Role of Matrix Metalloproteinases in Angiogenesis and Cancer

During angiogenesis, new vessels emerge from existing endothelial lined vessels to promote the degradation of the vascular basement membrane and remodel the extracellular matrix (ECM), followed by endothelial cell migration, and proliferation and the new generation of matrix components. Matrix metalloproteinases (MMPs) participate in the disruption, tumor neovascularization, and subsequent metastasis while tissue inhibitors of metalloproteinases (TIMPs) downregulate the activity of these MMPs. Then, the angiogenic response can be directly or indirectly mediated by MMPs through the modulation of the balance between pro- and anti-angiogenic factors. This review analyzes recent knowledge on MMPs and their participation in angiogenesis.


Epithelial-Mesenchymal Transition (EMT) in Metastasis and Migration
Currently, cancer research is focused on understanding the functional mechanisms underlying cell transformation and tumor progression that can be used to develop new markers and therapies (1). Cancer metastasis, the final step of tumor progression and the leading cause of cancer morbidity and mortality, involves the spread of cancer cells from the primary tumor to nearby tissues and distant organs; it is mediated by complex molecular changes of in cell cycle regulation (2,3). The molecular changes that regulate the cell morphology and functions of epithelial cells, that is epithelial-mesenchymal transition (EMT), include the destruction of intercellular relationships and cell-matrix adhesive characteristics, extracellular matrix (ECM) breakdown, and cleavage of basement membrane components by matrix metalloproteinase (MMP) activity modulation. For example, when epithelial cells lose their polarity through EMT, cell-cell tight junctions and adhesive connections are lost, resulting in infiltration and an enhanced migration ability of these cells (4,5). Therefore, EMT enables malignant cells to become motile and invasive, which constitutes a fundamental requisite for cancer metastasis (6).
On the other hand, angiogenesis, in which MMP participation is well-recognized, was found to be involved in cancer metastasis over 45 years ago. Interest in angiogenesis related to cancer arose in 1968 when it was highlighted that tumors secrete a diffusible substance that stimulates angiogenesis (7). It is now recognized that angiogenesis plays a crucial role in the establishment of cancer and is the rate-determining step in tumor progression (7,8). Numerous studies have demonstrated the key participation of MMPs along with EMT to promote angiogenesis, infiltration by cancer cells, and metastasis (9)(10)(11)(12)(13). MMPs are a family of zinc-binding metalloproteinases that participate in the degradation of ECM components, including the basement membrane and the tumor surface, resulting in tumor cell migration into the near tissue. Furthermore, MMPs promote tumor growth and spread through the capillary endothelium and neovascularization (14).
Given the relevance of MMPs in diseases such as cancer, this work presents the most representative studies on the subject. We emphasize the role of cytokines and growth factors inducing EMT in various types of cancer together with the role of MMPs. We also analyzed the carcinogenic and angiogenic processes, and with the participation of MMPs, cytokines, and immune system cells in these processes along with the regulation, activation, and signaling pathways of MMPs in cancer cells.

BIOCHEMICAL PROPERTIES OF MATRIX METALLOPROTEINASES
MMPs, also known as matrixins, are members of the metzincin protease superfamily of zinc-endopeptidases.They display a specific proteolytic activity against a broad range of substrates located on the ECM. Other members of the superfamily include A Disintegrin and metalloproteinases (ADAMs), and ADAMs with thrombospondin motifs (ADAMTSs), which contain a conserved methionine (Met or M) residue adjacent to the active site (15,16). The first MMP (collagenase/MMP-1) was identified more than five decades ago (17). Since then, a total of 28 members have been named MMPs and given a distinctive numbering, and have been identified in vertebrates. In humans, there are 23 paralogs of MMPs (including a duplicated MMP-23 gene that encodes two identical forms of MMP-23), out of which at least 14 can be found expressed in the vascular endothelium (18,19). The typical structure of MMPs consists of an N-terminal zymogenic propeptide domain (∼80 amino acids), a metaldependent catalytic domain (∼170 amino acids), a linker region (∼15-65 amino acids), and a C-terminal hemopexin-like domain (∼200 amino acids) (Figure 1) (19,21). MMP classification is traditionally centered on the substrate specificities observed and the common structural domain architecture. The MMP family can be divided into at least six subfamilies: (1) collagenases; (2) gelatinases, (3) stromelysins, (4) matrilysins, (5) MMP membrane-type (MT)-MMPs, and (6) other MMPs. However, since they present a wide range of substrates and different functions, many of these are similar but have a different biological function that has yet to be clarified. We used a classification related to their evolutionary origin to locate MMPs that have not been properly classified (21,22) (Figure 2).
All MMPs are produced as proenzymes and require a proteolytic cleavage under physiological conditions to promote the release of the propeptide domain (zymogen activation) and generate mature MMPs (22). This means that the activity of MMPs is regulated by a post-translational proteolytic cleavage and endogenous inhibitors (15,21). Nevertheless, efforts to define the substrate recognition profile by MMPs have resulted in substrate selectivity conferred by key subsite interactions (P3, P1 ′ , P2 ′ , and P3 ′ ) with a motif sequence specificity "P-X-X-|-L-X-X, " even though combined frequencies of subsites have been observed. It is known that subsite P3 maintains a high preference for Pro; still, many MMPs favor small residues (Ala/Val/Ile/Leu) and less frequent aliphatic residues. While subsite P1 ′ maintains hydrophobic residues with preferences for Leu/Ile/Val/Met, subsite P2 ′ maintains preference for Ile/Val, Glu/Asp, and Lys/Arg/His depending on the MMP. Finally, subsites P3 ′ and P2 ′ are inconsistent in all MMPs with any preference for Gly and Ala (26,27). Therefore, the ability to recognize a wide variety of substrates selected by profile signatures by MMPs involves the peptide hydrolysis of latent protein targets, located on the ECM and the surface of the cell membrane.
Moreover, the MMP catalytic domain of the metzincin clan of metalloendopeptidases shares a general zinc-binding signature as core of the catalytic reactivity; the signature conserved sequence is the H-E-X-X-H-X-X-G-X-X-H/D region. Additionally, the conserved M residue of the superfamily is located on the methionine containing turn (Met-turn) which is part of the catalytic region and likely has structuralstability functions; nevertheless, the strict conservation of this residue remains unclear (28,29) (Figure 1). All MMPs differ in expression, localization, substrate profile specificities, and structural organization. For further details about the structure and function of MMPs see (14,15,30).

CANCER AND ANGIOGENESIS
Angiogenesis is a process by which new blood vessels or capillaries grow from the preexisting vasculature, and it is necessary for diffusion of nutrients and delivery of oxygen for tissue metabolism or cells involved in wound healing, myeloid and stromal cells. New blood vessels require the dismantling of endothelial lined vessels via the "sprouting" of endothelial cells (ECs), expanding the vascular tree (31). Moreover, the neo-vessel networks play more complex roles in diverse tissues such as the endometrium during the menstrual cycle, implantation, and endothelial cell migration out of the existing blood vessels (32). Given the complexity of a process as angiogenesis, the vascular endothelial growth factor, VEGF (VEGF-A), plays a remarkable role in signaling through the VEGF receptor-2 (FLK1) which induces angiogenesis in both health and disease processes. VEGF activity is enhanced by VEGF co-receptors, such as NRP1 and NRP2. In contrast, the loss of VEGF results in the interruption of vascular development. Placental growth factor (PlGF) is a cytokine VEGF homolog that stimulates angiogenesis and various types of cells, such as myeloids and stromals cancers, in addition to activating tumor cells, while their inhibition improves cancer treatment (33).
Collagenases (MMP-1, −8, and −13) are proteins associated with angiogenesis, and their loss leads to the irreversible rupture of the matrix. Type IV collagen participates in cell endothelial migration in the interstitial stromal spaces. It is known that the tissue inhibitors of metalloproteinases (TIMP-1, TIMP-2, TIMP-3, and TIMP-4) regulate them, playing a key role in angiogenesis regulation by inhibiting neovascularization (34).
In adults, angiogenesis is initiated only under inflammation or hypoxic conditions (35). In the early proliferative stage, vascular repair must predominate to control bleeding by vasoconstriction and coagulation. During menstruation, the endometrium is expelled if the ovule is not fertilized. Women who suffer from  Figure 1E) with MMP-23A and MMP-23B proteins. MMP-23A gene is considered a pseudogene produced by duplication of the MMP-23B gene. Sources: "GeneCards: the human gene database" (24) and Uniprot databases (25). endometriosis show aggressive angiogenesis in the peritoneal cavity (36).
On the other hand, several studies have established the importance of transmembrane receptors and ligands participating in cell differentiation. Their role in endothelial sprouting during angiogenesis has recently been studied. ECs express several Notch receptors (such Notch1 and Notch4), as well as the Notch1 protein and Notch ligand delta-like 4 (DLL4), which are important signals for vascular development (37). In most of the healthy population, resting ECs showed long half-lives through VEGF activation, Notch signaling, and angiopoietin-1 (ANG-1) and fibroblast growth factors (FGFs) expression (33). Recent knowledge concerning the complexity of angiogenesis indubitably shows the role of the participants in this event and allows for finding applications in anti-angiogenic therapy.
As previously mentioned, angiogenesis is a normal development and part of the healing process; however, it is key to tumor branching and arborization under pathological conditions such as cancer. The formation of new vascular networks promotes the growth, maintenance, and spread of cancer (38). During angiogenesis in cancer, alterations have been described at the level of lymphangiogenesis and vasculogenesis, both processes are highly involved in the propagation of cancer cells and an unfavorable prognosis (39).
The accelerated growth of the tumor leads to hypoxic tumor microenvironment, interstitial hypertension, and acidosis. To reverse these adverse physicochemical changes, VEGF-C and VEGF-D are synthesized by the activation of VEGFR-3/2, triggering a rise in diameter and density of the peritumoral lymphatic vessels, favoring the propagation of tumor cells toward sentinel lymph nodes (40,41). It has been shown that the inhibition of these factors by the use of antibodies decreases lymphogenesis and metastasis in nearby ganglia (42)(43)(44). Then, angiogenesis maintains a constant and permanent supply of nutrients for cancer cells that leads to tumor growth. This aberrant revascularization begins after the loss of regulation of inhibitory factors (e.g., thrombospondin-1) and angiogenic promoters (VEGF) (45,46). Hypoxia-inducible factor (HIF) is one of the first growth factors to initiate the abnormal process of vascular growth and responds to the low oxygen tension in the tumor mass. Subsequently, a wave of growth factors such as EGF, basic and acidic FGF, estrogen, prostaglandin E1 and E2, IL-8, TGF, TNF, neuropilins, and VEGF promotes the formation of a vascular network that ensures the exchange of oxygen and nutrients with the tumor (5,31,47,48). This vascularization process is regulated primarily by VEGF-A/VEGF-1,2 and DLL4 signaling. The activation of ECs also triggers a branching process toward the central region of the tumor (49,50). This new supply and drainage network that supports the tumor allows the latter to maintain a favorable microenvironment for its growth and dissemination. At present, the tumor niche is considered an independent organ able to maintain itself (51). Additionally, integrin receptors are overexpressed in tumor ECs and play a key role connecting the cell cytoskeleton to the extracellular matrix protein ligands such as arginine-glycine-aspartic acid (RGD). This binding interaction between integrin and protein ligands is an important mechanism during the angiogenesis of tumor endothelial cells (52).

MMPs in Cancer Angiogenesis
It is well known that MMPs have been implicated in angiogenesis regulation as well as in the anomalous relationship between cancer and the related processes of angiogenesis, vasculogenesis and lymphangiogenesis. MMPs also have a role in the immune system action in cancer development and progression (Tables 1, 2). The pro-and anti-angiogenic effects of MMPs participate in crucial steps as the ability to degrade ECM or cleave several substrates. Specifically, MMP-2 and MMP-9 give rise to the modulation of the dynamic remodeling of ECM (editing aggrecan, collagens, elastin, fibronectin, laminins, and glycosaminoglycans, and latent signaling proteins), activating and deactivating by proteolytic cleavages releasing biological activities that induce cellular regulation (108,109). MMP activation can be induced by several angiogenic factors, such as VEGF, basic fibroblast growth factors (bFGF), TGF-α andβ, and angiogenin. Specifically, MMP-1 activity promotes the expression of the vascular endothelial growth factor receptor 2 (VEGFR2) and EC proliferation, stimulating serine/threonineprotein kinase MARK2 (PAR-1) and activating the transcription factor NF-κB, suggesting the existence of a mechanism by which MMP-1 stimulates vascular remodeling and angiogenesis (110). Similarly, MMP-7 modulates the VEGF pathway in human umbilical vein endothelial cells (HUVECs), degrading soluble VEGFR-1 and in turn promoting angiogenesis (111). TNF-α, IL-8 and other factors with a known pro-angiogenic capacity, stimulate the production of MMP-2,−8, and−9 in ECs and regulate the angiogenesis process (63,112).
Angiogenesis studies using MMP-8 and MMP-2 knock-out mice, show an in vitro reduction of cell proliferation and neocapillary network growth, as well as a decrease in HUVEC migration and poor in vivo angiogenesis. Interestingly, ischemiainduced neovascularization is also affected by a reduction in ECs and invasive, proliferative, or mobilizing activities of endothelial progenitor cells (EPCs) derived from bone marrow (113,114). Additionally, it is known that ECs secrete MMP-2 and−9-containing vesicles stimulated by VEGF and FGF-2 and thus regulate the proteolytic activity critical for the angiogenesis-related invasive and morphogenic processes (115). Furthermore, MMP-9 also generates the angiogenic and tumoral repressor, tumstatin by proteolysis of the non collagenous domain (NC1) from the collagen alpha-3(IV) chain. The antiangiogenic properties of tumstatin inhibit EC proliferation and induce apoptosis by interacting with alphaVbeta3 integrin (116).
Among the most studied MMPs participating in angiogenesis is the MMP-14 (MT1-MMP). It significantly contributes to angiogenesis regulation by cleaving ECM molecules as a matrixdegrading enzyme (Figure 3). This MMP also acts as a key effector in the production of pro-angiogenic factors such as VEGF. In addition, MT1-MMP interacts with cell surface molecules, such as CD44 and sphingosine 1-phosphate receptor 1 (S1P 1 ), to induce EC migration, and plays a critical role in the proteolytic degradation of anti-angiogenic factors as decorin. Furthermore, evidence shows that MT1-MMP is able to degrade pro-TGF-β and endoglin (TGF-β receptor), suggesting a pivotal role in vessel maturation and angiogenesis, respectively (117) (Figure 3A). In addition, MT1-MMP appears to be an essential molecule that determines ECM adhesion and human endothelial cell tube formation through the modulation of MMP-2 expression ( Figure 3B). This suggests an important role in regulating angiogenesis-related functions in human ECs (118).

Soluble MMPs in Cancer Angiogenesis
Soluble MMP expression and its effects on cancer stabilization/proliferation are intimately linked via vascular angiogenesis mechanisms that are now well recognized. In this regard, MMP-1 expression has been reported to contribute to the progression of Head and neck squamous cell carcinomas (HNSCC) and the suggest metastatic phenotype of human breast and colorectal cancers, among others (119)(120)(121). Interestingly, MMP-1/protease-activated receptor-1 (PAR1) signaling axis has been implicated in tumor angiogenesis and intravasation of carcinoma cells by inducing vascular permeability (122), as well as, hypoxia-regulated MMP-1 expression in metastatic bladder cancer cells, which could be associated to a reactive oxygen species (ROS)-related regulation of the spheroid metastatic phenotype and cell spread (123). The increased expression of MMP-1 in human chondrosarcoma is an important prognostic factor and its function in the spread of tumor cells has been evaluated by silencing assays in which cancer metastasis is impaired but local tumor growth and angiogenesis are enhanced (124). These findings strongly support a role for MMP-1 in the diverse proliferative outcomes of human cancer through angiogenic processes.
Many studies have been published describing the relationship between MMP-2 expression and tumor angiogenesis. One of the earliest reports indicates that IL-8, an angiogenic factor, induces MMP-2 expression and activity in melanoma cells, enhancing their invasion (125). A relationship between MMP-2 expression and stromal support, angiogenesis, invasiveness, and tumor Frontiers in Oncology | www.frontiersin.org      TNFα AMB cells stimulation with TNF-α increased IL-6 and MMP-9 mRNA expressions, via NF-kB activation. Furthermore, TGF-β and IFN-c increased TNF-α-mediated expressions of MMP-9 and IL-6 mRNA, while those responses were suppressed by NF-kB inhibitor Ameloblastoma cells (AMB) cultures from patients were inmortalized using hTERT vector MMP-9 (97) growth was demonstrated using an MMP-2-specific inhibitor in a mouse model of bladder cancer (126). Furthermore, an elevated expression of MMP-2 was correlated with VEGF expression in gastric cancer (127) which suggests that this MMP plays a critical role in the progression of cancer through ECM degradation, tumor neovascularization and metastasis. On the other hand, MMP-9 promotes endothelial cell migration and triggers the angiogenic switch by releasing VEGF during carcinogenesis (128). Decreased expression of VEGF and MMP-9 in medulloblastoma cells that overexpress osteonectin, also referred to as Secreted Protein Acidic and Rich in Cysteine (SPARC), leads to decreased angiogenesis and tumor growth, indicating the pro-angiogenic role of MMP-9 in cancer tissues (129). In contrast, the direct proteolytic cleavage of osteopontin (OPN) by MMP-9 contributes to cancer metastasis, most likely associated with angiogenesis via the regulation of VEGF and angiostatin secretion (130,131). This model suggests that cancer growth is accompanied by increased vascular permeability, due in part to the expression of MMP-9, leading to the regulation of angiogenic factors, and eventually, neovascularization in cancer tissue.
Studies have revealed that, both MMP-2 and MMP-9 can degrade type IV collagen and are frequently elevated in human cancer. Additionally, a cooperative effect of MMP-2 and MMP-9 was demonstrated in an in vivo experimental model establishing the angiogenic phenotype and invasiveness of tumor keratinocytes (132). The mechanism whereby MMP-2 and MMP-9 activity induces cancer angiogenesis involves the cleavage of latent TGF-β in a CD44-dependent manner, which can promote tumor growth and invasion (133). Together, these results confirm   the contribution of MMP-2 and MMP-9 to cancer angiogenesis through the degradation of ECM components and the activation of pro-angiogenic factors VEGF and TGF-β in diverse cancer tissues (134). The above findings may explain the central role of the metalloproteinases MMP-2 and MMP-9 in tumor angiogenesis through the induction of pro-angiogenic factors. Many other MMPs have also been implicated in the incipient establishment of cancer angiogenesis (Tables 1, 2). For example, MMP-3 and MMP-7 interact in vivo with osteopontin at tumor sites and may be related to the angiogenic process during tumor development (135). The interaction of diverse MMPs with another class of proteases also contributes to tumor angiogenesis. For example, the overexpression of the serine protease matriptase in human carcinoma cells regulates MMP-3 activity, promoting proliferation and angiogenesis of tumor tissues by degradation of surrounding ECM (136). MMP-13 has also been implicated in cancer angiogenesis promotion through tube formation and neocapillary network development mediated by stimulation of ERK-FAK signaling pathway stimulation. It also stimulates VEGF-A secretion, which contributes to the angiogenic process (137).
It has been widely accepted that MMPs likely play antagonistic roles in regulating cancer angiogenesis. MMP-7 and MMP-9 may be involved in the blockage of cancer angiogenesis by cleaving plasminogen and generating angiostatin molecules (138). Additionally, cross-talk between MMP-7 and MMP-9 leads to the cleavage of insulin-like growth factor-binding protein 2 (IGFBP-2), an angiogenic activator in major aggressive cancers via the transcriptional regulation of the VEGF gene, showing adverse effects in cancer angiogenesis in some tissues (139,140). It has also been described that MMP-19 is essential to the development of nasopharyngeal carcinoma due to its tumor suppressive and anti-angiogenic functions which can reduce secreted MMP-2 and VEGF (141).

Membrane-Type Metalloproteinases (MT-MMPs) in Cancer Angiogenesis
Membrane type 1 matrix metalloproteinase (MT1-MMP) is considered a key mediator of cancer progression and metastasis. The overexpression of MT1-MMP in malignant breast cells significantly enhances VEGF production via the Akt and mTOR signaling pathways activated by the MT1-MMP-VEGFR-2-Src complex, which promotes tumor growth and angiogenesis (142,143). Apparently, a similar mechanism could be involved in glioblastoma angiogenesis (144,145). Therefore, it is worth noting that this tumor phenotype appears to be associated with the dependence of Akt-mediated signaling pathway, which is stimulated by several angiogenic factors.
On the other hand, the proteolytic cleavage of semaphorin 4D into its soluble form by MT1-MMP provides a novel molecular mechanism to control tumor-induced angiogenesis in HNSCC (146). In addition, cross-talk between MT1-MMP, MMP-2, and laminin-5γ2 chain fragments contributes to the vasculogenic mimicry of melanoma cells (147). In contrast, the colorectal cancer cells that shed MT1-MMP-mediated endoglin fragments exhibit an anti-angiogenic effect (148). Other studies have demonstrated that MT1-MMP and membrane type 2 matrix metalloproteinase (MT2-MMP) work cooperatively as pro-invasive factors that directly lead to Snail1-triggered cell participation in cancer angiogenesis and metastasis (149). In addition, MT2-MMP is a potential EMT mediator in carcinomas that can degrade adherents and tight junction proteins (150). It has been reported that both MT1-MMP and membrane type 3 matrix metalloproteinase (MT3-MMP) modulate pro-MMP-2 activation, whose angiogenic role in cancer was mentioned above, through inhibition by TIMP-2 and TIMP-3 (151). These data show that a cooperative effect of MT-MMPs during cancer angiogenesis is required together with the angiogenic factors.
Moreover, MT4-MMP expression correlates with EGFR activation, which triggers an angiogenic switch through its catalytic activity and induces the dissemination of cancer cells by disturbing the vessel integrity of the primary breast tumor and promoting hematogenous but not lymphatic metastasis (152)(153)(154). Finally, it has been shown that a high MT6-MMP expression in cancer cells is associated with tumor growth; however, further experiments are necessary to determine the exact role of this MT-MMP in the angiogenic process (155).

MMPs and the Immune System in Cancer
It is known that uncontrolled angiogenesis, anomalous ECM turnover, decreased growth, and cell migration, as well as inflammatory response, are the result of an imbalance between MMPs activity and their inhibitors, which may be associated with different diseases. Several specific signals are responsible for coordinating the formation, growth, remodeling, and stabilization of blood vessels. It is recognized that excessive growth-promoting signal cues lead to pathological angiogenesis and cancer (15,156).
In the tumoral microenvironment, there is a complex and dynamically interacting areas involving stromal cells (fibroblasts, myofibroblasts, neuroendocrine cells and immune cells), blood vessels, lymphatic network, and ECM (157,158), resulting in a tremendous heterogeneity observed in cancer cells. This condition is because tumor cells express and modulate a broad group of signaling pathways including immune modulatory pathways of cytokines and chemokines, which participate in the progression and establishment of cancer cells (Tables 1, 2) (159)(160)(161)(162)(163)(164)(165)(166)(167)(168)(169). In this regard, both cytokines and chemokines induce the expression and activity of MMPs, which in turn allow for the activation of pro-inflammatory signaling pathways as well as the activating receptors, for example, expressed on the surface of T cells and NK cells. Of these molecules act as a powerful mechanism to regulate the immune response. We have analyzed and highlighted several studies showing the involvement of MMPs and their interactions with immune system proteins in angiogenesis and cancer processes as shown in Tables 1, 2.
The presence of secreted extra-cellular vesicles (exosomes) has recently gained importance within the tumor microenvironment (170)(171)(172). Exosomes are specific bearers of multiple modulating molecules, such as the antigens for cluster of differentiation (CD), cytokines and chemokines, growth factors (EGF, FGF, PIGS), adhesion proteins (L1/CD171), nucleotides (non-coding RNA, miRNA), and metalloproteinases. Exosomes activate and modify the activity of diverse proteins such as immune proteins and receptor ligands into the circulation by proteolytic cleavage, playing a role as effectors and regulators to promote crosstalk between cancer and stromal cells (53,157,(173)(174)(175)(176). In addition, in vitro studies revealed that a type of TGF-β mediated exosome derived from lung cancer cells increase the expression of MMP-2 (104), while immunosuppressive exosome secretion from lymphoblastoid cells induces apoptosis in CD4 + T cells (177). Therefore, exosomes could function as conversion markers of malignant cells.
Since they are diverse, MMPs influence multiple cellular processes, such as the inflammatory process regulating barrier function and the activity of inflammatory cytokines and chemokines. Chemoattractant proteins such as MCP-1, MCP-2, MCP-3, and MCP-4 are targets for MMP activity, as result the modified MCPs changing their activity from agonist to antagonist and causing inflammation. Inflammation produces immune tolerance and leads to specific micro-environment conditions, exploited by tumors to evade immune cells and enhance progression, angiogenesis and metastasis (78,171). In the inflammation process, FGF2 expression can facilitate induction of FGF-dependent angiogenesis by mononuclear phagocytes, Tlymphocytes, and mast cells. The mechanism mediated through FGF2 release induces pro-inflammatory molecules, such as IFNα, IL-2, IL1-β, and nitric oxide. During the first phases of the EC angiogenesis process, FGFs (1, 2, and 4) upregulate urokinasetype plasminogen activator (uPA) in vitro and transform plasmin into plasminogen, an activator of MMPs, triggering ECM degradation and the secretion of exosomes containing MMP-2, MMP-9, TIMP-1, and TIMP-2 (156,178).
Several studies, both clinical and experimental, have shown that elevated MMP (including MT1-MMPs) levels are associated with the modulation of tumor progression. In brain tumors, growth factors and cytokines modulate the activity of several MMPs. Additionally, it has been observed that MMP-2positive tumor cells in patients are correlated with low mean survival (54,179,180).
Furthermore, in the context of immune cells, there are tumor-associated cells that contribute to the synthesis and upregulation of MMPs (70,(181)(182)(183). Importantly, tumorassociated macrophages (TAMs) secrete membrane-bound or soluble proteases, such as MMP-2, MMP-9, and MMP-12, which are involved in ECM degradation and promote the infiltration of tumor-associated blood vessels (184). Macrophages are known to promote cancer initiation and tumor development in an inflammatory environment (185). Bone marrow-derived myeloid cells are also involved in the process, through active regulation of blood vessel formation and maintenance in tumors (186). Furthermore, accumulated evidence shows that primary tumors can recruit immune cells, such as MMP-9 positive neutrophils, B cells, and M2 polarized macrophages to produce tumor-associated immune cells, which are known to contribute to neovascularization by supplying MMP-9 and other MMPs (59-61, 90, 183, 187). Although most of the published studies consider that M2 macrophages produce high amounts of IL-10, IL-1β, VEGF and MMP, additional subsets have been described with different proportions of cytokines related to cancer microenvironment. A subset of high M1 and low M2 infiltration macrophages are associated with improved patient survival in non-small-cell lung cancer (188), while the activation of M2 macrophages is correlated with a negative prognosis in cancer progression (189). It is therefore important to reorient the associated functions of the M2 macrophage subset to stop and kill cancer cells.
Finally, the molecular role of MMPs in the immune system and cancer is to modulate a series of latent signaling proteins located in ECM, including cytokines and growth factors such as quiescent TGF-β forming a complex with TGF-β-binding protein-1 in ECM. Thus, TGF-β modulates MMP expression, resulting in a bidirectional regulatory loop enhancing TGF-β signaling and promoting cancer progression (133,(190)(191)(192)(193)(194). Another mechanism observed is MMP-9 activity, which truncates IL-8 (1-19, 21, 22, 26-52, 63, 108-135) into more active chains, altering the function of the receptor and improving its biological activity, resulting in greater chemotaxis for neutrophils than the intact form of theof cytokine (195,196).
Accordingly, immunomodulatory mechanisms of MMPs, cytokines, receptors, and growth drivers are involved in the development and progression of several types of cancer.

THERAPEUTIC PERSPECTIVE OF MMPS
MMPs and their inhibitors TIMP, control a wide variety of physiological processes. They constitute promising pharmaceutical targets for inhibition and other metastatic processes.
Currently, monoclonal antibodies are possible candidates to inhibit the activity of MMPs (MMP−14,−12,−9, and−2). However, studies have only managed to identify antibodies against MMP-9 activity, which has biological functions and not for the MMP−14,−12, and−2 (197). In prostate cancer, MMP-9 may amplify local angiogenesis by cleaving membranebound VEGF. Therefore, VEGF is a candidate to be blocked and controlled and to prevent the activation of the androgen receptor (AR)/phosphatidylinositol 4-phosphate 5-kinase type-1 alpha (PIP5K1α)/AKT/MMP-9/VEGF signaling axis required for cell survival and invasion of metastatic tumors (198). Similarly, the effect of phytochemicals on MMP-2, MMP-9, and their tissue inhibitors (TIMPs) has been tested in breast cancer, with no alterations observed in vitro (199). TIMP-3 has been another important target of study regarding the inhibition of cancer cell migration, invasion, and metastasis in vitro and in vivo by natural products (200).
Recently, the effect of MMP-2 gene silencing in normal and MCF-7 cells exposed to the irradiation has been studied. It is known that this MMP leads to the degradation of basement membranes; however, the differential response to DNA damage silencing the MMP-2 gene in normal and MCF-7 cells may be attributable to ROS generation (201). In addition, thrombospondin-2 (THBS2) is a target gene of microRNA-93-5p (miR-93-5p) and THBS2 is closely associated with ECM and MMP-2 and 9. This MicroRNA is involved in the progression of malignant tumors and is highly expressed in cervical cancer tissues and cells. Thus, the THBS2/MMP signaling pathway is relevant for more studies in clinical trials (202). Moreover, the combined therapy for glioma treatment using temozolomidemarimast (a specific alkylating agent and an MMP inhibitor, respectively) results in tumor cell progression and invasiveness. An alternative treatment proposed in an in vitro study uses a combination of temozolomide and compounds 1 and 2 of N-Oisopropyl sulfonamido-based hydroxamates (MMP-2 inhibitors) to inhibit cancer cell invasiveness and viability (203).
All these studies represent advances in cancer drug development and cancer therapy, with a focus on the control of MMPs and the proteins with which they form complex networks of multifunctional interactions to modulate the signaling pathways that deviate during the development of metastatic cancer. Importantly, the emerging combined clinical therapy mitigates the side effects of existing treatments and raises the anticancer efficacy of chemotherapeutic drugs.

CONCLUSION REMARKS
In this work, we have highlighted the role that MMPs play in the cancer and its interaction with growth factors, inhibitor proteins, and the EMT process. The activity of MMPs is involved in the degradation, remodeling, and exchange of ECM, which, under normal physiological conditions, contributes to homeostasis as part of an extensive network of extracellular tissue modulation. In cancer, homeostasis is modified, leading to localized abnormal physiological conditions that modify this extensive network of extracellular tissue modulation.
The increase in MMP activities, as an abnormal process, is a way of producing/inducing an erroneous metabolic cascade. Erroneous metabolic cascades are signals that trigger the emergence of complex abnormal cell pathways, which give rise to tumor/cancerous phenotype cells. In this regard, the transformation into tumor/cancerous phenotypes suggests an exacerbated adaptive survival process. MMPs are not the only elements of this extensive network of extracellular tissue modulation; others such as TIMP proteins, which modulate MMPs, ADAMs and ADAMTSs (15), play a role in this anomalous process as numerous regulatory branches of the network do, namely interleukins.
Although the information on the role of MMPs in cancer is very broad and the way these proteins are expressed is well known, we observed a significant lack of data at the fine level of the relationship between MMPs and the continuity of both the normal and altered signals that positively modulate the carcinogenic process. MMPs produce modulatory elements that remain unclear and, considering that the ECM is a complex array of proteins, fibers, and carbohydrates in different tissues, there may be several variants that generate the loss of homeostasis, causing the diverse cancerous processes observed.
In the angiogenesis process, MMPs are well-known key factors involved in ECM degradation that induce angiogenesis initiation in both physiological and pathological processes. However, the experimental evidence thus far demonstrates that MMPs also play a decisive role in the activation of pro-angiogenic and, in some cases, anti-angiogenic factors in cancer tissues. Thus, MMPs can be considered angiomodulators, which could control new vessel formation necessary for cancer growth, progression, and spread. Therefore, we speculate that MMPs participate in cancer angiogenesis in a cell context-dependent manner.
Most of the experimental data regarding MMP participation in cancer development, vascular endothelium processes, other epithelia (such as periodontal), and inflammatory processes allow us to assume that MMPs are proteins that carry out a type of external cellular regulation/signaling on the ECM. These proteins possess a different regulatory action mechanism that is complementary to other mechanisms such as ligand-receptor signaling pathways. The MMP mechanism is based on editing macromolecules by proteolysis, mainly anchored to the ECM.
It is evident that the proportion of MMPs and other macromolecules (cytokines, grown factors, fibers such as integrins, polysaccharides, and others) in the ECMs of different tissues in normal conditions are metabolically, microenvironmentally, and epigenetically balanced for their functions in each type of tissue. There are tissue-specific proportions, although ECMs have a high degree of heterogeneity.
Inspecting our concept of MMP participation in ECM in the literature, we found an excellent review of ECM with similar concepts (204). The ECM is a highly dynamic system in constant remodeling and is undoubtedly an extension of communication/modulation/signaling among cells located outside the plasma membrane, where MMPs participate in protein editing by providing post-translational modifications.
On the other hand, EMT is a biological process aimed producing mesenchymal phenotype cells from epithelial cells. Its inverse process, mesenchymal-epithelial transition (MET), is carried out with the participation of ECM elements. EMT and MET lead to normal tissue regeneration and fibrosis [EMT type 2, according to Kalluri (205)]. Regarding the participation of MMPs in EMT type 3 (abnormal type), it is evident that their participation is associated with errors in the communication, modulation, and signaling of this process, which is induced and directed by tumor cells.
Evidence suggests that tumor cells induce the uncontrolled upregulation of MMPs, producing a large number of stimulating factors that disrupt EMT and immunological processes that prevent tumor cell elimination and migration. The upregulation progresses to generating anomalous tissue-specific type signals.
It is known that tumor cells have extensive heterogeneity in their metabolism and phenotype relative to normal tissue across cancer types. Furthermore, these abnormal signals coming from the tumor cells are tissue-specific, leading to the adaptation to the microenvironment where they developed. Finally, an interesting observation of the MMP family is the large, robust specificity profile, which suggests that its role is controlled in a tissuespecific manner; that is, MMP types are expressed accordingly to the regulatory proteins needed for the tissue.
However, more research efforts are needed to determine when abnormal signals begin, what the determinants are, and how microenvironmental tissue-specific conditions can lead a cell to change its metabolism and phenotype. In addition, the question remains: When does the high expression problem of MMPs become a problem metabolically? Although MMPs do not seem to be the cause of the appearance of tumor cells, they induce tumor development because they are targets to regulate development, contributing to increased invasiveness and growth of metastatic tumors.