A Broad Application of CRISPR Cas9 in Infectious Diseases of Central Nervous System

Abstract

Virus-induced diseases or neurological complications are huge socio-economic burden to human health globally. The complexity of viral-mediated CNS pathology is exacerbated by reemergence of new pathogenic neurotropic viruses of high public relevance. Although the central nervous system is considered as an immune privileged organ and is mainly protected by barrier system, there are a vast majority of neurotropic viruses capable of gaining access and cause diseases. Despite continued growth of the patient population and a number of treatment strategies, there is no successful viral specific therapy available for viral induced CNS diseases. Therefore, there is an urgent need for a clear alternative treatment strategy that can effectively target neurotropic viruses of DNA or RNA genome. To address this need, rapidly growing gene editing technology based on CRISPR/Cas9, provides unprecedented control over viral genome editing and will be an effective, highly specific and versatile tool for targeting CNS viral infection. In this review, we discuss the application of this system to control CNS viral infection and associated neurological disorders and future prospects. Graphical Abstract CRISPR/Cas9 technology as agent control over CNS viral infection.

Keywords: CRISPR/Cas9; CRISPR/Cas9 delivery system; CRISPR/Cas9-mediated viral escape; Neurotropic viruses.

Fig 1.. CRISPR/Cas9 system.

The simplicity of CRISPR/Cas9 system is that a single protein, Cas9 guided by a short RNA sequence serves as a site-specific endonuclease. CRISPR/Cas9 consists of 3 components: two RNA and a Cas9 protein. One of the RNA (CRISPR RNA or crRNA) contains a short region of homology (20bp) that direct the CRISPR/Cas9 complex to target genetic locus. The second RNA (Trans activating crRNA or tracrRNA) base pairs with crRNA to form lops based RNA structure that leads Cas9 to the target locus where there is a PAM sequence in the target DNA to introduce double strand break (DSB) that is 3 base pair (bp) upstream of PAM. This DSB can be repaired either with NHEJ which is error-prone or homology-directed repair (HDR).

Fig.2. CRISPR/Cas9 mediated targeting of human pathogenic neurotropic viruses.

Cas9 and gRNAs, the main components of the CRISPR/Cas9 system form Cas9-gRNA complex. This complex target and cleaves the viral genome through double strand break which can be repaired by NHEJ. On the other hand, the creation of DSB result in the degradation of the virus genome. Other important outcomes like viral escape, viral DNA repair and off target effect on the host genome are shown.

Fig. 3. Future application and challenge of CRISPR-Cas9 editing approach.

Although CRISPR Cas9 system has tremendous potential for human genomic engineering application in vivo and in vitro, there are certain questions has to be addressed before its clinical use. One of the main concerns is the off-target effect of the system, in particular the CRISPR/Cas9 –mediated mosaic mutations. Moreover, the future therapeutic effect of the CRISPR/Cas9 system largely depend on the delivery of the CRISPR/Cas9 components to the target cells. The current available delivery system are not specific, efficient and have biosafety concern. Another major concern regarding future application of CRISPR Cas9 system is the recent finding of the presence of preexisting adaptive immune response in humans for a variety of Cas9 orthologs such as S. aureus and S. pyogenes. In summary, the future application of CRISPR/Cas9 for gene therapy has to improve, as CRISPR/Cas9 gene editing system has been shown powerful to target and eliminate viral infection associated with neurological complication.