HPV Serology and Circulating Viral DNA for Detection, Genotyping, and Measurement of Disease Burden in Oropharyngeal Cancer

The incidence of human papillomavirus-positive (HPV+) oropharyngeal cancer (OPC) has increased rapidly, and HPV early antigen serology has been proposed as a scalable and cost-effective early detection test. HPV seropositivity can precede clinical presentation of OPC by several years, so additional surveillance procedures may be necessary to optimize early cancer detection. The potential for HPV circulating tumor DNA (ctDNA) to confirm a diagnosis of OPC in seropositive individuals is poorly understood. Here, we assess the relationship between HPV serology and HPV ctDNA with disease burden in a large cohort of HPV+ OPC. We analyzed baseline blood samples from 262 patients using a multiplex serology assay and an HPV-targeted deep sequencing assay (HPV-seq), and HPV-seq results were validated by an orthogonal droplet digital PCR assay. Both assays identified HPV16 as the most prevalent genotype (84%), with serology results indicating a total of 4 HPV genotypes and HPV ctDNA results indicating a total of 6 HPV genotypes. HPV ctDNA results demonstrated higher sensitivity and lower cross-reactivity between HPV types compared with HPV serology results. Furthermore, HPV ctDNA but not HPV16 E6 antibody levels were positively associated with disease burden as determined by N-category, overall stage, and by tumor volume (ctDNA vs. volume, r = 0.48 p = 6.4e-12; E6 vs. volume, r = -0.079 p = 0.26). Overall, this is the largest cohort to compare HPV serology and HPV ctDNA results in OPC, and these findings highlight the potential for ctDNA to augment future strategies for blood-based early detection of HPV+ OPC.