New method boosts DNA editing efficiency and kills cancer cells in the lab

A universal approach for selection of edited cells and elimination of cells with cancer mutations

To the point

• DNA editing becomes more efficient: The new method increases the efficiency of genome editing by sevenfold and boosts the number of precisely modified cells.
• Mutation becomes the marker: In contrast to previous methods that add marker DNA, this new approach uses the edit itself as the marker, meaning that edited cells no longer match the cut site.
• Cancer mutations can be targeted: The new method can selectively kill cells with cancer mutations in a laboratory setting. However, much more research is needed before it can be used to treat cancer patients.

Genome editing enables scientists to alter the DNA sequence of cells. This technology has great potential for research and therapeutic applications. One challenge, however, is that editing often works only in a small fraction of cells. Researchers must therefore find ways to separate the successfully edited cells from all the others. In a new study, scientists at the Max Planck Institute for Evolutionary Anthropology in Leipzig present a new efficient strategy for selection, that uses programmable gene scissors to selectively kill unwanted cells. The team also demonstrates that this method can be used to precisely kill cells with cancer mutations in the lab. 

Introducing mutations into human cells by genome editing is a valuable approach to understanding diseases and developing new therapies. Genome editing works by cutting the DNA at a specific location using programmable ‘CRISPR-Cas’ gene scissors. In order to survive, cells must repair this break, and researchers can provide a DNA repair template that carries the mutation they want to introduce. However, the desired mutation is often only incorporated in a small subset of cells, making it necessary to enrich for the edited cells. Traditional selection methods depend on adding extra DNA sequences that produce fluorescence or confer resistance to certain drugs. While effective, these markers can unintentionally alter cell behavior and are not suitable for many applications.

Sevenfold increase in genome editing efficiency

To overcome these limitations, researchers at the Max Planck Institute for Evolutionary Anthropology developed a method in which the introduced mutation itself becomes the marker. After the first round of editing, the cells undergo a second round in which all DNA repair pathways are blocked. Cells that were successfully edited in the first step are protected as they no longer contain a matching target site and thus cannot be cut again. All unedited cells, however, are cut and die because they cannot repair the break.

Using this strategy, the team achieved a median sevenfold increase in editing efficiency, with some targets reaching up to 100 percent edited cells. The method was validated across 42 different genomic sites, various genome editing tools, and multiple cell types. “Our method provides an easy-to-use tool that greatly increases the number of precisely modified cells,” says Luise Fast, who led the study.

Targeting cancer-specific mutations

The researchers also demonstrated that the method can be used to selectively kill cells with cancer mutations in culture. Cancer cells carry specific mutations that distinguish them from healthy cells. By targeting these cancer-specific mutations, the new approach cuts only the DNA of the cancer cells. When repair pathways are inhibited, these cells die, but healthy cells remain intact. In some cases, cancer mutations are even repaired to the healthy state. “Our approach could offer an interesting alternative to traditional chemotherapy, which harms healthy cells as well,” says Stephan Riesenberg, senior researcher on the project. “Nevertheless, applying this method to kill cancer cells in patients will require a lot of further research, and the road from lab experiments to potential clinical use is still long.”