Unbiased assessment of genome integrity and purging of adverse outcomes at the target locus upon editing of CD4⁺ T-cells for the treatment of Hyper IgM1.

ABSTRACT: Hyper IgM1 is an X-linked combined immunodeficiency caused by CD40LG mutations, potentially treatable with CD4⁺ T-cell gene editing with Cas9 and a "one-size-fits-most" corrective template. Contrary to established gene therapies, there is limited data on the genomic alterations following long-range gene editing, and no consensus on the relevant assays. We developed drop-off digital PCR assays for unbiased detection of large on-target deletions and found them at high frequency upon editing. Large deletions were also common upon editing different loci and cell types and using alternative Cas9 and template delivery methods. In CD40LG edited T cells, on-target deletions were counter-selected in culture and further purged by enrichment for edited cells using a selector coupled to gen

SUBMITTER: Canarutto D 

PROVIDER: S-EPMC10690452 | biostudies-literature | 2023 Dec

REPOSITORIES: biostudies-literature