Senior Scientist AstraZeneca Mölndal, Vastra Gotaland, Sweden
Abstract: Genome editing through CRISPR/Cas9-induced DNA double-strand breaks offers promising opportunities to correct various genetic defects. However, the simultaneous activation of DNA repair pathways to repair the targeted lesion leads to a low efficiency of templated DNA integration and the generation of unwanted mutations, representing major limitations of genome editing applications. To overcome these challenges, we developed pharmacological treatments that modulate DNA repair outcomes and improve precise genome editing efficiency. Specifically, we identified potent and selective compounds that inhibit DNA-dependent Protein Kinase (DNA-PK) and DNA Polymerase ϴ (Polϴ), both involved in mutagenic end-joining repair pathways. The combined treatment, dubbed 2iHDR, significantly and robustly boosts homology-directed repair-mediated precise gene editing, resulting in up to 80% efficiency of templated insertions with minimal unintended Insertion-Deletions (InDels). Importantly, this strategy also reduces the risk of Cas9-related off-target effects, improving the performance and safety of this gene editing tool.
