CRISPR prime editing for unconstrained correction of oncogenicKRASvariants

ABSTRACT KRAS is the most commonly mutated RAS family gene and is a primary cause of the occurrence of several types of cancer. However, KRAS mutations have several unique and diverse molecular identities, making it difficult to find specific treatments. Here, we developed universal pegRNAs which can correct all types of G12 and G13 oncogenic KRAS mutations with CRISPR-mediated prime editors (PEs). The universal pegRNAs were found to be capable of correcting 12 types of KRAS mutations, accounting for 94% of all known KRAS mutations. Using our PE system, we successfully corrected 12 types of KRAS mutations with up to 54.7 % correction efficiency in HEK293T/17 cells. We also applied the universal pegRNA to correct endogenous KRAS mutations in pancreatic cancer cells and found that G12V and G12D KRAS mutations were successfully corrected to wild-type KRAS sequences with up to 18.7% correction efficiency without unwanted mutations. We propose prime editing with the universal pegRNA as a ‘one–to–many’ potential therapeutic strategy for KRAS oncogene variants.