Once cancer metastasizes it sets the course for a deadly chain of events with few treatment options. Understanding how cancer cells mutate and spread to new locations in the body may offer clues to stopping metastasis before it even starts.
Explore this interactive explainer article from Drug Discovery News to learn how cancer cells metastasize and spread, establishing new tumors in distant locations in the body.
Once cancer metastasizes, there is little that scientists and physicians can do to stop its spread. Understanding how cancer cells transform from stationary structures to mobile tissue-colonizing conquistadors may offer clues to stopping metastasis before it even starts.
BY TIFFANY GARBUTT, PHD, ILLUSTRATIONS BY SHANNON HERRING
Most cells tucked into the crevasses of mammalian tissues do not move, but cancer is different. Cancer possesses the unique ability to navigate throughout the body to establish new tumor sites in distant locations through metastasis. The biological toll of tumor growth in multiple locations is so immense that metastasis accounts for more than 90% of all cancer deaths (1). Despite this impact, only a few cells execute this process successfully. Cancer cells embarking on this odyssey must traverse the tumor microenvironment, endure the pressures of circulation, battle the immune system, and acclimate to a new cellular environment.
CAN ALL CANCER CELLS METASTASIZE?
Solid tumors are notorious for their cellular heterogeneity, but most solid tumors arise from exactly the same cell type: epithelial (2). Epithelial tissue is found throughout the body and lines most organs, from skin to the gastrointestinal tract. Accumulating genetic mutations in epithelial cells can cause them to aberrantly proliferate and form a tumor mass. As the cells continue to proliferate, cells at the center of the mass receive fewer nutrients. This couples with inadequate vascularization and increasing metabolic stresses such as hypoxia, resulting in the death of cells at the center of the tumor (3). Necrosis at the center of the tumor causes inflammation in the surrounding tissue, triggering the release of inflammatory cytokines that recruit immune cells to the presumed injury site (4).
Instead of remedying the situation, immune cell signaling lays the groundwork for metastasis. Growth factors and cytokines released by cancer-associated macrophages activate stem cell signaling pathways that convert select tumor cells into cancer stem cells (CSCs) (4). CSCs indefinitely self-renew and play a key role in tumor growth, recurrence, metastasis, and treatment resistance. Some scientists postulate that CSCs originate from resident adult stem cells or progenitor cells, but they are not the only key players in the story of cancer spread (5).
Myeloid and lymphoid cells secrete an assortment of cytokines that activate key developmental pathways and transcription factors that cause some epithelial cells to convert into mesenchymal cells (4). The epithelial to mesenchymal transition (EMT) of tumor cells is a hallmark of cancer metastasis. During EMT, select epithelial cells lose their cell-to-cell junctions and express elevated levels of vimentin, fibronectin, and N-cadherin, which facilitate dramatic remodeling of the cellular cytoskeleton, enabling these cells to traverse the tumor and extracellular microenvironments (5,6).
WHAT CHANGES HAPPEN IN THE MICROENVIRONMENT TO PROMOTE METASTASIS?
The extracellular microenvironment (ECM) consists of a network of collagen, elastin, fibronectin, laminin, proteoglycans/glycosaminoglycans, and other glycoproteins that bind to cells and add structure to tissues and organs (6). The ECM can be separated into two distinct layers: the basement membrane and the interstitial matrix. The basement membrane separates the epithelial cells from the underlying stroma. The interstitial matrix lies in the space between various cells (5). Tumor cells undergoing metastasis must penetrate both of these extracellular layers to navigate to new locations in the body. To accomplish this, tumor-derived factors initiate a cascade of events to reshape the ECM.
Tumor-secreted factors stimulate resident stromal cells to convert into cancerassociated fibroblasts (CAFs). Together with cancer cells, CAFs secrete large quantities of collagen and other ECM components that increase matrix stiffness around the tumor. The increasing density of ECM fibers places cell-surface receptors on tumor cells in close proximity to ECM components and promotes the physical connection of the ECM to tumor cells via integrins. Increased ECM density serves as a barrier that shields the tumor from immunosurveillance (7). The accumulating ECM also acts as a reservoir for transcription factors that support continued tumor growth (5).
Tumor cells and resident immune cells continue to secrete a compilation of chemokines and growth factors that recruit bone marrow-derived stem cells (BMDCs). In combination with tumor cells, CAFs, and BMDCs, neutrophils secrete matrix metalloproteinases (MMPs), which degrade the ECM. ECM degradation releases matrix-bound proteins, including vascular endothelial growth factor (VEGF), a critical protein needed for angiogenesis (7). The spouting of new blood vessels throughout the tumor lays the groundwork for the initiation of metastasis and provides a clear pathway to the rest of the body.
HOW DO CANCER CELLS BREAK FREE OF THEIR CONSTRAINTS?
Before cancer cells can migrate to the newly spouted vessels and ride the bloodstream to new organs, they must first break free of their cellular constraints and navigate through the unusually dense ECM.
In a tumor, cancer cells are held in place by various cell surface connections to other cells as well as to the basement membrane of the ECM. During the epithelial to mesenchymal transition, cancer cells lose their cell-to-cell junctions, freeing them of their connections to other cells, but they may still be held in place by integrins that connect them to the underlying stroma through the basement membrane (5, 6, 7).
To break free of this connection, tumor cells grow actin-rich foot-like projections called invadopodia. Integrins on the surfaces of invadopodia bind to ECM molecules and couple them intracellularly to contractile structures. Using these contractile structures, cancer cells pull ECM molecules apart. Cancer cell efforts are supported by CAFs that also possess invadopodia. Together, cancer cells and CAFs break the ECM fibers and apply force to the ECM basement membrane (7). Using this non-proteolytic approach, cancer cells breach the basement membrane and creep into the interstitial matrix.
Cancer cells can also break their connection to the basement membrane by secreting the potent ECM degrading proteases, MMPs. However, MMPs come in handy more for cancer cell navigation through the interstitial matrix. Both cancer cells and CAFs secret lysyl oxidase (LOX), a protein used to cross-link collagen fibers. Using LOX, cancer cells linearize and align collagen fibers in the surrounding interstitial matrix, creating smooth migratory tracks for efficient cell migration. MMP-facilitated ECM degradation further clears the path for cancer cells through the interstitial matrix (7).
As cancer cells arrive at newly formed blood vessels, they wiggle and slither their way through the tight junctions between endothelial cells that comprise vessel walls. In some cases, the structure of a cancer cell is so malleable that the cell passes directly through the middle of blood vessel cells without harming the cell. Cancer cells achieve this by pushing the cell surfaces of two endothelial cells together until they fuse. Cancer cells then pass through this fused surface almost like passing through the center of a donut (8, 9).
HOW DO CANCER CELLS SPREAD TO NEW LOCATIONS?
Once inside blood vessels, cancer cells face an arduous journey. They must endure the pressures of the circulatory system’s physical forces. For cells that are accustomed to a stationary environment, the 3-4 mph flow rate of the bloodstream is particularly stressful. Cancer cells have a maximum of three days to exit the blood vessel before they die (8, 10). Many cancer cells enter the bloodstream as single cells. However, they face a better chance of survival if they enter and travel through blood vessels as a clump of cells. In fact, clumps of cancer cells can make themselves thinner to better squeeze through capillaries during their travels (8). Clumps of cancer cells also interact with circulating platelets, which coat the cancer cells to prevent immune cell detection and help them better endure the stresses of circulation (6).
Not all cancer cells avoid immune cell detection. Macrophages and dendritic cells identify and eliminate a portion of circulating cancer cells. Cytotoxic T cells detect and destroy other cancer cells. To escape this fate, cancer cells express the cluster of differentiation 47 (CD47) protein on their surface. The signal regulatory protein alpha (SIRPa) on the surface of macrophages detects CD47 and interprets it as a “don’t eat me” signal, allowing tumor cells to proceed unharmed. To escape cytotoxic T cell detection, cancer cells modify the expression of their major histocompatibility complex one (MHC-I) proteins to almost undetectable levels. Circulating cancer cells also express the programmed death ligand (PD-L1) on their cell surface, which allows them to block key immune checkpoints. In the event that these defensive mechanisms fail, circulating tumor cells take a proactive approach and upregulate the FAS ligand (FASL), a transmembrane protein that triggers apoptosis, on their cell surface as a mechanism to kill patrolling T cells (11).
If circulating cancer cells can survive three days in the circulatory system and make it to the outer edge of the blood vessel, then they are protected. The outer edge of blood vessels serves as an oasis for traveling tumor cells, allowing them to rest and even lay dormant for extended periods of time. Exactly how long cancer cells can lay dormant at the outer edges of blood vessels is unclear, but many scientists suspect that this may underly the resurgence of metastatic cancer after remission. Some cancer cells may lie dormant for five, ten, or more years before finally reawakening and spreading to their new target tissues (8).
HOW DO CANCER CELLS TAKE ROOT AND GROW IN NEW TISSUES?
Although countless cancer cells embark on the journey through the bloodstream, fewer than 0.1% of them make it to their final destination to establish new tumors in distant organs. To account for such a treacherous journey, tumors have evolved mechanisms to ensure that these few successful cells land on fertile ground. First described in 1889 by Stephen Paget, a surgeon at the West London Hospital and the Metropolitan Hospital, the seed and soil hypothesis states that cancer cells from metastatic tumors can only grow in favorable microenvironments (6). In recent years, researchers have discovered that tumor cells send out signals ahead of metastasis to cultivate distant organ sites and establish a premetastatic niche.
Cancer cells are not the only factors that creep into the bloodstream. Tumors release exosomes, nano-sized membrane structures containing signaling molecules, and other biological contents from the parent tumor into extracellular spaces. These tiny vessels enter the bloodstream along with other tumor-derived transcription factors. If traveling cancer cells are the seeds, then exosomes and transcription factors are the fertilizer. As they enter blood vessels, they trigger a sequence of local changes that induce vascular permeability. These factors then seep out of the compromised vasculature and navigate to distant target locations (12).
For a long time, it baffled scientists as to why certain cancers repeatedly metastasized to particular organs. The findings from recent studies now suggest that integrins on the surface of exosomes guide circulating cancer cells to colonize specific tissue types (13). Once exosomes arrive at their target tissue, they activate resident stromal and fibroblast cells, converting them into CAFs. Similar to ECM remodeling near the parent tumor, CAFs begin to deposit new ECM components such as fibronectin that thicken the ECM in preparation for the arriving cancer cell. Exosomes also recruit BMDCs that contribute to immune response and angiogenesis. Exosomes can also deliver signaling information directly to endothelial cells to stimulate new vessel formation or to immunocytes to foster an immunosuppressive microenvironment. By the time the circulating cancer cell arrives at its target tissue, the landscape is already set to foster its continued growth (12).
GETTING AHEAD OF THE SPREAD
The challenge of preventing metastasis is that the process begins long before it is detected. Tumor-secreted exosomes may already be hard at work cultivating premetastatic niches in other organs, or circulating tumor cells may be lying dormant in hidden oases at the edges of blood vessels. While some scientists are tackling the problem of metastasis by stopping the spread of circulating tumor cells, others are trying to stop metastasis before it even begins by preventing the cultivation of pre-metastatic niches. Scientists are exploring the use of liquid biopsies to detect exosomes and predict metastasis. Once detected, research teams can then develop drugs to block the delivery of tumor exosomes to target cells. However, there are no know exosome biomarkers and the mechanism by which exosomes are taken up by cells remains unclear (13). Other strategies include increasing immune surveillance and regulating hypoxia in the primary tumor, which would decrease the release of exosomes and other tumor-derived factors that contribute to the establishment of premetastatic niches (14).
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