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Journal article
Subgenomic particles in rAAV vectors result from DNA lesion/break and non-homologous end joining of vector genomes
Molecular therapy. Nucleic acids, v 29, pp 852-861
13 Sep 2022
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Recombinant adeno-associated virus (rAAV) vectors have been developed for therapeutic treatment of genetic diseases. Current rAAV vectors administered to affected individuals often contain vector DNA-related contaminants. Here we present a thorough molecular analysis of the configuration of non-standard AAV genomes generated during rAAV production using single-molecule sequencing. In addition to the sub-vector genomic-size particles containing incomplete AAV genomes, our results showed that rAAV preparations were contaminated with multiple categories of subgenomic particles with a snapback genome (SBG) configuration or a vector genome with deletions. Through CRISPR and nuclease-based modeling in tissue culture cells, we identified that a potential mechanism leading to formation of non-canonical genome particles occurred through non-homologous end joining of fragmented vector genomes caused by genome lesions or DNA breaks present in the host cells. The results of this study advance our understanding of AAV vectors and provide new clues for improving vector efficiency and safety profiles for use in human gene therapy.
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Zhang et al. revealed that vector DNA lesion and NHEJ are a potential mechanism leading to subgenomic AAV particle generation in the rAAV vector production process. This finding will provide an alternate pathway for discovering novel methodology to improve the safety and efficacy of AAV vectors.
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Details
- Title
- Subgenomic particles in rAAV vectors result from DNA lesion/break and non-homologous end joining of vector genomes
- Creators
- Junping Zhang - Indiana University School of MedicinePing Guo - Huaqiao UniversityXiangping Yu - Huaqiao UniversityDylan A. Frabutt - Indiana University School of MedicineAnh K. Lam - Indiana University School of MedicinePatrick L. Mulcrone - Indiana University School of MedicineMatthew Chrzanowski - Temple UniversityJenni Firrman - United States Department of AgricultureDerek Pouchnik - Washington State UniversityNianli Sang - Drexel UniversityYong Diao - Huaqiao UniversityRoland W. Herzog - Indiana University School of MedicineWeidong Xiao - Indiana University School of Medicine
- Publication Details
- Molecular therapy. Nucleic acids, v 29, pp 852-861
- Publisher
- Elsevier
- Number of pages
- 10
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Biology
- Web of Science ID
- WOS:000852675600004
- Scopus ID
- 2-s2.0-85137294519
- Other Identifier
- 991019173546604721
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- Collaboration types
- Domestic collaboration
- International collaboration
- Web of Science research areas
- Medicine, Research & Experimental