In Situ Gene Therapy via AAV-CRISPR-Cas9-Mediated Targeted Gene Regulation.

Journal Article (Journal Article)

Development of efficacious in vivo delivery platforms for CRISPR-Cas9-based epigenome engineering will be critical to enable the ability to target human diseases without permanent modification of the genome. Toward this, we utilized split-Cas9 systems to develop a modular adeno-associated viral (AAV) vector platform for CRISPR-Cas9 delivery to enable the full spectrum of targeted in situ gene regulation functionalities, demonstrating robust transcriptional repression (up to 80%) and activation (up to 6-fold) of target genes in cell culture and mice. We also applied our platform for targeted in vivo gene-repression-mediated gene therapy for retinitis pigmentosa. Specifically, we engineered targeted repression of Nrl, a master regulator of rod photoreceptor determination, and demonstrated Nrl knockdown mediates in situ reprogramming of rod cells into cone-like cells that are resistant to retinitis pigmentosa-specific mutations, with concomitant prevention of secondary cone loss. Furthermore, we benchmarked our results from Nrl knockdown with those from in vivo Nrl knockout via gene editing. Taken together, our AAV-CRISPR-Cas9 platform for in vivo epigenome engineering enables a robust approach to target disease in a genomically scarless and potentially reversible manner.

Full Text

Duke Authors

Cited Authors

  • Moreno, AM; Fu, X; Zhu, J; Katrekar, D; Shih, Y-RV; Marlett, J; Cabotaje, J; Tat, J; Naughton, J; Lisowski, L; Varghese, S; Zhang, K; Mali, P

Published Date

  • July 5, 2018

Published In

Volume / Issue

  • 26 / 7

Start / End Page

  • 1818 - 1827

PubMed ID

  • 29754775

Pubmed Central ID

  • PMC6035733

Electronic International Standard Serial Number (EISSN)

  • 1525-0024

Digital Object Identifier (DOI)

  • 10.1016/j.ymthe.2018.04.017


  • eng

Conference Location

  • United States