CRISPR-cas gene-editing as plausible treatment of neuromuscular and nucleotide-repeat-expansion diseases: A systematic review

Abstract

Introduction The system of clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated proteins (cas) is a new technology that allows easier manipulation of the genome. Its potential to edit genes opened a new door in treatment development for incurable neurological monogenic diseases (NMGDs). The aim of this systematic review was to summarise the findings on the current development of CRISPR-cas for therapeutic purposes in the most frequent NMGDs and provide critical assessment. Methods and data acquisition We searched the MEDLINE and EMBASE databases, looking for original studies on the use of CRISPR-cas to edit pathogenic variants in models of the most frequent NMGDs, until end of 2017. We included all the studies that met the following criteria: 1. Peer-reviewed study report with explicitly described experimental designs; 2. In vitro , ex vivo , or in vivo study using human or other animal biological systems (including cells, tissues, organs, organisms); 3. focusing on CRISPR as the gene-editing method of choice; and 5. featured at least one NMGD. Results We obtained 404 papers from MEDLINE and 513 from EMBASE. After removing the duplicates, we screened 490 papers by title and abstract and assessed them for eligibility. After reading 50 full-text papers, we finally selected 42 for the review. Discussion Here we give a systematic summary on the preclinical development of CRISPR-cas for therapeutic purposes in NMGDs. Furthermore, we address the clinical interpretability of the findings, giving a comprehensive overview of the current state of the art. Duchenne’s muscular dystrophy (DMD) paves the way forward, with 26 out of 42 studies reporting different strategies on DMD gene editing in different models of the disease. Most of the strategies aimed for permanent exon skipping by deletion with CRISPR-cas. Successful silencing of the mHTT gene with CRISPR-cas led to successful reversal of the neurotoxic effects in the striatum of mouse models of Huntington’s disease. Many other strategies have been explored, including epigenetic regulation of gene expression, in cellular and animal models of: myotonic dystrophy, Fraxile X syndrome, ataxias, and other less frequent dystrophies. Still, before even considering the clinical application of CRISPR-cas, three major bottlenecks need to be addressed: efficacy, safety, and delivery of the systems. This requires a collaborative approach in the research community, while having ethical considerations in mind.

Therapeutic Strategies for Duchenne Muscular Dystrophy: An Update

“…To obtain permanent dystrophin expression in DMD patients, gene editing, particularly CRISPR/Cas9-based gene editing, has been adopted to correct genomic deficits. CRISPR/Cas9 system consists of the endonuclease Cas9 and a single-guide RNA (sgRNA) [95]. The Cas9 endonuclease produce DNA double-strand break at the editing sequence targeted by sgRNA.…”

“…The Cas9 endonuclease produce DNA double-strand break at the editing sequence targeted by sgRNA. Subsequent nonhomologous end-joining (NHEJ) leads to exon-skipping while homology-directed repair (HDR) could replace DMD mutations with correct sequences and generates normal dystrophin protein [95]. Two types of Cas9 endonucleases have been used for CRISPR gene editing, Staphylococcus aureus Cas9 (SaCas9) and Streptococcus pyogenes Cas9 (SpCas9).…”

“…In vivo CRISPR/Cas9 strategies have also been studied in DMD therapy, most of which choose NHEJ to achieve exon-skipping and subsequent dystrophin restoration [95]. Majority of the flanking regions are chosen within exon 45-55, while other regions like exon 21-23 have also been tested [95,99,101]. Administration routes include intramuscular or intravenous injection, pronuclear packaged nanoparticle microinjection and intramuscular electroporation of AAV or adenovirus (ADV) [95].…”

“…Majority of the flanking regions are chosen within exon 45-55, while other regions like exon 21-23 have also been tested [95,99,101]. Administration routes include intramuscular or intravenous injection, pronuclear packaged nanoparticle microinjection and intramuscular electroporation of AAV or adenovirus (ADV) [95]. Xu et al induced in-frame deletion of exon 21-23 in mice by ADV containing two sgRNA and spCas9 endonuclease [101].…”

Review of Zinc(II) Scaffolds: Efficient Role in Gene Delivery

“…Hence it is necessary to deliver the genetic material into the targeted cell to cure such diseases. As of now, different types of nano-vectors [2][3][4][5] are being in use to deliver the desired gene into the cytosol [3][4][5][6] effectively. From the recent past, cationic lipids are also emerged as a new choice and extensively have been using as vectors (vehicles) in order to replace the novel gene in place of such a defective gene.…”