Exosome-associated AAV vector as a robust and convenient neuroscience tool
Identifieur interne : 000285 ( Main/Merge ); précédent : 000284; suivant : 000286Exosome-associated AAV vector as a robust and convenient neuroscience tool
Auteurs : Eloise Hudry [États-Unis] ; Courtney Martin [États-Unis] ; Sheetal Gandhi [États-Unis] ; Bence György [États-Unis] ; Deborah I. Scheffer [États-Unis] ; Dakai Mu [États-Unis] ; Steven F. Merkel [États-Unis] ; Federico Mingozzi [France] ; Zachary Fitzpatrick [États-Unis] ; Hemi Dimant [États-Unis] ; Marissa Masek [États-Unis] ; Tim Ragan [États-Unis] ; Sisareuth Tan [France] ; Alain R. Brisson [France] ; Servio H. Ramirez [États-Unis] ; Bradley T. Hyman [États-Unis] ; Casey A. Maguire [États-Unis]Source :
- Gene therapy [ 0969-7128 ] ; 2016.
Abstract
Adeno-associated virus (AAV) vectors are showing promise in gene therapy trials and have proven to be extremely efficient biological tools in basic neuroscience research. One major limitation to their widespread use in the neuroscience laboratory is the cost, labor, skill, and time intense purification process of AAV. We have recently shown that AAV can associate with exosomes (exo-AAV) when vector is isolated from conditioned media of producer cells, and the exo-AAV is more resistant to neutralizing anti-AAV antibodies compared to standard AAV. Here we demonstrate that simple pelleting of exo-AAV from media via ultracentrifugation, results in high-titer vector preparations capable of efficient transduction of central nervous system (CNS) cells after systemic injection in mice. We observed that exo-AAV is more efficient at gene delivery to the brain at low vector doses relative to conventional AAV, even when derived from a serotype that does not normally efficiently cross the blood brain barrier. Similar cell types were transduced by exo-AAV and conventionally purified vector. Importantly, no cellular toxicity was noted in exo-AAV transduced cells. We demonstrated the utility and robustness of exo-AAV-mediated gene delivery by detecting direct GFP fluorescence after systemic injection, allowing 3-dimensional reconstruction of transduced Purkinje cells in the cerebellum using
Url:
DOI: 10.1038/gt.2016.11
PubMed: 26836117
PubMed Central: 4824662
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PMC:4824662Le document en format XML
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</author>
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<country xml:lang="fr">France</country>
<wicri:regionArea>UMR-CBMN CNRS-University of Bordeaux, Pessac</wicri:regionArea>
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<country xml:lang="fr">États-Unis</country>
<placeName><region type="state">Pennsylvanie</region>
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<wicri:cityArea>Department of Pathology and Laboratory Medicine, Temple University School of Medicine, Philadelphia</wicri:cityArea>
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<wicri:cityArea>Alzheimer Research Unit, The Massachusetts General Hospital Institute for Neurodegenerative Disease, Charlestown</wicri:cityArea>
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<affiliation wicri:level="3"><nlm:aff id="A2">Department of Neurology, The Massachusetts General Hospital, and NeuroDiscovery Center, Harvard Medical School, Boston, USA</nlm:aff>
<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Department of Neurology, The Massachusetts General Hospital, and NeuroDiscovery Center, Harvard Medical School, Boston</wicri:regionArea>
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<country xml:lang="fr">États-Unis</country>
<wicri:regionArea>Department of Neurology, The Massachusetts General Hospital, and NeuroDiscovery Center, Harvard Medical School, Boston</wicri:regionArea>
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<series><title level="j">Gene therapy</title>
<idno type="ISSN">0969-7128</idno>
<idno type="eISSN">1476-5462</idno>
<imprint><date when="2016">2016</date>
</imprint>
</series>
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<profileDesc><textClass></textClass>
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<front><div type="abstract" xml:lang="en"><p id="P1">Adeno-associated virus (AAV) vectors are showing promise in gene therapy trials and have proven to be extremely efficient biological tools in basic neuroscience research. One major limitation to their widespread use in the neuroscience laboratory is the cost, labor, skill, and time intense purification process of AAV. We have recently shown that AAV can associate with exosomes (exo-AAV) when vector is isolated from conditioned media of producer cells, and the exo-AAV is more resistant to neutralizing anti-AAV antibodies compared to standard AAV. Here we demonstrate that simple pelleting of exo-AAV from media via ultracentrifugation, results in high-titer vector preparations capable of efficient transduction of central nervous system (CNS) cells after systemic injection in mice. We observed that exo-AAV is more efficient at gene delivery to the brain at low vector doses relative to conventional AAV, even when derived from a serotype that does not normally efficiently cross the blood brain barrier. Similar cell types were transduced by exo-AAV and conventionally purified vector. Importantly, no cellular toxicity was noted in exo-AAV transduced cells. We demonstrated the utility and robustness of exo-AAV-mediated gene delivery by detecting direct GFP fluorescence after systemic injection, allowing 3-dimensional reconstruction of transduced Purkinje cells in the cerebellum using <italic>ex-vivo</italic>
serial 2-photon tomography. The ease of isolation combined with the high efficiency of transgene expression in the CNS, may enable widespread use of exo-AAV as a neuroscience research tool. Furthermore, the ability of exo-AAV to evade neutralizing antibodies while still transducing CNS after peripheral delivery is clinically relevant.</p>
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