AAV5 delivery of CRISPR-Cas9 supports effective genome editing in mouse lung airway

Shun-Qing Liang; Christopher J Walkey; Alexa E Martinez; Qin Su; Mary E Dickinson; Dan Wang; William R Lagor; Jason D Heaney; Guangping Gao; Wen Xue

doi:10.1016/j.ymthe.2021.10.023

AAV5 delivery of CRISPR-Cas9 supports effective genome editing in mouse lung airway

Mol Ther. 2022 Jan 5;30(1):238-243. doi: 10.1016/j.ymthe.2021.10.023. Epub 2021 Oct 23.

Authors

Affiliations

¹ RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, MA 01605, USA.
² Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX 77030, USA.
³ Viral Vector Core, University of Massachusetts Medical School, Worcester, MA 01605, USA.
⁴ Horae Gene Therapy Center and Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA 01605, USA; Li Weibo Institute for Rare Diseases Research, University of Massachusetts Medical School, Worcester, MA 01605, USA.
⁵ Department of Molecular and Human Genetics, Baylor College of Medicine, Houston, TX 77030, USA.
⁶ Horae Gene Therapy Center and Department of Microbiology and Physiological Systems, University of Massachusetts Medical School, Worcester, MA 01605, USA; Li Weibo Institute for Rare Diseases Research, University of Massachusetts Medical School, Worcester, MA 01605, USA. Electronic address: Guangping.Gao@umassmed.edu.
⁷ RNA Therapeutics Institute, University of Massachusetts Medical School, Worcester, MA 01605, USA; Li Weibo Institute for Rare Diseases Research, University of Massachusetts Medical School, Worcester, MA 01605, USA. Electronic address: wen.xue@umassmed.edu.

Abstract

Genome editing in the lung has the potential to provide long-term expression of therapeutic protein to treat lung genetic diseases. Yet efficient delivery of CRISPR to the lung remains a challenge. The NIH Somatic Cell Genome Editing (SCGE) Consortium is developing safe and effective methods for genome editing in disease tissues. Methods developed by consortium members are independently validated by the SCGE small animal testing center to establish rigor and reproducibility. We have developed and validated a dual adeno-associated virus (AAV) CRISPR platform that supports effective editing of a lox-stop-lox-Tomato reporter in mouse lung airway. After intratracheal injection of the AAV serotype 5 (AAV5)-packaged S. pyogenes Cas9 (SpCas9) and single guide RNAs (sgRNAs), we observed ∼19%-26% Tomato-positive cells in both large and small airways, including club and ciliated epithelial cell types. This highly effective AAV delivery platform will facilitate the study of therapeutic genome editing in the lung and other tissue types.

Keywords: AAV5; CRISPR; Cas9; ciliated cells; club cells; lung editing.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

Animals
CRISPR-Cas Systems*
Gene Editing* / methods
Lung
Mice
RNA, Guide, CRISPR-Cas Systems / genetics
Reproducibility of Results

Substances

RNA, Guide, CRISPR-Cas Systems

Abstract

Publication types

MeSH terms

Substances

Grants and funding