# Applying CRISPR Technologies for the Treatment of Human Herpesavirus Infections: A Scoping Review
## 1 Introduction
It functions by guiding the endonuclease Cas9 to specific DNA sequences using a guide RNA (gRNAA), resulting in targeted double-stranded breaks. These breaks are then repaired by either non-homologous end joining (NHE) or homology-directed repair (HDR) if a repair template is available [30; 31; 32]. NHEJ is faster and more efficient than HDR, but is inherently error-prone, often resulting in insertions or deletions (INDELs) at the cut site. INDEL formation in a gene of interest can result in loss of function, frameshift mutations, or the creation of a premature stop codon, leading to gene knock-out (KO). As an antiviral strategy, this approach is particularly useful for targeting essential viral genes, resulting in replication-defective virusesand ultimately the clearance of infection. However, there is a risk of mutational escape, whereby the virus accumulates sequence changes that prevent recognition by the original gRNA while retaining partial or full functionality. To mitigate this, multiple gRNAs can be introduced to target several sites within the viral genome, thereby increasing the frequency of double-strand breaks. This multiplexing strategy can lead to large genomic deletions and loss of viral genome stability, ultimately resulting in complete viral genome clearance. However, when applied to integrated viral genomes, such extensive genome editing may also pose a risk of chromosomal instability or large-scale host genomic rearrangements. The HDR pathway is more accurate, but less efficient than NHEJ. In the context of viral genome engineering, HDR is primarily employed for knock-in applications, such as the insertion of transgenes to generate viral vectors or gene drive viruses. Besides the CRISPR-Cas9 system, other CRISPR systems with different Cas enzymes have been developed, such as CRISPR-CasX, which also applies specific double-stranded breaks using the smaller endonuclease CasX [33]. Two different CRISPR systems, namely CRISPR interference (CRISPRRi) and CRISPR activation (CRISPRRa), do not create double-stranded breaks but transiently modulate transcription by using an endonucleolytically deactivated Cas9 (dCas9) protein [34]. More specifically, the dCas9 protein can be fused to transcriptional repressor or activator domains to temporarily inhibit or activate gene expression, enabling CRISPR or CRISPR, respectively [34]. Given its sequence-specific DNA-targeting ability, CRISPR-Cas represents an attractive approach for antiviral therapy, particularly for targeting and disrupting herpesviral genomes. The application of the CRISPR-Cas editing tool has already been explored in many fields, including biotechnology and medicine, targeting genomes in both prokaryotes and eukaryotes. Applications include disease modeling to investigate pathogenesis/oncogenesis, genome-wide loss-of-function screening, and gene therapy for genetic disorders [31]. This scoping review aimed to investigate the application of CRISPR-Cas systems as antiviral therapy against human herpesvirus. We summarize findings from 43 primary studies to provide an overview of the current state of research and mechanisms by which CRISPR systems are being applied to treat herpesvirus infections.