Discovery and development of novel HCMV inhibitors - ABSTRACT Infection with human Cytomegalovirus (HCMV) is nearly universal; seroprevalence rates reach 90% in individuals older than 80. Although usually asymptomatic in the normal host, HCMV causes severe morbidity and mortality in transplant recipients and patients with AIDS. Congenital HCMV is the leading infectious cause of childhood deafness and mental retardation. There is no approved vaccine for the prevention of HCMV. The nucleoside analog ganciclovir (GCV) and its oral formulation val-GCV have improved the outcome of transplantation. GCV may also prevent hearing deterioration in congenitally infected children. However, prolonged courses of GCV or val-GCV result in intolerable toxicities to the bone marrow and select for resistant viral strains. The alternative drugs for GCV-resistant HCMV, foscarnet, and cidofovir, also target the viral DNA polymerase. Both are nephrotoxic agents and can only be administered intravenously. The HCMV UL97 kinase inhibitor, maribavir, was recently approved for adults and children (> 12 years of age) with resistant/refractory HCMV disease, and the terminase inhibitor, letermovir, is approved for HCMV prophylaxis after hematopoietic stem cell transplantation. Resistance has already been reported to these two new agents. The limited HCMV therapeutic armamentarium and the emergence of drug-resistant mutants create an urgent need to identify new HCMV inhibitors with a novel mechanism of action. Our goal is to develop a new class of HCMV inhibitors that act through a novel viral target(s), the helicase-primase (H/P). As a prototype, compound MLS8091 inhibits HCMV, including GCV-resistant strains, and we have identified that resistance to MLSS8091 resides in the primase and primase-associated factor of the H/P complex. The following Specific Aims are proposed in this R01: 1) Delineate the mechanism(s) of action of MLS8091. 2) Define the interactions and activities within the H/P complex. 3) Design and synthesize MLS8091 analogs to identify a lead compound for in vivo studies. Our program is innovative and highly significant because the target for HCMV inhibition is novel and critical for virus replication, we already identified resistant mutants, and we assembled a multi-disciplinary investigative team to develop inhibitors of the H/P complex. Our team has combined expertise in virology, proteomics, structural biology, biophysics, biochemistry, and chemistry. At the end of the project, we expect to: 1) Confirm the HCMV H/P as a druggable target, 2) Provide purified proteins, biophysical and enzymatic data of the H/P complex for future HCMV drug discovery, and 3) Identify the most favorable small molecule HCMV inhibitor(s) through drug design and synthesis.
