Vector-delivered CRISPR/Cas as a cure for HSV-1-induced keratitis

DESCRIPTION provided by applicant Herpes simplex virus HSV is a ubiquitous viral pathogen that while rarely lethal nevertheless has the ability to cause severe morbidity in a subset of patients Among the more serious potential consequences of HSV infection is the development of HSV keratitis the most common cause of infectious blindness in the USA with new cases each year Currently available drugs do not effectively treat HSV keratitis and a new treatment modality is therefore clearly needed In this grant application we propose to optimize the bacterial CRISPR Cas mechanism of adaptive antiviral immunity in order to efficiently cleave and inactivate latent HSV genomes in neuronal cells in vivo Our initial goal is to develop vectors based on adeno associated virus AAV which has been widely used for gene therapy in humans to express the novel smaller Cas protein recently identified in Staphylococcus aureus Sau together with small guide RNAs sgRNAs specific for HSV After optimization and demonstration of function in cultured cells these AAV vectors will be introduced into mice containing trigeminal ganglia latently infected by HSV and the potential reduction in viral DNA load and loss of reactivation potential assayed In this way we hope to provide proof of principle for the idea that CRISPR Cas systems can be repurposed as a novel tool for the effective elimination of latent HSV DNA genomes in vivo PUBLIC HEALTH RELEVANCE Herpes simplex virus HSV is a ubiquitous pathogen that is responsible for substantial morbidity in a subset of patients including HSV keratitis the most prevalent form of infectious disease leading to blindness Here we propose using viral vectors to deliver the recently developed CRISPR Cas system of RNA guided DNA endonucleases to infected cells in order to cleave and destroy HSV DNA episomes in vivo