{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"Research Square"}],"indexed":{"date-parts":[[2025,5,14]],"date-time":"2025-05-14T06:40:49Z","timestamp":1747204849472,"version":"3.40.5"},"posted":{"date-parts":[[2023,9,6]]},"group-title":"In Review","reference-count":57,"publisher":"Springer Science and Business Media LLC","license":[{"start":{"date-parts":[[2023,9,6]],"date-time":"2023-09-06T00:00:00Z","timestamp":1693958400000},"content-version":"unspecified","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2023,7,31]]},"abstract":"<title>Abstract<\/title>\n        <p>Extracellular vesicles-associated adeno-associated viral vectors (EV-AAVs) emerged as a new opportunity for non-invasive gene therapy targeting the central nervous system (CNS). However, in previous reports, only AAV serotypes with known ability to cross the blood-brain barrier (BBB) have been used for EV-AAV production and testing through non-invasive strategies. In this work, we aimed at optimizing a size exclusion chromatography (SEC) protocol for the production and isolation of natural and biologically active brain-targeting EV-AAVs, that could be applied to any AAV serotype and further used for non-invasive gene delivery to the CNS. We performed a comparison between SEC and differential ultracentrifugation (UC) isolation protocols in terms of yield, contaminants, and transgene expression efficiency. We found that SEC allows a higher recovery of EV-AAVs, free of cell contaminating proteins and with less <italic>solo<\/italic> AAVs than UC. Remarkably, SEC-purified EV-AAVs also showed to be more potent at transgene expression than <italic>solo<\/italic> AAVs in neuronal cell lines. Brain-targeting EV-AAVs exhibited the ability to reach be brain upon intravenous administration. In conclusion, SEC-purified brain-targeting EV-AAVs show to be a promising gene delivery vector for therapy of brain disorders.<\/p>","DOI":"10.21203\/rs.3.rs-3220758\/v1","type":"posted-content","created":{"date-parts":[[2023,9,6]],"date-time":"2023-09-06T01:37:35Z","timestamp":1693964255000},"source":"Crossref","is-referenced-by-count":0,"title":["Isolation of Biologically Active Extracellular Vesicles-Associated AAVs for Gene Delivery to the Brain by Size Exclusion Chromatography"],"prefix":"10.21203","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5831-3307","authenticated-orcid":false,"given":"Luis Pereira","family":"de Almeida","sequence":"first","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Carina","family":"Henriques","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9869-0203","authenticated-orcid":false,"given":"Miguel","family":"Lopes","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Patr\u00edcia","family":"Albuquerque","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"David Rufino","family":"Ramos","sequence":"additional","affiliation":[{"name":"Massachusetts General Hospital and Harvard Medical School"}]},{"given":"Laetitia","family":"Gaspar","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Diana","family":"Lobo","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Kevin","family":"Leandro","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Ana","family":"Silva","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Rafael","family":"Baganha","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"S\u00f3nia","family":"Duarte","sequence":"additional","affiliation":[{"name":"CNC - Center for Neuroscience and Cell Biology, University of Coimbra, Portugal"}]},{"given":"Casey","family":"Maguire","sequence":"additional","affiliation":[{"name":"Massachusetts General Hospital"}]},{"given":"Magda","family":"Santana","sequence":"additional","affiliation":[{"name":"Massachusetts General Hospital"}]},{"given":"Rui","family":"Nobre","sequence":"additional","affiliation":[{"name":"Massachusetts General Hospital"}]}],"member":"297","reference":[{"issue":"1","key":"ref1","doi-asserted-by":"publisher","first-page":"7","DOI":"10.1177\/1535676019899502","article-title":"Viral Vector Systems for Gene Therapy: A Comprehensive Literature Review of Progress and Biosafety Challenges","volume":"25","author":"Ghosh S","year":"2020","unstructured":"Ghosh, S., Brown, A.M., Jenkins, C., Campbell, K.: Viral Vector Systems for Gene Therapy: A Comprehensive Literature Review of Progress and Biosafety Challenges. 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