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The secret signals tumor cell clusters use to drive metastasis

New study identifies 'nanolumina': intercellular chambers where tumor cells share growth signals, driving cancer spread
An image of a cluster of cancer cells with epigen, highlighted in bright green, being overexpressed between cells.
Pictured, a highly magnified image of a tumor cell cluster. Each purple area is a cell, green is the epigen concentrated between them in nanolumina. Epigen is a growth factor, which means it can tell cells near it to divide, grow, and form a new tumor. Stopping epigen from accumulating between cancer cells may slow or stop the growth of new metastatic tumors. Photo courtesy of Cheung Lab / Fred Hutchinson Cancer Research Center

It’s not the cancer in your breast that kills you, it’s the breast cancer that sets up shop in other organs like your liver or lungs or brain and starts growing uncontrollably.

Metastatic tumors, also called advanced or secondary cancer, are potent killers. In the U.S. alone, nearly 43,000 people die each year from breast cancer that’s metastasized. Metastatic prostate cancer kills around 33,000 a year and metastatic lung cancer claims a whopping 135,720.

And while science hasn’t yet figured out how to stop metastasis, a team of researchers at Fred Hutchinson Cancer Research Center have figured out that tumor cell clusters — clumps of anywhere from three to dozens of cells that break off from a primary tumor and travel through the lymph or bloodstream — are aggressive drivers of metastatic, or stage 4, disease.

Today, that team has a new paper out in the journal Cell outlining their discovery of a secret space between the cells of each microscopic cluster where the signal to grow is co-opted and miscommunicated to other cells. They’re now working on new ways to block that signal — call it growth factor “fake news” — in order to stop metastasis in its tracks.

“Metastasis forms in many ways,” said Dr. Kevin Cheung, a Hutch translational researcher who also treats breast cancer patients at Seattle Cancer Care Alliance. “In this paper we show that inducing cancer cells to become clusters gives them a 500-fold increase in the cells’ ability to form metastasis. We sought to understand why.”

Photograph of Fred Hutch researcher Emma Wrenn
Molecular biology grad student Emma Wrenn was first author on the new paper, just published in the journal, Cell. Fred Hutch file photo

A ‘nanoscale microenvironment’

For the past several years, the Cheung Lab, including the study’s first author and Fred Hutch / University of Washington molecular biology grad student Emma Wrenn, have been plumbing the biology of tumor cell clusters, trying to get to the heart of the mechanism that drives the formation of metastatic tumors, or “mets.”

They study the deadly process using mouse and patient-derived organoid models of breast cancer (in lab dishes), confirming their findings with high-resolution electron microscopy and other imaging techniques.

In this new paper, Cheung and team shared the first-ever views of a secret signaling chamber they call “nanolumina,” (lumina are “inside spaces”) where tumor cells communicate with each other as a group, a process called collective signaling.

“It’s sort of a nanoscale microenvironment, a localized signaling environment between cells,” Cheung said. “And it turns out cancer cells are communicating with each other via a signaling molecule called epigen that promotes the outgrowth of those cells in contact with it.”

“It’s very early days but we’re excited to think this could be a new means of treating breast cancer. That’s the dream.”

Fred Hutch metastatic breast cancer researcher Dr. Kevin Cheung

Epigen is a signal that’s received by a receptor on cells called epidermal growth factor receptor, or EGFR. It’s one of several molecular signals detected by EGFR, and “the most recently discovered,” Cheung said. These small molecules transmit signals between and within cells, working hand in glove with their receptors to regulate key processes of cell biology within organisms, big and small. They help our cells grow and develop, maintain optimal tissue functioning and, sometimes, create tumors.

Microscopic images of tumor cells and the nanoluminal spaces between them where epigen is overexpressesd.
Breast cancer tumor cell clusters with nanolumina highlighted in breakout box.  FAR LEFT: A breast cancer tumor cell cluster (around 5 cells). Highlighted in the yellow box (and enlarged center) is a nanolumina between two cells (the dark gray). FAR RIGHT: Cells pseudo-colored to highlight the nanolumina.  When tumor cell clusters form these nanolumina spaces, they concentrate special types of proteins called growth factors inside the nanolumina, which tell the cluster cells to divide and grow into a new metastasis. Photo courtesy of Cheung Lab / Fred Hutchinson Cancer Research Center

The Cheung Lab found that when clustered, tumor cells are somehow able to co-opt epigen signaling, using the molecule to broadcast bad information to its neighboring cells, much like an untrustworthy media outlet might transmit misinterpreted facts and science.

In both instances, it causes big problems.

“It acts like a switch between migration and proliferation,” Cheung said. “When epigen is present, they stay put. They start to divide rapidly and grow. We see this in [tissue] cultures; in human tumors; in in vivo and in metastatic mouse models. If you get rid of it — via epigen knockdown or suppression — the cells migrate more efficiently.”

Without epigen, “they still cluster — but instead of growing, those clusters migrate,” Wrenn said. “This suggests epigen helps cells decide whether to grow or move.”

Cheung said their findings definitely point to epigen being a molecule of interest.

“It suggests we could target collective signaling in patients,” he said. “It’s very early days but we’re excited to think this could be a new means of treating breast cancer. That’s the dream.”

‘Profound’ reduction in metastatic growth

For decades, scientists believed metastasis started with a single cancer cell that “crawled away and set up shop,” Wrenn said.

But Cheung Lab research shows tumor cell clusters are a prime metastatic culprit.

“Not only does cancer metastasize as clusters, clusters are more powerful than single cancer cells,” she said. “They travel through the body in a clump and resist hostile signals from the immune systems or the organs where they’re setting up. Within the nanolumina, though, they can share pro-growth signals with each other that have the potential to supersede the other hostile signals.”

To demonstrate how clusters flex their collective power, the team ran a number of experiments.

Using a mouse model of an aggressive breast cancer, they tracked and compared the metastatic growth of single cells versus tumor cell clusters. They divvied cancer cells into groups, letting each group grow for zero hours, six hours, 24 hours and, finally, 24 hours followed by separation back into single cells.

They then injected cells from the separate samples into mice.

Tumor cell clusters that grew for six hours formed 140-fold more lung mets than an equal number of single cells. And clusters that grew for 24 hours formed 536-fold more lung mets than an equal number of single cells. Tumor cell clusters also showed much higher capacity for survival than single cancer cells.

Using high-powered microscopes, they looked at the miniscule clusters and discovered tiny spaces between the cells. Follow-up experiments identified epigen as a key signaling molecule in these tiny spaces and found that suppressing it “profoundly” reduced primary tumor and metastatic outgrowth.

“Whether it’s the primary tumor or crawling through the body or within a metastatic tumor, they have a secret signal between themselves that says, ‘Keep growing,’” Wrenn said. “Obviously, we don’t want that.”

A high powered microscopic image of two slides sideby side, one with a cluster of tumor cells with epigen overexpressed, and another with epigen suppressed.
At left, a growth of metastasis in a mouse lung that is expressing epigen. Tumor cells are bright red and cell nuclei are blue. At right, growth of a metastasis when the cells are prevented from making the growth factor epigen. Photo courtesy of Cheung Lab / Fred Hutchinson Cancer Research Center

A few answers, many more questions

So far, the team has observed epigen-laden nanolumina in aggressive luminal B and triple-negative breast cancer cells. Basal-like 2 triple negative breast cancer, which is difficult to treat, had nanolumina with particularly high amounts of epigen.

Cheung and Wrenn hope to launch a clinical trial to identify drugs (ones that are already FDA-approved, if possible) that can tamp down the epigen.

“We’ll either break open the nanolumina so everything leaks out, or add drugs that can block the epigen signaling,” Wrenn said. “There’s not an anti-epigen drug, but we’re trying to make one. If we keep these clusters from having this secret, highly concentrated pro-growth signal, they should stop growing or grow less. And that would be beneficial for patients.”

This type of trial, of course, is still years away. For now, they need to answer basic questions.

“We know that breast cells have nanolumina, but we don’t know if prostate or liver cells do,” said Wrenn. “That’s an important question to answer. We’re talking about using this to treat a subset of a subset of cancers [basal-like 2 triple-negative breast cancer], but if we find it in another cancer, there would be that many more people this could help.”

Wrenn said she’s interested in getting "50 different patient tumors from different cancers and see if they have nanolumina, see how widespread these structures are.” She even encouraged researchers outside the Hutch to conduct their own investigations of this no-longer-secret intercellular space.

“That would tell us a lot about the underlying biology and who might benefit from this type of treatment,” she said.

Opening up the ‘locked box’

Cheung said it’s also crucial to discern normal cell function and signaling from normal growth signals that are being co-opted. Cancer has a reputation for “stealing” normal biological functions. Cheung thinks nanolumina may play a role in normal development that’s being co-opted to drive metastasis.

Wrenn explained it like this:

“It may be that when a normal mammary gland is growing, it has nanolumina and epigen which tells it in a normal healthy context, ‘You’re supposed to grow,’” she said. “But then the cancer cells take that and use it inappropriately. They use the same signals for normal growth but it’s not coordinated and regulated the right way. That’s when you get a tumor.”

In other words, cancer hijacks a mammary cell’s “puberty programming” and uses it to grow tumors instead of breasts.

Through patient tissue samples, Cheung has determined that nanoluminal spaces are different between patients, are different with metastatic progression and are different with therapy. And, he added, there may be other potential targets, other growth factors run amok, lurking within these unexplored spaces.

“It’s like a treasure chest or a locked box,” he said. “Molecules inside this compartment could be particularly important to tumor cells.”

Cheung will be delving into these intercellular signaling molecules further with a new grant award from the National Cancer Institute. He will receive $400,000 per year for an initial five-year stretch, after which he’ll have the opportunity to extend his project for two more years. 

“We’re still discovering totally new ways that cancer is growing and communicating, and every time we do, that means we have a new target and new tools that can be used to destroy it,” Wrenn said. “Some of those turn into dead ends, but we only need one to work in order to help a lot of people.”

This research was funded by the Department of Defense, the Burroughs Wellcome Fund, the Breast Cancer Research Foundation, the V Foundation, Phi Beta Psi, Seattle Translational Tumor Research and the National Institutes of Health / National Cancer Institute.

Scientists at Fred Hutch played a role in developing these discoveries, and Fred Hutch and certain of its scientists may benefit financially from this work in the future.

 

Diane Mapes is a staff writer at Fred Hutchinson Cancer Research Center. She has written extensively about health issues for NBC News, TODAY, CNN, MSN, Seattle Magazine and other publications. A breast cancer survivor, she blogs at doublewhammied.com and tweets @double_whammied. Email her at dmapes@fredhutch.org.

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Last Modified, September 30, 2020