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The ADAMTS (A Disintegrin and Metalloproteinase with Thrombospondin motifs) enzymes are secreted, multi-domain matrix-associated zinc metalloendopeptidases that have diverse roles in tissue morphogenesis and patho-physiological remodeling, in inflammation and in vascular biology. The human family includes 19 members that can be sub-grouped on the basis of their known substrates, namely the aggrecanases or proteoglycanases (ADAMTS1, 4, 5, 8, 9, 15 and 20), the procollagen N-propeptidases (ADAMTS2, 3 and 14), the cartilage oligomeric matrix protein-cleaving enzymes (ADAMTS7 and 12), the von-Willebrand Factor proteinase (ADAMTS13) and a group of orphan enzymes (ADAMTS6, 10, 16, 17, 18 and 19). Control of the structure and function of the extracellular matrix (ECM) is a central theme of the biology of the ADAMTS, as exemplified by the actions of the procollagen-N-propeptidases in collagen fibril assembly and of the aggrecanases in the cleavage or modification of ECM proteoglycans. Defects in certain family members give rise to inherited genetic disorders, while the aberrant expression or function of others is associated with arthritis, cancer and cardiovascular disease. In particular, ADAMTS4 and 5 have emerged as therapeutic targets in arthritis. Multiple ADAMTSs from different sub-groupings exert either positive or negative effects on tumorigenesis and metastasis, with both metalloproteinase-dependent and -independent actions known to occur. The basic ADAMTS structure comprises a metalloproteinase catalytic domain and a carboxy-terminal ancillary domain, the latter determining substrate specificity and the localization of the protease and its interaction partners; ancillary domains probably also have independent biological functions. Focusing primarily on the aggrecanases and proteoglycanases, this review provides a perspective on the evolution of the ADAMTS family, their links with developmental and disease mechanisms, and key questions for the future.

•  Introduction •  MMP‐3 •  MMP‐8 •  MMP‐9 •  MMP‐12 •  MMP‐19 •  MMP‐26 •  Conclusions The original notion that matrix metalloproteinases (MMPs) act as tumour and metastasis‐promoting enzymes by clearing a path for tumour cells to invade and metastasize has been challenged in the last decade. It has become clear that MMPs are involved in numerous steps of tumour progression and metastasis, and hence are now considered to be multifaceted proteases. Moreover, more recent experimental evidence indicates that some members of the MMP family behave as tumour‐suppressor enzymes and should therefore be regarded as anti‐targets in cancer therapy. The complexity of the pro‐ and anti‐tumorigenic and ‐metastatic functions might partly explain why broad‐spectrum MMP inhibitors failed in phase III clinical trials. This review will provide a focussed overview of the published data on the tumour‐suppressive behaviour of MMPs.

Transforming growth factor-β (TGF-β) promotes extracellular matrix deposition by down-regulating the expression of matrix degrading proteinases and upregulating their inhibitors. Tissue inhibitor of metalloproteinases (TIMP)-3 is an ECM-associated specific inhibitor of matrix degrading metalloproteinases. Here, we have characterized the signaling pathways mediating TGF-β-induced expression of TIMP-3. Basal and TGF-β-induced TIMP-3 mRNA expression was abolished in Smad4-deficient mouse embryonic fibroblasts and restoring Smad4 expression rescued the response. Inhibition of Smad signaling by expression of Smad7 and dominant negative Smad3 completely abolished TGF-β-elicited expression of TIMP-3 in human fibroblasts, whereas overexpression of Smad3 enhanced it. Inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) activation with PD98059 and p38 mitogen-activated protein kinase activity by SB203580 resulted in suppression of TGF-β-induced TIMP-3 expression, indicating that ERK1/2 and p38 MAPK mediate the effect of TGF-β on TIMP-3 expression. Specific activation of p38α and ERK1/2 by constitutively active mutants of MKK3b or MEK1, respectively, and simultaneous co-expression of Smad3 resulted in induction of TIMP-3 expression in the absence of TGF-β indicating that Smad3 co-operates with p38 and ERK1/2 in the induction of TIMP-3 expression. These results demonstrate the complex interplay between Smad3, p38α, and ERK1/2 signaling in the regulation of TIMP-3 gene expression in fibroblasts, which may play a role in inflammation, tissue repair, and fibrosis.

Key Points Matrix metalloproteinases (MMPs) and ADAMs (a disintegrin and metalloproteinase) are part of a larger family of structurally related zinc-dependent metalloproteinases called metzincins. Structurally, MMPs are divided in three domains: an amino-terminal propeptide region, an amino-terminal catalytic domain, and a carboxy-terminal domain that is involved in substrate binding. ADAMs have a prodomain, a metalloprotease region, a disintegrin domain for adhesion, a cysteine-rich region, epidermal-growth-factor repeats, a transmembrane module and a cytoplasmic tail. The activity of MMPs is tightly regulated in several ways: at the level of transcription, by post-translational modifications such as proteolysis, and through the action of endogenous tissue inhibitors of metalloproteinases. The regulation of ADAMs is less well understood, although there is some evidence that the same three levels of regulation might control ADAM activity. MMPs and ADAMs have been implicated in neuroinflammation and multiple sclerosis (MS), in the pathogenesis of malignant gliomas, and in other neurological conditions such as stroke, viral infections and Alzheimer's disease. In the case of ADAMs, their role in these pathological states has begun to be explored, but the available literature is still in its infancy. Although the detrimental roles of metalloproteinases are well documented, some of their functions in the central nervous system (CNS) might be beneficial. For example, some metalloproteinases are expressed in the CNS during development, pointing to a possible role in brain maturation. Similarly, metalloproteinases have been implicated in myelinogenesis and axon growth. Furthermore, metalloproteinases are upregulated after injury to the CNS, indicating a possible relevance to tissue repair. Several challenges remain in the study of metalloproteinases and their role in brain function. It will be necessary to understand the balance between the beneficial and detrimental roles of MMPs to determine whether they can be used as targets for therapeutic intervention. It will also be important to identify the physiological substrates of the different metalloproteinases, and to develop selective antagonists against the various members of the metalloproteinase families; the lack of such tools constitutes one of the main limitations to the growth of the field at present. Matrix metalloproteinases (MMPs) have been implicated in several diseases of the nervous system. Here we review the evidence that supports this idea and discuss the possible mechanisms of MMP action. We then consider some of the beneficial functions of MMPs during neural development and speculate on their roles in repair after brain injury. We also introduce a family of proteins known as ADAMs (a disintegrin and metalloproteinase), as some of the properties previously ascribed to MMPs are possibly the result of ADAM activity.

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity.

Elevated rates of mating and reproduction cause decreased female survival and lifetime reproductive success across a wide range of taxa from flies to humans. These costs are fundamentally important to the evolution of life histories. Here we investigate the potential mechanistic basis of this classic life history component. We conducted 4 independent replicated experiments in which female Drosophila melanogaster were subjected to 'high' and 'low' mating regimes, resulting in highly significant differences in lifespan. We sampled females for transcriptomic analysis at day 10 of life, before the visible onset of ageing, and used Tiling expression arrays to detect differential gene expression in two body parts (abdomen versus head+thorax). The divergent mating regimes were associated with significant differential expression in a network of genes showing evidence for interactions with ecdysone receptor. Preliminary experimental manipulation of two genes in that network with roles in post-transcriptional modification (CG11486, eyegone) tended to enhance sensitivity to mating costs. However, the subtle nature of those effects suggests substantial functional redundancy or parallelism in this gene network, which could buffer females against excessive responses. There was also evidence for differential expression in genes involved in germline maintenance, cell proliferation and in gustation / odorant reception. Interestingly, we detected differential expression in three specific genes (EcR, keap1, lbk1) and one class of genes (gustation / odorant receptors) with previously reported roles in determining lifespan. Our results suggest that high and low mating regimes that lead to divergence in lifespan are associated with changes in the expression of genes such as reproductive hormones, that influence resource allocation to the germ line, and that may modify post-translational gene expression. This predicts that the correct signalling of nutrient levels to the reproductive system is important for maintaining organismal integrity.

A Disintegrin and Metalloproteinase-15 (ADAM15) is a transmembrane protein involved in protein ectodomain shedding, cell adhesion and signalling. We previously cloned and characterised alternatively spliced variants of ADAM15 that differ in their intracellular domains and demonstrated correlation of the expression of specific variants with breast cancer prognosis. In this study we have created isogenic cell panels (MDA-MB-231 and MCF-7) expressing five ADAM15 variants including wild-type and catalytically inactive forms. The expression of ADAM15 isoforms in MDA-MB-231 cells led to cell clustering to varying degree, without changes in EMT markers vimentin, slug and E-cadherin. Analysis of tight junction molecules revealed ADAM15 isoform specific, catalytic function dependent upregulation of Claudin-1. The expression of ADAM15A, and to a lesser degree of C and E isoforms led to an increase in Claudin-1 expression in MDA-MB-231 cells, while ADAM15B had no effect. In MCF-7 cells, ADAM15E was the principal variant inducing Claudin-1 expression. Sh-RNA mediated down-regulation of ADAM15 in ADAM15 over-expressing cells reduced Claudin-1 levels. Additionally, downregulation of endogenous ADAM15 expression in T47D cells by shRNA reduced endogenous Claudin-1 expression confirming a role for ADAM15 in regulating Claudin-1 expression. The PI3K/Akt/mTOR pathway was involved in regulating Claudin-1 expression downstream of ADAM15. Immunofluorescence analysis of MDA-MB-231 ADAM15A expressing cells showed Claudin-1 at cell-cell junctions, in the cytoplasm and nuclei. ADAM15 co-localised with Claudin-1 and ZO1 at cell-cell junctions. Immunoprecipitation analysis demonstrated complex formation between ADAM15 and ZO1/ZO2. These findings highlight the importance of ADAM15 Intra Cellular Domain-mediated interactions in regulating substrate selection and breast cancer cell phenotype.

Anti-angiogenic treatments against αvβ3-integrin fail to block tumour growth in the long term, which suggests that the tumour vasculature escapes from angiogenesis inhibition through αvβ3-integrin-independent mechanisms. Here, we show that suppression of β3-integrin in mice leads to the activation of a neuropilin-1 (NRP1)-dependent cell migration pathway in endothelial cells via a mechanism that depends on NRP1's mobilisation away from mature focal adhesions following VEGF-stimulation. The simultaneous genetic targeting of both molecules significantly impairs paxillin-1 activation and focal adhesion remodelling in endothelial cells, and therefore inhibits tumour angiogenesis and the growth of already established tumours. These findings provide a firm foundation for testing drugs against these molecules in combination to treat patients with advanced cancers.

Mycobacterium tuberculosis can cause lung tissue damage to spread, but the mechanisms driving this immunopathology are poorly understood. The breakdown of lung matrix involves MMPs, which have a unique ability to degrade fibrillar collagens at neutral pH. To determine whether MMPs play a role in the immunopathology of tuberculosis (TB), we profiled MMPs and their inhibitors, the tissue inhibitor of metalloproteinases (TIMPs), in sputum and bronchoalveolar lavage fluid from patients with TB and symptomatic controls. MMP-1 concentrations were significantly increased in both HIV-negative and HIV-positive patients with TB, while TIMP concentrations were lower in HIV-negative TB patients. In primary human monocytes, M. tuberculosis infection selectively upregulated MMP1 gene expression and secretion, and Ro32-3555, a specific MMP inhibitor, suppressed M. tuberculosis-driven MMP-1 activity. Since the mouse MMP-1 ortholog is not expressed in the lung and mice infected with M. tuberculosis do not develop tissue destruction equivalent to humans, we infected transgenic mice expressing human MMP-1 with M. tuberculosis to investigate whether MMP-1 caused lung immunopathology. In the MMP-1 transgenic mice, M. tuberculosis infection increased MMP-1 expression, resulting in alveolar destruction in lung granulomas and significantly greater collagen breakdown. In summary, MMP-1 may drive tissue destruction in TB and represents a therapeutic target to limit immunopathology.

Approximately 80% of patients diagnosed with acute myeloid leukemia (AML) die as a consequence of failure to eradicate the tumor from the bone marrow microenvironment. We have recently shown that stroma-derived interleukin-8 (IL-8) promotes AML growth and survival in the bone marrow in response to AML-derived macrophage migration inhibitory factor (MIF). In the present study we show that high constitutive expression of MIF in AML blasts in the bone marrow is hypoxia-driven and, through knockdown of MIF, HIF1α and HIF2α, establish that hypoxia supports AML tumor proliferation through HIF1α signaling. In vivo targeting of leukemic cell HIF1α inhibits AML proliferation in the tumor microenvironment through transcriptional regulation of MIF, but inhibition of HIF2α had no measurable effect on AML blast survival. Functionally, targeted inhibition of MIF in vivo improves survival in models of AML. Here we present a mechanism linking HIF1α to a pro-tumoral chemokine factor signaling pathway and in doing so, we establish a potential strategy to target AML.