The German-born PhD candidate conducted his doctoral research under the supervision of Wilbert Zwart, a full professor at the TU/e Department of Biomedical Engineering and the leader of the Hormone-associated Cancer research group at the Netherlands Cancer Institute (NKI). “In this group we focus on hormone-dependent cancers, primarily breast cancer and prostate cancer, but in recent years we've been finding that hormones also play a role in some other types of cancers, such as lung cancer,” Linder says. Conducting his doctoral research within this research group, his own particular area of study was prostate cancer. This is a very prevalent hormone-driven cancer; in the Netherlands alone about 12,000 men are diagnosed with the disease every year, and more than 3,000 die. “So there is still much we can do to improve the therapy available to these patients.”

Resistance

“If the disease is contained within the organ, we can remove the prostate surgically - with a prostatectomy as it is called - or we can give radiotherapy,” he continues. “Around 70% of patients are cured by this intervention, which is great. But in around 30% of patients the disease returns.” These patients are usually treated with hormonal therapy. This treatment aims to inhibit the body's production of male sex hormones, called androgens, which fuel prostate cancer growth.

“In prostate cancer cells, hormones activate a certain pathway to cell growth and survival. If you either block the biosynthesis (assembly of biomolecules, ed.) of these hormones or inhibit the molecule that needs the hormones to activate the pathway, you can successfully inhibit this process”, Linder explains. “These therapies are very effective at first, but the big problem is that the tumors become resistant to them pretty soon.”

Smart cancer cells

“In a way, cancer cells are very smart,” he continues. “If you block the pathway they usually take and depend on, after a while they find byways that enable them eventually to accomplish the same goal. By which I mean they activate something else that can carry and drive the disease.” Unfortunately it is inevitable that sooner or later these tumors will become resistant. And it is this resistance that poses the biggest challenge in treating both prostate cancer and other hormone-driven diseases.

The important question is this: how do these tumor cells adapt to hormonal therapies? If we are to prevent cancer cells from developing resistance, a more fundamental understanding of these processes is required. “We need to outsmart them,” Linder says. During his research at the NKI, in association with the Antoni van Leeuwenhoek clinic, he focused on patients with primary, localized prostate cancer in order to gain greater insight into the behavior of cancer cells. “We gave these patients hormonal therapy three months before they had surgery. Biopsies were taken before and after the hormonal therapy which enabled us to address an important question: what does hormone therapy do to cancer cells?”

The bigger picture

In this work, Linder used what is known as the multi-omics approach. This involves analyzing a large amount of biochemical data to gain new and deeper insights. Omics refers to different fields of molecular biology that end in ‘-omics’, such as genomics (the study of DNA), transcriptomics (the study of RNA) or proteomics (the study of proteins). “With multi-omics, we're looking at all the molecules of the same type, let's say DNA, in the same sample,” he explains. “So we don't look at only one DNA sequence, but at all of them. And we do the very same thing for the RNA - a kind of recipe for how to make the protein later on - and for the proteins themselves.”

“With this broad multi-omic analysis, you have a lot of small parts that you need to put together to form the bigger picture. It's like a jigsaw puzzle with lots of little pieces that add up to something much bigger.” Linder analyzed all these factors within the samples taken before and after hormonal treatment to see what was different and to understand what was driving the resistance. The ultimate goal would be to find ways to block this and prevent resistance, or even to re-sensitize cells that have become resistant to the therapy.

Surprising discovery

After extensive analysis, he discovered that a protein that normally plays a role in a very general physiological process had suddenly taken on a whole new role in driving this resistance. "This protein normally plays a role in the circadian rhythm, meaning the day and night rhythm that is present in every cell of our body and causes us to be more awake in the morning and sleepier at night," he explains. "The interesting thing is that the circadian rhythm is often lost in cancer cells and suddenly this protein was given a new role in prostate cancer. We were all very surprised by this, because it has never been described in prostate cancer before, especially not in driving resistance to hormonal therapy."

Linder decided to delve deeper into this and then conducted more analysis to confirm this surprising discovery. No longer on patient samples, but in vitro, using a disease model that he grew in the lab to see how the cells respond to therapy. "This way we can both model therapies and also try to better understand the biology of these tumors," he explains. This analysis work showed that resistant cells become sensitive to therapy again when you prevent the production of this specific circadian rhythm protein, confirming the role of this protein in the resistance mechanism. "If you block the main pathway, the cancer cells find a workaround to achieve their goal. But now if we also block this bypass option, the cancer cells can't do anything and eventually they die."

Outside the box

"It was very cool to see that these initially very broad analysis with lots of patient samples and lots of data eventually led to one driver that could be validated in these models," Linder says. Ultimately, he believes these findings could lead to a new therapeutic approach for patients with prostate cancer. "Not today, but in the future," he adds, since specific drugs that target this circadian rhythm protein first need to be developed.

“From this research project I learned that you should never rule out anything in advance,” he continues. “If you had asked me beforehand what drives resistance in prostate cancer, the circadian rhythm would never have been something I'd have put my money on, just because it’s such a normal physiological pathway. But now we know that cancer cells can hijack even proteins that normally play a role in a very normal process and use them to achieve their ultimate goal, which is to grow and survive. So if we want to advance therapies and improve patients' lives, we need to do more thinking outside the box."