Off-target genomic DNA sites with partial sgRNA complementarity (CRISPR off-targets)

Off-target genomic DNA sites with partial sgRNA complementarity refer to unintended locations in the genome where a CRISPR-Cas9 complex binds and introduces double-strand breaks (Hsu et al., 2013, Nature Biotechnology). These sites possess DNA sequences that are similar, but not identical, to the intended target sequence defined by the single guide RNA (sgRNA). Because the Cas9 enzyme can tolerate a limited number of mismatches between the sgRNA and the genomic DNA, it may catalyze cleavage at these non-target loci (Tsai et al., 2015, Nature Methods). The subsequent repair of these breaks by cellular mechanisms like non-homologous end joining (NHEJ) can result in permanent insertions, deletions, or translocations (Stadtmauer et al., 2020, Science). In a clinical context, such off-target effects pose significant safety risks, including the potential for oncogenic transformation or the disruption of vital cellular functions (Kosicki et al., 2018, Nature Biotechnology). Consequently, identifying and minimizing these interactions using tools like GUIDE-seq or CIRCLE-seq is a critical component of developing safe CRISPR-based therapeutics like Exagamglogene autotemcel (FDA, 2023).

Other names

CRISPR off-target effectsNon-specific cleavage sitesMismatched DNA sequencesOff-target mutationsOff-target effects (OTEs)

Unintended binding and cleavage of DNA by CRISPR-Cas9 complexes due to sequence homology with the guide RNA, leading to double-strand breaks and subsequent error-prone repair (Hsu et al., 2013).