA detection. Data were synthesized narratively because of substantial heterogeneity in p16 thresholds, HPV assays, and study populations.

Results

Fourteen studies were included. p16 positivity thresholds varied widely, ranging from ≥30% moderate/strong staining to ≥70-75% diffuse nuclear and cytoplasmic staining. HPV reference standards also differed substantially across studies. Overall, p16 IHC showed inconsistent concordance with molecular HPV testing. Studies using RNA-based reference standards showed that transcriptionally active HPV was uncommon in LSCC and that p16-positive tumours often lacked evidence of active HPV transcription.

Conclusions

p16 IHC should not be used as a standalone surrogate marker for high-risk HPV-driven carcinogenesis in invasive LSCC. When HPV attribution is clinically or analytically important, p16-positive cases should be confirmed using HPV-specific molecular testing, preferably RNA-based assays. Future prospective studies using standardized p16 protocols and transcriptionally active HPV reference standards are needed.