CRISPR-Cas9 Used to Uncover Immunotherapy-Resistant Genes

Human metastatic melanoma cells, stained and magnified to 320 times. (Dr. Lance Liotta Laboratory/NCI)

Immunotherapy -- using a patient’s immune system to recognize and destroy cancer cells -- has been one of the more exciting developments in cancer treatment.  Where once there was little hope for some patients, now they can be treated, and with a lot fewer side effects than conventional treatments like chemotherapy and radiation. 

But not every patient responds to immunotherapy. For example, the drug Keytruda, used to treat advanced melanoma, does not work in around 60 percent of patients.

In a new study, published this month in Nature, a group of scientists from the National Cancer Institute set out to find why immunotherapy fails in so many people. The researchers used the gene‐editing tool CRISPR‐Cas9 to uncover 554 genes that may, when mutated, cause advanced melanoma tumors to be resistant to the treatment. If scientists can find which of these genes are the culprits, drugs could potentially be created to correct them, making the cancers responsive to immunotherapy.

A Gene Implicated

Finding these 554 genes was like finding a needle in a haystack. The researchers started out with a cancer cell that responds to immunotherapy. They then disabled, one at a time in the cell, almost all of the known 19,050 human genes, cutting each in different spots. After incapacitating an additional 3,000 parts of the DNA that don't fit the classical definition of a gene, the scientists ended up generating 100,000 different cancer cells, each varying by only a single DNA change. Of these 100,000 cells, 554 showed resistance to immunotherapy.

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The researchers then showed mutation of one those genes, called APLNR, was most likely responsible for immunotherapy resistance in some cancer patients.

When the scientists disabled APLNR in a cancer cell, the cell became resistant. The scientists then added a working copy of APLNR, and the cell's resistance disappeared.

Next the researchers searched through databases with the DNA sequences of patients' immunotherapy-resistant tumors. The scientists discovered that some did have mutations in the APLNR gene.

Doing the experiment in petri dishes is only the first step. Unlike in this process, real-world cancers can’t be made sensitive to immunotherapy by adding back a working gene, because too few cells would adopt it. Instead, a drug would have to be developed. Researchers will also need to experiment with the other 553 genes to see which behave like APLNR.

CRISPR-Cas9 -- A Killer App

This gargantuan effort would have been much more difficult even a few years ago. It's only with the advent of the CRISPR‐Cas9 gene‐editing system that it could be done so efficiently.

The big advantage of the enzyme Cas9, the workhorse of the CRISPR‐Cas9 system, is how easily it can be programmed to precisely cut the right spot in the over six feet of DNA each of us has packed into every cell. It's that ease of use that allowed these scientists to program Cas9 to specifically travel to more than 100,000 different spots in the cancer cell’s DNA.

This is a really exciting use for CRISPR‐Cas9 that a lot of people have not heard of. While most news stories focus on using the system to directly cure a disease or even to make designer babies, this gene- editing tool is revolutionizing the kind of fundamental research that could help us discover new drugs.

To say nothing of what it is teaching us about basic biology.

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Dr. Barry Starr is a scientist in Stanford University's Department of Genetics. He runs both the Stanford at The Tech program and the Understanding Genetics website with The Tech Museum of Innovation in San Jose, California. He earlier worked as a research scientist in the biotechnology field.

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