Comprehensive analyses of therapeutic strategy against chronic active Epstein-Barr virus(EBV) infection

本研究的目的是通过利用显性阴性EBNA 1（DNE 1; Mal. Ther.，11（4）：578 - 90，2005），其可以从细胞中根除EBV附加体。我们通过DNE 1转导制备了EBV阳性和阴性NK和T细胞系的同基因对，并比较了它们的恶性程度。同时，用多细胞因子测定系统全面评估这些细胞对中细胞基因表达的改变，从而阐明EBV在恶性转化中的作用。此外，我们通过使用3种EBV抗原的四聚体测定分析了EBV特异性细胞毒性T细胞（EBV-CTL），即，BRLF1、BMLF1和EBNA3。研究结果如下：1.腺病毒载体介导的DNE 1转导成功地根除了大多数自然EBV阳性T细胞和NK细胞淋巴瘤和上皮肿瘤中的EBV附加体， ...更多信息 r细胞株的生长情况。与亲本EBV阳性T细胞和NK细胞肿瘤相比，DNE 1诱导的EBV缺失的T细胞和NK细胞显示出对其恶性表型的显著抑制，例如倍增时间延长和锚定非依赖性生长潜力的丧失。在部分EBV缺失的T细胞和NK细胞中，病毒基因组的缺失也导致细胞死亡.与亲代EBV阳性T细胞和NK细胞肿瘤相比，EBV丢失的T细胞和NK细胞显示出一些细胞因子如白细胞介素-9的产生显著降低。这些结果表明，T细胞和NK细胞中的EBV依赖性恶性表型以及可能还在上皮细胞中的EBV依赖性恶性表型，如在B细胞淋巴瘤中，至少部分地与EBV依赖性恶性表型的上调相关（即，自分泌机制）和/或某些细胞因子的下调。我们正计划探索细胞因子基因控制的机制和对EBV肿瘤发生的特异性影响。少

The aim of this study was to identify the cellular genes associated with or responsible for the oncogenesisi of Epstein-Barr virus (EBV) in EBV-positive T, MC and epithelial tumor cells, by utilizing dominant-negative EBNA1 (DNE1; Mal. Ther., 11(4): 578-90, 2005)that can eradicate EBV episomes from cells. We prepared isogenic pairs of EBV-positive and -negative NK- and T-cell lines by DNE1 transduction and compared their malignant grade. Concomitantly alterations of cellular gene expression in those cell pairs were comprehensively assessed with the multi-cytokine assay system, thereby elucidating the role(s)of EBV in malignant conversion. Further, we analysed EBV-specific cytotoxic T-cells (EBV-CTL) by a tetramer assay using 3 EBV-antigens, i.e., BRLF1, BMLF1 and EBNA3. The results obtained are as follows.1. Adenovirus vector-mediated transduction of our DNE1 successfully eradicated EBV episomes from the majority of naturally EBV-positive T-cell and NK-cell lymphoma and epithelial tumo … More r cell lines in a few days.2. The DNE1-induced EBV-lost T-cells and NK-cells showed a striking suppression of their malignant phenotypes, such as prolongation of doubling time and loss of anchorage-independent growth potential, compared with parental EBV-positive T-cell and NK-cell tumors. In a part of the EBV-lost T-cells and NK-cells, loss of viral genome also brought about cell death.3. The EBV-lost T-cells and NK-cells showed significantly decreased production of some cytokines, such as interleukin-9, compared with the parental EBV-positive T-cell and NK-cell tumors. Conversely, the production levels of another cytokine was also found to be upregulated in the EBV-lost T-cells.These results indicate that the EBV-dependent malignant phenotypes in T- and NK-cells and perhaps also in epithelial cells, as in B-cell lymphomas, are associated, at least partly, with upregulation (i.e., the autocrine mechanism) and/or down regulation of certain cytokines. We are planning to explore precisely the mechanisms of cytokine gene control and specific effects on EBV oncogenesis. Less