A couple of decades after the discovery of systems that could selectively target DNA, we’re starting to see the first therapies based on gene editing. One challenge these developments have faced is safety. While we can make them pretty specific to the gene we want edited, the human genome is very large, and even rare DNA sequences can appear a couple of times by chance.

As a result, all the original gene-editing systems had known rates of what are called off-target effects, in which they simply edit the wrong sequence. This may be a low-probability event, but edit enough cells—and therapies generally have to edit many—and errors become inevitable.

A lot of effort has gone into finding ways to minimize or eliminate off-target edits. In a recent issue of Nature, researchers described modifying the AI protein-folding software AlphaFold to help identify key areas of gene-editing proteins responsible for off-target effects. Those areas were then modified to reduce the problems.

Gene editing and off-target effects depend on three key components. The first is guide RNA, which can base-pair with the targeted genome sequence. It’s possible to design many guide RNAs that all target the same gene, so one approach to making the system safer is to pick sequences that don’t share much similarity with any other genomic locations. This is now widely used as part of the basic design process for a gene-editing system.

The second component is a Cas protein named after the CRISPR system’s Cas9 protein. It interacts with both the guide RNA and genomic RNA and helps enforce the specificity of the interactions. If there are too many mismatches in the base pairing between the RNA and DNA, Cas9 (or other Cas family members) won’t stick there. Various approaches have led to improved Cas family members that have reduced the tendency to enable off-target edits.