Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.24.3 (collagenase)
 18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The authors, including an academic-inventor, the director of the University's tech-transfer office, and the CEO of a "start-up" pharmaceutical company based on the professor's (and his colleague's) inventions, relate the history and current status of their interactions. To start, the basic research leading to the "eureka" experiment (such things really do happen) is summarized: namely, the discovery of (i) the surprising ability of the tetracycline antibiotics to inhibit mammalian tissue-destructive proteinases (collagenase, gelatinase) during a variety of disease processes, e.g., periodontitis, the arthritides, osteopenia/osteoporosis, sterile corneal ulcers, tumor invasion/metastasis/angiogenesis, and (ii) a series of chemically-modified, non-antibacterial analogs of tetracyclines to inhibit these enzymes without producing typical antibiotic side effects. The role of the University in obtaining the services of patent attorneys, and its assistance in developing the strategy to deal with an industrial partner, is addressed. Above all, the authors stress the need for close cooperation and collegial interactions between all three groups in this high-risk (but potentially high-benefit-to-the-public) enterprise.
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PMID:New therapeutic uses for an old family of drugs: travels of a dental researcher from the lab to the university's office of technology transfer and beyond. 893 Dec 41

A seminal experiment involving a germ-free rat model of connective tissue breakdown (followed soon thereafter by a series of in vitro studies) identified an unexpected non-antimicrobial property of tetracyclines (TCs). This ability of TCs to inhibit matrix metalloproteinases (MMPs) such as collagenase was found to reflect multiple direct and indirect mechanisms of action, and to be therapeutically useful in a variety of dental (e.g., adult periodontitis) and medical (e.g., arthritis, osteoporosis, cancer) diseases. The site on the TC molecule responsible for its MMP-inhibitory activity was identified which led to the development of a series of chemically modified non-antimicrobial analogs, called CMTs, which also have therapeutic potential but do not appear to induce antibiotic side-effects. Longitudinal double-blind studies on humans with adult periodontitis have demonstrated that a sub-antimicrobial dose of doxycycline (previously reported to suppress collagenase activity in the periodontal pocket) is safe and effective and has recently been approved by the FDA as an adjunct to scaling and root planing.
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PMID:Tetracyclines inhibit connective tissue breakdown by multiple non-antimicrobial mechanisms. 997 17

Estrogen deficiency in the postmenopausal (PM) female is the major cause of osteoporosis and may contribute to increased periodontal disease, including alveolar bone loss, seen in these women. In the current study, an animal model of PM osteoporosis, the OVX adult female rat, was studied to determine: (i) the relationship between periodontal breakdown and skeletal bone loss, and (ii) the effect of CMT-8 on gingival collagenase and bone loss. OVX rats were daily gavaged with CMT-8 (1, 2, or 5 mg/rat) for 28 or 90 days; non-OVX rats and those gavaged with vehicle alone served as controls. Elevated collagenase activity, assessed using [3H-methyl] collagen as substrate in the presence or absence of APMA, was seen in the gingiva of the OVX rats, and CMT-8 therapy suppressed this effect. Western blot revealed a similar pattern for MMP-8 and MMP-13 concentrations. The changes in the gingival collagenase activity paralleled changes in periodontal bone loss, which, in turn, reflected trabecular bone density changes. Preliminary studies on PM humans administered sub-antimicrobial tetracycline as a matrix metalloproteinase inhibitor are under way.
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PMID:A chemically modified nonantimicrobial tetracycline (CMT-8) inhibits gingival matrix metalloproteinases, periodontal breakdown, and extra-oral bone loss in ovariectomized rats. 1041 37

Bisphosphonates are a group of drugs capable of inhibiting bone resorption, and are thus used for the treatment of bone diseases, such as Paget's disease, osteoporosis, and for bone metastases of malignant tumors. Their primary cellular target is considered to be the osteoclast. The molecular mechanisms responsible for the downregulation of bone resorption by bisphosphonates have remain unclear. We have discovered that various matrix metalloproteinases (MMPs) are inhibited in vitro by several bisphosphonates. This novel finding may, in part, explain the efficacy of bisphosphonates in their current indications in humans. In enzyme activity tests using purified and recombinant enzymes, we have observed the inhibition of MMP-1, -2, -3, -7, -8, -9, -12, -13, and -14 by clondronate, alendronate, pamidronate, zolendronate, nedrinate, and clodrinate. The IC50s range from 50 to 150 microM. We have also shown that clodronate can downregulate the expression of MT1-MMP protein and mRNA in several cell lines. Additionally, several bisphosphonates decrease the degree of invasion of malignant melanoma (C8161) and fibrosarcoma (HT1080) cells through artificial basement membrane (Matrigel) in cell cultures at IC50s of 50-150 microM and below. Having low toxicity and proven to be well tolerated after several years in human use, bisphosphonates have the potential to become one of the main MMP-inhibitors for MMP-related human soft and hard tissue-destructive diseases in the near future.
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PMID:MMP inhibition and downregulation by bisphosphonates. 1041 48

It is well documented that glucocorticoid excess causes bone loss, but the mechanisms of these effects remain poorly defined. To understand further the mechanisms of glucocorticoid-induced osteoporosis, we investigated the effects of glucocorticoids on bone formation and bone resorption by examining the proliferation, functional activities, and cytokine secretion of cultured human bone marrow stromal cells (hBMSC). Treatment with dexamethasone for 24 h at the concentration of 10(-8) M significantly suppressed [(3)H]thymidine incorporation and further inhibition was observed with longer treatment (8 days) or higher concentration (10(-7) M). Alkaline phosphatase activity of hBMSC was markedly stimulated with addition of dexamethasone (10(-8) M), to 191 +/- 22% (after 4 days) and 317 +/- 46% (after 7 days) of control. Dexamethasone (10(-8) M) treatment for 48 h decreased the incorporation of [(3)H]proline into collagenase-digestible protein (CDP; 43.7+/-7.9% of control) and non-collagen protein (65.2+/-8.4% of control), with a greater effect on CDP. Northern blot analysis indicated that alpha1(I)-collagen mRNA level was decreased by dexamethasone to 27.6 +/- 9.0% of the control value after 1 day of exposure, and to 55.2 +/- 6.2% after 7 days. Dexamethasone markedly suppressed basal production of interleukin (IL)-6 and IL-11 and that stimulated by parathyroid hormone (PTH), IL-1alpha, or tumour necrosis factor-alpha in a dose-dependent manner. These results suggest that the glucocorticoid-induced bone loss is derived at least in part via inhibition of bone formation, which includes the suppression of osteoblast proliferation and collagen synthesis. As both basal and PTH-stimulated production of IL-6 and IL-11 are decreased by dexamethasone, the increased bone resorption observed in glucocorticoid-induced osteopenia does not appear to be mediated by IL-6 or IL-11.
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PMID:Effects of dexamethasone on proliferation, activity, and cytokine secretion of normal human bone marrow stromal cells: possible mechanisms of glucocorticoid-induced bone loss. 1046 28

A human in vitro resorption assay has been developed using osteoclastoma-derived osteoclasts and used to evaluate novel antiresorptive agents including antagonists of the alphavbeta3 integrin, and inhibitors of cathepsin K and the osteoclast ATPase. The potency of novel compounds in the in vitro resorption assay correlates with functional assays for each class of inhibitor: the human alphavbeta3-mediated cell adhesion assay for the vitronectin receptor antagonists (r2 = 0.82), the chick osteoclast vacuolar ATPase enzyme assay for the H+-ATPase inhibitors (r2 = 0.77) and the recombinant human cathepsin K enzyme assay for the cathepsin K inhibitors (r2 = 0.80). Cell suspensions, rich in osteoclasts, are prepared by collagenase digestion of the tumor tissue. These cells can be stored long-term in liquid nitrogen and upon thawing maintain their bone-resorbing phenotype. The cryopreserved cells can be cultured on bovine cortical bone for 24-48 h and resorption can be measured by either confocal microscopy or biochemical assays. The resorptive activity of osteoclasts derived from a number of tumors can be inhibited reproducibly using a number of mechanistically unique antiresorptive compounds. In addition, the measurement of resorption pits by laser confocal microscopy correlates with the release of type I collagen C-telopeptides or N-telopeptides, as measured by enzyme-linked immunosorbent assay. Resorption can be measured reproducibly using a 48-h incubation of osteoclasts on bone slices, or a 24-h incubation with bone particles. This in vitro human osteoclast resorption assay provides a robust system for the evaluation of inhibitors of osteoclastic function that may be developed for the treatment of metabolic bone diseases such as osteoporosis.
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PMID:Development and characterization of a human in vitro resorption assay: demonstration of utility using novel antiresorptive agents. 1046 85

Dehydroepiandrosterone (DHEA) and its sulfate (DHEA-S) are the most abundant steroids in humans whose low levels are related to aging, greater incidence of various cancers, immune dysfunction, atherosclerosis, and osteoporosis. It has been shown that collagen and collagenase gene expression decreases in fibroblasts taken from more aged donors. In this paper, to investigate the relationship between DHEA and skin aging, we examined the effects of DHEA on the regulation of collagen, collegians and stromelysin-1 genes in cultured human skin fibroblasts. In collagen assay, DHEA slightly increased collagen production in a dose-related fashion, its maximal effect occurred at 10(-5) M DHEA (P>0.05). In the presence of DHEA, steady-state levels of alpha1 (I) procollagen mRNA increased to 1. 6-fold of the non-treated group, while those of fibronectin were not. Interestingly, DHEA differently regulated collagenase and stromelysin-1 gene expression. The steady-state levels of collagenase mRNA decreased in response to DHEA by 40%, whereas those of stromelysin-1 mRNA increased up to 2.4-fold, compared to controls. Similar results were obtained for chloramphenicol acetyltransferase assay (CAT); maximal promoter activation of stromelysin-1 gene occurred at 10(-6) M DHEA, 4.5-fold higher than control. CAT assay revealed that treatment with 10(-5) M DHEA resulted in a strong ( approximately 70%) inhibition of the collagenase promoter activity. In our experiments, the effects of DHEA on these gene expressions were higher at pharmacologic concentration (>/=10(-5) M) than those at physiologic concentration (10(-8)-10(-6) M). This study suggests that the level of DHEA may be related to the process of skin aging through the regulation of production and degradation in extracellular matrix.
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PMID:Effects of dehydroepiandrosterone on collagen and collagenase gene expression by skin fibroblasts in culture. 1080 27

Bisphosphonates inhibit bone resorption by reducing osteoclastic cell number and activity. Alendronate is a nitrogen-containing bisphosphonate analog used in the treatment of postmenopausal osteoporosis. The effects of alendronate in osteoclasts are well documented; however, there is limited information on the actions of alendronate in osteoblasts (Ob's). In this study, we investigated the effects of alendronate at concentrations of 1-100 microM on the synthesis of collagenase 3 or matrix metalloproteinase 13 (MMP-13) and tissue inhibitors of MMPs (TIMPs) 1, 2, and 3 in primary Ob-enriched cells from 22-day-old fetal rat calvariae. Alendronate at concentrations higher than 10 microM markedly stimulated the synthesis of collagenase messenger RNA (mRNA) and immunoreactive protein in Ob's. Alendronate did not stimulate the transcriptional rate of the collagenase 3 gene. However, in transcriptionally arrested cells, alendronate prolonged the half-life of collagenase transcripts. Alendronate did not alter the expression of TIMP 1 and 2, but modestly stimulated the expression of TIMP 3. The actions of alendronate in Ob's suggest potential additional effects in bone remodeling.
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PMID:Alendronate stimulates collagenase 3 expression in osteoblasts by posttranscriptional mechanisms. 1112 99

The cholesterol-lowering drug, simvastatin, is a pro-drug of a potent 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor and inhibits cholesterol synthesis in humans and animals. In addition, the bone effects of statins including simvastatin are being studied. We assessed the effects of simvastatin on osteoblastic differentiation in nontransformed osteoblastic cells (MC3T3-E1) and rat bone marrow cells. Simvastatin enhanced alkaline phosphatase (ALP) activity and mineralization in a dose- and time-dependent fashion. This stimulatory effect of the statin was observed at relatively low doses (significant at 10(-8) M and maximal at 10(-7) M). Northern blot analysis showed that the statin (10(-7) M) increased in bone morphogenetic protein-2 as well as ALP mRNA concentrations in MC3T3-E1 cells. Simvastatin (10(-7) M) slightly increased in type I collagen mRNA abundance throughout the culture period, whereas it markedly inhibited the gene expression of collagenase-1 between days 14 and 22 of culture. These results indicate that simvastatin has anabolic effects on bone through the promotion of osteoblastic differentiation, suggesting that it could be used for the treatment of common metabolic bone diseases such as osteoporosis.
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PMID:Simvastatin promotes osteoblast differentiation and mineralization in MC3T3-E1 cells. 1116 4

Recent studies have demonstrated that tetracyclines can reduce bone loss in the ovariectomized (OVX) rat model of osteoporosis. In the current study, a non-antimicrobial, chemically modified doxycycline (CMT-8), alone or in combination with a bisphosphonate (Clodronate), was evaluated in this model. Forty-two, 6month old, female rats were randomly assigned to the following groups, (6/ group): a) sham/vehicle, b) OVX/vehicle; c) OVX/1 mg/day CMT-8; d) OVX/2 mg/day CMT-8, e) OVX/1 mg/week Clodronate; and f) OVX/1 mg/day CMT-8 + 1 mg/week Clodronate, CMT-8 was administered by oral gavage, Clodronate injected S/C. Following sham surgery or OVX, the rats were treated for 90 days with CMT-8 or vehicle alone, injected at three different times with fluorochrome labels, the rats were sacrificed, and the tibiae excised for analysis by dynamic bone histomorphometry. Femurs were aseptically removed and analyzed for collagen, collagenase and osteopontin mRNAs by Northern and dot blot analysis. As expected, OVX decreased trabecular bone volume (BV/TV by 73.8% vs. sham p<.01), and also reduced trabecular thickness, numbers, and increased spacing. Bone loss in the OVX animals was partially prevented with either 2 mg/day CMT-8 or 1 mg/wk Clodronate (p<.01), while the 1 mg/day CMT-8 had no effect. Interestingly, the efficacy of the combination therapy of CMT-8 and Clodronate was significantly better than either treatment by itself, maintaining bone mass and structural indices at levels identical to sham values. OVX rats mRNA for collagen, collagenase and osteopontin were elevated indicating high-turnover bone loss. Only COMBO therapy significantly reduced the collagenase and osteopontin mRNA. In summary, CMT-8 mono-therapy (2 mg) alone partially inhibited bone loss in this animal model of osteoporosis. However, 1 mg/day (CMT-8) monotherapy had no effect on bone loss or bone mRNA levels and when combined with Clodronate, interacted to increase efficacy. Thus, a combination of a suboptimal dose of CMT-8 and a bisphosphonate appears to increase the amount of bone by suppressing resorption in a model of osteoporosis.
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PMID:A combination of subtherapeutic doses of chemically modified doxycycline (CMT-8) and a bisphosphonate (clodronate) inhibits bone loss in the ovariectomized rat: a dynamic histomorphometric and gene expression study. 1117 84


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