The researchers found that adding an excitable form of the protein Rac2 to macrophages, the immune cells that engulf pathogens, caused the macrophages to engulf T cells. The new technology has the potential to improve the effectiveness of an emerging cancer treatment.
Rac protein has a long history. This protein is well conserved during evolution and is thought to be present in the earliest nucleated cells. Despite their age, scientists continue to unravel their mysteries. In a new study, researchers at the University of California, Santa Barbara (UCSB) have discovered more about how Rac proteins work and how they could potentially improve cancer treatments.
The human genome encodes three Rac proteins. Rac1 is widely expressed, Rac2 is mainly expressed in cells that produce blood components (hematopoietic cells), and Rac3 is mainly expressed in brain tissue. As early as 1996, researchers studying fruit flies discovered that these proteins play an important role in cell movement, and that a hyperactive form of Rac1, expressed in only a few cells in the fly egg chamber, destroyed the entire tissue.
"Expressing this active Rac in just six to eight cells killed an entire tissue consisting of about 900 cells," said Abhinava Mishra, first author of the study.
This is the research result achieved by researchers in the 1990s. It was only a few years ago that research began to suggest that phagocytosis might be responsible for this tissue destruction.
In 2019, a study published in the journal Blood reported that three unrelated people with recurrent infections and severe deficiencies in T cells, specialized white blood cells critical to the immune system, all had the same mutation that overactivated Rac2. The study also observed that many patients had enlarged neutrophils (cells that capture and ingest invading microorganisms), suggesting that they were consuming large amounts of cellular material.
After reading the study, Denise Montell, who participated in the 1996 study and is the corresponding author of this study, wondered whether the disappearance of T cells was due to the phagocytosis of T cells by Rac2-active innate immune cells, as was the case in the Drosophila study. So Monteil and other researchers turned to macrophages, the voracious counterparts of neutrophils. The researchers cultured human macrophages with and without hyperactive Rac2 alongside T cells and found that macrophages with hyperactive Rac engulfed more cells, confirming their hypothesis.
The researchers then cultured the bone marrow stem cells into macrophages using bone marrow samples from mice with the same Rac2-boosting mutation as the patients in the 2019 study. The researchers conducted similar experiments, but this time mixed macrophages and T cells with cells that had a Rac2 mutation and cells without the Rac2 mutation. They found that not only were macrophages with active Rac depleted significantly more T cells, but T cells with active Rac2 were also more likely to be depleted, regardless of whether they had mutations.
Next, they cultured macrophages with active Rac2 from the bone marrow of normal and mutant mice. Macrophages from each group of mice expressed a pseudoreceptor or chimeric antigen receptor (CAR) designed to target B cells, another type of immune cell. They found that macrophages with the pseudoreceptor did not eat as many B cells; however, macrophages with both hyperactive Rac and CAR ate twice as many B cells as the macrophage group with only CAR. Activated Rac2 also appears to increase the number of so-called "super-predators" - voracious macrophages - that eat and kill multiple cancer cells.
"If you add active Rac but don't have the right receptor, you can't do anything," Montel said.
CAR-T is a type of cancer immunotherapy currently used to treat cancer. It uses the patient's own T cells and modifies them, adding CAR to help the cells attach to specific cancer cell antigens, thereby targeting the cells to attack cancer cells. This therapy is very effective against some cancers, such as leukemias and lymphomas, but some solid tumor cancers do not respond to this therapy. CAR-M, a new type of CAR-T therapy that relies on macrophages rather than T cells, has recently been used in clinical trials and is considered safe so far. However, one limitation of CAR-M is that CAR macrophages tend to "gnaw" cells rather than phagocytes. Therefore, enhancing phagocytosis of entire target cells is an important goal.
The researchers are interested in using Rac-enhanced CAR macrophages (which they call "RaceCAR-M" in the patent provided) to increase the effectiveness of CAR-M therapy.
They also plan to continue research investigating whether the new technique they have proven effective in the lab can be applied to new collections of human immune cells and animal cancer models. They are also exploring how Rac2 works at the molecular level.
"We found this 25-year-old mystery in fruit flies and solved it," Montel said. "That helped us solve the mystery of human immune deficiency. We then used that knowledge to enhance a potential cancer immunotherapy. It was just one mystery after another, and Luck turned out to be the answer to each one."
The research was published in the Proceedings of the National Academy of Sciences (PNAS).