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Mol Ther Methods Clin Dev. 2019 Jan 18;12:202-222. doi: 10.1016/j.omtm.2019.01.003. eCollection 2019 Mar 15.

Novel Chimeric Gene Therapy Vectors Based on Adeno-Associated Virus and Four Different Mammalian Bocaviruses.

Author information

1
Department of Infectious Diseases/Virology, Heidelberg University Hospital, Heidelberg, Germany.
2
BioQuant Center, University of Heidelberg, Heidelberg, Germany.
3
Translational Research Unit, Thoraxklinik at Heidelberg University Hospital, Heidelberg, Germany.
4
German Center for Lung Research (DZL), Translational Lung Research Center Heidelberg (TLRC), Heidelberg, Germany.
5
Hubrecht Institute and Oncode Institute, Royal Netherlands Academy of Arts and Sciences (KNAW), Utrecht, the Netherlands.
6
Research Group "Cellular Polarity of Viral Infection", German Cancer Research Center (DKFZ), Heidelberg, Germany.
7
Institute for Pathology, Kliniken der Stadt Köln gGmbH, Hospital of the Private University Witten/Herdecke, Cologne, Germany.
8
University Medical Center (UMC) Utrecht, Utrecht, the Netherlands.
9
Princess Máxima Centre, Utrecht, the Netherlands.
10
Department of Anatomy and Cell Biology, Center for Gene Therapy, The University of Iowa, Iowa City, IA, USA.
11
Department of Microbiology, Molecular Genetics and Immunology, University of Kansas Medical Center, Kansas City, KS, USA.
12
German Center for Infection Research (DZIF), partner site Heidelberg, Heidelberg, Germany.

Abstract

Parvoviruses are highly attractive templates for the engineering of safe, efficient, and specific gene therapy vectors, as best exemplified by adeno-associated virus (AAV). Another candidate that currently garners increasing attention is human bocavirus 1 (HBoV1). Notably, HBoV1 capsids can cross-package recombinant (r)AAV2 genomes, yielding rAAV2/HBoV1 chimeras that specifically transduce polarized human airway epithelia (pHAEs). Here, we largely expanded the repertoire of rAAV/BoV chimeras, by assembling packaging plasmids encoding the capsid genes of four additional primate bocaviruses, HBoV2-4 and GBoV (Gorilla BoV). Capsid protein expression and efficient rAAV cross-packaging were validated by immunoblotting and qPCR, respectively. Interestingly, not only HBoV1 but also HBoV4 and GBoV transduced pHAEs as well as primary human lung organoids. Flow cytometry analysis of pHAEs revealed distinct cellular specificities between the BoV isolates, with HBoV1 targeting ciliated, club, and KRT5+ basal cells, whereas HBoV4 showed a preference for KRT5+ basal cells. Surprisingly, primary human hepatocytes, skeletal muscle cells, and T cells were also highly amenable to rAAV/BoV transduction. Finally, we adapted our pipeline for AAV capsid gene shuffling to all five BoV isolates. Collectively, our chimeric rAAV/BoV vectors and bocaviral capsid library represent valuable new resources to dissect BoV biology and to breed unique gene therapy vectors.

KEYWORDS:

AAV; BoV; adeno-associated virus; bocavirus; capsid engineering

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