Longitudinal liquid biopsy alongside positron emission tomography‐computed tomography surveillance in p16‐positive oropharyngeal squamous cell carcinoma: A translational case study

Abstract Background Post‐treatment surveillance of p16‐positive oropharyngeal squamous cell carcinoma (OPSCC) relies on clinical assessment and fluorodeoxyglucose positron emission tomographycomputed tomography (FDG PET‐CT). Longitudinal liquid biopsy may provide complementary biological information, but the relationship between oral‐rinse human papillomavirus (HPV) DNA, circulating tumour cells (CTCs), and imaging findings remains incompletely defined. Methods A 77‐year‐old man with p16‐positive OPSCC underwent serial liquid‐biopsy sampling within an ongoing prospective cohort. Peripheral blood and oral‐rinse specimens were collected before treatment, at week 4 of chemoradiotherapy, and approximately 13 weeks after treatment completion. CTCs were enriched from 15 mL peripheral blood using marker‐independent spiral microfluidics and characterised by immunofluorescence. Oral‐rinse high‐risk HPV DNA was assessed by multiplex quantitative polymerase chain reaction and reported qualitatively. Results Baseline PET‐CT demonstrated a base‐of‐tongue primary tumour and cervical nodal disease without distant metastases. Oral‐rinse HPV16 DNA was detected, and CTCs were identified at 3 CTCs/mL, predominantly with cytokeratin‐positive phenotypes. During chemoradiotherapy, HPV16 DNA remained detectable and CTCs persisted at 1 CTC/mL, with a shift towards cell‐surface‐vimentin‐positive phenotypes. At post‐treatment assessment, PET‐CT showed complete locoregional metabolic response but identified a new right iliac crest lesion subsequently confirmed as metastatic p16‐positive squamous cell carcinoma. Oral‐rinse HPV16 DNA was no longer detected, whereas CTCs persisted at 4 CTCs/mL, including cell‐surface‐vimentin‐positive and CD44‐positive cells and a CTC‐white‐blood‐cell cluster. The patient subsequently developed widespread osseous progression and died approximately eight months after chemoradiotherapy. Conclusions Oral‐rinse HPV16 DNA clearance coincided with locoregional metabolic response, whereas persistent CTCs coincided with biopsy‐confirmed distant metastasis. This hypothesis‐generating observation supports prospective evaluation of complementary imaging and liquid‐biopsy surveillance but does not establish earlier metastatic detection or discrimination between locoregional and distant recurrence.