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Retained chromosomal integrity following CRISPR-Cas9-based mutational correction in human embryos

Retained chromosomal integrity following CRISPR-Cas9-based mutational correction in human embryos

Authors :
Bekaert, Bieke
Boel, Annekatrien
De Witte, Lisa
Vandenberghe, Winter
Popovic, Mina
Stamatiadis, Panagiotis
Cosemans, Gwenny
Tordeurs, Lise
De Loore, Athina-Maria
Chuva de Sousa Lopes, Susana Marina
De Sutter, Petra
Stoop, Dominic
Coucke, Paul
Menten, Björn
Heindryckx, Björn
Source :
Molecular Therapy; August 2023, Vol. 31 Issue: 8 p2326-2341, 16p
Publication Year :
2023

Abstract

Human germline gene correction by targeted nucleases holds great promise for reducing mutation transmission. However, recent studies have reported concerning observations in CRISPR-Cas9-targeted human embryos, including mosaicism and loss of heterozygosity (LOH). The latter has been associated with either gene conversion or (partial) chromosome loss events. In this study, we aimed to correct a heterozygous basepair substitution in PLCZ1, related to infertility. In 36% of the targeted embryos that originated from mutant sperm, only wild-type alleles were observed. By performing genome-wide double-digest restriction site-associated DNA sequencing, integrity of the targeted chromosome (i.e., no deletions larger than 3 Mb or chromosome loss) was confirmed in all seven targeted GENType-analyzed embryos (mutant editing and absence of mutation), while short-range LOH events (shorter than 10 Mb) were clearly observed by single-nucleotide polymorphism assessment in two of these embryos. These results fuel the currently ongoing discussion on double-strand break repair in early human embryos, making a case for the occurrence of gene conversion events or partial template-based homology-directed repair.

Details

Language :
English
ISSN :
15250016 and 15250024
Volume :
31
Issue :
8
Database :
Supplemental Index
Journal :
Molecular Therapy
Publication Type :
Periodical
Accession number :
ejs63483968
Full Text :
https://doi.org/10.1016/j.ymthe.2023.06.013