New CRISPR tool selectively cuts tumor DNA while sparing healthy cells

Cancer cells excel at evading detection, but subtle chemical differences set them apart from healthy cells. Now, a team of scientists from Wageningen University & Research and Van Andel Institute has identified a way to exploit this distinction. Using a variant of CRISPR, a modern tool for editing DNA, they distinguished tumor DNA from healthy DNA and selectively cut only the former. The study, published today in Nature, is an early but promising step toward a cancer therapy that targets and destroys tumor cells with high precision.

The new method relies on methyl groups, small chemical tags attached to DNA that regulate whether genes are on or off. This process, called DNA methylation, is altered in cancer cells and can act as a molecular "fingerprint" that differentiates malignant cells from healthy ones.

Precision gene editing with ThermoCas9

The team conducted the study using ThermoCas9, a CRISPR variant discovered in bacteria several years ago by Wageningen's John van der Oost, Ph.D. Like other CRISPR systems, researchers can program ThermoCas9 to locate and cut specific sections of DNA within a cell. VAI's Hong Li, Ph.D., and her lab analyzed ThermoCas9's structure and found that it can distinguish between unmethylated and methylated genes.

The team then introduced ThermoCas9 into human cells grown in culture dishes: healthy cells in one set of dishes and tumor cells in another set. This approach worked: ThermoCas9 cut DNA in tumor cells while leaving healthy DNA intact. The system therefore proved capable of detecting the subtle chemical differences between healthy and tumor cells and acting on them.

ThermoCas9 is the first CRISPR-associated enzyme to respond to differences in the most abundant type of DNA methylation in human and other eukaryotic cells. This means we now have a system that we can target specifically toward tumor cells."

John van der Oost, Ph.D., Wageningen University & Research

The study represents the first time a CRISPR-based method has relied on methylation to target human cancer cells.

"ThermoCas9 uses methylation like an address to precisely target cancer cells while leaving healthy cells untouched," Li said. "The findings could be a game changer." 

A precise molecular fit

The explanation for ThermoCas9's selective behavior lies in the way it binds to DNA. Before a CRISPR system cuts DNA, it must first attach to a short recognition sequence next to its target, known as the PAM (Protospacer Adjacent Motif). ThermoCas9 is unique in that its PAM sequence includes a human methylation site, meaning it can contain a methyl group.

"The CRISPR system binds very precisely to this recognition code," Van der Oost explained.

Compare it to a screwdriver that fits perfectly into a matching screw head. If there is a protrusion inside the groove, the screwdriver no longer fits, nor is it capable of performing its job. In the same way, a methyl group disrupts the fit between ThermoCas9 and the DNA, preventing binding and leaving the DNA sequence untouched. 

"ThermoCas9 is a perfect example of the value of fundamental research; you have to know how these individual pieces work together," Li said. "We used biochemistry and structural biology to discover a mechanism that we one day hope will lead to more precise, effective cancer treatment."

Steps toward clinical research

There is still a long way to go before the technology can be translated into a potential cancer treatment. The new study demonstrates selective DNA cleavage but does not yet show that this effect can kill tumor cells. The next step focuses on damaging tumor DNA sufficiently to trigger cell death.

Aberrant methylation patterns also play a role in many other diseases, including childhood cancers such as neuroblastoma and autoimmune disorders. In the future, ThermoCas9 or a similar CRISPR tool may evolve into a versatile molecular strategy that recognizes diseased cells by their chemical "signature" and selectively disables them.