T Cell-Resistant Cancer Genes Identified by Genome-Scale CRISPR Screen

Nicholas Restifo, MD

A new study published in the August 7 issue ofNatureidentified genes that are necessary in cancer cells for immunotherapy to work, addressing the problem of why some tumors do not respond to immunotherapy or respond initially but then stop as tumor cells develop resistance to immunotherapy. Nicholas Restifo, MD, and colleagues identified all the protein-encoding genes that must be present in a tumor for T cells to recognize and destroy it.

They used the CRISPR-Cas9 system for gene editing to systematically eliminate genes in a melanoma cell line and, as a result, were able to test every gene in the human genome for its impact on the T-cell response. Surprisingly, this screen identified dozens of new genes that potentially influence tumor cells’ susceptibility to T-cell attack. When these genes were “knocked out,” tumor cells were significantly more likely to survive and continue to multiply after exposure to T cells that the investigators had genetically engineered to recognize tumor-associated antigens.

“Many of the genes identified in our screen are involved in previously known strategies used by T cells to enhance their own ability to find and destroy their targets,” said Restifo, senior investigator with the Center for Cancer Research of the National Cancer Institute (NCI).

“There is a great deal of interest in cancer immunotherapy, especially for patients who have metastatic cancer,” continued Restifo. “The response to immunotherapy can be fantastic, but understanding why some patients don’t respond will help us improve treatments for more patients.”

It’s a frustrating problem, said Restifo, especially for many of the most common types of cancer. “So far, immunotherapies have not been highly effective against lung cancer, and they barely impact cancers of the breast, ovary, and colon,” he said. “To fully harness the potential of the body’s immune system to treat cancer, we still need to overcome major hurdles involving tumor escape.”

Cancer immunotherapy relies on T cells to destroy tumors. Restifo and his colleagues have previously shown that the infusion of large numbers of T cells can trigger complete regression of cancer in patients. They, along with other investigators, have also shown that T cells can directly recognize and kill tumor cells.

However, some tumor cells are resistant to the destruction unleashed by T cells. To research the basis for this resistance, the investigators sought to identify the genes in cancer cells that are necessary for them to be killed by T cells.

Working with a melanoma tumor cell line, the investigators used CRISPR to stop the expression of individual genes in cancer cells. By knocking out every known protein-encoding gene in the human genome and then testing the ability of the gene-modified melanoma cells to respond to T cells, they found more than 100 genes that may play a role in facilitating tumor destruction by T cells.

Once the team identified these “candidate” genes, they sought additional evidence that these genes play a role in susceptibility to T cell—mediated killing. To this end, they examined data on cytolytic activity in more than 11,000 patient tumors from The Cancer Genome Atlas, a collaboration between NCI and the National Human Genome Research Institute, also part of the National Institutes of Health. They found that a number of the genes identified in the CRISPR screen as being necessary for tumor cells to respond to T cells were indeed associated with tumor cytolytic activity in patient samples.

One such gene is APLNR. The product of this gene is the protein called the apelin receptor. Although it had been suspected to contribute to the development of some cancers, this was the first indication of a role in the response to T cells. Further investigation of tumors from patients resistant to immunotherapies showed that the apelin receptor protein was nonfunctional in some of them, indicating that the loss of this protein may limit the response to immunotherapy treatment.

What was most surprising to the investigators was that several of the genes whose loss had the greatest effect on tumor cells’ survival have not previously been linked to T cells’ ability to eliminate their targets, leading the team to new signaling pathways that will be important to explore.

The investigators wrote in the article that this gene list could serve as a blueprint for studying the emergence of tumor resistance to T cell—based cancer therapies. Restifo noted that if this set of genes is validated in clinical trials, then these data could eventually lead to more-effective treatments for patients.

“If we can truly understand mechanisms of resistance to immunotherapy, we might be able to develop new therapeutics,” he said. “In fact, in the future, this knowledge could speed the development of a new category of drugs that can circumvent these escape mechanisms of tumor cells and help patients experience complete responses.”

Reference:

Patel SJ, Sanjana NE, Kishton RJ, et al. Identification of essential genes for cancer immunotherapy. Nature. 2017;548(7669):537-542. doi: 10.1038/nature23477.