In Vivo Target Gene Activation via CRISPR/Cas9-Mediated Trans-epigenetic Modulation

Abstract

SUMMARY Current genome-editing systems generally rely on the creation of DNA double-strand breaks (DSBs). This may limit their utility in clinical therapies, as unwanted mutations caused by DSBs can have deleterious effects. The CRISPR/Cas9 system has recently been repurposed to enable target gene activation, allowing regulation of endogenous gene expression without creating DSBs. However, in vivo implementation of this gain-of-function system has proven difficult. Here we report a robust system for in vivo activation of endogenous target genes through trans-epigenetic remodeling. The system relies on recruitment of Cas9 and transcriptional activation complexes to target loci by modified single guide RNAs. As proof-of-concept, we used this technology to treat several mouse models of human diseases. Results demonstrate that CRISPR/Cas9-mediated target gene activation can be achieved in vivo, leading to observable phenotypic changes, and amelioration of disease symptoms. This establishes new avenues for developing targeted epigenetic therapies against human diseases.

Epigenetic editing: How cutting-edge targeted epigenetic modification might provide novel avenues for autoimmune disease therapy

“…Finally, several examples of epigenetic editing in vivo have recently been published [21, 40, 59, 60]. One recent example by Lei and colleagues [40] leveraged a dCas9-MQ1 fusion.…”

In vivo epigenome editing and transcriptional modulation using CRISPR technology

“…Compared with AAV, these viruses are inefficient for in vivo applications owing to high immunogenicity. AAV8 (Thakore et al 2018; Zhou et al 2018) and AAV9 (Chew et al 2016; Liao et al 2017) were used to package and deliver CRISPR components in all the previous studies. For example, a fusion of dSaCas9 and KRAB repressor domain has a transgene size that falls within the AAV packaging capacity.…”

“…In another elegant study, trans-epigenetic remodeling approach was developed to achieve a potent in vivo target gene activation and phenotypic change (Liao et al 2017). In this strategy, a truncated guide RNA (dgRNA) and an MS2-P65-HSF1 (MPH) transcriptional activation complex were packaged into two separate AAV vectors for co-delivery into Cas9- or dCas9-expressing mice.…”

Clinical applications of CRISPR‐based genome editing and diagnostics

“…Researchers have used CRISPR‐based editing to correct the genetic basis of many diseases in isolated cells or animal models . The first wave of clinical trials using CRISPR enzymes to treat inherited disorders in humans involve removing a patient's cells, editing ex vivo, and reinfusing the corrected cells .…”