Intrathymic AAV Delivery Results in Therapeutic Site-specific Integration at TCR Loci in Mice
Overview
Authors
Affiliations
Adeno-associated virus (AAV) vectors have been successfully exploited in gene therapy applications for the treatment of several genetic disorders. AAV is considered an episomal vector, but it has been shown to integrate within the host cell genome after the generation of double-strand DNA breaks or nicks. Although AAV integration raises some safety concerns, it can also provide therapeutic benefit; the direct intrathymic injection of an AAV harboring a therapeutic transgene results in integration in T-cell progenitors and long-term T-cell immunity. To assess the mechanisms of AAV integration, we retrieved and analyzed hundreds of AAV integration sites from lymph node-derived mature T cells and compared these with liver and brain tissue from treated mice. Notably, we found that although AAV integrations in the liver and brain were distributed across the entire mouse genome, >90% of the integrations in T cells were clustered within the T-cell receptor α, β, and γ genes. More precisely, the insertion mapped to DNA breaks created by the enzymatic activity of recombination activating genes (RAGs) during variable, diversity, and joining recombination. Our data indicate that RAG activity during T-cell receptor maturation induces a site-specific integration of AAV genomes and opens new therapeutic avenues for achieving long-term AAV-mediated gene transfer in dividing cells.
Singh K, Fronza R, Evens H, Chuah M, VandenDriessche T Mol Ther Methods Clin Dev. 2024; 32(4):101365.
PMID: 39655309 PMC: 11626537. DOI: 10.1016/j.omtm.2024.101365.
Modulation of AAV transduction and integration targeting by topoisomerase poisons.
Kasimsetty A, Hwang Y, Everett J, McFarland A, Zolnoski S, Lu T Mol Ther Methods Clin Dev. 2024; 32(4):101364.
PMID: 39618424 PMC: 11607603. DOI: 10.1016/j.omtm.2024.101364.
AAV-mediated genome editing is influenced by the formation of R-loops.
Puzzo F, Crossley M, Goswami A, Zhang F, Pekrun K, Garzon J Mol Ther. 2024; 32(12):4256-4271.
PMID: 39369271 PMC: 11638834. DOI: 10.1016/j.ymthe.2024.09.035.
Bazick H, Mao H, Niehaus J, Wolter J, Zylka M Mol Ther. 2024; 32(11):4122-4137.
PMID: 39367606 PMC: 11573598. DOI: 10.1016/j.ymthe.2024.09.032.
T-cell specific in vivo gene delivery with DART-AAVs targeted to CD8.
Demircan M, Zinser L, Michels A, Guaza-Lasheras M, John F, Gorol J Mol Ther. 2024; 32(10):3470-3484.
PMID: 39113357 PMC: 11489536. DOI: 10.1016/j.ymthe.2024.08.002.