Light-Fingered Macrophages Caught Red Handed at Checkpoint

A macrophage (red) removing PD-1-blocking antibodies (yellow) from the surfaces of neighboring CD8 T cells (blue), cutting short the drug’s activity. [Center for Systems Biology, MGH]

While pickpocketing is becoming a lost art in our cities, it is still practiced at the cellular level by macrophages. These cells, which do good work as part of the immune system, sometimes misbehave, plucking checkpoint antibody drugs away from T cells. Macrophages have even been caught in the act, thanks to a surveillance system operated by scientists at Massachusetts General Hospital (MGH).

Using in vivo imaging, the MGH scientists, led by Mikael Pittet, Ph.D., found that an antibody-based drug designed to block the immunosuppressive molecule programmed cell death protein 1 (PD-1) is removed from its target T cells by macrophages within minutes of administration. This observation was confirmed in several mouse models of cancer. The MGH team also identify the molecular mechanism behind this drug capture, which could lead to ways to prevent the process.

Just as better surveillance of public places led to more focused policing of pickpockets, the in vivo imaging carried out by the MGH scientists could lead to improved checkpoint inhibition therapies. For example, therapies could combine checkpoint inhibitor drugs and agents that could discourage macrophages from treating T cells as “marks.”

Detail of the MGH investigation appeared May 10 in the journal Science Translational Medicine, in an article entitled “In Vivo Imaging Reveals a Tumor-Associated Macrophage–Mediated Resistance Pathway in Anti–PD-1 Therapy.” The article describes how high-resolution molecular imaging was used to track immune checkpoint drugs, monoclonal antibodies (mAbs), in real time. The imaging helped the MGH scientists “discover what was happening,” said Dr. Pittet, and “devise ways to extend the time the drug binds to its target and improve treatment efficacy in our models.”

“We show that aPD-1 [anti-PD-1] mAbs effectively bind PD-1⁺ tumor-infiltrating CD8⁺ T cells at early time points after administration,” the article’s authors wrote. “However, this engagement is transient, and aPD-1 mAbs are captured within minutes from the T cell surface by PD-1⁻ tumor-associated macrophages.”

These observations suggest how it is that checkpoint blockade, a promising immunotherapy against cancer, does not always work. That is, this therapy works with some, but not all, patients and has improved long-term survival in just a minority of patients.

Immune checkpoint molecules like PD-1 are expressed on the surface of CD8 T cells—the immune system's “killer cells” that attack cells that are damaged or diseased, including cancer cells—and act to suppress an inappropriate T-cell response. mAbs that block pathways controlled by checkpoint molecules are the basis of current checkpoint blockade drugs.

The MGH team used intravital microscopy—which examines biological processes in living animals through tiny implanted windows—to track the activity of an aPD-1 drug in mouse models of colon cancer. As expected, the labeled antibody was observed to bind to PD-1 molecules on CD8 T cells within a few minutes. But as little as 20 minutes later, the drug had been taken up by macrophages within the tumors.

The same process of rapid antibody binding to PD-1 molecules on CD8 T cells, followed by macrophage uptake, was observed in models of melanoma and lung cancer. To determine how the antibodies were being removed from T cells, the researchers first confirmed that the macrophages neither expressed PD-1 molecules nor did they take up antibody not bound to T cells.

“We further show that macrophage accrual of aPD-1 mAbs depends both on the drug’s Fc domain glycan and on Fcγ receptors (FcγRs) expressed by host myeloid cells and extend these findings to the human setting,” the authors continued. “Finally, we demonstrate that in vivo blockade of FcγRs before aPD-1 mAb administration substantially prolongs aPD-1 mAb binding to tumor-infiltrating CD8⁺ T cells and enhances immunotherapy-induced tumor regression in mice.”

Essentially, administering an Fc receptor inhibitor prior to anti-PD-1 treatment both extended the binding of the drug to CD8 T cells and led to complete tumor disappearance in a mouse model. Whether a similar strategy could improve the results of immune checkpoint blockade in human patients may be answered by current clinical trials that combine immune checkpoint blockers with drugs targeting macrophages, which have number of detrimental effects in cancer.

“Our observations would not have been possible without a method of dynamically imaging drug action on a cellular level,” noted Dr. Pettit. “Our platform for imaging anti-PD-1 in live animals can easily be adapted to study additional checkpoint blockade agents, so we are building a program to track the cellular interactions that will allow us to decipher drug mechanisms and hopefully leverage knowledge into engineering better therapeutics.”