HIV STUDIES: Rockefeller's Melissa Pope is investigating dendritic cell-T cell fusions with an eye toward potential strategies for interrupting HIV replication.
Dendritic cells are found in almost every type of tissue, including lymphatic, blood, and skin, albeit in small numbers in each type. They make up between 0.5 percent and 4 percent of the cells at various sites. A category of antigen-presenting cells (APCs), dendritic cells display antigens; when T cells recognize these antigens, an immune response is set into motion.
"What dendritic cells do is pick up bacteria, whole proteins, and viruses; engulf them; chop [them] up into peptides; and present the peptides to T cells to activate the T cells," explains Drew Weissman, an assistant professor of medicine at the University of Pennsylvania.
"They are 20 to 100 times more potent on a cell-for-cell basis in stimulating T cells than macrophages are," says Frank J. Hsu, an assistant professor of medicine at Yale University School of Medicine. Macrophages are immune-system cells that also recognize antigens and present them to T cells.
Dendritic cells' power in presenting antigens suggests they could be used in treating cancer. Researchers also are encouraged by studies finding a positive link between the presence of dendritic cells and prognosis in cancer patients (M.J. Alavaikko et al., American Journal of Clinical Pathology, 101:761-7, 1994; T. Shunichi et al., Cancer, 75:1478-83, 1995).
EXPLORING THE POTENTIAL: Pittsburgh's Michael Lotze plans clinical trials of dendritic cell vaccines for head, neck, and colorectal cancers.
GMCSF is essentially basic nourishment-it makes sure the cells survive, according to Lotze. IL-4, when added to monocyte cultures, helps direct these cells to develop as dendritic cells. Similarly, TNF-alpha, when mixed with undifferentiated stem cells, increases the likelihood the stem cells will develop as dendritic cells instead of macrophages.
In one of the early attempts to use dendritic cells as tumor therapy, Hsu, who then was at Stanford University, and his colleagues found that dendritic cells could successfully stimulate T cells to attack B cell lymphoma (F.J. Hsu et al., Nature Medicine, 2:52-8, 1995).
The researchers created a vaccine by "pulsing," or culturing immature dendritic cells with idiotype protein from lymphoma cells. The protein is a unique tumor marker that can be readily recognized by dendritic cells.
The dendritic cells were autologous-the patient's own. Hsu used immature cells from the peripheral blood. Immature cells can process whole proteins, an ability they lose as they mature. As the cells grow, they develop costimulatory molecules, such as CD 80 and CD 86, which stimulate T cells to generate an immune response to the antigens presented to them by the dendritic cells. When the dendritic cells were injected into the patients, "they were able to stimulate T cells," he says.
Of the original four patients in the pilot study, all produced measurable T-cell responses against the tumors. One had a complete regression, and two others experienced some tumor regression. While the tumor initially shrank in the fourth patient, the disease later continued to develop.
As the Stanford trials continue, researchers at other institutions are planning to test dendritic cell vaccines in cancer patients. At Pittsburgh, Lotze is planning clinical trials exploring the value of dendritic cells in treating head, neck, and colorectal cancers. Brian J. Czerniecki, an assistant professor of surgery at the University of Pennsylvania, plans to test a dendritic cell vaccine this spring in melanoma patients. His trial will involve taking monocytes from the blood and treating them with calcium. It had been thought, he says, that these precursor cells would eventually become macrophages.
"What we found is that by treating them with calcium agents, such as calcium ionophore, a monocyte could . . . become a dendritic cell. The monocyte pool can uniformly be activated to express everything that a dendritic cell does essentially overnight by treatment with calcium," he says (B.J. Czerniecki et al., Journal of Immunology, 159:3823-37, 1997).
Calcium ionophore brings calcium into cells. Calcium turns on genes that express CD 80 and CD 86.
In in-vitro experiments, Czerniecki succeeded in co-culturing dendritic cells with melanoma antigens, having the dendritic cells interact with T cells and having those T cells initiate an immune response against melanoma cells.
While Czerniecki sees promise in this approach, he explains that there are possible side effects, one of which is triggering an autoimmune response. There is concern because melanoma tumor cells share antigens with normal melanocytes-cells that produce pigment coloring skin, hair, and eyes.
Paul Sondel, a professor of human oncology at the University of Wisconsin, has other concerns about such efforts. "Some of the antigens being looked at on colon cancer or on breast cancer are also molecules that are also expressed on certain normal tissues," he says. "It's a formidable hurdle to turn on these immune responses against these autoantigens because the immune system doesn't want to recognize them. So we might not be able to turn them on as easily as people are hoping."
Tumors themselves also may pose obstacles. The fact that people get cancer means that tumor cells have their own proteins that are toxic to the body's immune-system cells.
"Tumor cells secrete such factors as IL-10 and TGF-beta, which can inhibit immune-system cells," notes Lotze. "There must at least 50 ways in which tumors can escape immune recognition," he adds.
If using dendritic cells in cancer therapy works, Sondel contends, "the people doing it will be heroes. If it doesn't, we'll learn something and move on to the next thing."
A PROMISING APPROACH: Penn's Brian Czerniecki plans to test melanoma patients to see whether treating monocytes with calcium will produce dendritic cells.
Bearing a load of HIV, dendritic cells bind to T cells, thus forming syncytia. These are fusions of the two types of cell, where HIV can replicate and destroy T cells (S.S. Frankel et al., Science, 272:115-7, 1996). Consequently, researchers are working to understand what happens when dendritic cells meet HIV.
"What we have found is when HIV encounters a dendritic cell, some of the HIV is taken up and processed, but some of the HIV remains stuck to the surface as an infectious virion," says Penn's Weissman (D. Weissman et al., J. Immunol., 155: 4111-7, 1995). The reason for this abnormal binding of a virus to the surface appears to lie in the presence of a variety of binding sites on the dendritic cells' surface. These include the CD-4 molecule and proteins known as mannose binding proteins, which nonspecifically bind foreign antigens such as viruses and bacteria to the cell surface.
Blocking mannose binding sites may offer a possible treatment for individuals exposed to HIV (D. Weissman, A.S. Fauci, Clinical Microbiology Reviews, 10:358-67, 1997).
Melissa Pope, an assistant professor in the Laboratory of Cellular Physiology and Immunology at Rockefeller University, is focusing on another target: the dendritic cell-T cell syncytia. "What we're trying to do is identify the molecules that are interacting between the two cell types. We're trying to see if there are any signals provided by that interaction that are important in the binding and in supporting viral replication." Understanding that basis, she notes, could provide strategies aimed at interrupting HIV replication.
| The Fifth International Symposium on Dendritic Cells in Fundamental and Clinical immunology will be held Sept. 24-28 in Pittsburgh. For information, contact Diane Applegate, (412) 647-8263. Fax: (412) 647-8222. E-mail: firstname.lastname@example.org|
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Meanwhile Lotze, who is co-organizing an international conference on dendritic cells, is exploring a different approach to therapy. He and Cara Wilson, an assistant professor of medicine at the University of Pittsburgh, have planned trials for the coming year using dendritic cells as therapeutic vaccines in HIV patients.
A pilot study to assess the safety of such a treatment and see if it enhances T-cell responses against HIV is planned. It would involve patients receiving highly active anti-retroviral therapy (HAART), the combined use of protease inhibitors and reverse transcriptase inhibitors to control HIV infection. These patients, Wilson explains, would be treated with autologous dendritic cells cultured with an HIV protein (p24) and tetanus toxoid. (The toxoid is a control to see if the patients' immune systems respond.) The treated cells would be injected into the patients.
"We're going to follow them, look at changes in their HIV viral load, and look at immune responses to the particular HIV protein," says Wilson.
The study is designed to assess the safety of such a treatment and see if T-cell responses against HIV are enhanced. In a second planned study, the researchers would use liposomal preparations to culture stem cell-derived dendritic cells with three proteins unique to HIV-gag, pol, and env proteins. Stem cells are precursors of dendritic cells. The mature dendritic cells would then be injected into patients receiving HAART.
"We want to generate as broad [an immune] response as possible," says Wilson, explaining why the researchers plan to use a combination of HIV proteins in the trial. The eventual goal is to develop therapeutic vaccines for patients receiving HAART who still have some HIV in their systems. "We hope this immunotherapy can target those latently infected cells," she says. Harvey Black is a freelance science writer in Madison, Wis.