High Throughput Screening for Identification of Human Cytomegalovirus Inhibitors

用于鉴定人类巨细胞病毒抑制剂的高通量筛选

基本信息

批准号：
8870334
负责人：
Ravit Boger
金额：
$ 34.08万
依托单位：
JOHNS HOPKINS UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-13 至 2017-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8870334
关键词：
Acquired Immunodeficiency Syndrome Adult Adverse effects Antibodies Area Artemisinins Binding Biological Biological Assay Cardiac Glycosides Cells Chemicals Child Chronic Cidofovir Classification Clinical Collaborations Cytomegalovirus Cytomegalovirus Infections Cytomegalovirus Vaccines DNA Polymerase Inhibitor DNA biosynthesis DNA-Directed DNA Polymerase Deterioration Development Dose Drug Combinations Drug Formulations Drug resistance Exclusion Family Foscarnet Future Ganciclovir Goals HIV Health Hepatitis C Hour Immunocompetent Individual Infection Investigation Laboratories Lead Libraries Luciferases Measures Mental Retardation Morbidity - disease rate Oral Organ Transplantation Pathway interactions Patients Pharmaceutical Preparations Population Pregnant Women Proteins Recombinants Reporter Reporting Resistance Solid Specificity Staging System Techniques Therapeutic Uses Time Toxic effect Transplant Recipients United States United States National Institutes of Health Valganciclovir Viral Virus Virus Diseases Virus Replication Western Blotting artemisinine base burden of illness cohort congenital cytomegalovirus congenital infection cost cytotoxicity deafness experience follow-up hearing impairment high throughput screening indexing inhibitor/antagonist luminescence maribavir miniaturize mortality mutant neoplastic novel novel therapeutics pathogen prevent promoter prophylactic recombinant virus response salinomycin screening small molecule libraries terminase treatment strategy viral DNA

项目摘要

DESCRIPTION: Infection with human Cytomegalovirus (CMV) continues to be a major threat for pregnant women, solid organ transplant recipients and patients with HIV-AIDS. Congenital CMV is the leading infectious cause of mental retardation and deafness. Although therapies for CMV can suppress virus replication, their prolonged use results in serious side effects and the emergence of resistant viruses. The available CMV inhibitors share the same mechanism of action - inhibition of viral DNA polymerase. There is an absolute need to identify new CMV inhibitors and to develop treatment strategies for CMV. The development of novel therapies for CMV could reduce morbidity and mortality in congenitally-infected children and the solid organ transplant population. Identification of novel compounds for CMV therapy is now possible because of the availability of large chemical libraries formatted as high-throughput screening (HTS) systems. The overall goals of this application (PAR-12-058) are to identify compounds that inhibit CMV replication using a quantitative HTS and a luciferase-recombinant CMV assay, to validate the specificity of the compounds for CMV using secondary and tertiary assays and to discover combination of anti-CMV agents by HTS to understand their mechanism of CMV inhibition. During the last several years we developed and validated sensitive and reproducible assays for quantification of CMV replication inhibition including a luciferase-recombinant CMV, luciferase-recombinant ganciclovir-resistant CMV, virus yield and DNA replication based on real-time PCR. These techniques have already been used extensively in our studies of CMV inhibitors such as artemisinins, and cardiac glycosides, two families of CMV inhibitors that differ in their mode of action and salinomycin. Our assays can be applied to screen the largest library of chemical probes in collaboration with the NIH and to identify chemical probes for the stage of virus replication in which these compounds act secondary and tertiary assays. The proposed investigations are likely to succeed because of our established experience in anti-viral assays and the expertise of NIH in HTS. The information derived from this application will have critical impact on CMV therapy. It may lead to the development of new concepts in CMV therapeutics by using compounds that inhibit a target different from the viral DNA polymerase and by implementing combination anti-CMV therapy.

描述：人类巨细胞病毒（CMV）的感染仍然是孕妇，固体器官移植受者和HIV-AID患者的主要威胁。先天性CMV是智力低下和耳聋的主要感染原因。尽管CMV的疗法可以抑制病毒复制，但它们的延长使用会导致严重的副作用和抗性病毒的出现。可用的CMV抑制剂具有相同的作用机理 - 抑制病毒DNA聚合酶。绝对需要识别新的CMV抑制剂并制定CMV的治疗策略。新型CMV疗法的发展可以降低先天感染儿童和固体器官移植人群的发病率和死亡率。现在可以识别用于CMV治疗的新型化合物，因为将大型化学文库的可用性作为高通量筛选（HTS）系统的可用性。该应用程序的总体目标（PAR-12-058）是识别使用定量HTS和荧光素酶 - 重组CMV测定法抑制CMV复制的化合物，以使用二级和第三级分析和第三级化合物的特异性来验证CMV的特异性，并通过HTI-CMV的组合来理解HTS的组合，以了解他们的机制。在过去的几年中，我们开发了并验证了敏感和可重复的测定法，以定量CMV复制抑制，包括荧光素酶 - 成分CMV，基于实时PCR的荧光素酶 - 成分CMV，荧光素酶 - 重组抗性CMV，耐药CMV，病毒率和DNA复制。这些技术已经广泛用于我们对CMV抑制剂（例如青蒿素和心脏糖苷）的研究，这是两个不同的CMV抑制剂家族以其作用方式和盐霉素。我们的测定方法可用于与NIH合作筛选最大的化学探针文库，并确定这些化合物在其中作用于二级和第三次分析的病毒复制阶段的化学探针。拟议的调查很可能会成功，因为我们在抗病毒测定法和NIH在HTS中的专业知识的既定经验。从该应用程序得出的信息将对CMV疗法产生关键影响。它可能通过使用抑制与病毒DNA聚合酶不同的靶标并实施组合抗CMV治疗的化合物，从而导致CMV疗法中新概念的发展。