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Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain

Deverman, Benjamin E. and Pravdo, Piers L. and Simpson, Bryan P. and Ravindra Kumar, Sripriya and Chan, Ken Y. and Banerjee, Abhik and Wu, Wei-Li and Yang, Bin and Huber, Nina and Pasca, Sergiu P. and Gradinaru, Viviana (2016) Cre-dependent selection yields AAV variants for widespread gene transfer to the adult brain. Nature Biotechnology, 34 (2). pp. 204-209. ISSN 1087-0156. PMCID PMC5088052. doi:10.1038/nbt.3440.

[img] PDF - Accepted Version
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[img] Image (JPEG) (Supplementary Figure 1: Capsid library fragment generation and Cre-dependent capsid sequence recovery) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 2: The most enriched variants recovered from in vivo selections in GFAP-Cre mice) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 3: AAV-PHP.B transduces several interneuron cell types and endothelial cells but does not appear to transduce microglia) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 4: Long-term eGFP expression in the brain following gene transfer with AAV-PHP.B) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 5: Representative images of native GFP fluorescence and IHC for several neuron and glial cell types following transduction by AAV-PHP.B:CAG-NLS-GFP) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 6: AAV9, AAV-PHP.A and AAV-PHP.B transduce human iPSC-derived cortical neurons and astrocytes in dissociated cultures and intact 3D cortical cultures) - Supplemental Material
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[img] Image (JPEG) (Supplementary Figure 7: AAV-PHP.B and AAV-PHP.A capsids localize to the brain vasculature after intravenous injection and transduce cells along the vasculature by 24 hours post-administration) - Supplemental Material
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[img] Video (MPEG) (Video 1: Supplementary Movie 1) - Supplemental Material
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[img] Video (MPEG) (Video 3: Supplementary Movie 3) - Supplemental Material
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[img] PDF (Supplementary Figures 1–7) - Supplemental Material
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Recombinant adeno-associated viruses (rAAVs) are commonly used vehicles for in vivo gene transfer. However, the tropism repertoire of naturally occurring AAVs is limited, prompting a search for novel AAV capsids with desired characteristics. Here we describe a capsid selection method, called Cre recombination–based AAV targeted evolution (CREATE), that enables the development of AAV capsids that more efficiently transduce defined Cre-expressing cell populations in vivo. We use CREATE to generate AAV variants that efficiently and widely transduce the adult mouse central nervous system (CNS) after intravenous injection. One variant, AAV-PHP.B, transfers genes throughout the CNS with an efficiency that is at least 40-fold greater than that of the current standard, AAV9, and transduces the majority of astrocytes and neurons across multiple CNS regions. In vitro, it transduces human neurons and astrocytes more efficiently than does AAV9, demonstrating the potential of CREATE to produce customized AAV vectors for biomedical applications.

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URLURL TypeDescription ReadCube access CentralArticle
Deverman, Benjamin E.0000-0002-6223-9303
Ravindra Kumar, Sripriya0000-0001-6033-7631
Chan, Ken Y.0000-0002-8853-5186
Banerjee, Abhik0000-0001-9127-5461
Wu, Wei-Li0000-0003-2610-1881
Pasca, Sergiu P.0000-0002-3216-3248
Gradinaru, Viviana0000-0001-5868-348X
Additional Information:© 2015 Macmillan Publishers Limited. Received 09 January 2015. Accepted 23 November 2015. Published online 01 February 2016. This article and the naming of the novel AAV clones are dedicated to the memory of Paul H. Patterson (P.H.P.), who passed away during the preparation of this manuscript. We wish to thank L. Rodriguez and P. Anguiano for administrative assistance, A. Balazs and S. Cassenaer and the entire Gradinaru and Patterson laboratories for helpful discussions, and A. Choe for helpful comments on the manuscript. We thank the University of Pennsylvania vector core for the AAV2/9 Rep-Cap plasmid, A. Balazs and D. Baltimore for the AAV genome plasmid, and the Biological Imaging Facility, supported by the Caltech Beckman Institute and the Arnold and Mabel Beckman Foundation, for use of imaging equipment. This work was supported by grants from the Hereditary Disease Foundation and the Caltech–City of Hope Biomedical Initiative (to P.H.P.) and from the National Institutes of Health (NIH) Director's New Innovator 1DP2NS087949; NIH/National Institute on Aging (NIA) 1R01AG047664; Beckman Institute for CLARITY, Optogenetics and Vector Engineering Research; and the Gordon and Betty Moore Foundation through grant GBMF2809 to the Caltech Programmable Molecular Technology Initiative (to V.G.). Work in the Gradinaru laboratory is also funded by the following awards (to V.G.): NIH BRAIN 1U01NS090577; NIH/National Institute of Mental Health (NIMH) 1R21MH103824-01; Pew Charitable Trust; Sloan Foundation; Kimmel Foundation; Human Frontiers in Science Program; Caltech-GIST; Caltech–City of Hope Biomedical Initiative. Work in the Pasca laboratory is supported by a NIMH 1R01MH100900 and 1R01MH100900-02S1, the NIMH BRAINS Award (R01MH107800), the California Institute of Regenerative Medicine (CIRM), the MQ Fellow Award and the Donald E. and Delia B. Baxter Foundation Scholar Award (to S.P.P.). Contributions: B.E.D. designed and performed experiments, analyzed data, prepared figures and wrote the manuscript. P.L.P., B.P.S., S.R.K., A.B. and K.Y.C. performed experiments, virus production and characterization. W.-L.W. performed tissue processing and IHC. B.Y. assisted with tissue clearing and imaging. N.H. and S.P.P. performed the experiments with human cells, analyzed the data, and prepared the associated figure and text. V.G. helped with study design and data analysis, manuscript and figure preparation and supervised the project. All authors edited and approved the manuscript. Competing financial interests: The California Institute of Technology has filed patent applications related to this work with B.E.D, B.Y. and V.G. listed as inventors. Accession codes. GenBank: AAV capsid sequences, KU056473, KU056474, KU056475 and KU056476. Primary accessions NCBI Reference Sequence KU056473 KU056474 KU056475 KU056476 Referenced accessions NCBI Reference Sequence AF085716.1
Funding AgencyGrant Number
Caltech Beckman InstituteUNSPECIFIED
Arnold and Mabel Beckman FoundationUNSPECIFIED
Hereditary Disease FoundationUNSPECIFIED
Caltech–City of Hope Biomedical InitiativeUNSPECIFIED
Beckman Institute for CLARITY, Optogenetics and Vector Engineering ResearchUNSPECIFIED
Gordon and Betty Moore FoundationGBMF2809
Pew Charitable TrustUNSPECIFIED
Alfred P. Sloan FoundationUNSPECIFIED
Sidney Kimmel Foundation for Cancer ResearchUNSPECIFIED
Human Frontier Science ProgramUNSPECIFIED
National Institute of Mental Health (NIMH)1R01MH100900
National Institute of Mental Health (NIMH)1R01MH100900-02S1
National Institute of Mental Health (NIMH)R01MH107800
California Institute for Regenerative Medicine (CIRM)UNSPECIFIED
Donald E. and Delia B. Baxter FoundationUNSPECIFIED
Issue or Number:2
PubMed Central ID:PMC5088052
Record Number:CaltechAUTHORS:20160201-131537838
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Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:64130
Deposited By: George Porter
Deposited On:01 Feb 2016 21:47
Last Modified:12 May 2022 18:26

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