Exposing synthetic lethal vulnerabilities in EBV-positive AIDS-NHL through novel replication dependency factors

项目总结

PROJECT SUMMARY Virus-associated lymphomas cause significant morbidity and mortality in HIV-infected individuals – indeed, the oral pathogen Epstein-Barr virus (EBV) contributes to up to 90% of diffuse large B-cell lymphomas (DLBCL) and 40% of Burkitt lymphomas (BL). Although combined antiretroviral therapy (cART) and chemotherapy have improved outcomes for AIDS lymphomas, challenges remain particularly with virus-associated AIDS lymphomas, prompting efforts to better understand virus-associated factors and pathways. In particular, EBV-driven cellular genome replication which is essential to lymphoma proliferation remains underexplored. Upon infection of B cells, EBV drives host DNA replication which is essential for establishment of viral latency as well as proliferation of cancer cells. However, such viral oncoprotein-driven DNA replication is plagued with physical and functional obstacles, resulting in replication stress. Such replication stress is a barrier to cancer. And yet, how EBV-cancer cells overcome such stress at replication forks to successfully proliferate is not well understood. In addressing this knowledge gap, we combined isolation of proteins on nascent DNA (iPOND) and mass spectrometry to discover novel fork proteins. This revealed a critical role for ZC3H18 (or ZC3) as a replication dependency factor that EBV upregulates to ensure host genome replication and lymphoma cell proliferation; notably, ZC3 had not been previously linked to DNA replication. Indeed, EBV+ DLBCL from AIDS patients have elevated ZC3 expression compared to EBV- lymphomas. An intrinsically disordered protein, ZC3 has the potential to concentrate a variety of proteins at replication forks. We find a direct interaction between ZC3 and MCM7 (a core component of the replicative helicase complex), further pointing to ZC3’s influential role in proliferation of EBV transformed cells. Importantly, ZC3’s partnership with other replication dependency factors exposes EBV-lymphoma cells to synthetic lethality – such therapies exploit the property that cancer cells tolerate perturbation of a single gene but succumb to co-disruption of multiple genetic events. In this application, we will test the hypothesis that EBV modulates the DNA replication machinery, ensuring proliferation of transformed cells in the face of replication stress and enhancing the potential for susceptibility to synthetic lethality. We will perform the following aims using ex vivo models and translate our results to patient- derived EBV+ AIDS lymphomas obtained from the AIDS and Cancer Specimen Resource (ACSR). Aim 1. Investigate how novel dependency factors unmask synthetic lethal vulnerabilities in EBV- transformed cells & Aim 2. Investigate mechanisms of ZC3 upregulation, replication machinery rewiring, and contribution of replication dependency factors to EBV+ AIDS lymphomas. These studies will identify mechanisms and generate new paradigms that reveal how an opportunistic virus modulates the host replication machinery to maintain the transformed state. Our long-term goal is to identify novel druggable targets that demonstrate synthetic lethality against EBV+ lymphomas in persons with HIV/AIDS.