Identification of novel inhibitors targeting entry of human cytomegalovirus

描述(申请人提供)：人类巨细胞病毒(HCMV)是一种机会病原体，能够与其宿主建立和维持终身关系，这种持续状态称为潜伏期。初次感染和潜伏的HCMV的重新激活会导致高病毒载量，免疫功能低下的患者通常会导致病毒复制失控，并导致多器官疾病和功能障碍。事实上，巨细胞病毒对新生儿、器官移植受者、艾滋病患者、癌症患者、心血管疾病患者、自身免疫性疾病患者以及老年人等免疫功能低下的人的发病率和死亡率有很大影响。在发达国家，巨细胞病毒是影响1-4%新生儿的出生缺陷的主要原因，也是造成移植失败的重要原因。巨细胞病毒还会增加心血管疾病、自身免疫性疾病和各种癌症的严重程度。目前针对HCMV的治疗以病毒DNA聚合酶为靶点，但长期治疗会导致严重的副作用(如血小板减少症)和耐药病毒株。我们的中心假设是，通过靶向HCMV生命周期的不同步骤来抑制HCMV的增殖将有效地限制HCMV相关疾病。因此，我们建议利用表达融合到黄色荧光蛋白的病毒蛋白(立即早期蛋白2)的HCMV变异体来识别抑制HCMV感染早期步骤的新疗法。NIH化学基因组核心将进行高含量筛选，以确定阻止病毒感染早期步骤的化合物。将进行高通量二次分析，通过在各种高含量、纳摩尔范围内的有效浓度和低毒性的筛选中验证先导化合物来识别它们。此外，三级分析将定义先导化合物的作用模式。针对早期感染的抗巨细胞病毒试剂的发现代表了一类新的化合物，它们将针对不同于目前治疗方法的蛋白质，目的是减轻免疫受损个体的巨细胞病毒增殖和巨细胞病毒相关疾病。

DESCRIPTION (provided by applicant): Human cytomegalovirus (HCMV) is an opportunistic pathogen with the ability to establish and maintain a lifelong relationship with its host, a persistent state termed latency. Primary infections and reactivation of latent HCMV results in a high viral load in patients with inadequate immune function commonly results in uncontrolled virus replication, and multi-organ disease and dysfunction. In fact, HCMV significantly contributes to morbidity and mortality in immunocompromised individuals such as newborns, organ transplant recipients, AIDS patients, cancer patients, patients with cardiovascular diseases, autoimmune disease patients, as well as the elderly. HCMV is the leading cause of birth defects affecting 1-4% of newborns in developed countries and significantly contributes to transplant failure. HCMV also increases the severity of cardiovascular diseases, autoimmune diseases, and various cancers. Current therapy against HCMV targets the viral DNA polymerase, but induces severe side effects (e.g. thrombocytopenia) and drug-resistant viral strains upon prolonged treatment. Our central hypothesis is that inhibiting HCMV proliferation by targeting diverse steps of the HCMV life cycle would effectively limit HCMV-associated diseases. Thus, we propose to identify novel therapeutics that inhibit the early steps of HCMV infection utilizing a HCMV variant that expresses a viral protein (immediate-early protein 2) fused to yellow fluorescent protein. A high-content screen will be performed at the NIH Chemical Genomics Core to identify compounds that block the early steps of virus infection. High- throughput secondary assays will be performed to identify lead compounds through their validation in a diverse high-content screen, an effective concentration in the nanomolar range and low toxicity. Also, tertiary assays will define the mode of action of the lead compounds. The discovery of anti-HCMV reagents against the early phase of infection represents a novel class of compounds that will target different proteins than the current therapeutics with the objective o attenuating HCMV proliferation and HCMV-associated diseases in immunocompromised individuals.