Longitudinal measurement of HPV copy number in cell-free DNA is associated with patient outcomes in HPV-positive oropharyngeal cancer

Sophie J Warlow, Martyna Adamowicz, John P Thomson, Robert A Wescott, Christelle Robert, Lara M Carey, Helen Thain, Kate Cuschieri, Lucy Q Li, Brendan Conn, Ashley Hay, Iain J Nixon*, Timothy J Aitman*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


INTRODUCTION: Oropharyngeal squamous cell carcinoma (OPSCC) is increasing in global prevalence and is divided into two types dependent on association with human papillomavirus (HPV). Assay of HPV copy number in plasma cell-free DNA (cfDNA) provides a minimally invasive method for detecting and monitoring tumour-derived HPV, with potential for enhancing clinical care.

MATERIALS AND METHODS: In a prospectively recruited cohort of 104 OPSCC patients, we evaluate the utility of cfDNA droplet digital PCR (ddPCR) as a method for characterisation and longitudinal monitoring of patients with OPSCC.

RESULTS: ddPCR assay of pre-treatment plasma cfDNA for five HPV types showed overall 95% concordance with p16 immunohistochemistry and PCR analysis of tumour tissue. Longitudinal sampling in 48 HPV+ve patients, with median follow-up of 20 months, was strongly associated with patient outcomes. Persistently elevated cfDNA-HPV post-treatment was associated with treatment failure (2/2 patients) and an increase of cfDNA-HPV in patients whose HPV levels were initially undetectable post-treatment was associated with disease recurrence (5/6 patients). No recurrence was observed in patients in whom cfDNA-HPV was undetectable in all post-treatment samples. In two patients, sequential HPV measurement could have avoided surgical intervention which did not confirm recurrence.

CONCLUSION: The high concordance of pre-treatment plasma cfDNA-HPV analysis with tissue-based assays, together with the clinical associations of sequentially measured post-treatment cfDNA-HPV copy number add to a growing body of evidence that suggest utility of cfDNA-HPV ddPCR in management of OPSCC. Standardised clinical trials based on these data are now needed to assess the impact of such testing on overall patient outcomes.

Original languageEnglish
Pages (from-to)1224-1234
Number of pages11
JournalEuropean Journal of Surgical Oncology (EJSO)
Issue number6
Early online date6 Apr 2022
Publication statusPublished - 1 Jun 2022


  • oropharyngeal squamous cell carcinoma
  • human papillomavirus
  • liquid biopsy
  • cell-free DNA
  • droplet digital PCR


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