OptiPrime, a mechanistic machine learning model that incorporates the biological mechanism of prime editing through differential equations governing pseudo-rates of individual biochemical steps, achieves superior prediction accuracy compared to existing models and enables rapid optimization of therapeutic prime editing strategies, including achieving >40% in vivo correction efficiency in mouse brain with streamlined pegRNA design requiring only 15 candidates rather than hundreds typically needed.

Although prime editing (PE) can effect virtually any specified local change to genomic DNA in living systems, its efficient application currently requires extensive optimization of prime editing guide RNA (pegRNA) sequences. We present OptiPrime, a machine learning model of PE efficiency based on our current understanding of the mechanism of prime editing. OptiPrime achieves state-of-the-art accuracy on PE efficiency prediction and also enables prediction of nicking guide RNA (PE3) and dual pegRNA (twinPE) outcomes. We validated that OptiPrime has learned the determinants of mammalian mismatch repair (MMR), and is therefore well suited for nominating MMR-evasive silent edits that improve PE efficiency. We demonstrate the utility of OptiPrime in a variety of prospective therapeutic contexts, including in primary human and mouse cells. Finally, we show how OptiPrime can be used to achieve highly streamlined and efficient in vivo correction of a pathogenic mutation in the brain of a mouse model of KIF1A-associated neurological disorder.