How Golgi stress affects T-cells' tumor-fighting ability

It's a core function, but little studied in the setting of cancer immunology, especially when compared to other organelles like the mitochondria or endoplasmic reticulum.

"So we were interested in looking a little bit more at the Golgi apparatus. It's obviously an important organelle. How is it being changed or what is its role in T-cells in terms of fighting cancer?" said Nathaniel Oberholtzer, an M.D./Ph.D. student who worked in the lab of Shikhar Mehrotra, Ph.D., co-leader of the Cancer Biology & Immunology Research Program at MUSC Hollings Cancer Center and scientific director of the Center for Cellular Therapy in the MUSC College of Medicine.

As it turns out, the healthy function of the Golgi apparatus has a lot to do with how well T-cells function in killing cancer cells. Understanding how a signaling axis mitigates Golgi stress, enabling it to perform properly, points to a possible new therapeutic target for researchers to pursue to strengthen T-cells. Not only that, but Oberholtzer's research shows how the Golgi could be used as a biomarker to select the strongest T-cells for immunotherapy.

Oberholtzer, as first author, and Mehrotra, as senior author, along with a team of Hollings scientists published the research this month in Science Advances.

T-cells, part of the immune system, can kill cancer cells. CAR-T cells are T-cells that have been modified in the lab to home in on proteins on the surface of an individual's cancer cells. CAR-T cells are custom-made for each patient.

Both T-cells and CAR-T cells can become "exhausted" in the hostile tumor microenvironment. Mehrotra's lab looks at ways to boost these cells so that they can fight cancer for a longer time.

"The whole tumor microenvironment is conducive for the tumor itself, but not for the other cells which are trying to get in there," Mehrotra said.

Just like people, cells are constantly subjected to stress -- stress from biochemical reactions that have become unbalanced and mechanical stress from moving. Transient stress can be good. Stressing your muscles through exercise strengthens them, and transient stress on cells can prompt them to a response that ultimately strengthens them.

"But if this stress stays there, which it does in the tumor microenvironment, the cells are just in continuous stress, and that will then lead to a very different phenotype and death," Mehrotra said.

However, the researchers found that treating the Golgi apparatus with hydrogen sulfide created T-cells that could take more stress.

"Hydrogen sulfide is a gaseous signaling molecule present in pretty much all mammalian cell types. Typically, it's a byproduct of different cellular processes, but it's actually been shown to have really important signaling roles as well," Oberholtzer said.

"It can modify proteins through a process called sulfhydration, where it modifies cysteine residues and can change their activity."

In this project, Oberholtzer found that this sulfhydration process, in modifying a protein called Prdx4 within the Golgi apparatus, confers protection in an oxidative setting.

"When you have the stressors that the tumor microenvironment puts on T-cells, you get a disruption, or fragmentation, of the Golgi apparatus where it essentially isn't able to do its job. Hydrogen sulfide protects against that disruption," Oberholtzer said.

Looking into this protective effect then led the researchers to look more closely at the Golgi apparatus by itself.

"Essentially, if you just use the Golgi apparatus as a simple marker, if T-cells have a lot of Golgi versus less, the ones that have more Golgi are much more robust at killing tumor cells and controlling tumors," Oberholtzer explained.

Using cell sorting technology at the Flow Cytometry & Cell Sorting Shared Resource at Hollings, the researchers sorted T-cells according to the amount of Golgi they contained. The top 30% were labeled Golgi-hi and the bottom 30% were labeled Golgi-lo.

"Basically, all the cells which are expressing high Golgi have a very different phenotype. They are less exhausted, and they are much more potent in controlling tumors," Mehrotra said.

This pre-clinical work suggests that sorting T-cells into Golgi-hi and Golgi-lo and reinfusing only the Golgi-hi cells into a patient would create a better chance of controlling the tumor.

"Right now, we're working on doing some validation studies in the Center for Cellular Therapy to potentially be able to start a clinical trial to see if that has a translational ability as well," Oberholtzer said.

More work is also needed to understand the role of Golgi stress when all of the organelles in a cell are under stress because of the tumor microenvironment.