Inhibitors of Epstein-Barr Virus Nuclear Protein 1 Mediated Latent Infection

爱泼斯坦-巴尔病毒(EBV)潜伏感染导致几乎所有EBV相关疾病 和死亡率，包括传染性单核细胞增多症早期的淋巴母细胞增殖， 艾滋病和其他免疫受损患者的淋巴增生性疾病 与EB病毒相关的淋巴瘤、霍奇金氏病和鼻咽癌 癌症。EBV基因组在所有潜伏感染的细胞中作为非整合的 多个副本的圣公会。EB病毒上皮体在细胞分裂中的持久性依赖于 EBV编码核抗原1蛋白(EBNA1)。EBNA1结合到特定的位置 EB病毒Episome和增强Episome的初始复制、转录和持久性。 由于EBNA1对于EBV附粒在所有分裂和 恶性细胞，这项提议的中心目标是鉴定能够 抑制EBNA1介导的Episome持久性。为达致这个目标，我们建议： (1)进行筛选以确定中断EBNA1-ORIP依赖的化合物 Episome转录和体内持久性，干扰EBNA1的化合物 体外与同源DNA的二聚化和结合，以及与EBNA1结合的化合物 在硅胶里。(2)鉴定已鉴定化合物的生物和生化效应 B淋巴母细胞中EBNA1-ORIP依赖的Episome转录和持久性 EB病毒转化的淋巴母细胞(LCL)生长及对LCL诱导淋巴瘤的影响 以及裸鼠体内的鼻咽癌。(3)确定生物活性化合物的作用部位 EBNA1结合，使用生化、生物物理和结构方法。用这个 最有效地进行结构活动修改以改进的知识 化合物活性和特异性。(4)利用反向遗传学识别关键基因 EBNA1 DBD中的残留物可提高筛查灵敏度并在硅胶中提供信息 口袋选择、化合物修饰和化合物相互作用分析。

Epstein-Barr Virus (EBV) latent infections cause almost all EBV associated morbidity and mortality including lymphoblast proliferation early in Infectious Mononucleosis, Lymphoproliferative Diseases in people with AIDS and other immune compromised states, and EBV associated Lymphomas, Hodgkin's Disease, and Nasophryngeal Carcinoma. The EBV genome persists in all latently infected cells as a non-integrated multi-copy episome. The persistence of EBV episomes in dividing cells is dependent on the EBV encoded nuclear antigen 1 protein (EBNA1). EBNA1 binds to a specific site in the EBV episome and enhances episome initial replication, transcription, and persistence. Since EBNA1 is essential for the persistence of EBV episomes in all dividing and malignant cells, the central objective of this proposal is to identify compounds that can inhibit EBNA1 mediated episome persistence. To achieve that objective, we propose to: (1) Undertake screens to identify compounds that interrupt EBNA1-oriP dependent episome transcription and persistence in vivo, compounds that interrupt EBNA1 dimerization and binding to cognate DNA in vitro, and compounds that bind to EBNA1 in silico. (2) Identify the biological and biochemical effects of the identified compounds on EBNA1-oriP dependent episome transcription and persistence in B lymphoblasts, on EBV transformed lymphoblastoid cell (LCL) growth, and on LCL induced Lymphoma and NPC tumors in nude mice. (3) Determine the sites of bioactive compound effects in EBNA1 binding, using biochemical, biophysical, and structural approaches. Use this knowledge to most effectively undertake structure activity modifications to improve compound activity and specificity. (4) Use reverse genetics to identify the critical residues in EBNA1 DBD that can improve screening sensitivity and inform in silico pocket selection, compound modification, and compound interaction analyses.