Ultra-sensitive detection and quantification of HPV DNA in the plasma of patients with oropharyngeal squamous cell carcinoma (OPSCC) enrolled in the OPTIMA 2 treatment de-escalation trial.
Authors: Hillary Sloane, Evgeny Izumchenko, Austin Mattox, Rifat Hasina, Aashay Patel, Frederick Jones, Hannah Quinn, Kirsten Keyser, Johannes Fredebohm, Frank Holtrup, Alexander T. Pearson, Everett E. Vokes, Tanguy Y. Seiwert, Daniel L. Edelstein, Ari Rosenberg, Nishant Agrawal
Journal: Journal of Clinical Oncology (2021)

6048 Background: Human papillomavirus (HPV) infection is a primary factor driving the increasing incidence of OPSCC. As patients with HPV+ OPSCC show significantly improved treatment response and prognosis, there is an urgent need to de-escalate treatment of HPV+ OPSCC that optimizes oncologic control while minimizing treatment-related toxicity. Cell-free HPV DNA (cfHPV-DNA) from plasma specimens represents a promising noninvasive surrogate of disease burden in these patients. To enable cfHPV-DNA analysis as a strategy to monitor response to therapy and guide treatment de-escalation, we developed a highly sensitive assay for HPV16/18 detection and quantification in plasma, based on the SafeSEQ next-generation sequencing (NGS) technology. Methods: Longitudinal plasma samples were collected from patients with locoregional HPV+ OPSCC treated on our institutional de-escalation protocol of induction chemoimmunotherapy followed by risk/response stratified de-escalated locoregional therapy, OPTIMA 2 (NCT03107182). Neck CT or MRI was obtained for all patients at baseline and following induction chemoimmunotherapy; radiographic response to induction therapy was assessed per RECIST 1.1 criteria. cfHPV-DNA was quantified in plasma samples collected at baseline and at the end of induction therapy. Changes in cfHPV-DNA levels were correlated with radiographic response. Results: The SafeSEQ HPV assay demonstrates high analytical sensitivity, with ability to detect a single copy of HPV DNA. Replicate testing of contrived samples containing HPV 16/18 DNA at defined levels revealed robust quantitative detection across a dynamic range over 5 orders of magnitude. The assay showed a low level of background signal ( &lt; 0.04 copies per sample) across 20 healthy donor samples, indicating high specificity. In plasma samples collected at baseline from patients enrolled in OPTIMA 2, cfHPV-DNA was detected at levels ranging from 1 to &gt; 30,000 copies/ml. A high correlation was observed between dynamic changes in patients’ cfHPV-DNA levels and radiographic responses following induction therapy. Furthermore, in samples collected longitudinally during induction therapy, changes in cfHPV-DNA levels accurately tracked radiographic responses to therapy. Conclusions: We have developed a highly sensitive and specific cfHPV-DNA detection assay based on SafeSEQ NGS technology and have successfully applied it to monitor therapeutic response in HPV+ OPSCC patients. The assay exhibits robust quantitative detection of HPV across a broad range of levels, even when only a few copies are present, enabling high-resolution molecular monitoring. Prospective studies are underway to further evaluate the kinetics of cfHPV-DNA as a predictor of response to therapy in order to more precisely guide the management of patients with HPV+ OPSCC.