HPV-16 E6 mutation and viral integration related host DNA methylation implicate the development and progression of cervical cancer.

Background: HPV-16 infection and viral-host integration are the most important risk factors for cervical cancer (CC). The aim of this study is to develop a new molecular strategy integrated both the viral and host genome variations identifying and monitoring CC.

Method: A total of 312 methylation and 538 RNA-seq datasets were collected from public databases to identify differentially methylated and expressed genes. HPV associated virus integration sites (VISs) were analysed using the ViMIC database. From September 2020 to August 2021, the 70 HPV-16 positive cases retrospectively collected from multi-centre cohorts were subjected to HPV-16 E6 deep sequencing and PCR-based host gene (ASTN1, DLX1, ITGA4, RXFP3, SOX17, ZNF671) methylation detection. RNAseq and expression validation (NNF671) were performed in C-33A cell line harbouring HPV D32E. Lasso and logistic regression algorithm were used to construct the CC diagnostic model.

Results: A positive correlation was observed between the average methylation level of CC patients and their pathological features including tumour stage (p = 0.0077) and HPV subtype (p < 0.001). ZNF671 was identified as a CC-specific methylation marker, with an impressive 93% sensitivity. Both HPV-16 D32E mutation and integration of HPV-16 down-regulated the ZNF671 expression. Finally, a CC diagnostic nomogram was developed by integrating ZNF671 methylation level and HPV E6 mutation feature, yielding an exceptional AUC of 0.997 (95% CI: 0.934-1.000).

Conclusions: Our study demonstrated HPV viral mutations are closely related to host gene epigenetic alterations in CC. Integration of the viral and host genetic information might be a new promising strategy for CC screening.