Targeting the Accomplices:
Homing in on Immune Cells that Aid Tumors
Cancer researchers are increasingly expanding their focus beyond malignant cancer cells to other players within and near tumors, in what is often called the tumor microenvironment. Many of these studies have implicated a versatile immune cell, the macrophage, as a common tumor accomplice.
Macrophages in and around tumors, often referred to as tumor-associated macrophages (TAMs), can help combat cancer, killing tumor cells as well as calling in the big guns of the immune system, killer T cells, to help rid the body of malignant cells.
But accumulating evidence from studies performed in cancer cell lines and animal models of different cancer types indicate that, more often than not, TAMs aid tumors in numerous ways and at numerous points, from the earliest development of a malignant mass to the formation of metastases.
Data in humans are far more limited but mostly support the cell line and animal model data, with studies consistently showing a correlation between elevated TAM levels and poor prognosis. Researchers working in this area believe that TAMs may not only be valuable prognostic markers that can help in the selection of therapy, but also an important new target to attack tumors.
The concept, explained Dr. Jeffrey Pollard of the Albert Einstein Cancer Center at Yeshiva University, is straightforward. “We need to do more than just attack the tumor cells,” he said. “We need to target the support system, by pulling the foundation out from under tumor cells as well.”
A Diversity of TAMs
More recently, researchers have been studying TAMs more closely to learn what these immune cells are doing in and around the tumor and how they do it. It has become clear, for instance, that specific macrophage subtypes provide the heavy lifting tumors need from their surrounding environment to survive and thrive, such as suppressing the immune response or growing new blood vessels.
“Macrophages are very plastic,” explained Dr. Johanna Joyce of Memorial Sloan-Kettering Cancer Center, whose research focuses on the tumor microenvironment. The functions that TAMs perform, Dr. Joyce continued, can vary tremendously depending on the molecular markers they express, the signals they receive from the tumor itself and other components of the microenvironment, and their location—for instance, in the oxygen-depleted core or at the “invading front” of a tumor.
Even though the field has made great strides, she cautioned that “we’re really just scratching the surface of what we understand about these cells.”
Macrophages are often split into two subtypes—M1, or “classically activated” macrophages, which do things like help heal wounds and present antigens to killer T cells, and M2, or “alternatively activated” macrophages, which can promote tumor growth and metastasis.
That classification system is probably too simplistic, explained Dr. Pollard, who has been studying the tumor microenvironment for more than two decades. “I think it can blind people to the complexity of the macrophage response,” he said. In mouse models of breast cancer, for example, Dr. Pollard’s lab has identified at least six different macrophage subtypes.
The M1/M2 classification is primarily based on differentiating macrophages in tissue culture, explained Dr. Lisa Coussens of the University of California San Francisco (UCSF).
“You don’t ever see a whole tumor populated by one species of macrophage,” she said. “You see a broad spectrum of macrophage phenotypes.” The macrophage types that predominate in a given tumor are determined in large part by the immune microenvironment around it, Dr. Coussens continued, such as the presence of different types of helper T cells.
Drs. Pollard and Coussens have a collaborative grant to identify markers or patterns of markers that more clearly identify TAM subtypes that are associated with different pro- and antitumor activities, she noted.
Partners in Crime
In the meantime, studies have begun to unravel the co-dependent relationship between tumor cells and macrophages.
Using a mouse model of metastatic breast cancer, for example, Dr. Pollard and his colleagues recently showed how macrophages can promote metastasis. Tumor cells that had traveled to blood vessels around the lungs, they found, secreted a chemokine—a hormone-like protein that attracts and mobilizes immune cells—called CCL2 (also known as MCP-1). This action, in turn, drew macrophages to the tumor cells, at which point the macrophages secreted a growth factor called VEGF, which made the blood vessels leaky, allowing the tumor cells to move into the lungs.
TAMs have also been implicated in disease recurrence. In a study using a mouse model of glioblastoma multiforme (GBM), Dr. Martin Brown and his colleagues at the Stanford University School of Medicine found that regrowth of tumors that initially shrank after radiation therapy was due in part to macrophages and similar cells that were recruited to the tumor site. This recruitment was abetted in large part by a chemokine called SDF-1 (or CXCL12) produced by tumor cells.
SDF-1 “is not stimulated by the radiation itself, it’s stimulated by the product of the radiation,” Dr. Brown said. As the tumor shrinks, its blood supply is diminished and it becomes oxygen depleted, much like tissue in a wound, setting off a chain of events, beginning with the production of the protein HIF-1. This, in turn, induces the production of SDF-1, which mobilizes the macrophage precursor cells from the bone marrow to come into the tumor. Once in the tumor, explained Dr. Brown, the macrophages are retained or “captured” by another protein, CXCR4.
Dr. Brown said much of his work suggests that “what these macrophages are doing [following radiation treatment] is promoting a blood supply. They’re very pro-angiogenic, provasculature.”
Transition to the Clinic?
TAM research has yet to have an impact on patient care. But it’s beginning to move in that direction.
Several recent studies, for example, strongly suggest that TAM levels can be used as prognostic markers.An excess of TAMs has been linked with adverse outcomes in a growing list of cancers, including breast, thyroid, liver, melanoma, lung, glioma, and several blood cancers.
A study published last year in the New England Journal of Medicine (NEJM), for instance, found that in tumor samples from patients with Hodgkin lymphoma, higher levels of TAMs—identified by the expression of the CD68 marker—were associated with poorer patient outcomes. In fact, there were no lymphoma-related deaths in patients who had limited-stage disease and low TAM levels. And when the initial treatment failed—which happens in about 20 percent of patients—second-line treatment with a stem-cell transplant was far more likely to succeed in patients with low TAM levels than in those with high levels.
“I think there is overwhelming evidence now that the number of TAMs in tissue biopsies is a valuable prognostic marker in Hodgkin lymphoma,” said Dr. Christian Steidl of the British Columbia Cancer Agency, the first author of the NEJM study.
More than 10 studies have confirmed that high levels of TAMs are correlated with poor primary treatment outcomes and lower overall survival in Hodgkin lymphoma, he continued. “The next steps would be to incorporate this information into clinical trials, and look in a prospective setting to establish evidence that it improves patient management,” he said.
In another study published earlier this year, Dr. Coussens and her colleagues found that patients with breast cancer whose tumor samples had what they called a specific “immune signature”—high levels of TAMs (identified with CD68) and low levels of another immune cell, killer T cells—had poorer outcomes than patients with low TAM levels and elevated killer and helper T cells. The presence of the immune signature was most strongly associated with outcomes in women whose tumors were classified as HER2-positive or basal-type/triple-negative, two aggressive breast cancer subtypes.
In the same study, using a mouse model of breast cancer, the investigators found that an investigational drug called PLX3397 hinders the recruitment of macrophages by the cytokine colony-stimulating factor 1 (elevated levels of which have also been associated with poor outcomes in breast cancer), reducing the cancer’s spread to the lungs and increasing survival.
Dr. Coussens has a grant from Susan G. Komen for the Cure, in conjunction with Drs. Shelley Hwang and Hope Rugo from UCSF, to build on this work, including conducting a multicenter phase I/II clinical trial of women with triple-negative breast cancers that have recurred after treatment. With the trial and related studies, she hopes to validate the TAM-based immune signature and evaluate the safety and potential efficacy of PLX3397 in combination with chemotherapy.
Results from Drs. Pollard’s and Brown’s recent studies also used agents that target TAMs, showing that these agents could reduce recurrences and improve survival in mice. Human trials testing these agents may not be far off, they said.
If these therapies prove to be effective in humans, in some cases it may be because they are simply killing macrophages. In other cases, it may be because they are depleting certain macrophage populations that aid the tumor, essentially “reprogramming” the tumor microenvironment in a way that can enhance the effectiveness of other therapies and the body’s own natural defenses, Dr. Coussens said.
“It’s such new biology,” she continued. “I really think that by supporting this kind of research, we’ll be able to improve clinical responses going forward.”