An engineered xCas12i with high activity, high specificity, and broad PAM range
Key findings
Engineered hfCas12Max (high-fidelity Cas12Max) through arginine substitutions at N243R, E336R, and D892R positions in the xCas12i scaffold. hfCas12Max recognizes broad 5'-TN and 5'-TNN PAM sequences and achieves average 70% editing efficiency at TTR loci, superior to SpCas9, LbCas12a, and Ultra AsCas12a. Demonstrated robust in vivo editing via LNP delivery (~70% efficiency at 0.3-0.5 mpk) and ex vivo T cell editing via RNP delivery (~90% efficiency), with minimal off-target activity.