| Persistent infection of high-risk human papillomavirus(HPV) is the important risk factor of developing cervical cancer. Hepatitis B virus(HBV) is the major cause of hepatitis and closely associated with liver cancer. From biochemical perspective, cervical cancer and liver cancer are closely related to the infection of virus, expression of viral protein and host immune response. From the aspect of genome, the integration of HPV and HBV had been found in human genome and could trigger genome instability and structural variation. However, the characteristic of HPV and HBV integration were delayed to explore due to lacking high-throughput, accurate and cost-effective detecting technology. Next generation sequencing technology has obvious advantages in these aspects. As a consequence, this technology has gained huge popularity. Without doubt, next generation sequencing is the first-choice tool for detecting characteristic of viral integration. Based on these factors, we developed a novel method for detecting the regularity of HPV and HBV integration by combination of liquid chip capture, next generation sequencing and bioinformatics.Firstly, we obtained the chimeric DNA fragments of human and virus through liquid chip capture technology. Then, we carried on the bioinformatics analysis to detect viral integration in human genome after applying next generation sequencing. We reassessed the HBV integration sites of 28 hepatic carcinoma samples previously assayed by whole genome sequencing(WGS) approach. The obtained breakpoints contained 89% of previous detected HBV integration sites, and many novel sites.We then investigated the characteristic of HBV integration in liver cancer samples. Our data indicated HBV integration was prone to occur in TERT, KMT2 B, CCNA2 genes. Moreover, it was noted that the survival of patients was negatively correlated with HBV integration in TERT. Overall, these results implicated the regularity of HBV integration which may be valuable to consider as novel clinical index. We also detected the HPV integration sites in cervical cancer samples using similar method. HPV integration hotspots on human(FHIT gene) and HPV genome(E1 gene) had been found. We also tried to explore the correlation between HPV integration and protein expression. The result revealed that expression of FHIT gene was affected by HPV integration. In addition, we carried out pathway analysis on genes of HPV and HBV integration. The results revealed that there were significant enrichment in pathway of cancer among these genes which were integrated by HBV or HPV, although there were different breakpoints between HPV and HBV integration. These results suggested that there were close correlation between tumorigenesis and viral integration.In addition, we developed a method which could detect the integration of three kinds of viruses once by improvement on technology of liquid chip capture. Our data showed this method could also detect viral integration sites from cervical exfoliated cells, or cfDNAs of hepatoma carcinoma patients.In summary, we developed a novel method to detect viral integration with low cost, high sensitivity and accuracy. Then, we investigated the regularity of HBV and HPV integration in liver and cervical cancer. Our study provided theoretical and technical bases for the clinical application of viral integration. |
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