**6. MMPs**

MMPs belong to the family of zinc-dependent endopeptidases collectively referred to as metzincins. The metzincins can be subdivided into four families: seralysins, astacins, ADAMs/ adamalysins, and MMPs [130, 136]. So far to date, 28 members of the MMP family have been identified [130, 135, 136] which are primarily responsible for most of the ECM degradation observedduringthe invasiveprocesses.MMPs areproducedbyskincells (fibroblasts,keratino‐ cytes, melanocytes) as well as macrophages, endothelial cells and mast cells [10, 25, 81, 137].

the Golgi compartment where the prodomain was removed by a furin-type proprotein convertase [77, 84, 85]. As increased amounts of mature TACE are detected in furin overexpressing cells, it appears that proTACE is a better substrate for furin than it is for PC7 [85]. A similar observation has been seen in cells overexpressing TACE [58, 83, 99] where furin was shown to be responsible for its cleavage [83, 99]. This finding was confirmed using cell permeable furin inhibitors CMK and PDX in Cos7 cells [83] and keratinocytes [98] where

Furin mRNA, protein and enzyme activity has been observed in human epidermal keratino‐ cytes [29, 98, 111, 128, 129]. Skiba *et al*. [29] found that UVA and UVB radiation immediately increased furin mRNA levels in HaCaT cells. UVB irradiation induced higher levels of furin mRNA expression [29]. The time course for furin mRNA levels in cells irradiated with low dose of UVA or high dose of UVB was similar to that for TNFα, whereas maximal mRNA induction of both genes were detected 8 h post-irradiation [29]. Although UV irradiation does appear to have an effect on furin gene expression, no direct relationship was apparent between TACE and furin mRNA induction. A recent study has shown that following exposure to UVA and UVB, furin levels in HaCaT cells fell with respect to time [49, 129]. However, it was unknown whether this was due to the loss of the pro or mature form of the enzyme. Through its effect on stimulating MMPs, as well as activating TACE and the resultant effect this has on TNFα released by the cell, furin activity has an influence on the inflammation seen in the skin

Furin/PC processing of substrates has been shown to also contribute to tumour progression, aggressiveness, metastasis, and angiogenesis [23, 24, 104-106]. Tumour invasion and metasta‐ sis represent a multistep process that depends on the activity of many proteins [46, 101, 104, 130]. Proteolytic degradation of the ECM components is a central event of this process. Several classes of proteases, including MMPs, serine proteases and cysteine proteases have been implicated in the tumour cell invasive process [104, 130, 131]. Of these, MMPs appear to be primarily responsible for much of the ECM degradation observed during invasive processes [111, 130, 132-134]. They can contribute to tumour growth not only by degradation of the ECM but by the release of sequestered growth factors or the generation of bioactive fragments VEGF, bFGF or TGFβ, the suppression of tumour cell apoptosis and the destruction of immunemodulating chemokine gradients [131, 132, 135]. Furin also cleaves a number of MMPs from

MMPs belong to the family of zinc-dependent endopeptidases collectively referred to as metzincins. The metzincins can be subdivided into four families: seralysins, astacins, ADAMs/ adamalysins, and MMPs [130, 136]. So far to date, 28 members of the MMP family have been identified [130, 135, 136] which are primarily responsible for most of the ECM degradation observedduringthe invasiveprocesses.MMPs areproducedbyskincells (fibroblasts,keratino‐ cytes, melanocytes) as well as macrophages, endothelial cells and mast cells [10, 25, 81, 137].

reduced levels of mature TACE were formed.

280 Highlights in Skin Cancer

following exposure to UV radiation as seen in Figure 2.

their proform, and activating them as a result [23, 86, 102, 105].

**6. MMPs**

**Figure 2.** The role furin plays in the maturation of TACE and MMPs in skin cells. Furin cleaves and activates TACE, which in turn can process TNFα from its proform. Keratinocytes secrete TNFα following exposure to UVB radiation, and this is enhanced if IL-1α is present. Furin also cleaves MMPs from their respective proforms, and the expression and activity of these proteases are elevated when the cells have been exposed to UVB radiation, and they are enhanced if either IL-1α (MMP-9) or TNFα (MMP-2) is present. The effect of UVB radiation on the expression of the enzymes and pTNFα in the cell is represented by dashed lines, if it is enhanced it is represented by (+), and if it is unknown (?)

MMPs are also implicated in cell migration, proliferation, and tissue remodelling and thereby may also play a role in growth and development, angiogenesis, and atherosclerosis [138, 139].

Structurally MMPs consist of a signal peptide followed by pro, catalytic, hemopexin and cytoplasmic domains, respectively [130]. MMPs cleave peptides and proteins, which have a myriad of functions that are independent of their proteolytic activity [140]. They have distinct but often overlapping substrate specificities, hence leading to the absence of distinct pheno‐ types in most genetically-engineered mice with knockdown of specific MMPs [140].

MMPs are generally expressed in very low amounts and their transcription is tightly regulated either positively or negatively by cytokines and growth factors such as IL-1, IL-4, IL-6, TGFβ, or TNFα [130, 135, 141, 142]. Some of these regulatory molecules can be proteolytically activated or inactivated by MMPs (via a feedback loop). MMPs are synthesized as latent proenzymes, which are converted into mature, catalytically active forms in the TGN by PCs [111, 139]. Activation of MMPs following secretion from the cell depends on disruption of the prodomain interaction with the catalytic site, which may occur either by conformational changes or proteolytic removal of the prodomain. With the exception of MMP-2, the mecha‐ nism for *in vivo* activation of secreted MMPs is not well understood [135].

In normal skin, MMPs are not constitutively expressed but can be induced temporarily in response to exogenous signals such as UVR [10, 25, 143]. Elevated levels of MMP activity in human skin, as a result of prolonged periods of sun exposure, confirm that it plays a major role in photoageing [10, 25, 28]. Onoue *et al*. [144] suggested that MMP-9 secreted from keratino‐ cytes afterUVBirradiationmightresultfromapoptotic events.UVradiationisknowntoelevate the expression of MMP-1, MMP-3 (stromelysin-1) and MMP-9 in human skin *in vivo* [25]. All three MMPs (1, 3 and 9) can degrade most of the proteins found in the extracellular matrix [25]. MMP-1,whichisproducedbybothdermalfibroblastsandepidermalkeratinocytes,cleavestype 1 collagen into specific fragments. These fragments can then be further hydrolysed by MMP-2 and MMP-9 [137, 145]. Steinbrenner *et al*. [137] found that UVA irradiation dose-dependently decreased the steady-state mRNA levels of MMP-2 and MMP-9 and lowered the gelatinolytic activityofbothenzymesincellculturesupernatants.Ofinterestisthat*invivo*,followingexposure to UV radiation only keratinocytes express large levels of MMP on the cell membrane [146], but when fibroblasts grown in culture are irradiated they express higher levels of MMP on their plasmamembrane [147].The reasons forthediscrepanciesbetweenthe responsesofhumanskin cells under *in vivo* and *in vitro* conditions are not known.

result of UV exposure or other factors, these skin cancer cells may become metastatic as a result

The Role of Furin in the Development of Skin Cancer

http://dx.doi.org/10.5772/55569

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In the progression of a mutated cell to that of a tumorigenic or metastatic cell, PCs have been shown to cleave a range of precursors of growth factors, their receptors, adhesion molecules, proteases and MMPs. Some of these molecules include cadherins, TGFβ, platelet derived growth factor as well as insulin-like growth factor 1 and its receptor [23, 101, 106, 117, 155]. Tumour progression and metastasis may enhanced by a number of factors such as (a) hypoxiainduced upregulation of furin activity within the solid tumour mass [105], (b) changes in cell adhesion through PC cleavage and activation of integrins and related adhesion molecules [156], (c) furin processing of vascular endothelial growth factor resulting in increased angio‐

Furin/PC expression and processing can increase the incidence and severity of the cancer phenotype [104, 111]. Aberrant furin expression has been observed in a number of tumours including those from the breast [159], ovary [160], liver [125], brain [161], skin [111, 132] and from other tissues [23, 128, 162, 163]. Bassi *et al*. [101] observed that PACE4 transgenic mice were more susceptible to epidermal carcinogenesis and tumour progression compared to controls. These transgenic keratinocytes had higher rates of processing of MT1-MMP and MT2-

MMP-2 (gelatinase A) and MMP-9 (gelatinase B) have been frequently associated with the invasive and metastatic potential of tumour cells [10, 104, 111, 130, 132, 133, 137]. The expres‐ sion of MMP-2, is regulated independently of MMP-9 [144]. The close correlation observed between MMP-2 activation and metastatic progression in various tumours suggests that it may act as a "master switch" triggering tumour spread [114]. The expression of MMP is low in keratinocytes but elevated in BCC and SCC [49, 164, 165]. In SCC, MMP proteins (1, 2, 3 and MT1-MMP) are expressed both in tumorous and stromal cells [165], while MMP proteins (1, 2, 3 and 9) are observed in BCCs and melanomas [165, 166]. This expression correlates with the progression and the metastatic potentials of these tumours [106, 135, 165, 167]. UVR can participate to the development of skin cancer by the activation of MMPs. Two molecular mechanisms contribute to the UV-induced MMPs expression. First, the activation of cellsurface receptors with subsequent activation of mitogen activated protein kinase (MAPK) cascade that in turn contributes to the transcriptional up-regulation of MMPs [168]. Second, through the expression of pro-inflammatory cytokines, which induce the expression of MMPs [46, 133, 142, 143]. The role of UV in the induction of MMPs is supported by two experimental findings. First, UV-irradiation of SCC cell lines results in an increased secretion of MMPs [49, 132]. Second, UV-induced phosphorylation of extracellular signal-regulated kinase (ERK) and stress kinases precedes the rapid stimulation of MMPs in SCC cells [10]. If there are cells in the skin which become cancerous as a result of DNA damage, some may go onto become metastatic due to increased MMP activity [111]. This increase in levels of activated MMPs on the surface of the cell could be due to either increased expression of proMMP protein and/or increased furin activity. The role furin plays in the development of skin cancer suggests that it could be significant, and as such the development of specific inhibitors may offer a new therapy to treat

genic activity [157], and (d) furin enhanced expression of MMPs [84, 158].

of epithelial to mesenchymal transition (EMT) [154].

MMP resulting in increased collagen degradation.

such tumours.

TNFα has been shown to induce proMMP-2 in human dermal fibroblasts [8], while IL-1α induced proMMP-9 levels in fibroblasts and keratinocytes [148]. In mesenchymal cells TNFα was shown to stimulate MMP-2 activity by activating a proteolytic cascade involving furin and MT1-MMP [139]. It is not known if TNFα activates MMP activity in epidermal cells via a similar mechanism. MMP-9 [144], has been shown to play an important role in the patho‐ physiologies of many skin conditions such as wound healing [145], and angiogenesis [87]. Activation of pro-MMP2 takes place at the cell surface and involves interactions with active MT1-MMP, which is itself activated through rapid trafficking to the cell surface and proteolytic processing [139]. Maquoi *et al*. [114] demonstrated that furin-inhibitor reduces the level of mature MT1-MMP, which is paralleled by a decrease in pro-MMP-2 activation as well as in cell invasiveness, confirming the furin plays a role in this process. Direct cleavage of proMMP-2 by furin in the TGN has shown to inactivate this matrix metalloprotease [149]. Therefore changes to the level of MMP-2 activity on the surface of the cell can be directly or indirectly regulated by furin either through cleavage in the TGN to reduce activity or indirectly via MT1- MMP, which increases activity. The mechanism by which this is regulated is not clear.
