Thermostable Cas9 Enhances RNP Performance in Lung and Liver

The study focused on overcoming challenges related to delivering CRISPR-Cas9 ribonucleoproteins (RNPs) for in vivo editing by developing a more stable version of the enzyme. Cas9 enzymes are prone to denaturation under the conditions typically used in LNP formulations, which often involve organic solvents, so a more stable variant would be expected to offer more effective genome editing when delivered as an RNP complex.

The researchers at the University of California Berkeley engineered a thermostable Cas9 from Geobacillus stearothermophilus (GeoCas9) to generate iGeoCas9 variants with expanded PAM compatibility and optimised activity in mammalian cells. The researchers used LNPs to deliver iGeoCas9 RNPs in vivo to mouse liver and lung tissues and achieved 16-37% editing efficiency in single-dose treatments.

The method achieved successful editing of the disease-causing SFTPC gene in lung tissue with a 19% efficiency, marking a substantial improvement over traditional delivery methods like viral vectors.

Niren Murthy and Jennifer Doudna at the University of California, Berkeley, led the study, which was published yesterday in Nature Biotechnology.