Purification of recombinant adeno-associated virus type 8 vectors by ion exchange chromatography generates clinical grade vector stock

Journal of Virological Methods
Andrew M DavidoffAmit C Nathwani

Abstract

Recombinant vectors based on the recently isolated AAV serotype 8 (rAAV-8) shows great promise for gene therapy, particularly for disorders affecting the liver. Transition of this vector system to the clinic, however, is limited by the lack of an efficient scaleable purification method. In this report, we describe a simple method for purification of rAAV-8 vector particles based on ion exchange chromatography that generates vector stocks with greater than 90% purity. The average yield of purified rAAV-8 from five different vector preparation was 41%. Electron microscopy of these purified stocks revealed typical icosohedral virions with less than 10% empty particles. Liver targeted delivery of ion-exchange purified rAAV-8 vector encoding the human factor IX (hFIX) gene, resulted in plasma hFIX levels approaching 30% of normal in immunocompetent mice, which is 20-fold higher than observed with an equivalent number of rAAV-5 ion exchange purified vector particles. The method takes less then 5 h to process and purify rAAV-8 vector from producer cells and represents a significant advance on the CsCl density centrifugation technique in current use for purification of rAAV-8 vector systems and will likely facilitate the transition of ...Continue Reading

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Citations

Aug 15, 2008·Gene Therapy·A C NathwaniA M Davidoff
Oct 19, 2005·Gene Therapy·R Morenweiser
May 27, 2005·Human Gene Therapy·S Zolotukhin
Jul 8, 2005·Human Gene Therapy·Corinna BurgerRonald J Mandel
Jul 18, 2008·Briefings in Functional Genomics & Proteomics·Alejandro Negrete, Robert M Kotin
May 3, 2011·Human Molecular Genetics·Robert M Kotin
Nov 15, 2014·Human Gene Therapy·Amit C NathwaniAndrew M Davidoff
Feb 26, 2016·Molecular Therapy. Methods & Clinical Development·Taro TomonoTakashi Okada
Jul 14, 2010·Journal of Pediatric Surgery·Regan F WilliamsAndrew M Davidoff
Mar 27, 2010·International Journal of Radiation Oncology, Biology, Physics·Mackenzie C McGeeAndrew M Davidoff
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Oct 7, 2015·Molecular Therapy : the Journal of the American Society of Gene Therapy·Joshua C GriegerRichard Jude Samulski
Mar 26, 2016·Molecular Therapy. Methods & Clinical Development·Nathalie Clément, Joshua C Grieger
Sep 15, 2006·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·María de las Mercedes SeguraAlain Garnier
Feb 2, 2006·Biotechnology Advances·María de Las Mercedes SeguraAlain Garnier
Oct 13, 2011·Expert Review of Vaccines·Michael W Wolf, Udo Reichl
Oct 7, 2006·Biotechnology Progress·David L GrzeniaS Ranil Wickramasinghe
Nov 22, 2007·Experimental Neurology·Ayse UlusoyDeniz Kirik
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Jul 11, 2006·Molecular Therapy : the Journal of the American Society of Gene Therapy·Zhijian WuR Jude Samulski
Nov 26, 2015·Molecular Therapy. Methods & Clinical Development·Qiang WangJames M Wilson

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