Empty Virions In AAV8 Vector Preparations Reduce Transduction Efficiency And May Cause Total Viral Particle Dose-Limiting Side-Effects

Kai Gao; Mengxin Li; Li Zhong; Qin Su; Jia Li; Shaoyong Li; Ran He; Yu Zhang; Gregory Hendricks; Junzhi Wang; Guangping Gao

doi:10.1038/mtm.2013.9

Empty Virions In AAV8 Vector Preparations Reduce Transduction Efficiency And May Cause Total Viral Particle Dose-Limiting Side-Effects

Mol Ther Methods Clin Dev. 2014;1(9):20139. doi: 10.1038/mtm.2013.9.

Authors

Kai Gao¹, Mengxin Li², Li Zhong³, Qin Su⁴, Jia Li⁵, Shaoyong Li⁵, Ran He⁴, Yu Zhang⁶, Gregory Hendricks⁷, Junzhi Wang⁸, Guangping Gao⁹

Affiliations

¹ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; National Institute for Food & Drug Control, Beijing, China.
² Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Viral Vector Core, University of Massachusetts Medical School, Worcester, MA 01605 ; Department of Microbiology and Physiology Systems, University of Massachusetts Medical School, Worcester, MA 01605.
³ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Viral Vector Core, University of Massachusetts Medical School, Worcester, MA 01605 ; Division of Hematology/Oncology, Department of Pediatrics, University of Massachusetts Medical School, Worcester, MA 01605.
⁴ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Viral Vector Core, University of Massachusetts Medical School, Worcester, MA 01605.
⁵ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Department of Microbiology and Physiology Systems, University of Massachusetts Medical School, Worcester, MA 01605.
⁶ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Department of Oncology, Guizhou People's Hospital, Guiyang, Gizhou, China.
⁷ Department of Cell and Developmental Biology & Core Electron Microscopy Facility, University of Massachusetts Medical School, Worcester, MA 01605.
⁸ National Institute for Food & Drug Control, Beijing, China.
⁹ Gene Therapy Center, University of Massachusetts Medical School, Worcester, MA 01605 ; Viral Vector Core, University of Massachusetts Medical School, Worcester, MA 01605 ; Department of Microbiology and Physiology Systems, University of Massachusetts Medical School, Worcester, MA 01605 ; Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, Sichuan, China.

Abstract

Empty virions are inadvertent by-products of recombinant adeno-associated virus (rAAV) packaging process, resulting in vector lots with mixtures of full and empty virions at variable ratios. Impact of empty virions on the efficiency and side-effects of rAAV transduction has not been well characterized. Here, we generated partially and completely empty AAV8 virions, fully packaged rAAV8 lots as well as mixtures of empty and fully packaged virions with variable ratios of empty virions (REVs). The aforementioned dosing formulations of rAAV8 expressing either cellular (EGFP or nuclear-targeted (n) LacZ) or secreted (human α1-antitrypsin, hA1AT) reporter genes were intravenously injected into two different mouse strains, followed by analyses of transgene expressions and serum alanine aminotransferase (ALT) levels at different time points. We found that addition of empty particles to the fixed doses of rAAV8 preparations repressed liver transduction up to 64% (serum hA1AT) and 44% (nLacZ) in C57BL/6 mice, respectively. The similar trend in inhibiting EGFP expression together with concurrent elevations of serum ATL levels were observed in the BALB/c mice, indicating that empty particles may also exacerbate side-effects of rAAV8EGFP transduction. Our results suggest that removal of empty particles from rAAV preparations may improve efficacy and safety of AAV in clinical applications.

Keywords: empty virions; liver gene transfer; rAAV; side-effect.

Abstract

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