Search

The purpose of this research is to evaluate the potential of the CRISPR-derived class of prime editors for precise correction of the E342K mutation in Alpha-1 Antitrypsin Deficiency (A1AD) and to discover genetic dependencies of the prime editing mechanism. A1AD is thus far a disease with unmet need that remains challenging to target with genome editors which would benefit from an expanded set of editing tools. Several versions of modified prime editors targeting NGA and NGC PAMs were tested with a library of pegRNAs and nickRNAs generated through PrimeDesign, a program published by the lab which originated prime editors. In doing so, we have shown that A1AT E342K is accessible for precise correction via two NGA PAMs. As prime editing technology is new, the molecular interactions with cellular repair are as of yet only putative and require elucidation that could serve to further bolster success of this and other therapeutic applications. An siRNA library of DNA repair factors was compiled and evaluated for the ability to up- or down- regulate prime editing efficiency. Through this screen, we have discovered a gene, TREX2, which influences prime editing and have validated this through an orthogonal overexpression experiment.