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Fabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmid

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dc.contributor.authorJo, Amien
dc.contributor.authorRingel-Scaia, Veronica M.en
dc.contributor.authorMcDaniel, Dylan K.en
dc.contributor.authorThomas, Cassidy A.en
dc.contributor.authorZhang, Ruien
dc.contributor.authorRiffle, Judy S.en
dc.contributor.authorAllen, Irving C.en
dc.contributor.authorDavis, Richey M.en
dc.contributor.departmentBiomedical Sciences and Pathobiologyen
dc.contributor.departmentChemical Engineeringen
dc.contributor.departmentChemistryen
dc.contributor.departmentMacromolecules Innovation Instituteen
dc.contributor.departmentVirginia Tech Carilion School of Medicineen
dc.date.accessioned2020-02-03T12:59:21Zen
dc.date.available2020-02-03T12:59:21Zen
dc.date.issued2020-01-20en
dc.date.updated2020-01-26T04:12:46Zen
dc.description.abstractBackground The clustered regularly interspaced short palindromic repeats (CRISPR) and Cas9 protein system is a revolutionary tool for gene therapy. Despite promising reports of the utility of CRISPR–Cas9 for in vivo gene editing, a principal problem in implementing this new process is delivery of high molecular weight DNA into cells. Results Using poly(lactic-co-glycolic acid) (PLGA), a nanoparticle carrier was designed to deliver a model CRISPR–Cas9 plasmid into primary bone marrow derived macrophages. The engineered PLGA-based carriers were approximately 160 nm and fluorescently labeled by encapsulation of the fluorophore 6,13-bis(triisopropylsilylethynyl) pentacene (TIPS pentacene). An amine-end capped PLGA encapsulated 1.6 wt% DNA, with an encapsulation efficiency of 80%. Release studies revealed that most of the DNA was released within the first 24 h and corresponded to ~ 2–3 plasmid copies released per nanoparticle. In vitro experiments conducted with murine bone marrow derived macrophages demonstrated that after 24 h of treatment with the PLGA-encapsulated CRISPR plasmids, the majority of cells were positive for TIPS pentacene and the protein Cas9 was detectable within the cells. Conclusions In this work, plasmids for the CRISPR–Cas9 system were encapsulated in nanoparticles comprised of PLGA and were shown to induce expression of bacterial Cas9 in murine bone marrow derived macrophages in vitro. These results suggest that this nanoparticle-based plasmid delivery method can be effective for future in vivo applications of the CRISPR–Cas9 system.en
dc.description.versionPublished versionen
dc.format.mimetypeapplication/pdfen
dc.identifier.citationJournal of Nanobiotechnology. 2020 Jan 20;18(1):16en
dc.identifier.doihttps://doi.org/10.1186/s12951-019-0564-1en
dc.identifier.urihttp://hdl.handle.net/10919/96659en
dc.language.isoenen
dc.rightsCreative Commons Attribution 4.0 Internationalen
dc.rights.holderThe Author(s)en
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/en
dc.titleFabrication and characterization of PLGA nanoparticles encapsulating large CRISPR–Cas9 plasmiden
dc.title.serialJournal of Nanobiotechnologyen
dc.typeArticle - Refereeden
dc.type.dcmitypeTexten

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