Identification of novel inhibitors targeting entry of human cytomegalovirus

描述（由申请方提供）：人巨细胞病毒（HCMV）是一种机会致病菌，能够与其宿主建立并维持终身关系，这种持续状态称为潜伏期。原发感染和潜伏HCMV的再活化导致免疫功能不足的患者的高病毒载量，通常导致病毒复制不受控制，以及多器官疾病和功能障碍。事实上，HCMV显著地导致免疫功能低下个体如新生儿、器官移植受者、AIDS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的发病率和死亡率。HCMV是出生缺陷的主要原因，影响发达国家1-4%的新生儿，并显著导致移植失败。HCMV还增加心血管疾病、自身免疫性疾病和各种癌症的严重程度。目前针对HCMV的治疗靶向病毒DNA聚合酶，但在长期治疗后诱导严重的副作用（例如血小板减少症）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增殖将有效地限制HCMV相关疾病。因此，我们提出利用表达与黄色荧光蛋白融合的病毒蛋白（即早期蛋白2）的HCMV变体来鉴定抑制HCMV感染的早期步骤的新疗法。NIH化学基因组学核心将进行高含量筛选，以确定阻断病毒感染早期步骤的化合物。将进行高通量二次试验，以通过在多种高含量筛选中验证先导化合物、纳摩尔范围内的有效浓度和低毒性来鉴定先导化合物。此外，三级试验将确定先导化合物的作用模式。针对感染早期的抗HCMV试剂的发现代表了一类新的化合物，其将靶向与当前治疗剂不同的蛋白质，目的是在免疫受损个体中减弱HCMV增殖和HCMV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.