High Throughput Screening for Identification of Human Cytomegalovirus Inhibitors

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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.