Sleeper cells: the science of cancer dormancy

The path to finding dormant cancer cells began, like so many scientific discoveries, with a happy accident. Lewis Chodosh, MD, PhD, chair of the department of Cancer Biology at Penn Medicine and associate director of basic science at the Abramson Cancer Center at the University of Pennsylvania, is an endocrinologist by training, and became interested in breast cancer because of its close association with hormonal risk factors.

More than 20 years ago, he developed a genetically engineered mouse model of breast cancer to study how pregnancy hormones affect cancer risk. For that question, the model turned out to be a bust. But he found that some of the animals had cancers recur months after their primary tumors had been eradicated. “That told us there had to be residual tumor cells that survived, maybe in a dormant state,” he said.

He recognized the significance at once. Nearly a third of all human breast cancer survivors have a relapse of their illness. Those recurrent tumors have the same molecular features as the original tumor, suggesting that some dormant cells are left behind after treatment, lying in wait before eventually growing again. But there was no sign of the cells in the bloodstream, no evidence on a scan—no clear way to find, let alone study, the mysterious sleeper cells. “Now, with the mice, we had a window into the biology of dormant tumor cells,” he said.

Thus began a years-long scientific chase to find and eliminate the cells in his mouse models. Marking tumor cells with fluorescent tags, he could isolate and track any cells that remained after the primary tumor was gone. The cells, known as dormant disseminated tumor cells (DTCs), sometimes stayed near the original tumor site. But often, a reservoir of the cells settled in the bone marrow. Chodosh was able to prove that those dormant cells were the source of recurrent cancers in mice.

He and his team made other important discoveries about these strange sleeper cells. For one thing, they didn’t behave like typical cancer cells. They didn’t divide, nor did they use standard cellular energy sources.

“It was crystal clear that these dormant cells were in a biological state that was completely different from an actively growing primary tumor, and also completely different from an actively growing recurrent tumor,” Chodosh said.

What’s more, the dormant cells all looked and behaved similarly to one another, even when they came from mice with different types of breast cancer. “It was striking to us that these dormant cells were in some special biological state,” he added.

Chodosh and his team continued their efforts, mapping several cellular processes that the DTCs use to survive in their simmering dormant state. The dormant cells, for instance, often turned to a process known as autophagy, in which they digest parts of themselves to generate energy. The mTOR pathway, known to regulate growth and metabolism, also seemed to play a role in keeping the cells alive.

Chodosh found that traditional chemotherapy agents didn’t do much to damage the dormant cells. But treatments aimed at blocking autophagy or the mTOR pathway were a different story. “By treating mice with those agents, we both decreased the number of residual tumor cells, and improved recurrence-free survival rates,” he said.

Needles in a haystack

It was a thrilling breakthrough. Yet even after the concept was proven in mice, moving the research to humans required a big leap.

The first hurdle was simply to find the dormant cells.

“Looking for these cells in the bone marrow, you really are looking for a needle in a haystack,” said Jonni S. Moore, PhD, a professor of Pathology and Laboratory Medicine at Penn Medicine.

The traditional method involved taking a bone marrow biopsy, then staining the sample with an antibody that detects proteins on the surface of cancer cells. But the assay wasn’t terribly sensitive, and it destroyed the DTCs in the process of finding them. “You’d be looking for one brown cancer cell in a sea of blue bone marrow cells,” Chodosh said. “And once you find it, you can’t do any genomics on the cell, or study what other proteins it might express.”

So Chodosh’s team set about creating a new test using flow cytometry, a laser-based technique that can detect various chemical and physical differences between cells in a sample. The project was led by Elizabeth Chislock, PhD, a senior scientist in Chodosh’s lab, in collaboration with Moore, who has longstanding expertise in flow cytometry, and DeMichele, who designed and actualized the complex process of obtaining bone marrow aspirates from patients. Advances in technology made the goal of finding DTCs feasible, but the task remained daunting. Bone marrow contains a complex mix of many cell types, including various immature cells at different stages of development—some of which express biomarkers that mimicked potential markers that could have been used to pinpoint DTCs.

Over the better part of a decade, Chislock worked to differentiate the DTCs from those many other cell types. The result of that painstaking work was a next-generation assay, DTC-Flow, that can pick out a single dormant DTC among 15 million cells, while also preserving the cell for further analysis. That technology was instrumental in moving forward with clinical trials in breast cancer survivors, in a series of studies led by Angela DeMichele, MD, MSCE, the Mariann T. and Robert J. MacDonald Professor in Breast Cancer Care Excellence and co-leader of the Breast Cancer Research Program at the Abramson Cancer Center at the University of Pennsylvania, and her clinical colleagues.

Now, the team is working to refine the complicated assays, developing a standardized test that could be FDA-approved for use in clinical settings.

“It started with a mouse and a simple assay. Now we can isolate one cell in 15 million. There’s no other technology that can do that,” Moore said. “But it’s not just having the right technology, it’s also about having the right infrastructure, interactions, and opportunities—and that’s why I’m so excited about this collaboration.”

That collaboration, accelerated through the Translational Center of Excellence for this work at the Abramson Cancer Center, is already bearing fruit. DeMichele is moving forward with clinical trials while Chodosh continues exploring the mechanisms that keep dormant cancer cells alive, and how to target those lifelines to wipe out the cells for good. He’s also working to understand what factors trigger the sleeper cells to reawaken from their zombie state. “That’s the critical event that will lead to a recurrence,” he said.

Once seen as a long shot, the idea of preventing cancer recurrence no longer seems so implausible. “We were told for years that this was a crazy idea,” Chodosh said. “Not only is it not crazy, I’m convinced this is the future.”