Targeting Breast Cancer Metastasis

Artistic rendering of a cancer cell in red with round, blue accents

Researchers used GTPase inhibitors to attenuate the migration of cancer cells and macrophages and to reduce the levels of inflammatory cytokines driving metastasis.

Breast cancer is the most frequently diagnosed cancer and contributes to 15 percent of all cancer-related deaths in women worldwide. Though 20–30 percent of patients with early-stage breast cancers eventually develop metastatic cancer, few effective treatments for preventing rapid metastatic progression exist.¹

Tumor cell motility and invasion are essential drivers of metastasis, as tumor cells must migrate away from the primary tumor and invade new sites. Tumor cells at the primary cancer site promote metastasis by recruiting immunosuppressive inflammatory cells, such as activated macrophages, neutrophils, and myeloid-derived suppressor cells, which facilitate tumor cell migration and survival.

Researchers previously showed that inhibitors preventing the activation of the Rac and Cdc42 GTPases inhibited cancer cell migration and metastasis in breast cancer cell lines.^2-4 These proteins regulate both tumor and immune cell migration, as well as the communication between them in the tumor microenvironment. However, scientists had only explored the inhibitors’ effects on tumor cells alone. In their recent paper in Frontiers in Oncology, Suranganie Dharmawardhane, Stephanie Dorta-Estremera, and their team from the University of Puerto Rico showed that EHop-016 and MBQ-167, inhibitors that prevent the activation of either Rac or both Rac and Cdc42, respectively, promoted an anti-tumor environment by blocking not only metastatic cancer cells but also immunosuppressive myeloid cells.⁵

To investigate the effects of EHop-016 and MBQ-167 on immune cells, the researchers first treated differentiated macrophages and breast cancer cells in vitro with either drug. They found that EHop-016 reduced Rac activation and MBQ-167 reduced both Rac and Cdc42 activation in the cancer cells as expected. Additionally, drug-treated macrophages had similar outcomes.

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Next, the researchers tested the drugs’ effects on macrophage migration. Activated Rac and Cdc42 promote cell migration and invasion by phosphorylating p21-activated kinase (PAK), their common downstream effector. Activated PAK then induces actin cytoskeleton reorganization into lamellipodia, filopodia, and invadopodia. The researchers found that compared to vehicle treatment, these inhibitors both reduced the macrophages’ PAK activity, cell surface actin structures, cell migration, and ability to phagocytize fluorescently-labeled particles in vitro.

To determine whether the Rac and Cdc42 inhibitors reduced myeloid cell activation and tumor infiltration in vivo, Dharmawardhane and Dorta-Estremera’s team treated mice bearing tumors derived from breast cancer cells with either drug. Compared to controls, macrophage and neutrophil infiltration was reduced in mammary tumors from mice treated with EHop-016, while macrophage infiltration was reduced in the spleens, but not in the tumors, of mice treated with MBQ-167.

The researchers also analyzed cytokine production in the mouse tumor microenvironment in response to drug treatment. “One surprising finding was that only IL-6 was reduced from the other six cytokines tested,” said Dharmawardhane and Dorta-Estremera in an email. IL-6 is a pro-inflammatory cytokine largely produced by macrophages. “The mechanism by which this occurs needs to be investigated,” they added.

Taken together, these results suggest the dual role played by these inhibitors in preventing metastasis: they inhibit the migration of cancer cells and macrophages and reduce the levels of inflammatory cytokines important for driving metastasis. Some questions, however, remain. “What is the pharmacokinetics and pharmacodynamics of the drug? Does it get where they expect it to be? Does it get inside the cell appropriately?” asked Danny Welch, a cancer biologist from the University of Kansas School of Medicine who was not involved in this study.

The answers to these questions may come from the FDA-approved phase I clinical trial that is underway. “We will know for sure whether these Rac/Cdc42 inhibitors will be safe in the upcoming clinical trial that has been initiated this month,” said Dharmawardhane and Dorta-Estremera. “This will be the first cancer drug made in Puerto Rico and tested in clinical trials in Puerto Ricans.”