Detection of human papillomavirus type 16 integration in pre-neoplastic cervical lesions and confirmation by DIPS-PCR and sequencing

BackgroundPersistent infections with high-risk types of human papillomavirus (HR-HPV) favour integration of viral DNA into the host cells and are associated with cervical carcinoma. HPV16 is the prevalent HR-type worldwide associated to cervical cancer. Integration of viral DNA promotes a selective cell growth advantage, resulting a risk factor for cancer development.ObjectivesTo test physical status of HPV16 infection in pre-neoplastic cervical lesions using a quantitative real time-PCR (QRT-PCR) based method. To investigate reliability of this method in identification of HPV16 integrated sequences, by detection of integrated papillomavirus sequences (DIPS-PCR) assay and sequencing.Study designOne hundred and seventy HR-HPV positive archival cervical specimens were tested for presence of HPV16 DNA. In HPV16 positive samples, viral load and physical status were evaluated.ResultsHPV16 DNA was detected in 74/170 (43%) HR-HPV positive specimens. In 52/74 a QRT-PCR was performed, and 3 integrated, 13 mixed and 36 episomal forms were detected. Presence of integrated forms was confirmed by DIPS-PCR and sequencing.ConclusionsPresence of HPV integrated forms was detected and confirmed in pre-neoplastic cervical lesions. The QRT-PCR method we used is sensitive and specific for identification of HPV integration in cervical samples, and may be suitable for large scale investigations with prognostic and clinical implications in management of cervical cancer.