Abstract 1875: HPV16 detection by PCR in serum of HNSCC patients and comparison with primary tumour tissue
Authors: Vivenza Daniela , Monteverde Martino , Denaro Nerina , Fortunato Mirella , Comino Alberto , Merlano Marco C. , Nigro Cristiana Lo 
Published: May 2015

Abstract
Background: HPV16 in Head and Neck (HN) cancer is a prognostic marker for positive oropharynx tumor (OT) patients. A lot of methods for direct and indirect HPV16 detection in tissue and body fluids have been analysed. Here we sought to determine the presence of HPV16 in serum collected at diagnosis in locally advanced HN tumors and to compare serum with primary tumor.
Aim: Final aim is to establish the prognostic role of genomic HPV16 detection by PCR in serum of OT and in non-oropharynx tumor (non-OT) patients and to compare sensitivity with overall survival (OS).
Methods: We analyzed HPV16 status in 18 OT and 32 non-OT patients (5F/45M; median age 62, range 47-81) by qualitative PCR on genomic DNA extracted from Formalin Fixed Paraffin Embedded tissues and serum samples, using specific primer pairs for E1, E6 and L1 fragments. Pos and neg control cell lines were added at each PCR session and amplicons were visualized on 2% agarose gel. Serum and tissue DNAs were extracted using the QIAmp Blood Midi Kit according to manufacturer's protocol and by proteinase K and phenol, respectively. Overall survival was performed by Kaplan-Meier curves using SPSS version 13.
Results: HPV16 positive data on primary tumors were 44.4% (N= 8) in OT and 3.1% (N= 1) in non-OT for E1, 50% (N= 9) and 21.9% (N= 7) for L1 and 61.2% (N= 11) and 53.1% (N= 17) for E6 fragments, respectively. We determined that in OT patients all the HPV16 negative tissues showed a full concordance with the corresponding serum samples, on the contrary in non-OT patients we found 5 positive serum samples with a correspondent HPV16 negative tissue (7% of discordance). Concordance in HPV16 positive samples (tissue and serum) was higher in OT, with 75% for E1 (6/8), 66.7% for L1 (6/9) and 54.5% for E6 (6/11) fragments, than in non-OT, with 0% (0/1), 14.3% (1/7) and 41.2% (7/17), respectively. Moreover, our previously data performed on 66 OT patients reported that HPV16 pos tissues (13/66) for E1 showed a higher OS than in the negative ones (p = 0.016; median OS = 161.8 in pos vs 15 months in neg).
Conclusions: The high concordance between sera and tissues for E1 could enable its determination on serum as an emerging marker for prognosis in OT patients. Indeed, analysis on additional 100 serum samples is ongoing with the aim of reaching statistical significance in the correlation with OS. Moreover we are collecting sequential sera taken at different times during patients' follow up in order to study HPV16 dynamical changes relevant in HN tumors.
Citation Format: Daniela Vivenza, Martino Monteverde, Nerina Denaro, Mirella Fortunato, Alberto Comino, Marco C. Merlano, Cristiana Lo Nigro. HPV16 detection by PCR in serum of HNSCC patients and comparison with primary tumour tissue. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1875. doi:10.1158/1538-7445.AM2014-1875