Off-target genomic DNA site and bystander adenine

Off-target genomic DNA sites and bystander adenines represent the unintended consequences and safety liabilities associated with genome editing technologies, particularly base editors and CRISPR-Cas systems. Off-target effects occur when the editing machinery, guided by a programmable RNA, binds to and modifies DNA sequences that possess high homology but are not the intended target (Gaudelli et al., 2017, Nature). This can lead to deleterious mutations across the genome, potentially disrupting functional elements or coding sequences. Bystander adenines specifically refer to non-target adenine residues located within the active catalytic window of an Adenine Base Editor (ABE). Because these editors often lack single-nucleotide precision within this window, they may convert multiple adenines to guanine, leading to unintended amino acid changes or splice site disruptions (Komor et al., 2016, Nature). These phenomena are critical hurdles in the clinical translation of genetic therapies, as they can result in permanent genomic alterations (Rees & Liu, 2018, Nature Reviews Genetics). Such alterations may include the inactivation of tumor suppressor genes or the creation of oncogenic drivers. Current research focuses on engineering high-fidelity variants and optimizing guide RNA design to narrow the editing window and increase specificity (Anzalone et al., 2020, Nature Biotechnology). Mitigating these risks is essential for ensuring the genotoxic safety of precision medicine applications. Ultimately, characterizing these sites is a requirement for regulatory approval of any CRISPR-based therapeutic.

Other names

Off-target effectsBystander editingUnintended genomic modificationsCollateral mutationsOff-target adenine deaminationNon-target DNA modificationsBystander effect

Unintended enzymatic deamination of adenine to inosine (read as guanine) by deoxyadenosine deaminase domains or Cas-mediated cleavage at non-homologous genomic loci.