权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Basic research on novel Epstein-Barr virus(EBV)-specific gene therapy against uncontrollable EBV- associated diseases

针对无法控制的 EBV 相关疾病的新型 Epstein-Barr 病毒（EBV）特异性基因治疗的基础研究

基本信息

批准号：
15590421
负责人：
IMAI Shosuke
金额：
$ 2.24万
依托单位：
Kochi University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

项目摘要

Epstein-Barr virus(EBV) is associated with a variety of human malignant disorders, which are clinically aggressive and often less responsive to the conventional cancer therapy, and currently available anti-herpesvirus drugs are ineffective treatments. EBV nuclear antigen 1(EBNA1), a latent viral protein consistently expressed in all infected proliferating cells, is essentially required in trans to maintain EBV episomes in cells. In this research project, to exploit a novel therapeutic strategy specifically acting against uncontrollable EBV diseases, we constructed a mutant(mt) EBNA1 lacking most of the N-terminal-half, relative to wild-type(wt) EBNA1, and examined whether the mtEBNA1 exerted dominant-negative effects on maintenance of the viral episome thereby leading to abrogation of EBV-infected tumor cell growth. The results obtained are as follows.1.Using lymphocyte and epithelial cell lines converted with neomycin-resistant recombinant EBV(rEBV) as models, adenovirus vector-mediat … More ed transduction of mtEBNA1 brought about rapid and striking reductions of rEBV-derived wtEBNA1 expression levels and viral genomic loads in converted cell lines of three major viral latencies. This outcome was further validated at the single cell level by cellular loss of viral genomic signals in situ, thereby indicating that mtEBNA1 can function as a dominant-negative(dn) EBNA1.2.The dnEBNA1 transduction significantly impaired growth of naturally EBV-harboring Burkitt's lymphoma cells and T/NK lymphoma cells in vitro and in vivo, in association with the eradication of viral episomes.3.Expression of dnEBNA1 per se caused no detectable cytotoxicity in EBV-uninfected cells including normal human fibroblasts.4.The tumor suppressive effects by dnEBNA1 was achieved by three mechanisms : 1) competitive inhibition of wtEBNA1 binding to oriP, 2) decrease of de novo wtEBNA1 synthesis due to suppression of Qp, and 3) direct induction of apoptosis in tumor cells.These results indicate that our dnEBNA1 can efficiently impedes the EBV-dependent malignant phenotypes in cells regardless of viral latency, tissue origin or resident viral strain. Therefore, the mutant will afford an additional therapeutic approach specifically targeting EBV-associated malignancies. Moreover, dnEBNA1 may facilitate analysis of possible oncogenic role(s) for wtEBNA1. Less

EB病毒（Epstein-Barr virus，EBV）与多种人类恶性疾病相关，临床上具有侵袭性，对常规的癌症治疗反应较差，目前可用的抗疱疹病毒药物治疗效果不佳。EBV核抗原1（EBNA 1）是一种在所有感染的增殖细胞中一致表达的潜伏病毒蛋白，其基本上是维持细胞中EBV游离体所必需的。在这项研究项目中，为了开发一种新的治疗策略，专门针对无法控制的EBV疾病，我们构建了一种突变体（mt）EBNA 1，相对于野生型（wt）EBNA 1缺乏大部分的N-末端一半，并检查mtEBNA 1是否对病毒附加体的维持产生显性负效应，从而导致消除EBV感染的肿瘤细胞生长。主要结果如下：1.以新霉素抗性重组EBV（rEBV）转化的淋巴细胞和上皮细胞为模型，腺病毒载体介导的重组腺病毒载体介导的逆转录病毒介导的逆转录病毒介导的逆转 ...更多信息艾德转导mtEBNA 1导致三种主要病毒乳杆菌转化细胞系中rEBV衍生的wtEBNA 1表达水平和病毒基因组载量的快速和显著降低。这一结果在单细胞水平上通过原位病毒基因组信号的细胞丢失进一步验证，从而表明mtEBNA 1可以作为显性阴性（dn）EBNA 1起作用。2. dnEBNA 1转导在体外和体内显著损害天然携带EBV的伯基特淋巴瘤细胞和T/NK淋巴瘤细胞的生长，dnEBNA 1本身的表达在未感染EBV的细胞（包括正常人成纤维细胞）中没有引起可检测的细胞毒性。4. dnEBNA 1的肿瘤抑制作用通过三种机制实现：1）竞争性抑制wtEBNA 1与oriP的结合，2）由于抑制Qp而减少从头wtEBNA 1合成，以及3）直接诱导肿瘤细胞中的凋亡。这些结果表明，我们的dnEBNA 1可以有效地阻止细胞中的EBV依赖性恶性表型，而不管病毒潜伏期、组织来源或驻留病毒株如何。因此，突变体将提供特异性靶向EBV相关恶性肿瘤的额外治疗方法。此外，dnEBNA 1可能有助于分析wtEBNA 1可能的致癌作用。少