Boosting cancer-killing cells through oxygen starvation

New research has suggested that an immunotherapy type that cultures patients' killer T cells in a laboratory before fine-tuning their cancer-destructive powers and returning them could be made more effective by growing the killer T cells in a low-oxygen culture.

Writing in the journal Cell Reports, researchers from the Weizmann Institute of Science in Israel suggest that their finding could increase the power of immunotherapy to fight solid tumors, which pose a particular challenge to killer T cells.

Killer T cells, which are also known as cytotoxic T lymphocytes (CTLs), are a specialized type of white blood cell called CD8+ T cells that are considered to be the "foot soldiers of the immune system."

CTLs directly kill damaged cells, cancer cells, and cells infected with viruses and other pathogens.

Senior author Guy Shakhar, who is a professor at the Weizmann Institute, explains that while killer T cells are the main agents of cancer immunotherapy, "they don't always manage to eliminate the malignancy."

However, he says that "by growing these T cells in an oxygen-poor environment, we can turn them into more effective killers."

Tumor cells can withstand low oxygen

At present, cancer immunotherapy that boosts patients' own killer T cells works best against certain leukemias and lymphomas. However, it is less effective at eliminating solid tumors, in which oxygen levels are very low.

Oxygen levels inside solid tumors, expressed as a percentage of gas dissolved in the fluid surrounding cells, range from 0.5 to 5 percent. This is lower than in most healthy organs and considerably lower than the 20 percent normally set for growing cells in the laboratory incubator.

Low oxygen does not appear to hamper tumor cells - for example, it does not stop them from taking in and metabolizing glucose, which is their main source of fuel.

But the low-oxygen environment of the tumor is not favorable to killer T cells. The researchers refer to several studies that offer evidence of this.

For example, it appears that tumor areas with low oxygen attract other cells, such as "M2-like tumor-associated macrophages" and "regulatory T cells," that can disrupt the function of killer T cells.

In addition, low-oxygen pockets in tumors favor anerobic metabolism of glucose, resulting in an acid environment that dampens killer T cell activity.

'Better tumor rejection and survival'

Previous studies have shown that growing killer T cells under conditions of low oxygen enhances their ability to kill other cells in the laboratory. But until this new work in Prof. Shakhar's laboratory, their ability to kill cancer had not been studied.

Thus, for their own investigation, the team experimented with growing killer T cells in culture in the laboratory at just 1 percent oxygen and compared them with cells grown at 20 percent, as before.

They found that the oxygen-starved killer T cells were more effective at attacking cultured melanoma cancer cells than killer T cells grown under the usual oxygen conditions.

In the meantime, the team implanted mice with tumor cells under their skin. Then, once tumors had formed 7 days layer, they injected some of the mice with oxygen-starved killer T cells, some with killer T cells grown under the usual conditions, and left the rest untreated.

They found that the mice treated with oxygen-starved killer T cells, or "hypoxic CTLs," showed "better tumor rejection and survival" than any of the others.

Low oxygen deals deadlier punches

When the researchers looked more closely at the oxygen-starved killer T cells, they found that they were not necessarily penetrating the tumors more deeply, but they had better weaponry for killing the tumor cells.

Killer T cells have a number of weapons for attacking cancer cells. One of these is a destructive enzyme called granzyme-B that they package into "granules." They release the granules into the cancer cells after they have punched holes in them using a protein named "perforin."

The team found that while the oxygen-starved killer T cells contained the same number of granules as the non-starved cells and released them as efficiently, they "packaged more granzyme-B in each granule without producing more perforin."

In other words, although the oxygen-starved killer T cells delivered the same number of punches as non-starved cells, each punch was deadlier.

Prof. Shakhar likens the oxygen-starved killer T cells to mountaineers who become acclimatized to lower oxygen levels. "Just as altitude training increases endurance in humans, so putting killer T cells through a 'fitness regimen' apparently toughens them up," he explains.

He and his colleagues point out that their method now needs to be confirmed in humans. If it is, then it offers an immediate way to improve the effectiveness of immunotherapy for solid tumors.

"In cellular immunotherapy, T cells need to be removed and grown outside the body in any event. Growing them under low oxygen pressure is relatively simple, but this small adjustment to existing clinical protocols may significantly improve the therapy's effectiveness."

Prof. Guy Shakhar