Plasma human papillomavirus (HPV) DNA as a potential tool for tumor detection and monitoring response in HPV-related oropharyngeal carcinoma (OP).

Abstract

5529 Background: There is a rising incidence of HPV-related OP, which has a distinct prognosis. Previous studies in other viral-related cancers such as nasopharyngeal carcinoma have shown that the level of circulating instigating virus DNA in the blood can be used to monitor treatment response and tumor relapse. The main objective of this study is to develop a similar assay for HPV(+) OP. Methods: We designed a semiquantitative PCR approach to detect circulating HPV DNA using the L1 Gp5/6 degenerate primer as well as HPV16/18 E6 and E7 nested primers. We used this approach to detect HPV DNA in culture media of SCC90, a HPV(+) OP cell line as well as plasma samples of SCC90 bearing SCID mice. Control mice were those implanted with non-HPV tumors. HPV DNA was also assessed in pretreatment plasma samples of 32 HPV(+) OP patients and 4 HPV(-) control patients with other head and neck squamous cell carcinoma. HPV status within the tumor was confirmed by both p16 staining and HPV16/18 PCR (N = 30) or in-situ hybridization (n = 2). Plasma HPV DNA levels were also quantified by a fluorescent probe multiplex real-time PCR (QRTPCR) assay, targeting the HPV16/18 E6 region, in 6 patients with HPV(+) OP and serial plasma samples obtained during radiotherapy (RT). Results: Cell-free HPV DNA was detected in the culture media of SCC90 and the plasma of SCC90 bearing mice but not in control mice. It was detected in pretreatment plasma of 84% HPV(+) patients and in none of the HPV(-) controls using the L1 primers and conventional PCR. However using less sensitive primer sets (nested E6 and E7), plasma HPV DNA was positive in only 50% of patients with detectable E6/E7 DNA in the tumors. QRTPCR was able to detect as low as 10 copies of HPV DNA in each reaction. In 6 patients with serial plasma samples during RT, QRTPCR measurement showed a rapid decline in HPV DNA level, which became undetectable at RT completion. Conclusions: The xenograft studies indicate that plasma HPV DNA is released directly from HPV-related tumors. The high sensitivity of this method allows detection of HPV DNA in plasma even with low viral load. Plasma HPV DNA may be a valuable tool for monitoring therapeutic response and disease progression in HPV(+) OP. No significant financial relationships to disclose.