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Arpad Palfi, Naomi Chadderton, Alex G. McKee, Alfonso Blanco Fernandez, Carol J. Loscher, Paul F. Kenna, G. Jane Farrar; Transgene Co-delivery Via Aav Co-transduction. Invest. Ophthalmol. Vis. Sci. 2011;52(14):1401.
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Recombinant adeno-associated virus (AAV) represents a highly efficient vector system for neuronal transduction in vivo. Additionally, AAV has many convenient qualities making the technology an ideal choice for gene delivery to the central nervous system. For example, it is suggested that the AAV genome is episomal in post-mitotic cells. Immune reaction against AAV is minimal in most cases. Moreover, different AAV capsid serotypes have been engineered, which can provide trophisms for different neuronal types. For example, in the retina, AAV2/5 efficiently transduces photoreceptor cells while AAV2/2 has a broader target range encompassing most retinal cell types. As a result, AAV is a common vector choice for neuronal, especially retinal, gene therapy. However, a potential drawback of AAV is a limited packaging capacity of approximately 4.5 kb. While ongoing research has been focused on achieving increases in the size limit, co-administration of two AAV vectors may provide an alternative solution in certain situations. For example, when delivering two or more transgenes, in theory, it is possible to administer the components using two separate vectors, thereby potentially ‘doubling’ packaging capacity. In this study, we investigate the feasibility of this approach in vivo.
AAV2/5 vectors carrying CMV promoter driven DS-Red-Express2 and EGFP reporter genes were produced. Mixtures of the two AAVs were co-administered to the subretinal space or the dorsal hippocampus in vivo. Marker expression was followed up by flow cytometry and fluorescent microscopy.
Significant co-transduction with DS-Red-Express2 and EGFP was detected in both the retina and the hippocampus using fluorescent microscopy. In photoreceptor cells, flow cytometric analysis demonstrated approximately 70% co-transduction efficiency by the two markers.
In this study, efficient co-delivery of two genes using two separate AAV vectors has been demonstrated in vivo. These results indicate that co-administration of two AAVs could deliver approximately 9.0-10.0 kb if the cargo consists of two or more genes which could be divided between the two vectors. The approach could be particularly useful for delivering gene therapies consisting of two components, such as gene suppression and replacement or gene replacement combined with growth factor supplementation.
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