Antiviral Targeting of Varicella Zoster Virus Replication and Neuronal Reactivation Using CRISPR/Cas9 Cleavage of the Duplicated Open Reading Frames 62/71.

Betty W. Wu,Michael B. Yee,Paul R. Kinchington,Ronald S. Goldstein

doi:10.3390/v14020378

Betty W. Wu, Michael B. Yee + Show 2 more

Open Access

https://doi.org/10.3390/v14020378

Copy DOI

Journal: Viruses	Publication Date: Feb 12, 2022
Citations: 2	License type: CC BY 4.0

Affiliation: University of Pittsburgh, Bar-Ilan University

Abstract

Varicella Zoster Virus (VZV) causes Herpes Zoster (HZ), a common debilitating and complicated disease affecting up to a third of unvaccinated populations. Novel antiviral treatments for VZV reactivation and HZ are still in need. Here, we evaluated the potential of targeting the replicating and reactivating VZV genome using Clustered Regularly Interspaced Short Palindromic Repeat-Cas9 nucleases (CRISPR/Cas9) delivered by adeno-associated virus (AAV) vectors. After AAV serotype and guide RNA (gRNA) optimization, we report that a single treatment with AAV2-expressing Staphylococcus aureus CRISPR/Cas9 (saCas9) with gRNA to the duplicated and essential VZV genes ORF62/71 (AAV2-62gRsaCas9) greatly reduced VZV progeny yield and cell-to-cell spread in representative epithelial cells and in lytically infected human embryonic stem cell (hESC)-derived neurons. In contrast, AAV2-62gRsaCas9 did not reduce the replication of a recombinant virus mutated in the ORF62 targeted sequence, establishing that antiviral effects were a consequence of VZV-genome targeting. Delivery to latently infected and reactivation-induced neuron cultures also greatly reduced infectious-virus production. These results demonstrate the potential of AAV-delivered genome editors to limit VZV productive replication in epithelial cells, infected human neurons, and upon reactivation. The approach could be developed into a strategy for the treatment of VZV disease and virus spread in HZ.

Highlights

Varicella-zoster virus (VZV) is the human alphaherpesvirus that causes varicella during primary infection and herpes zoster (HZ; commonly called “shingles”) when the virus reactivates from the latent state, often decades after the initial infection [1]
The goal of the studies was to assess the potential of using gene editing as an anti-viral strategy targeting VZV, in lytic-infected cells that are permissive for VZV, and in reactivating neuron cultures
Gene editing strategies targeting herpes simplex virus type 1 (HSV-1) have suggested that two sites of double stranded breaks (DSBs) are more efficient in reducing the HSV genome load and progeny virus production compared to single genome DSB sites, which can repair

Summary

Introduction

Varicella-zoster virus (VZV) is the human alphaherpesvirus that causes varicella (chickenpox) during primary infection and herpes zoster (HZ; commonly called “shingles”) when the virus reactivates from the latent state, often decades after the initial infection [1]. The first HZ licensed vaccine (used since 2005) was based on a higher dose version of the live-attenuated VZV strain used in the varicella vaccine, and it reduced HZ incidence by half and the disease burden by two-thirds [6]. It was contraindicated in immunocompromised patients who could develop vaccine virus-induced disease [7,8]. A more recent Federal Drug Administration (FDA)-approved vaccine is based on the novel AS01B adjuvant and purified VZV glycoprotein E (gE) This subunit vaccine has higher efficacy against HZ, but requires two doses, has frequent side effects or injection-site reactions and is not used worldwide [9,10,11]. VZV diseases still impact millions of people worldwide and there is a need for improved HZ treatments/prevention

Objectives

Methods

Results

Conclusion