The effects of hepatitis B virus integration into the genomes of hepatocellular carcinoma patients

Abstract

Hepatitis B virus (HBV) infection is a leading risk factor for hepatocellular carcinoma (HCC). HBV integration into the host genome has been reported, but its scale, impact and contribution to HCC development is not clear. Here, we sequenced the tumor and nontumor genomes (>803 coverage) and transcriptomes of four HCC patients and identified 255 HBV integration sites. Increased sequencing to 2403 coverage revealed a proportionally higher number of integration sites. Clonal expansion of HBV-integrated hepatocytes was found specifically in tumor samples. We observe a diverse collection of genomic perturbations near viral integration sites, including direct gene disruption, viral promoterdriven human transcription, viral-human transcript fusion, and DNA copy number alteration. Thus, we report the most comprehensive characterization of HBV integration in hepatocellular carcinoma patients. Such widespread random viral integration will likely increase carcinogenic opportunities in HBV-infected individuals.[Supplemental material is available for this article.] . HBV integration into the host genome has been reported both in tumors (Gozuacik et al. 2001;Murakami et al. 2005;Saigo et al. 2008) and in nontumor liver tissue from HBV-infected individuals (Mason et al. 2010), although such integration is not essential for HBV replication. The relative extent, mutation model, and the functional impact of HBV integration in host genomes is not clear due to the lack of an unbiased approach to identify and quantify genome-wide HBV integration sites. Recent advances in sequencing technologies (Meyerson et al. 2010) provide an opportunity to investigate the global extent, mutation model, and functional impact of viral integration in the host genome. Recently, a primary hepatitis C virus-infected HCC patient has been subjected to whole-genome sequencing, and many somatic mutations were reported (Totoki et al. 2011). However, as an RNA virus, HCV never integrates into the host genome during its life cycle; therefore, liver cancer with HCV infection is not an optimal model to study viral-human genomic interactions. To that end, sequencing the genome and transcriptome of an HBV-positive HCC patient provides a great opportunity to reveal the functional impact of viral integration on the host genome.

Genome-wide analysis of HPV integration in human cancers reveals recurrent, focal genomic instability

“…Intriguing recent data showed that hepatitis B virus (HBV) integrants in hepatocellular carcinoma similarly can be associated with regional CNVs, viral-host fusion transcripts and alterations in host genome expression (Jiang et al 2012;Sung et al 2012;Toh et al 2013), analogous to our findings on HPV integrants. However, although CNVs were identified near HBV integrants in ;25% of cases, they and other structural variants were not fully resolved.…”

Targeted sequencing of cancer‑associated genes in hepatocellular carcinoma using next‑generation sequencing

“…Various types of mutation spectrum have been found in various tumor with different rates of transitions and transversions (22). In the present study, C:G>T:A transition (50%) that somatic mutation patterns are significantly distinct from the expected spectrum (24,(26)(27)(28). In HCC, the C:G>T:A transition, T:A>C:G transition and C:G>A:T transversion are common mutations; however, in the present study, the C:G>G:C transversion was observed at a high frequency.…”

C-terminal-truncated hepatitis B virus X protein enhances the development of diethylnitrosamine-induced hepatocellular carcinogenesis

“…Recent studies using next-generation sequencing technologies have highlighted the fact that, following the integration of HBV into the cellular genome, the 39-end of the HBx gene is often deleted and this event is observed more frequently in tumour tissues (Jiang et al, 2012;Sung et al, 2012;Toh et al, 2013). Furthermore, truncated HBx human chimeric transcripts may be seen and expressed as chimeric proteins (Sirma et al, 1999;Toh et al, 2013;Tu et al, 2001).…”

“…These include hot-spot mutations in HBx at aa 130 and 131, which have been associated with the development of HCC (Iavarone et al, 2003;Kuang et al, 2004;Lee et al, 2011). Furthermore, the random integration of HBV DNA into the host genome frequently leads to 39-end-deleted sequences of the HBx gene and the observation of chimeric transcripts which often carry deletions at the 39-end of HBx gene, resulting in C-terminal-truncated HBx protein (Iavarone et al, 2003;Jiang et al, 2012;Ma et al, 2008;Sirma et al, 1999;Sung et al, 2012;Sze et al, 2013;Toh et al, 2013;Tu et al, 2001). Interestingly, a recent study with human HCC samples reports an association between the presence of C-terminal truncation of HBx and venous invasion (Sze et al, 2013).…”