High-Throughput Genotyping of Oncogenic Human Papilloma Viruses with MALDI-TOF Mass Spectrometry (Molecular Diagnostics and Genetics‪)‬

Infection with oncogenic types of human papillomavirus (HPV) [4] is the principal cause of invasive cervical cancer and its precursor lesion, cervical intraepithelial neoplasia (CIN) (1, 2); HPV DNA is found in almost all cervical cancers (3). HPV types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 68, 73, and 82 are found in cervical cancer, whereas HPV 6, 11, 40, 42, 43, 44, 54, 61, 70, 72, and 81 are not associated with cervical cancer (4). Genotyping of HPV infections is important, because different HPV genotypes confer distinctly different risks for development of cervical disease (5). The addition of HPV testing to cytology in primary screening produces a higher sensitivity for the detection of CIN than cytology alone (6, 7). A knowledge of the prevalence and associated risks for each specific HPV genotype also has been essential for the development of vaccines against HPV. High-throughput HPV genotyping will be essential not only for cervical-screening programs but also for monitoring the effectiveness of HPV vaccination programs, i. e., for documenting decreasing prevalence of the HPV types for which vaccines are available and possibly for changing the occurrence of HPV types without vaccines via type replacement or cross-protection (8). Although several primarily PCR-based methods have been used to type HPV (9), the massive scale of the monitoring and primary-screening efforts likely to be launched in the near future will require more efficient methods. In the present study, we developed a high-throughput multiplex analysis of 14 distinct HPV high-risk genotypes on the Sequenom MassARRAY matrix-assisted laser desorption/ionization time-offlight (MALDI-TOF) mass spectrometry (MS) system. We compared its performance with that of another method for HPV genotyping, reverse dot blot hybridization (RDBH) of PCR amplimers, as well as with histopathologic diagnoses.