WASHINGTON —

A National Institutes of Health (NIH)-funded research team has discovered an enhanced CRISPR gene-editing system that could enable targeted delivery inside the human body, marking a key step toward broader clinical use.

Overcoming Delivery Limitations

Researchers identified a naturally occurring enzyme, Al3Cas12f, that is small enough to fit into adeno-associated virus (AAV) vectors, a leading targeted delivery method for gene therapies. They then engineered an enhanced version that dramatically improved gene-editing performance in human cells. This advance addresses a major limitation in CRISPR technology, as commonly used gene-editing proteins are too large for targeted delivery systems, restricting clinical applications to cells modified outside the body, such as blood and bone marrow.

Structural Stability and Enhanced Efficiency

Using imaging and machine learning tools, researchers at the University of Texas at Austin analyzed the enzyme’s structure. They found it forms a more stable and tightly connected complex than other enzymes of a similar size, allowing it to function more effectively in human cells. "The expanded interface means the enzyme is much more stable," said corresponding author David Taylor, Ph.D., a molecular bioscience professor at UT Austin. "Compared to the others we looked at, Al3Cas12f basically comes preassembled and ready to go shortly after its pieces are produced."

The team engineered a variant, known as Al3Cas12f RKK, which significantly improved editing efficiency from less than 10% to more than 80% across tested targets. In a commonly edited region of the genome, efficiency reached an unprecedented 90%. The team introduced instructions for RKK directly into a line of human cells originally isolated from a patient with leukemia. Mutations in several of the genes they aimed to edit were associated with diseases such as cancer, atherosclerosis, and amyotrophic lateral sclerosis (ALS).

Future Clinical Implications

The authors expect to build on their encouraging results by conducting tests of the nuclease’s performance when packaged into AAV vectors. If successful, this could bring gene editing therapy for many hereditary and acquired diseases much closer to clinical reality.