DURHAM, N.C. -- Researchers at Duke University Medical Center have discovered that an ancient protein can perform a decidedly modern function: stimulating the immune system to fight tumors.
The protein, called calreticulin, belongs to a family of proteins called chaperones that help fold other proteins into their globular working shapes after they are synthesized as string-like molecules inside the cell. These evolutionarily ancient proteins are found in all mammalian cells and have relatives in organisms as simple as bacteria and yeast.
But calreticulin can perform another vital function, the researchers discovered. Through experiments in mice, scientists found it can act as an informant to the immune system, stimulating it to destroy skin and thymus cancers.
The two disparate functions appeared to be difficult to reconcile. But now Duke cell biologist Christopher Nicchitta has begun to resolve the mystery, demonstrating that at least two chaperone proteins appear to have evolved a dual function: assisting protein folding inside the cell and alerting the immune system if they are released from the cell, as happens when tissue is under assault from inflammation, wounding or autoimmune attack.
The discovery is described and explained in two papers in the June 1 issue of the Journal of Immunology and the July 1 issue of the Journal of Cell Science.
"Calreticulin and its relatives are complex molecules that appear to have evolved a second function," Nicchitta said. "Since they normally are found only inside the cell, when they are released they can act as a danger signal to the immune system, which responds by trying to eliminate the insult at the precise location in the body signaled by these proteins."
The work, which was supported by the National Institutes of Health, is part of Duke's Center for Genetic and Cellular Therapies continuing effort to coax the body's own immune system to fight cancer. If further research proves the proteins act the same way in humans, it may help scientists tailor specific cancer therapies, said Nicchitta.
So far, the scientists' experiments reveal that calreticulin and a related protein called GRP94 both can alert the immune system by informing it about where in the body an insult has occurred. The proteins act as informants by carrying small pieces of cellular proteins called peptides as a type of calling card, with an address describing where in the body the injury is; for example, the skin or the thymus.
"The peptides appear to act as a fingerprint of the cell," Nicchitta said. "They tell the immune system where to marshal its forces. We are now using that property of these chaperone proteins to educate the immune system to fight cancer."
These molecular calling cards act as immune stimulating substances called antigens and are recognized by specialized immune system cells, either macrophages or dendritic cells. When these immune cells come into contact with calreticulin or GRP94, they appear to swallow the protein, release the peptides and display them on their cell surface. This display of a foreign peptide can trigger a strong response from immune system fighters known as killer T cells.
The Duke researchers showed that when they injected mice with dendritic cells that had been mixed with calreticulin or GRP94 purified from melanoma (skin cancer) or thymoma (thymus tumors), the animals responded by generating large numbers of killer T cells directed at the specific tumor type from which it had come.
"The immune response mediated by calreticulin is impressive," said Dr. Eli Gilboa, research director of Duke's Center for Genetic and Cellular Therapies. "This result shows the promise of this class of proteins to elicit a targeted cytotoxic T lymphocyte (killer T cell) response to tumor tissue."
Tumor cells have evolved ways of effectively hiding from patrolling dendritic cells, making cancer virtually invisible to the immune system, which can only mount a very weak response at best. The Duke researchers, led by Gilboa, have devised a way to engineer dendritic cells and grow them in the lab, and they have been testing various methods to prompt the cells to recognize cancerous tumors, thus targeting them for destruction.
Upon hearing that researchers at other institutions had successfully used GRP94 to mount an immune response, Gilboa and his colleagues launched a collaboration with Nicchitta, who had been studying the chaperone proteins for many years. Before long, the researchers had discovered that calreticulin is even better than GRP94 in stimulating the immune system. That work is the subject of the June 1 Journal of Immunology paper.
The finding prompted Nicchitta and his colleagues James Wassenberg and Cameron Dezfulian to examine just how a protein-folding chaperone might stimulate the immune system. Their findings are detailed in the July 1 issue of the Journal of Cell Science.
"To our surprise, we found that macrophages have a specific receptor on their cell surface that recognizes GRP94 and internalizes it," said Nicchitta. "The question is why would macrophages have a receptor on the exterior of the cell for a protein that is normally found only inside cells?"
Nicchitta speculates that the researchers have come across a previously unknown role for chaperone proteins.
The scientists are now trying to determine the pathway that antigen-presenting cells like macrophages and dendritic cells use to convert the molecular flags carried by chaperone proteins into a specific immune response.
Meanwhile, Gilboa and his colleague immunologist Smita Nair are beginning to explore the use of chaperone proteins as a basis for a new type of cancer vaccine.
"We are enthusiastic about the response we have seen to our vaccine in animals," Gilboa said. However, he cautions that such a vaccine is still far from ready for trials in people.
Pamela Wearch and Duane Mitchell also contributed to the work.
Note to editors: Graphic available in color at http://ed-media.mc.duke.edu/aved/avart/hede/index.htm.