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Abstract
AAV vectors poorly transduce Dendritic cells (DC), a feature invoked to explain AAV’s low immunogenicity. However, the reason for this non-permissiveness remained elusive. Here, we performed an in-depth analysis using human monocyte-derived immature DC (iDC) as model. iDC internalized AAV vectors of various serotypes, but even the most efficient serotype failed to transduce iDC above background. Since AAV vectors reached the cell nucleus, we hypothesized that AAV’s intracellular processing occurs suboptimal. On this basis, we screened an AAV peptide display library for capsid variants more suitable for DC transduction and identified the I/VSS family which transduced DC with efficiencies of up to 38%. This property correlated with an improved vector uncoating. To determine the consequence of this novel feature for AAV’s in vivo performance, we engineered one of the lead candidates to express a cytoplasmic form of ovalbumin, a highly immunogenic model antigen, and assayed transduction efficiency as well as immunogenicity. The capsid variant clearly outperformed the parental serotype in muscle transduction and in inducing antigen-specific humoral and T cell responses as well as anti-capsid CD8+ T cells. Hence, vector uncoating represents a major barrier hampering AAV vector-mediated transduction of DC and impacts on its use as vaccine platform.
Highlights
Adeno-associated viral (AAV) vectors have become standard tools for in vivo gene transfer[1]
Due to the key function of Dendritic cells (DC) in inducing and shaping adaptive immune responses, the DC-AAV interaction determines vector and transgene-product immunogenicity with a major impact on long-term transgene expression when AAV are used in gene therapy and, on vaccination efficiency when used as vaccines[14,20]
The entry and the transduction efficiency were monitored in parallel (Supplementary Fig. S6). These analyses indicated the presence of higher amounts of episomes in immature DC (iDC) and mDCpi transduced with VSSTSPR compared to AAV2 at 24 hrs p.i. (Fig. 5C)
Summary
Adeno-associated viral (AAV) vectors have become standard tools for in vivo gene transfer[1]. Due to the key function of DC in inducing and shaping adaptive immune responses, the DC-AAV interaction determines vector and transgene-product immunogenicity with a major impact on long-term transgene expression when AAV are used in gene therapy and, on vaccination efficiency when used as vaccines[14,20]. By in vitro high-throughput selection screening of our AAV2 peptide display library on iDC we identified a family of capsid variants (I/VSS group) that showed improved transduction of human DC This property correlated with improved uncoating capacity as compared to the parental serotype. Inefficient intracellular processing of AAV particles and, inefficient vector uncoating are barriers that limit AAV vector-mediated transduction of DC Such barriers, can be overcome by capsid modifications, such as those selected for the VSSTSPR variant, which significantly improved transgene expression levels and help advance this promising vector system for novel applications
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