Research Project
9255235
ABSTRACT Infection of the eye by Herpes Simplex Virus-1 (HSV-1) can result in Herpes Keratitis (HK), which is the leading cause of corneal blindness worldwide. In the U.S., nearly 500,000 individuals experience ocular herpes infections that are often recurrent and culminate in progressive corneal scarring and loss of vision. The gold standard of care for HK is treatment with Acyclovir (ACV) that targets HSV-1 thymidine kinase (TK). Although ACV is extremely effective in both oral and genital herpes with negligible drug failure, the emergence of viral resistant mutants in 7-14% of ocular HK patients is compelling. Significantly, the mechanism of drug resistance is directly related to mutational alterations in the TK gene of HSV-1 isolated from HK patients unresponsive to ACV. This high level of resistance can be prevented if ACV is combined with a second antiviral directed against a different target to block mutants that escape either drug. One new class of antiviral targets is the processivity factors (PFs) that are essential to keep their cognate viral polymerases (Pols) tethered to the template for continuous viral DNA synthesis. Our objective is to develop early lead antiviral compounds that are directed against the HSV-1 processivity factor (PF). We have now identified a potent early Lead Z9445, which blocks infection of HSV-1 with an EC50 of 280 nM. We have also identified structurally diverse backup leads. With the future requirement of animal model validation, we are in a unique position to treat multiple feline patients (Phase 2) from the UPenn Vet School who are infected with the homologous Feline Herpes Virus-1 (FHV-1). These feline patients provide a close and natural model of human herpes keratitis that can be followed for extended periods for recurrent infections. We have now cloned the FHV-1 PF/Pol genes which share considerable homologies to those of HSV-1 and have validated that the majority of our compounds that block HSV-1, also block FHV-1 infection with similar potencies. Even though the feline model is highly attractive, our experimental design is focused, such that analogs that are singly superior for HSV-1 (but not FHV-1) will be developed further (Phase 2) using traditional models. The Aims of this proposal are tightly linked, employing medicinal chemistry as an iterative process to improve the therapeutic index of Z9445 by generating new analogs that have increased potency with no detectable toxicities. Rational design based on docking of Z9445 to the known crystal structure of the PF target protein will be one means on producing new Leads. Backup Leads will also be optimized should properties such as solubility or stability need to be enhanced (Phase 2). Analogs will be tested for blocking processive DNA synthesis in vitro, physical binding to the PF target protein, toxicity and cell proliferation, antiviral activities in primary feline corneal epithelial cells and in the human 3D corneal tissue. The human 3D corneal tissue will be tested further for permeability and histology. .
