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Structure-Function Characterization of Non-Primate AAV Capsids for Their Usage as Therapeutic Gene Delivery Vectors

Mietzsch, Mario; Fielding, Victoria E.; Pénzes, Judit J; Chipman, Paul; McKenna, Robert; DiPasquale, Giovanni; Chiorini, John A; Li, Ya; Yang, Lin; Yu, Qian; Zolotukhin, Sergei; Tijssen, Peter; Agbandje-McKenna, Mavis . Structure-Function Characterization of Non-Primate AAV Capsids for Their Usage as Therapeutic Gene Delivery Vectors In: ASGCT 21st Annual Meeting, 16-19 mai 2018, Chicago.

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Résumé

All adeno-associated virus (AAV) vectors currently used in gene therapy trials or approved human gene therapy biologics are based on primate AAVs. One hurdle for AAV vectors in human gene therapy are preexisting neutralizing antibodies (NAb) that target virus capsids, leading to vector inactivation and to a loss of treatment efficacy. To overcome this issue the capsid surface of these AAVs can be mutated to escape from Nabs. An alternative to this strategy is the utilization of AAVs that do not exist in the primate population, but have the ability to infect mammalian cells and exhibit low or no reactivity with human sera. Here we present the capsid structures of bovine AAV (BAAV), BatAAV and snake AAV (SAAV) determined by cryo-electron microscopy to a resolution of 2.57 Å, 2.98 Å, and 3.15 Å, respectively. These viruses show very low capsid sequence identities (~51-59%) to AAV2,to other primate AAVs, and to each other, as has been reported for the sequence and structurally diverse AAV4 and AAV5 serotypes. While the core structure of the capsids of these non-primate AAVs are conserved, their surface loops display unique structural features different to the primate AAVs and each other. These differences are located in the previously defined capsid variable regions (VR). Significant structural differences include a large insertion in VR-V of BAAV, large insertions in VR-III and VR-VII of BatAAV, and distinct truncations of VR-IV of BatAAV and SAAV. Many of the surface loops have previously been identified as binding sites for NAbs in the primate AAVs. Native immuno-dot-blots confirm the lack of antigenic reactivity of BAAV, BatAAV, and SAAV towards different anti-AAV antibodies or human sera. Similar to the primate AAVs, vectors packaging a recombinant genome into the non-primate AAVs can be generated by triple transfection of HEK 293 cells. While the transduction of different mammalian cell lines by BAAV vectors have previously been reported, including the identification of sialic acid-carrying gangliosides and the Chitotriose trisaccharide as receptor, nothing is known about the receptor or tropism of BatAAV or SAAV. Here we pinpoint the region of BAAV required for receptor attachment and show that SAAV also requires terminal sialic acids for cell attachment. Furthermore, SAAV-GFP vectors are capable of transducing CHO Pro5 cells leading to cellular GFP expression. This structure-function characterization of the non-primate AAVs will be beneficial to further expand the current repertoire of AAV vectors in human gene therapy applications

Type de document: Document issu d'une conférence ou d'un atelier
Informations complémentaires: Molecular Therapy 26 (5):435
Mots-clés libres: -
Centre: Centre INRS-Institut Armand Frappier
Date de dépôt: 16 août 2018 03:00
Dernière modification: 22 mars 2019 15:24
URI: http://espace.inrs.ca/id/eprint/7494

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