Comparison of molecular assays for HPV testing in oropharyngeal squamous cell carcinomas: A population-based study in Northern Ireland

Stephanie G. Craig, Lesley A. Anderson, Michael Moran, Laura Graham, Keith Currie, Keith Rooney, Max Robinson, Victoria Bingham, Kate S. Cuschieri, Stephen McQuaid, Andrew G. Schache, Terry M. Jones, Dennis McCance, Manuel Salto-Tellez, Simon S. McDade, Jacqueline A. James*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

13 Citations (Scopus)


Background: Determination of human papillomavirus (HPV) status has become clinically relevant for patient stratification under UICC TNM8 staging. Within the United Kingdom, a combination of p16 IHC and HPV DNA-ISH is recommended for classifying HPV status. This study will assess a series of clinically applicable second-line molecular tests to run in combination with p16 IHC to optimally determine HPV status. Methods: The ability of HPV RNA-ISH, HPV DNA-ISH, and HPV DNA-PCR to identify p16-positive/HPV-positive patients was investigated in a population-based oropharyngeal squamous cell carcinoma (OPSCC) cohort of patients diagnosed in Northern Ireland from 2000 to 2011. Results: Only 41% of the Northern Irish OPSCC patient population was associated with HPV-driven carcinogenesis. Both ISH assays were more specific than the DNA-PCR assay (100% and 95% vs. 67%) and were less likely to be affected by preanalytic factors such as increasing block age. A pooled HPV genotype probe for RNA-ISH was found to be the most accurate molecular assay assessed (95% accuracy) when compared with p16 positivity. Conclusions: Our study demonstrates the advantage of tissue-based molecular assays when determining HPV status in retrospective samples. Specifically, we demonstrate the enhanced sensitivity and specificity of ISH techniques compared with PCR-based methodology when working with formalin-fixed paraffin-embedded tissue, and found HPV RNA-ISH to be the most effective assay for determining HPV status. Impact: As p16 IHC is a relatively inexpensive, accessible, and sensitive test for stratifying patients by HPV status, this study finds that more patients would benefit from first-line p16 IHC followed by specific HPV testing using HPV RNA-ISH to confirm HPV status.

Original languageEnglish
Pages (from-to)31-38
Number of pages8
JournalCancer Epidemiology Biomarkers and Prevention
Issue number1
Early online date30 Oct 2019
Publication statusPublished - 9 Jan 2020

Bibliographical note

This work received funding from the Medical Research Council (R1310CNR; to D.
McCance and J.A. James), the Health and Social Care Research and Development
Division of the Northern Ireland Public Health Agency (R4556CNR; to D. McCance, J.A. James, and M. Moran), Cancer Research UK (R2111CNR; to M. Salto-Tellez and J.A. James), the Wellcome Trust through the Wellcome-FDS Research Training Fellowship, the Faculty of Dental Surgery of the Royal College of Surgeons of England (WT093893MA; to A.G. Schache), and GlaxoSmithKline Ltd (to T.M. Jones). The Northern Ireland OPSCC TMAs used in this research were received from the Northern Ireland Biobank, which has received funds from Health and Social Care Research and Development Division of the Public Health Agency in Northern Ireland and the Friends of the Cancer Center. The Northern Ireland Cancer Registry, which receives funding from the Northern Ireland Public Health Agency, carried out collection of clinical data for the Northern Ireland OPSCC patients.


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