cathepsin-g and Lung-Diseases

Polymorphonuclear neutrophils are the first cells recruited to inflammatory sites and form the earliest line of defense against invading microorganisms. Neutrophil elastase, proteinase 3, and cathepsin G are three hematopoietic serine proteases stored in large quantities in neutrophil cytoplasmic azurophilic granules. They act in combination with reactive oxygen species to help degrade engulfed microorganisms inside phagolysosomes. These proteases are also externalized in an active form during neutrophil activation at inflammatory sites, thus contributing to the regulation of inflammatory and immune responses. As multifunctional proteases, they also play a regulatory role in noninfectious inflammatory diseases. Mutations in the ELA2/ELANE gene, encoding neutrophil elastase, are the cause of human congenital neutropenia. Neutrophil membrane-bound proteinase 3 serves as an autoantigen in Wegener granulomatosis, a systemic autoimmune vasculitis. All three proteases are affected by mutations of the gene (CTSC) encoding dipeptidyl peptidase I, a protease required for activation of their proform before storage in cytoplasmic granules. Mutations of CTSC cause Papillon-Lefèvre syndrome. Because of their roles in host defense and disease, elastase, proteinase 3, and cathepsin G are of interest as potential therapeutic targets. In this review, we describe the physicochemical functions of these proteases, toward a goal of better delineating their role in human diseases and identifying new therapeutic strategies based on the modulation of their bioavailability and activity. We also describe how nonhuman primate experimental models could assist with testing the efficacy of proposed therapeutic strategies.

Topics: Animals; Catalytic Domain; Cathepsin G; Humans; Leukocyte Elastase; Lung Diseases; Molecular Targeted Therapy; Myeloblastin; Neutropenia; Papillon-Lefevre Disease

The proteinase-antiproteinase hypothesis still receives support from clinical and experimental observations in a range of inflammatory lung diseases. The function of these molecules appears to be broader than originally believed and further research is likely to lead to an improved understanding of their role in the regulation of both the beneficial and detrimental effects in inflammatory response and the maintenance of the homeostasis in the normal lung. Thus the potential for the development as therapeutic tools is likely to become more attractive as improved drug development and delivery mechanisms appear.

Topics: alpha 1-Antitrypsin; Cathepsin G; Cathepsins; Genetic Therapy; Humans; Leukocyte Elastase; Lung Diseases; Myeloblastin; Proteinase Inhibitory Proteins, Secretory; Proteins; Serine Endopeptidases

Mast cells and neutrophils are key contributors to the pathophysiological inflammatory processes that underpin asthma and chronic obstructive pulmonary disease, partly through the release of noxious serine proteases, including cathepsin G (Cat G) and chymase. From this standpoint, a dual inhibitor of neutrophil Cat G and mast cell chymase could protect against these disease-related inflammatory responses.. We examined the antiinflammatory pharmacology of RWJ-355871, a dual inhibitor of Cat G and chymase, in animal models of inflammation that evince pathophysiological pathways relevant to asthma and chronic obstructive pulmonary disease to determine the therapeutic potential of this compound.. In an ovalbumin (OVA)-sensitized rat model, RWJ-355871 was administered to block the mast-cell-mediated increase in paw volume caused by OVA injection. In a sheep asthma model, antigen-induced airway responses were assessed with and without aerosol treatment with RWJ-355871. In a murine tobacco-smoke model of airway inflammation, the effect of RWJ-355871 on smoke-induced neutrophilia was determined.. Intravenous treatment of OVA-sensitized rats with RWJ-355871 provided dose-dependent reduction in the increase in rat paw volume. In allergic sheep, aerosol pretreatment with RWJ-355871 showed dose-dependent inhibition of the antigen-induced early response, late response, and post-antigen-induced airway hyperreponsiveness. In tobacco-smoke-exposed mice, nebulized RWJ-355871 significantly reduced the smoke-induced neutrophilia from the levels observed in untreated mice.. The preclinical antiinflammatory effects of RWJ-355871 in these animal models of inflammation indicate that this dual inhibitor may have therapeutic utility for treating airway inflammatory diseases involving mechanisms that depend on Cat G and/or chymase.

Topics: Animals; Biomarkers; Bronchoalveolar Lavage Fluid; Cathepsin G; Chymases; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Female; Injections, Intravenous; Lung Diseases; Mice; Organophosphonates; Piperidines; Pulmonary Disease, Chronic Obstructive; Rats; Sheep; Treatment Outcome

The secretory leukocyte protease inhibitor (SLPI), elafin, and its biologically active precursor trappin-2 are endogeneous low-molecular weight inhibitors of the chelonianin family that control the enzymatic activity of neutrophil serine proteases (NSPs) like elastase, proteinase 3, and cathepsin G. These inhibitors may be of therapeutic value, since unregulated NSP activities are linked to inflammatory lung diseases. However SLPI inhibits elastase and cathepsin G but not proteinase 3, while elafin targets elastase and proteinase 3 but not cathepsin G. We have used two strategies to design polyvalent inhibitors of NSPs that target all three NSPs and may be used in the aerosol-based treatment of inflammatory lung diseases. First, we fused the elafin domain with the second inhibitory domain of SLPI to produce recombinant chimeras that had the inhibitory properties of both parent molecules. Second, we generated the trappin-2 variant, trappin-2 A62L, in which the P1 residue Ala is replaced by Leu, as in the corresponding position in SLPI domain 2. The chimera inhibitors and trappin-2 A62L are tight-binding inhibitors of all three NSPs with subnanomolar K(i)s, similar to those of the parent molecules for their respective target proteases. We have also shown that these molecules inhibit the neutrophil membrane-bound forms of all three NSPs. The trappin-2 A62L and elafin-SLPI chimeras, like wild-type elafin and trappin-2, can be covalently cross-linked to fibronectin or elastin by a tissue transglutaminase, while retaining their polypotent inhibition of NSPs. Therefore, the inhibitors described herein have the appropriate properties to be further evaluated as therapeutic anti-inflammatory agents.

Topics: Amino Acid Sequence; Cathepsin G; Cathepsins; Dose-Response Relationship, Drug; Drug Discovery; Elafin; Humans; Lung Diseases; Membrane Proteins; Models, Molecular; Molecular Sequence Data; Myeloblastin; Neutrophils; Pancreatic Elastase; Protease Inhibitors; Protein Engineering; Recombinant Fusion Proteins; Secretory Leukocyte Peptidase Inhibitor; Sequence Alignment; Serine Endopeptidases; Static Electricity; Transglutaminases

Patients with cystic fibrosis (CF) of the same age differ significantly in their degree of pulmonary disease. Based on preliminary observations, we postulated that the activity of myeloperoxidase would be significantly increased in patients with greater structural lung damage than in those with less lung damage. Acid extracts of weighed sputum samples were assayed for lactoferrin concentrations by ELISA. Activities of peroxidase, cathespsin G, and elastase (with and without proteinase 3) were determined by kinetic analysis using chromogenic substrates. The patients were divided into quartiles based on their Brasfield chest-radiograph score. Patients in the first quartile (least amount of structural lung abnormality) were compared to those in the fourth quartile. The concentration of lactoferrin, a specific (secondary) granule protein of neutrophils, did not differ between the two patient groups. However, the activities of the neutrophil primary granule proteins, peroxidase, elastase, and elastase plus proteinase 3, were significantly elevated in the group with the most structural lung abnormality. Sputum albumin concentration was used to estimate leakages of plasma proteins into the airways. Peroxidase activity, but not the activity of cathepsin G, of elastase, or of elastase plus proteinase 3, correlated significantly with albumin/g sputum in both quartile groups. To confirm the association of sputum peroxidase activity with differences in lung structure and to test its correlation with lung function, spirometry was performed in a second group of patients during the week prior to the time of sputum sampling. In this second group, increased sputum peroxidase activity was associated with worse Brasfield scores and with decreased percent-predicted forced expiratory volume in 1 sec.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adult; Analysis of Variance; Cathepsin G; Cathepsins; Cystic Fibrosis; Enzyme-Linked Immunosorbent Assay; Female; Humans; Lactoferrin; Lung Diseases; Male; Pancreatic Elastase; Peroxidase; Respiratory Function Tests; Serine Endopeptidases; Severity of Illness Index; Sputum

The sulfated polymer MDL 101,028 was found to be a potent-inhibitor of both human neutrophil elastase (HNE) and human neutrophil cathepsin G (CatG). Cleavage of synthetic substrate by HNE was inhibited by MDL 101,028 with an IC50 of 40 nM, while CatG was inhibited with an IC50 of 80 nM. Degradation of a macromolecular connective tissue substrate (cartilage proteoglycan) by HNE or CatG was inhibited by MDL 101,028 with an IC50 of approximately 10 microM. MDL 101,028 at concentrations of 4, 10 and 25 microM inhibited degradation of cartilage proteoglycan by human neutrophil lysate or stimulated human neutrophils by 54%, 70% and 79%, and 31%, 47% and 73%, respectively. Acute pulmonary injury resulting from the intratracheal (i.t.) instillation of HNE in rats was inhibited by 48%, 90% and 90% at concentrations of MDL 101,028 of 1.1 mg/kg, 2.8 mg/kg and 11 mg/kg. The duration of action of the compound after i.t. instillation was between 2 and 4 h. These results suggest that sulfated polymers such as MDL 101,146 may be useful as inhibitors of HNE-mediated lung injury.

Topics: Amino Acid Sequence; Animals; Biphenyl Compounds; Cartilage; Cathepsin G; Cathepsins; Cattle; Connective Tissue; Extracellular Matrix; Hemorrhage; Humans; Leukocyte Elastase; Lung Diseases; Male; Molecular Sequence Data; Neutrophils; Pancreatic Elastase; Proteoglycans; Rats; Rats, Sprague-Dawley; Serine Endopeptidases

The pulmonary tree is exposed to neutrophil-derived serine proteinases and matrix metalloproteinases in inflammatory lung diseases, but the degree to which these enzymes participate in tissue injury remains undefined, as does the therapeutic utility of antiproteinase-based interventions. To address these issues, an in vivo rat model was examined in which the intrapulmonary deposition of immune complexes initiates a neutrophil-mediated acute alveolitis. In vitro studies demonstrated that rat neutrophils can release neutrophil elastase and cathepsin G as well as a neutrophil progelatinase, which was subsequently activated by either chlorinated oxidants or serine proteinases. Based on structural homologies that exist between rat and human neutrophil proteinases, rat neutrophil elastase and cathepsin G activities could be specifically regulated in vitro by recombinant human secretory leukoproteinase inhibitor, and rat neutrophil gelatinase activity proved sensitive to inhibition by recombinant human tissue inhibitor of metalloproteinases 2. When either of the recombinant antiproteinases were instilled intratracheally, in vivo lung damage as assessed by increased permeability or hemorrhage was significantly reduced. Furthermore, the coadministration of the serine and matrix metalloproteinase inhibitors almost completely prevented pulmonary damage while effecting only a modest decrease in neutrophil influx. These data support a critical role for neutrophil-derived proteinases in acute lung damage in vivo and identify recombinant human secretory leukoproteinase and recombinant human tissue inhibitor of metalloproteinases 2 as potentially efficacious interventions in inflammatory disease states.

Topics: Animals; Antigen-Antibody Complex; Cathepsin G; Cathepsins; Cytochalasin B; Enzyme Precursors; Gelatinases; Humans; Leukocyte Elastase; Lung Diseases; Male; Metalloendopeptidases; N-Formylmethionine Leucyl-Phenylalanine; Neoplasm Proteins; Neutrophils; Oxidants; Pancreatic Elastase; Proteinase Inhibitory Proteins, Secretory; Proteins; Pulmonary Alveoli; Rats; Recombinant Proteins; Serine Endopeptidases; Serine Proteinase Inhibitors; Tetradecanoylphorbol Acetate; Tissue Inhibitor of Metalloproteinase-2