Comprehensive Profiling of Epstein-Barr Virus MicroRNAs in Nasopharyngeal Carcinoma

Department of Pathology, Tufts University School of Medicine, Jaharis Building, Boston, Massachusetts 02111, USA.
Journal of Virology (Impact Factor: 4.44). 12/2008; 83(5):2357-67. DOI: 10.1128/JVI.02104-08
Source: PubMed


Epstein-Barr Virus (EBV) establishes a long-term latent infection and is associated with a number of human malignancies that are thought to arise from deregulation of different stages of the viral life cycle. Recently, a large number of microRNAs (miRNAs) have been described for EBV, and it has been suggested that their expression may vary between the different latency states found in normal and malignant tissue. To date, however, no technique has been utilized to comprehensively and quantitatively test this idea by profiling expression of the EBV miRNAs in primary infected tissues. We describe here a multiplex reverse transcription-PCR assay that allows the profiling of 39 of the 40 known mature EBV miRNAs from as little as 250 ng of RNA. With this approach, we present a comprehensive profile of EBV miRNAs in primary nasopharyngeal carcinoma (NPC) tumors including estimates of miRNA copy number per tumor cell. This is the first comprehensive profiling of EBV miRNAs in any EBV-associated tumor. In contrast to previous suggestions, we show that the BART-derived miRNAs are present in a wide range of copy numbers from < or =10(3) per cell in both primary tumors and the widely used NPC-derived C666-1 cell line. However, we confirm the hypothesis that the BHRF1 miRNAs are not expressed in NPC. Lastly, we demonstrate that EBV miRNA expression in the widely used NPC line C666-1 is, with some caveats, broadly representative of primary NPC tumors.

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    • "In EBV-positive Burkitt lymphoma , the level of miR-BART-6-3p was high, and it affected the regulation of cell growth and immune response (Imig et al. 2011). The levels of miR-BART17-5p, -7, -16, -14*, and -22 were high in nasopharyngeal carcinoma and miR-BART7 might be a potential biomarker for monitoring this cancer type (Cosmopoulos et al. 2009; Zhang et al. 2015). However, EBV-miRNAs in EBV-HLH have never been profiled. "
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    ABSTRACT: Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) is a life-threatening complication of EBV infection. MicroRNAs (miRNAs) were small non-coding RNA, and EBV could encode miRNAs that are involved in the progression of infection. However, the profiles of EBV-miRNAs in EBV-HLH were unknown. Here, we aimed to profile the expression of EBV-miRNAs in children with EBV-HLH by analyzing 44 known EBV-miRNAs, encoded within the BamHI fragment H rightward open reading frame 1 (BHRF1) and the BamHI-A region rightward transcript (BART), in plasma and cellular targets by real-time quantitative PCR. The study included 15 children with EBV-HLH, 15 children with infectious mononucleosis (IM), and 15 healthy controls. CD8+ T cells were found to be the cellular target of EBV infection in EBV-HLH, while CD19+ B cells were infected with EBV in IM. We also found the greater levels of several miRNAs encoded by BART in EBV-HLH, compared to those in IM and healthy controls, whereas the levels of BHRF1 miRNAs were lower than those in IM. The profile and pattern of EBV-miRNAs in EBV-HLH indicated that EBV could display type II latency in EBV-HLH. Importantly, the level of plasma miR-BART16-1 continued decreasing during the whole chemotherapy, suggesting that plasma miR-BART16-1 could be a potential biomarker for monitoring EBV-HLH progression. The pathogenesis of EBV-HLH might be attributed to the abundance of EBV-miRNAs in EBV-HLH. These findings help elucidate the roles of EBV miRNAs in EBV-HLH, enabling the understanding of the basis of this disease and providing clues for its treatment.
    Preview · Article · Oct 2015 · The Tohoku Journal of Experimental Medicine
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    • "There is abundant expression of BARTs and BARF1 mRNA in NPC samples, suggesting their crucial role in the pathogenesis.56,78,79 Multiple EBV-encoded miRNAs have been detected in NPC, encoded in the BART region.80 The virus actively employs its miRNAs to manipulate various viral and cellular functions. "
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    ABSTRACT: Nasopharyngeal carcinoma (NPC) is highly endemic in certain regions including the People's Republic of China and Southeast Asia. Its etiology is unique and multifactorial, involving genetic background, epigenetic, and environment factors, including Epstein-Barr virus (EBV) infection. The presence of EBV in all tumor cells, aberrant pattern of antibodies against EBV antigens in patient sera, and elevated viral DNA in patient circulation as well as nasopharyngeal site underline the role of EBV during NPC development. In NPC tumors, EBV expresses latency type II, where three EBV-encoded proteins, Epstein-Barr nuclear antigen 1, latent membrane protein 1 and 2 (LMP1, 2), are expressed along with BamH1-A rightward reading frame 1, Epstein-Barr virus-encoded small nuclear RNAs, and BamH1-A rightward transcripts. Among all encoded proteins, LMP1 plays a central role in the propagation of NPC. Standard treatment of NPC consists of radiotherapy with or without chemotherapy for early stage, concurrent chemoradiotherapy in locally advanced tumors, and palliative systemic chemotherapy in metastatic disease. However, this standard care has limitations, allowing recurrences and disease progression in a certain proportion of cases. Although the pathophysiological link and molecular process of EBV-induced oncogenesis are not fully understood, therapeutic approaches targeting the virus may increase the cure rate and add clinical benefit. The promising results of early phase clinical trials on EBV-specific immunotherapy, epigenetic therapy, and treatment with viral lytic induction offer new options for treating NPC.
    Full-text · Article · Sep 2014 · Therapeutics and Clinical Risk Management
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    • "EBV-miRNAs are separated into three clusters of the viral genome: BHRF1, and cluster1 and cluster 2 BARTs [13] [14], which are abundantly expressed in EBV-associated tumours and EBV-transformed lymphoblastoid cell lines (LCLs) [13] [15]. Comprehensive studies in vitro indicate that their expression pattern is linked to viral latency stage [13] [14] [16]. Accumulating evidence suggests that miRNAs play important roles in tumourigenesis, modulating apoptosis and host innate defence mechanisms [17] [18]. "
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    ABSTRACT: EBV is a human herpesvirus associated with a number of malignancies. Both lymphoblastoid cell lines (LCLs), and EBV-infected nasopharyngeal carcinoma (NPC) cells have been demonstrated to release exosomes containing the EBV-encoded latent membrane protein 1 (LMP1), and mature micro-RNAs (EBV-miRNAs). Here we analyze the EBV protein and nucleic acid content of exosomes from different EBV-infected cells (LCL, 721 and Daudi) and we show for the first time that exosomes released from LCLs and 721 also contain EBV-encoded latent phase mRNAs. This confirms and strengthens exosomes pathogenetic potential, and might provide insights for development of novel diagnostic and therapeutic strategies.
    Full-text · Article · May 2013 · Cancer letters
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