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Self-inactivating helper virus for the production of high-capacity adenoviral vectors
Autor(es) : Gonzalez-Aparicio, M. (Manuela)
Mauleon, I. (Itsaso)
Alzuguren, P. (Pilar)
Buñuales, M. (M.)
Gonzalez-Aseguinolaza, G. (Gloria)
Sanmartin-Grijalba, C. (Carmen)
Prieto, J. (Jesús)
Hernandez-Alcoceba, R. (Rubén)
Palabras clave : Helper adenovirus
Cre recombinase
High-capacity adenovirus
Tet-on system
Fecha incorporación: 2011
Editorial : Nature Publishing Group
Versión del editor: http://www.nature.com/gt/journal/vaop/ncurrent/full/gt201158a.html
ISSN: 1476-5462
Cita: Gonzalez-Aparicio M, Mauleon I, Alzuguren P, Buñuales M, Gonzalez-Aseguinolaza G, San Martin C, et al. Self-inactivating helper virus for the production of high-capacity adenoviral vectors. Gene Ther 2011 Apr 28.
Resumen
Standard methods for producing high-capacity adenoviral vectors (HC-Ads) are based on co-infection with a helper adenovirus (HV). To avoid HV encapsidation, its packaging signal (Ψ) is flanked by recognition sequences for recombinases expressed in the producing cells. However, accumulation of HV and low yield of HC-Ad are frequently observed, due in part to insufficient recombinase expression. We describe here a novel HV (AdTetCre) in which Ψ is flanked by loxP sites that can be excised by a chimeric MerCreMer recombinase encoded in the same viral genome. Efficient modulation of cleavage was obtained by simultaneous control of MerCreMer expression using a tet-on inducible system, and translocation to the nucleus by 4-hydroxytamoxifen (TAM). Encapsidation of AdTetCre was strongly inhibited by TAM plus doxycicline. Using AdTetCre and 293Cre4 cells for the production of HC-Ads, we found that cellular and virus-encoded recombinases cooperate to minimize HV contamination. The method was highly reproducible and allowed the routine production of different HC-Ads in a medium-scale laboratory setting in adherent cells, with titers >1010 infectious units and <0.1% HV contamination. The residual HVs lacked Ψ and were highly attenuated. We conclude that self-inactivating HVs based on virally encoded recombinases are promising tools for the production of HC-Ads.
Enlace permanente: http://hdl.handle.net/10171/17859
Aparece en las colecciones: DA - CIMA - Terapia génica y Hepatología - Terapia génica hepatitis virales - Artículos de revista
DA - CIMA - Terapia génica y Hepatología - Terapia génica del cáncer - Artículos de revista

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