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Sites in the AAV5 capsid tolerant to deletions and tandem duplications |
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| Authors: | Kaoru Hida Giovanni Di Pasquale John A Chiorini |
| Institution: | a Department of Biomedical Engineering, The Johns Hopkins University, Baltimore, MD 21218, USA b Department of Chemical and Biomolecular Engineering, The Johns Hopkins University, Baltimore, MD 21218, USA c Gene Therapy and Therapeutics Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA d Institute for NanoBioTechnology, The Johns Hopkins University, Baltimore, MD 21218, USA |
| Abstract: | Gene therapy vectors based on adeno-associated virus (AAV) have shown much promise in clinical trials for the treatment of a variety of diseases. However, the ability to manipulate and engineer the viral surface for enhanced efficiency is necessary to overcome such barriers as pre-existing immunity and transduction of non-target cells that currently limit AAV applications. Although single amino acid changes and peptide insertions at select sites have been explored previously, the tolerance of AAV to small deletions and tandem duplications of sequence has not been globally addressed. Here, we have generated a large, diverse library of >105 members containing deletions and tandem duplications throughout the viral capsid of AAV5. Four unique mutants were identified that maintain the ability to form viral particles, with one showing improved transduction on both 293T and BEAS-2B cells. This approach may find potential use for the generation of novel variants with improved and altered properties or in the identification of sites that are tolerant to insertions of targeting ligands. |
| Keywords: | Directed evolution AAV Viral gene delivery |
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