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Cancer immunotherapy: Combining approaches to achieve checkmate

CD8 stain for the extensive tumor necrosis

CTLA-4 antibody blockade evokes CD8+ cytotoxic T cells that destroy melanoma cells.

Like a clever chess player, cancer has evolved strategies to escape capture by its opponent, the immune system—a coordinated network of specialized cells that normally protects the body against attacks from foreign invaders. “These strategies are quite effective at disarming or suppressing an immune response,” says Glenn Dranoff, MD (DFCI), leader of the DF/HCC Cancer Immunology Program. But as research in the lab advances understanding of immune response pathways, investigators are gaining crucial insights into the mechanisms that regulate the antitumor response. Basic and clinical scientists in the program are collaborating across multiple DF/HCC institutions to translate these insights into immunotherapies in the clinic that will overcome tumor immune escape.

Many of the program’s clinical trials are testing a key hypothesis: whether improving one regulatory point in a specific pathway will enhance its function and trigger the rest of the interconnected immune cells into action, explains Dranoff. The trials highlighted below combine two or more of the program’s immunologic approaches—bone marrow transplantation, adoptive cellular therapies, and cancer vaccines—to increase the antitumor immune response, much as a player positions two or more chess pieces to achieve checkmate.

Delivering a vaccine to a fresh immune system

For over a decade, Robert Soiffer, MD, chief of the Division of Hematologic Malignancies in DFCI’s Department of Medical Oncology, has conducted a series of trials attempting to induce antitumor immunity using a therapeutic cancer vaccine called GVAX. Pioneered by Dranoff, the vaccine is made of patient-derived cancer cells that have been genetically modified to secrete GM-CSF, a factor that stimulates the immune system. GVAX induces dendritic cells, sentinels that convey information about an invader, to evoke a T cell response by “educating” T cells to recognize and kill the cancer cells. Since GVAX contains the patient’s own cancer cells (which have been irradiated) and the GM-CSF booster, it theoretically strengthens the body’s natural defenses without harming normal cells. But, says Soiffer, vaccines alone do not always work because the immune system of a cancer patient may already be compromised or the patient may have a large volume of disease.

Meanwhile, allogeneic transplantation (from a donor) has been found to cure a number of patients with hematologic malignancies, though success has been limited by complications such as graft-versus-host disease (GVHD), in which the donor’s immune cells react against the patient’s tissues. In the latest iteration of Soiffer’s immunotherapy work, he and Vincent Ho, MD (DFCI) proposed combining therapies: administering GVAX to patients who had already undergone an allogeneic bone marrow transplant—thereby capitalizing on chemotherapy-reduced disease as well as a fresh immune system from the transplant donor.

In this trial, GVAX is given to patients with advanced leukemia or myelodysplastic syndrome (MDS) 30 days after a reduced-intensity transplant—a procedure using low-dose chemotherapy that usually results in higher relapse rates but lower toxic side effects than a full transplant. “It is hypothesized that the vaccine will attract and stimulate immune cells at the site of injection, and those cells will circulate and kill any residual leukemia cells that might persist after the transplant,” explains Soiffer. “The ultimate goal of the trial is to improve patient outcomes by inducing an immune-mediated antileukemic effect without GVHD,” he says. To date, of the 21 patients who received the vaccine post transplant, eight remain in remission after one year—very encouraging results in patients with active leukemia, says Soiffer.

What made this trial possible is the Connell and O’Reilly Families Cell Manipulation Core, under the direction of Jerome Ritz, MD (DFCI). The facility, which produced the vaccine, assists DF/HCC investigators in developing new cell-based therapies that meet FDA requirements for clinical research studies. “The core is truly a gem of a facility,” notes Dranoff. “None of these trials could have been done without this precious resource.” The Cancer Vaccine Center, at DFCI, also provides much needed support for conducting complex clinical studies and measuring antitumor immune responses in patients.

Helping dendritic cells grow up

“One of the major defects in cancer is that dendritic cells remain immature and therefore unable to present tumor antigens to T cells,” says Steven Balk, MD, PhD, associate professor and staff physician, Division of Hematology-Oncology, at BIDMC. Scientists like Balk believe that the maturation of DCs is the job of iNKT cells (a subset of natural killer T cells), which are decreased or deficient in certain tumors such as melanoma. What’s more, like a double attack in chess—where a knight might simultaneously threaten two or more of the opponent’s pieces—iNKT cells appear to activate multiple effector cells of the immune system including dendritic cells, T cells, and NK cells. Thus Balk is co-leading a trial to test whether infusions of purified iNKT cells can restore strong cellular antitumor responses in patients with melanoma. In preparation for the trial, Balk’s lab created an antibody specific to iNKT cells to separate them from the patient’s other white cells.

“When you develop cells that may have the potential for therapeutic use,” says Balk, “you go to the Center for Human Cell Therapy to translate that work to the clinic.” The CHCT figured out how much antibody to use, how to obtain the best yield, and how to stimulate growth in culture, he explains. “Once the methods were perfected, we transferred the project to the Cell Manipulation Core, where we went through the entire protocol in a ‘trial run’: harvesting the white cells of two healthy donors (by a process called leukapheresis), purifying the iNKT cells, expanding them in culture, and making sure the process actually worked.” Patients could begin receiving the new immunotherapy in the next couple of months. Infusions will be administered in three equal doses, two weeks apart; if three patients do not experience any toxic effects, they will also receive GM-CSF at their second and third infusions.

“It’s a race,” cautions Balk, “to see whether we can infuse a reasonable number of iNKT cells to stimulate antitumor responses before the cells become inactivated.”

Exploiting the unique sensitivity of antibodies

Antibodies are famed for their exquisite sensitivity, each one binding only to its matching antigen. When an antibody latches to its counterpart, it signals powerful cells of the immune system to neutralize, block, or destroy the antigen. This sensitivity can be exploited for therapeutic purposes, which is precisely what principal investigator F. Stephen Hodi, MD, assistant professor of medicine and clinical director of melanoma at DFCI, is attempting to do.

In this phase I trial, Hodi is testing an antibody therapy in patients who have previously been vaccinated with GVAX. The antibody, provided by the National Cancer Institute, binds with a molecule called CTLA-4, a receptor on the surface of T cells. Normally, CTLA-4 acts as Mother Nature’s “brake”—stopping the T cell response, when appropriate, to prevent autoimmunity. (Each component of the immune system has positive signals that activate a response and negative signals that deactivate it.) Since, in most cases, cancer is unchecked due to a deficient or suppressed immune system, temporarily turning off this negative signal is a way to increase the immune response. The antibody releases the brake by blocking CTLA-4.

“The strategy of the trial is to outmaneuver the tumor,” explains Hodi, “by first educating the immune system with the vaccine and then giving just enough antibody.” Patients receive the antibody by intravenous infusion every two months, a dosing schedule chosen “because it was biologically active, had synergy with the vaccine, and was safe,” says Hodi. Since 2003, the CTLA-4 antibody has been administered to about 30 patients with ovarian cancer, melanoma, acute myeloid leukemia, MDS, or non-small cell lung cancer. “Patients tolerated the therapy well,” says Hodi, “and a dramatic response was seen in several melanoma patients.”

Creating cytotoxic T lymphocytes in the lab

Dendritic cells, which activate the T cell response, are a type of antigen-presenting cell. An APC engulfs foreign substances such as tumor cells, digests them into smaller fragments, and displays the resulting antigens on its surface, thereby training the T cells to attack the tumor. These T cells are often called cytotoxic T lymphocytes, or CTLs.

For the last several years, Marcus Butler, MD, an instructor at BWH and DFCI, has been working on laboratory methods for creating CTLs that could be infused into patients to stimulate antitumor effects. Recently, he and his colleagues hit on the novel idea of creating an artificial APC to generate the desired T cells. After establishing a cell line for an aAPC in his own lab, Butler worked with the Harvard Gene Therapy Initiative to develop a clinical-grade product. The Center for Human Cell Therapy advanced the project by developing the protocols to be used in the Cell Manipulation Core, which is now conducting the project’s final trial run. “Our strategy has passed FDA scrutiny,” says Butler, “and we will soon be ready to begin the clinical trial.”

The protocol calls for white blood cells to be taken from a melanoma patient via leukapheresis, purified for CTL precursor cells, and cultured with the aAPC for three weeks to generate sufficient antitumor CTLs – those that specifically target a tumor antigen expressed on melanoma cells. “The aAPCs are used only to stimulate CTLs,” assures Butler. “They survive for a few days in culture and are then irradiated, so they do not get infused into the patient.” The study will measure the feasibility of this unique adoptive cell transfer approach and the safety of two different dose levels, with patients receiving two infusions two weeks apart. If successful, Butler will collaborate with BWH to combine infusion of antitumor CTLs with radiation.

Treating virus-associated tumors with CTLs

“Some malignancies are caused by persistent virus infections,” says Fred Wang, professor of medicine in the Infectious Diseases Division at BWH. Nasopharyngeal carcinoma (NPC), for example, is linked to the Epstein-Barr virus (EBV), a common virus that is normally kept in check by CTLs. In EBV-associated NPC, says Wang, EBV infection combined with some chromosomal abnormality and perhaps a subtle immune defect drives the development of tumors. Once tumors emerge, they express viral proteins, or antigens, which act as foreign markers on the malignant cells. “Everybody develops EBV-specific CTLs when they are first infected with the virus,” says Wang. “We can amplify these cells, and then use them to target the tumor.”

That’s the logic behind a new EBV immunotherapy trial in which Wang is collaborating with co-investigators Lori Wirth, MD, and Marshal Posner, MD, in the Head and Neck Oncology Program, and BWH viral immunologist Mark Fogg, PhD. “The key question we’re trying to answer,” explains Wang, “is whether we can treat these tumors in patients with relapsed or metastatic NPC by enhancing the immune response to the viral antigens.”

To amplify CTLs, the patient’s lymphocytes (B cells and T cells) are separated from the rest of the blood cells. One aliquot is exposed to the EBV in culture, where the virus infects the B lymphocytes (immune cells that make antibodies), causing them to proliferate and become a permanently growing virus-infected B cell line, matched to the patient. The EBV-infected B lymphocytes are then irradiated and mixed with the patient's lymphocytes, exposing the T lymphocytes to the viral antigens and stimulating them to grow. After a week, another aliquot of irradiated EBV-infected B lymphocytes is used to restimulate the T cells. This process is repeated several times, expanding the number of T lymphocytes that recognize EBV antigens. After this population has been enriched, interleukin 2 (a chemical messenger of the immune system) is added to help the T cells grow faster. The entire process results in the clinical-grade EBV-specific T cell product that will be infused into patients.

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Clinical trials like these - supported by the Cell Manipulation Core, the Center for Human Cell Therapy, the Cancer Vaccine Center, and other facilities across DF/HCC institutions - represent a fraction of the novel immunotherapies arising from the Cancer Immunology Program and underscore the complex, collaborative nature of immunologic approaches. The most effective trials, says Dranoff, are likely to be those that elicit the coordination of several components of the immune system. “Our broader vision is that immunologic approaches to cancer treatment will complement and extend other therapies that attack the cancer cell or its microenvironment,” predicts Dranoff. “Drugs are likely to work much better when the host has an adequate immune response.”

- Lonnie Christiansen