AAV's anatomy: roadmap for optimizing vectors for translational success

Abstract

Adeno-Associated Virus based vectors (rAAV) are advantageous for human gene therapy due to low inflammatory responses, lack of toxicity, natural persistence, and ability to transencapsidate the genome allowing large variations in vector biology and tropism. Over sixty clinical trials have been conducted using rAAV serotype 2 for gene delivery with a number demonstrating success in immunoprivileged sites, including the retina and the CNS. Furthermore, an increasing number of trials have been initiated utilizing other serotypes of AAV to exploit vector tropism, trafficking, and expression efficiency. While these trials have demonstrated success in safety with emerging success in clinical outcomes, one benefit has been identification of issues associated with vector administration in humans (e.g. the role of pre-existing antibody responses, loss of transgene expression in non-immunoprivileged sites, and low transgene expression levels). For these reasons, several strategies are being used to optimize rAAV vectors, ranging from addition of exogenous agents for immune evasion to optimization of the transgene cassette for enhanced therapeutic output. By far, the vast majority of approaches have focused on genetic manipulation of the viral capsid. These methods include rational mutagenesis, engineering of targeting peptides, generation of chimeric particles, library and directed evolution approaches, as well as immune evasion modifications. Overall, these modifications have created a new repertoire of AAV vectors with improved targeting, transgene expression, and immune evasion. Continued work in these areas should synergize strategies to improve capsids and transgene cassettes that will eventually lead to optimized vectors ideally suited for translational success.

Fig. (1)

Past and Current Clinical Trials Utilizing rAAV. The percentage of clinical trials using rAAV (A) in different categories of disease, (B) in different phases, and (C) using capsids of different serotypes of AAV. Other serotypes includes AAV8, AAV2.5, AAV6, and AAVrh.10. Clinical trial data was compiled from the Gene Therapy Clinical Trials Worldwide Database (http://www.wiley.co.uk/genetherapy/clinical).

Fig. (2)

Representation of various strategies to modify AAV capsids for tissue targeting and immune evasion. A. Example of a mosaic virus, where plasmids encoding for the capsids of two different serotypes are combined in a molar ratio of 1:5 , AAVx; , AAVy. B. Example of a chimeric virus (adapted from chimeric-1829 [195]). , AAV1; , AAV8; , AAV2; , AAV9. C. Combinatorial representation of human and murine antibody and cell mediated epitopes of AAV serotypes displayed on an AAV2 background. , , , [243]; , [246]; ○, [244]; , [238]; , [245]; , [247]; , AAV2 background. D. Capsid modifications rendering ability to escape neutralization displayed on an AAV2 background. , [264]; , [162]; , [208]; , [263]; , AAV2 background. The available structure of the AAV2 monomer (PDB accession no. 1LP3) was supplied as a template for capsid generation. Capsid features were rendered using PyMOL (http://pymol.sourceforge.net). For figure 2a, monomers were structurally aligned to an AAV2 template using PyMOL. For figure 2b-d, AAV2 models were generated with VIPER [287].