Infectious agents and cancer: A look into EBV pathways involved in the transition from infection to lymphomagenesis

Epidemiological and biological studies have conclusively proved that infectious agents are among the main causes of cancer worldwide. Approximately 18% of all human cancer have been linked to persistent infections from RNA or DNA viruses that include Epstein-Barr virus (EBV), human papilloma virus and Human T-lymphotropic virus type 1. Although each virus has its own specific mechanism for promoting carcinogenesis, the most common outcome for virus-induced reprogramming is genomic instability, including accumulation of mutations, aberrations and DNA damage. The progression to cancer as result of infection is usually a rare event and when it occurs it requires years to decades from the initial infection to tumour development. EBV, classified as class I carcinogen by WHO for its ability of transforming B-cell and functioning as oncogenic factor, is one of the many factors that can be linked to human malignancies. It is estimated that it accounts for more than 200,000 cases of cancer each year and that 1.8% of all cancer deaths are due to EBV. Here, we described several molecular mechanisms underlying the virus-induced carcinogenesis, expecially in Burkitt’s and Plasmablastic lymphoma. We performed RNA-Seq on 20 eBL cases from Uganda and we showed that the mutational and viral landscape of eBL is more complex than previously reported. First, we found the presence of other herpesviridae family members in 8 cases (40%), in particular human herpesvirus 5 and human herpesvirus 8 and confirmed their presence by immunohistochemistry in the adjacent non-neoplastic tissue. Second, we identified a distinct latency program in EBV involving lytic genes in association with TCF3 activity. Third, by comparing the eBL mutational landscape with published data on sporadic Burkitt lymphoma (sBL), we detected lower frequencies of mutations in MYC, ID3, TCF3 and TP53, and a higher frequency of mutation in ARID1A in eBL samples. Recurrent mutations in two genes not previously associated with eBL were identified in 20% of tumors: RHOA and cyclin F (CCNF). We also observed that polyviral samples showed lower numbers of somatic mutations in common altered genes in comparison to sBL specimens, suggesting dual mechanisms of transformation, mutation versus virus driven in sBL and eBL respectively. We also identified a subset of cellular and viral microRNAs differentially expressed between Epstein-Barr-positive and Epstein-Barr-negative Burkitt lymphoma cases. Of these, we characterized the effects of viral BART6-3p on regulation of cellular genes. In particular, we analyzed the IL-6 receptor genes (IL-6Rα and IL-6ST), PTEN and WT1 expression for their possible relevance to Burkitt lymphoma. By means of immunohistochemistry, we observed a down-regulation of the IL-6 receptor and PTEN specifically in Epstein-Barr-positive Burkitt lymphoma cases, which may result in the impairment of key cellular pathways and may contribute to malignant transformation. On the contrary, no differences were observed between Epstein-Barr-positive and Epstein-Barr-negative Burkitt lymphoma cases for WT1 expression. The oncogenic role of EBV was also investigated in Plasmablastic lymphoma. Our analysis revealed a non-canonical latency program with the partial expression of some proteins characterizing latency II and the activation of an abortive lytic cycle. We also performed microRNAs expression profiling through next generation sequencing to investigate the cellular and viral pattern of Plasmastic lymphoma. We identify a subset of viral miRNAs differentially expressed and showed an important role of EBV-miRNAs-Bart-19 in affecting many cellular pathways including lipid metabolism and cell proliferation. Finally, we considered the fact EBV might have contributed to lymphomagenesis in more samples than those remaining EBV positive by exploting a “hit and run” mechanism. We investigated a total of 10 cases and we found that all the samples (n=6) diagnosed as EBV negative by immunohistochemistry and EBER-ISH demonstrated the presence of EBV-microRNAs and EBV genome. This points at the possibility that EBV might have contributed to lymphomagenesis in all our patients, and propose microRNAs detection as the most specific and sensitive tool to recognize EBV vestiges. Collectively, our preliminary results point at an active role for the Epstein-Barr virus in lymphomagenesis and suggest new possible mechanisms used by the virus in determining dysregulation of the host cell physiology. Moreover, our data would have considerable implications for EBV-related diseases control and development of novel EBV-detection strategies.