Metastasis formation is a multi-step process that involves the journey of a cancer cell from a primary tumor site to another localization. On the invasion site, cancer cells nest, grow and rebuild a secondary tumor, the metastasis. The metastatic process is clearly inefficient. It is estimated that a tumor of 1cm size, corresponding to one billion cells, can infiltrate the circulatory system with one million cells per day. The number of natural defenses and the high number of non- compatible invasion sites limit to less than 0.1% the number of cells actually developing a metastasis. Unfortunately for us, it still represents thousands of cells and 90% of all cancer deaths are associated to the presence of metastasis [Van Zijl et al., 2001; Valastyan and Weinberg, 2011]. For a carcinoma, the metastatic process is divided into 7 steps:
Local invasion of the surrounding tissue. This first step is crucial
and differs by the type of invasion induced. Cancer cells may move collectively, as a tissue, or by single cell progression via mesenchymal or amoeboid transition and invasion. During cancer progression, tumor cells modify their physiological and morphological characteristics to get around obstacles. Among those transformations, there is the epithelial to mesenchymal transition (EMT : “molecular and cellular program by which epithelial cells shed their differentiated characteristics, including
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cell-cell adhesion, planar and apical-basal polarity, and a lack of motility, and acquire instead mesenchymal features, including motility, invasiveness and a heightened resistance to apoptosis.” [Polyak and Weinberg, 2009]), the collective to amoeboid transition (CAT: this specifies the detachment of a specific carcinoma cell from the tumor by the use of amoeboid migration. Cells undergoing CAT show little connection with the extracellular matrix (ECM) and migrate through “holes” in the ECM without degrading it) and the mesenchymal to amoeboid transition (MAT : Mesenchymal cells produce matrix metalloproteases (MMPs) that help them to move through the ECM. If, for any reason, the extracellular proteolysis is blocked, MAT transition allows the cancer cell to continue its progression through amoeboid progression) [Van Zijl et al., 2001].
Intravasation in the lumen of lymphatic or blood vessels.
Although lymphatic dissemination is regularly observed, it seems that blood dissemination represents the main path of metastatic dissemination [Gupta and Massagué, 2006]. Blood vessels possess a basement membrane usually inhibiting cell movement. However, as mentioned earlier, cancer capillaries networks are tortuous and prone to leakiness. Moreover, weak interactions between endothelial cells facilitate the intravasation of cancer cells. Finally, intravasation of carcinoma cells may be assisted by cytokine such as TGF-β [Giampieri et
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al., 2009] or by cells like tumor activated macrophages (TAMs) [Hernandes et al., 2009].
Survival in the circulation. In the circulation, carcinomas cells are
renamed “circulating tumor cells” (CTCs). The circulation entails many stresses that cells have to deal with to survive. Firstly, lack of anchorage to the ECM is supposed to trigger anoikis, an apoptotic process triggered by the loss of anchorage to the substratum. However, since the average diameter of a carcinoma cell is 30µm and since average capillaries possess a diameter of less than 8µm, carcinoma cells are rapidly blocked in some capillaries, re-enter the surrounding ECM and evade anoikis before it even starts [Valastyan and Weinberg, 2011].
In the bloodstream, CTCs also suffer from predation by circulating immune cells and from physical stresses (pressure, movement…). A rather unexpected mechanism of defense consists in the expression at their surface of the membrane bound tissue factor (Tf) which is the receptor for coagulation factors VIIa and X. As consequence, CTCs shield themselves in platelets aggregation allowing them to escape physical stresses or immune destruction [Van Zijl et al., 2011; Valastyan and Weinberg, 2011].
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Arrest at a distant organ site. It is common knowledge that cells
originating from a particular cancer do not nest and create metastasis anywhere in the body. Some organs seem more appropriate than others. Already in 1889, Stephen Paget described an organ specific pattern of metastasis. For example, breast carcinoma rather invades bones, lungs and brain while colorectal cancers show a preference for the liver. This last example may be explained by the liver vascularization acting as a trap. CTCs, coming from the guts, are outgushed in the portal vein and rapidly arrive in liver capillaries with a diameter smaller than their size [Valastyan and Weinberg, 2011]. It remains to be determined whether CTCs are able to actively select the best environment for them, or if passive trapping in the capillaries is the sole mechanism for site selection.
Extravasation. Here, several hypothesis stand and may be
applied in parallel by CTCs. First, Al Medhi et al. proposed a mechanism in which CTCs trapped in microcapillaries continue to grow and develop a microtumor [Al-Mehdi et al., 2000]. Tumor growth may eventually break endothelial tight junctions, creating a hole in the capillaries allowing cells to invade the stroma. This hypothesis does not take into account anoikis, nevertheless, it is likely that specific CTCs are able to downregulate anoikis due to the spectrum of mutations they contain. In
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addition, CTCs could undergo extravasation, the transfer of the cell from the lumen of the capillaries to the ECM of the colonized organ. Mechanistically, evidences suggest differences with intravasation, even if TAMs also help CTCs to cross the endothelial barrier [Qian and Pollard, 2010].
Micrometastasis formation. The new environment differs from
the initial tumor site. Characteristics of the invaded organ, such as the type of stromal cells, ECM constituents or growth factors available may not be suitable for the newly arrived carcinoma cells. Some recent studies raised the hypothesis that cancer cells may prepare the invasion site before actually leaving the primary tumor. Systemic signals may be released in the blood to modify the normal behavior of fibroblasts in the future invaded organ. For example, on the invasion site, cells expressing MMP9 were shown to be recruited, remodel the ECM and release cytokines. This hypothesis is currently questioned and under intense debate [Psaila and Lyden, 2009]. Another hypothesis to CTCs adaptation to a new environment is simply its autonomous program. Cells express proteins and use pathways that dodge the harmful conditions imposed by the foreign environment.
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Metastatic colonization. Once survival on the new site is
achieved, proliferation is not guaranteed. Indeed, cells may enter a quiescent state and fail to replicate. In breast cancer, this has been linked to a lack of adhesion with the surrounding ECM. Moreover, if cells effectively begin to divide, the apoptotic rate may be so intense that they will never be able to develop a metastasis. Mechanisms responsible for apoptosis, in this case, are poorly understood but failure to induce neoangiogenesis may be a key to understand the high apoptotic rate [Chambers et al., 2002].
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