@article{f0f1388a52c24ee8a8ed11f44589c041,
title = "Co-opting regulation bypass repair as a gene-correction strategy for monogenic diseases",
abstract = "With the development of CRISPR-Cas9-mediated gene-editing technologies, correction of disease-causing mutations has become possible. However, current gene-correction strategies preclude mutation repair in post-mitotic cells of human tissues, and a unique repair strategy must be designed and tested for each and every mutation that may occur in a gene. We have developed a novel gene-correction strategy, co-opting regulation bypass repair (CRBR), which can repair a spectrum of mutations in mitotic or post-mitotic cells and tissues. CRBR utilizes the non-homologous end joining (NHEJ) pathway to insert a coding sequence (CDS) and transcription/translation terminators targeted upstream of any CDS mutation and downstream of the transcriptional promoter. CRBR results in simultaneous co-option of the endogenous regulatory region and bypass of the genetic defect. We validated the CRBR strategy for human gene therapy by rescuing a mouse model of Wolcott-Rallison syndrome (WRS) with permanent neonatal diabetes caused by either a large deletion or a nonsense mutation in the PERK (EIF2AK3) gene. Additionally, we integrated a CRBR GFP-terminator cassette downstream of the human insulin promoter in cadaver pancreatic islets of Langerhans, which resulted in insulin promoter regulated expression of GFP, demonstrating the potential utility of CRBR in human tissue gene repair.",
author = "Jingjie Hu and Bourne, {Rebecca A.} and McGrath, {Barbara C.} and Alice Lin and Zifei Pei and Cavener, {Douglas R.}",
note = "Funding Information: Human pancreatic islets were provided by the NIDDK -funded Integrated Islet Distribution Program (IIDP) (RRID:SCR _014387) at City of Hope, NIH grant no. 2UC4DK098085 . We thank the Harvard Genome Modification Facility for providing the mouse zygote microinjection service (job 1772 for Perk C528X allele generation and job 2182 for Perk rPERK-CRBR allele generation). We thank the Genomics Core Facility, Flow Cytometry Facility, and Microscopy Core Facility at the Huck Institutes of the Life Sciences at Penn State University for providing the Sanger sequencing service, cell sorting service, and histology lab equipment for tissue processing. We thank the Makova Lab at Penn State University for providing the ddPCR system and Dr. Tomaszkiewicz for her help with optimizing the assay. This work was supported by NIH grant no. R01 DK88140. Funding Information: Human pancreatic islets were provided by the NIDDK-funded Integrated Islet Distribution Program (IIDP) (RRID:SCR _014387) at City of Hope, NIH grant no. 2UC4DK098085. We thank the Harvard Genome Modification Facility for providing the mouse zygote microinjection service (job 1772 for PerkC528X allele generation and job 2182 for PerkrPERK-CRBR allele generation). We thank the Genomics Core Facility, Flow Cytometry Facility, and Microscopy Core Facility at the Huck Institutes of the Life Sciences at Penn State University for providing the Sanger sequencing service, cell sorting service, and histology lab equipment for tissue processing. We thank the Makova Lab at Penn State University for providing the ddPCR system and Dr. Tomaszkiewicz for her help with optimizing the assay. This work was supported by NIH grant no. R01 DK88140., Conceptualization, J.H. R.A.B. B.C.M. and D.R.C.; Methodology, J.H. R.A.B. and D.R.C.; Validation, J.H. R.A.B. B.C.M. and D.R.C.; Formal analysis, J.H. A.L. and R.A.B.; Investigation, J.H. R.A.B. and A.L.; Resources, Z.P.; Writing ? original draft, J.H.; Writing ? review & editing, J.H. R.A.B. B.C.M. and D.R.C.; Visualization, J.H.; Funding acquisition, J.H. R.A.B. B.C.M. and D.R.C. J.Hu, R. A. Bourne, B.C. McGrath, A. Lin, and D.R. Cavener have submitted a provisional patent related to this work. Publisher Copyright: {\textcopyright} 2021 The Authors",
year = "2021",
month = nov,
day = "3",
doi = "10.1016/j.ymthe.2021.04.017",
language = "English (US)",
volume = "29",
pages = "3274--3292",
journal = "Molecular Therapy",
issn = "1525-0016",
publisher = "Nature Publishing Group",
number = "11",
}