Proteases & Inhibitors

Serine proteases (plasminogen activator systems and their inhibitor, the Plasminogen Activator Inhibitor-1, PAI-1), soluble matrix metalloproteinases (MMPs), membrane-type MMPs (MT-MMPs), A Disintegrin and Metalloproteases (ADAMs), A Disintegrin and Metalloproteases with Thrombospondin motif (ADAMTS) are major effectors in tissue remodelings.

The plasminogen-plasmin system

Plasminogen (Plg) is converted into its active form, plasmin (Pln) by two plasminogen activator (PA), urokinase-type plasminogen activator (uPA) or tissue associated PA (t-PA). uPA itself is secreted as an inactive precursor form (pro-uPA) that binds with high affinity to a specific cell surface glycosylphosphatidylinositol-anchored receptor designated uPAR. The binding of pro-uPA to uPAR activates uPA and enhances the generation of plasmin at the cell surface, promoting matrix degradation, the activation of matrix metalloproteinases (MMP) and growth factors, and the release of matrix- or cell surface-anchored growth factor. PAI-1 is the physiological inhibitor of PA and therefore controls the activation of Plg into Pln.

In addition to interacting with uPA, PAI-1 binds to vitronectin leading to its stabilization. In addition, binding of PAI-1 to vitronectin interferes with the binding of integrins of the alphav family to the RGD domain of vitronectin and thereby regulate cell migration.

Paradoxical function of PAI-1:

The 2 inhibitory functions of PAI-1 on proteolysis and on cell attachment initially led to the prediction that PAI-1 would have an anti-angiogenic function. It was therefore surprising to find that high, rather than low, levels of PAI-1 were predictive of poor outcome in patients suffering from several types of cancers. Consistent with a positive effect of PAI-1 on cancer progression, a defect of tumor and choroidal vascularization in mice implanted with malignant keratinocytes was observed in the absence of host-derived PAI-1

The effect of PAI-1 on in vivo and in vitro angiogenesis is dose dependent, with a stimulatory role at low physiological concentrations and an inhibitory role at pharmacological concentrations.

Matrix Metalloproteinases (MMPs)

Zn-dependent endopeptidases degrade extracellular matrix components and contribute to the fine tuning of diverse biological processes through limited proteolysis of specific targets including growth factors, chemokines/cytokines and cell surface receptors. They are produced by tumor cells themselves and/or by several host cells. The evidence of a key implication of proteolytic events in cancer progression is based on

- the use of physiological and synthetic inhibitors in various in vivo cancer models,

- profiling of protease production and expression in human cancer samples,

- overexpression of MMPs in cancer cells,

- genetic approaches using MMP-deficient mice.

MMP-2 and MMP-9 appear as key regulators of tumor and choroidal angiogenesis. Their combined deficiency results in an abrogation of tumor vascularization. In sharp contrast, host MMP-19 deficiency was associated with an increased early angiogenic response and increased tumor invasion. Therefore, in contrast to most MMPs which promote tumor progression, MMP-19 is a negative regulator of early steps of tumor angiogenesis and invasion.

In conclusion, proteases can now be viewed as sophisticated modulators, positive or negative regulators of cancer development and metastatic dissemination.

These data provide at least partly, an explanation for the disappointing results of MMP inhibitors in clinical trials. At the time that these initial clinical trials were designed, only few MMPs were studied, and the MMPs related families (ADAM and ADAMTS) were almost unknown. The synthetic broad spectrum MMP inhibitors used were targeting a set of proteases, both positive and negative regulators of cancer progression. Although the contribution of various MMPs in cancer growth and dissemination is now well accepted, the role of related ADAM and ADAMTS in the different steps of cancer progression is poorly known. In particular the function of a recently identified ADAMTS-12 is completely unknown.

Membrane type MMP

Although most MMPs are secreted as soluble enzymes, 6 of them are membrane-type MMPs (MT-MMPs) which are associated with the cell membrane by either a transmembrane domain (MT1-, MT2-, MT3-, MT5-MMPs) or a glycosylphosphatidyl-inositol (GPI) anchor (MT4-, MT6-MMPs).

MT1-MMP

MT1-MMP, the first discovered member of this subfamily, was initially described as an activator of pro-MMP2. MT1-MMP plays critical roles in cell migration, cell invasion, physiological and pathological matrix remodeling. It is a multifunctional protease for which an increasing number of substrates have been identified, including pro-MMP2, pro-MMP13, extracellular matrix components, cell surface receptors, growth factors, chemokines, adhesion and signaling molecules. Its implication in tumor growth, invasion and angiogenesis is now well documented. A pro-angiogenic effect of MT1-MMP has been linked, at least in part, to an up-regulation of vascular endothelial growth factor (VEGF)

MT4-MMP

Although much emphasis has been placed on understanding the biochemical features and functions of MT1-MMP (the first identified Membrane type MMPs), little attention has been given to the implication of other MT-MMPs in cancer progression. However, MT4-MMP over-expression in human breast cancer cells promotes primary tumor growth and lung metastasis. The implication of MT4-MMP in metastatic dissemination of human breast cancers is underlined by its strong expression in metastatic cells in human lymph nodes. MT4-MMP appears therefore as an unexpected interesting novel target to design anti-cancer agents.