The effect of FUT-175 (nafamstat mesilate) on C3a, C4a and C5a generation in vitro and inflammatory reactions in vivo

doi:10.1016/0192-0561(90)90062-R

International Journal of Immunopharmacology

Volume 12, Issue 1, 1990, Pages 1-9

International Journa...

https://doi.org/10.1016/0192-0561(90)90062-R Get rights and content

Abstract

FUT-175 is a synthetic protease inhibitor and an inhibitor of the classical and alternate pathways of complement activation. In human serum, FUT-175 inhibited C3_a, C4_a and C5_a generation induced by heat aggregated IgG, zymosan and Cobra venom factor with IC₅₀ values in the range of 3–43 μM depending on the stimulus and the fragments. To assess in vivo anti-inflammatory activity, inflammatory reactions induced in the skin of rabbits were quantitated by using ¹²⁵I-albumin extravasation, ⁵¹Cr-labelled leukocyte accumulation and ⁸⁶RbCl accumulation as a measure of hyperemia. Infusion of FUT-175 at 2 mg/kg/h inhibited all three parameters by 50–80% in dermal reactions induced by killed E. coli, zymosan, immune complexes, the reversed Arthus reaction, zymosan activated plasma (ZAP), f-norleu-leu-phe (FNLP) and LTB₄. In contrast, the response to endotoxin (0.1 μg) was not effected by FUT-175 treatment. The effect of FUT-175 was comparable to that of local or systemic therapy with indomethacin, but unlike indomethacin, the effect of FUT-175 was not reversed by local PGE₂ administration. Furthermore, indomethacin and FUT-175 had additive anti-inflammatory effects. These results suggest that although FUT-175 is a potent inhibitor of C3_a, C4_a and C5_a generation, it has novel and broad anti-inflammatory effects, possibly through actions in addition to complement inhibition as indicated by inhibition of FNLP-, LTB₄- and ZAP-induced reactions.

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Cited by (37)

Nafamostat mesilate, a serine protease inhibitor, suppresses interferon-gamma-induced up-regulation of programmed cell death ligand 1 in human cancer cells
2018, International Immunopharmacology
Citation Excerpt :
Also, in other two human pancreatic cancer cell lines, Capan-1 and Capan-2, IFN-gamma-induced PD-L1 up-regulation was significantly suppressed by NM treatment (Fig. 5-B, C). Regarding the immunological aspects of NM treatment, NM inhibits the complement molecules C3a, C4a and C5a and elicits anti-inflammatory effects [27]. T cell auto-reactivity was suppressed by NM in experimental autoimmune encephalomyelitis by decreasing granzyme activity and CTL cytolysis [28,29].
Programmed cell death ligand-1 (PD-L1) plays a pivotal role in the suppression of antitumour immunity by binding to programmed cell death-1 (PD-1) on tumouricidal cytotoxic T lymphocytes (CTLs), rendering them inactive. As blockade of PD-1/PD-L1 interaction by the monoclonal antibodies induced effective T cell-mediated antitumour response, suppression of PD-L1 expression in tumour cells by the chemical agent might contribute to treatment against malignant tumours. Nafamostat mesilate (NM), a serine protease inhibitor that is frequently used in the clinic, potently suppressed interferon-gamma (IFN-gamma)-induced up-regulation of PD-L1 in cultured human lung cancer cells (HLC-1) at both the messenger RNA (mRNA) and protein levels. Interestingly, suppression of IFN-gamma-induced up-regulation of human leukocyte antigen (HLA)-ABC by NM was limited, suggesting that NM did not block CTL responses to tumour cells. NM treatment did not affect the activation status of signal transducer and activator of transcription (STAT) 1 or the induction of interferon regulatory factor (IRF)-1 expression in IFN-gamma-treated HLC-1 cells. Although NM treatment promoted the phosphorylation of extracellular signal-regulated kinases (Erk) 1/2, an Erk inhibitor, U0126, could not reverse the suppression of PD-L1 up-regulation by IFN-gamma. Suppression of IFN-gamma-induced up-regulation of PD-L1 by NM was not associated with the inhibition of nuclear factor kappa B (NF-kB) or protease-activated receptor (PAR)-1 pathway. Besides HLC-1 cells, NM suppressed IFN-gamma-induced PD-L1 up-regulation in three human pancreatic cancer cell lines. NM could potentiate the antitumour effect of cancer vaccines or immune checkpoint inhibitors by preventing IFN-gamma-induced PD-L1 up-regulation and blocking immune checkpoint suppression.
Effect of synthetic protease inhibitor gabexate mesilate on attenuation of coagulant activity and cytokine release in a rat model of islet transplantation
2011, Transplantation Proceedings
Citation Excerpt :
Actually, the level of IL-6 or HMGB1 in the present study was much lower than that following lipopolysaccharide administration or puncture of duodenal wall.7,11 Serine proteases are involved in a broad range of physiologic and inflammatory processes, including complement activation.12 To elucidate the effect of GM on the activation of complement cascades, we sought to examine the deposits of C5b-9 on the islet grafts by immunohistochemical analyses.
The instant blood-mediated inflammatory reaction (IBMIR), in which the activation of coagulation cascade plays a key role, is one of the serious obstacles to successful islet engraftment. Gabexate mesilate (GM) is well known to elicit anticoagulant and antiinflammatory effects. The aim of this study was to evaluate the effect of GM on syngeneic IBMIR.
Syngeneic rat islet grafts (2.5 IEQ/g) were transplanted intraportally into 2 groups (control group and GM group; n = 10–11) of streptozotocin-induced diabetic rats. The GM group was injected intravenously with GM for 30 minutes before islet infusion to 1 hour after. The control group was injected with equivalent amount of saline solution. Plasma samples were collected before and 0.5, 1, 3, 6, and 24 hours after transplantation, and several proinflammatory mediators, including interleukin-6 and high-mobility group Box 1 were measured. Curative rate, intravenous glucose tolerance test, and insulin amount in the recipients' livers were also evaluated.
Little difference was observed in any proinflammatory mediators. Whereas none of the animals in the control group became normoglycemic, 2 of 6 rats transplanted with the same number of islets in the GM group became normoglycemic during the study period. The glucose tolerance response was significantly ameliorated in the GM group compared with the control group (P < 0.001). The insulin amount in the liver of the recipients was considerably higher in the GM group (5.6 ± 4.1 vs 12.6 ± 5.3 ng/IEQ; P < .05).
These data suggest that GM improves islet engraftment not through suppressing the proinflammatory cytokines but as an anticoagulant. We therefore think that GM could be a useful anticoagulant to control IBMIR induced in clinical islet transplantation, although antiinflammatory reagents are considered to be needed for the ideal regimen.
Inhibiting the C5-C5a receptor axis
2011, Molecular Immunology
Citation Excerpt :
For example the synthetic protease inhibitor, FUT-175, prevents C5 convertase activity (Inagi et al., 1991) and has been widely used to inhibit complement-mediated immunopathology (La Bonte et al., 2008; Li et al., 2009). However, this protease inhibitor also prevents C3 cleavage, as well as a variety of other serine proteases (Inagi et al., 1991; Issekutz et al., 1990). Thus, any serine protease inhibitor may indeed inhibit C5 convertase activity, but would, by necessity, have numerous non-C5 specific activities, which may hamper their clinical utility.
Activation of the complement system is a major pathogenic event that drives various inflammatory responses in numerous diseases. All pathways of complement activation lead to cleavage of the C5 molecule generating the anaphylatoxin C5a and, C5b that subsequently forms the terminal complement complex (C5b-9). C5a exerts a predominant pro-inflammatory activity through interactions with the classical G-protein coupled receptor C5aR (CD88) as well as with the non-G protein coupled receptor C5L2 (GPR77), expressed on various immune and non-immune cells. C5b-9 causes cytolysis through the formation of the membrane attack complex (MAC), and sub-lytic MAC and soluble C5b-9 also possess a multitude of non-cytolytic immune functions. These two complement effectors, C5a and C5b-9, generated from C5 cleavage, are key components of the complement system responsible for propagating and/or initiating pathology in different diseases, including paroxysmal nocturnal hemoglobinuria, rheumatoid arthritis, ischemia–reperfusion injuries and neurodegenerative diseases. Thus, the C5–C5a receptor axis represents an attractive target for drug development. This review provides a comprehensive analysis of different methods of inhibiting the generation of C5a and C5b-9 as well as the signalling cascade of C5a via its receptors. These include the inhibition of C5 cleavage through targeting of C5 convertases or via the C5 molecule itself, as well as blocking the activity of C5a by neutralizing antibodies and pharmacological inhibitors, or by targeting C5a receptors per se. Examples of drugs and naturally occurring compounds used are discussed in relation to disease models and clinical trials. To date, only one such compound has thus far made it to clinical medicine: the anti-C5 antibody eculizumab, for treating paroxysmal nocturnal hemoglobinuria. However, a number of drug candidates are rapidly emerging that are currently in early-phase clinical trials. The C5–C5a axis as a target for drug development is highly promising for the treatment of currently intractable major human diseases.
Nafamostat mesilate attenuates colonic inflammation and mast cell infiltration in the experimental colitis
2011, International Immunopharmacology
Citation Excerpt :
In the present study, treatment with NM significantly attenuated colonic MPO, TNF-α, COX-2 and iNOS level associated with ulceration. There have been a number of reports suggesting anti-inflammatory activities of NM [16,17,21]. Noguchi et al. reported that NM suppressed the expression of iNOS in RAW264.7 murine macrophages treated with lipopolysaccharide [27].
Serine proteases are important in the pathogenesis of intestinal inflammation. Recent studies have shown that nafamostat mesilate (NM) can inhibit the colonic mucosal inflammation induced by TNBS in rats. The aim of this study was to investigate the anti-inflammatory effects of NM on a DSS-induced colitis. Colitis was induced in female BALB/c mice by 5% dextran sulfate sodium (DSS) for 6 days. NM (2 or 20 mg/kg body weight) was orally administered once a day for 6 days during treatment of the mice with DSS. The inflammatory response of the colon was assessed 1 week after DSS treatment. NM at a high dose, but not at a low dose significantly decreased disease activity index (DAI) and myeloperoxidase (MPO) induced by DSS. Furthermore, NM (20 mg/kg) inhibited the production of tumor necrosis factor (TNF)-α, cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in the colonic tissues treated with DSS. The increase in chymase activity by DSS treatment was also attenuated by the administration of NM (20 mg/kg). NM (20 mg/kg) significantly decreased the colonic mucosal injury and the infiltrated mast cell number induced by DSS. These results indicate that NM might inhibit the colonic inflammation through inhibition of both chymase activity and mast cell infiltration in colon tissues of DSS-induced colitis.
Inhibition of classical pathway of complement activation with negative charged derivatives of bisphenol a and bisphenol disulphates
2005, Bioorganic and Medicinal Chemistry
In order to obtain strong inhibitors of classical pathway of complement activation the low weight negative charged compounds have been investigated. On the basis of bisphenol A anionic derivatives with one or two carboxylic, sulphate and phosphate groups the critical role of negative charged groups for complement-inhibiting activity has been established. It was determined that two sulphate or phosphate groups in the molecule provide the most inhibiting effect. At the next stage a set of bisphenol disulphates of varying structures has been synthesized and investigated. Bulky hydrophobic groups (cyclohexyliden, fluorenyliden, anthronyliden) at the central part of the bisphenol molecule it was found to increase complement-inhibiting activity markedly. The replacement of the ortho-positions to the charged group by halogens or alkyl groups (allyl, propyl) increases the inhibiting effect. It was showed by ELISA that several compounds studied interact with C1q, $C 1 \bar{r}$ / $C 1 \bar{s}$ components of complement. For the set of bisphenol disulphates the QSAR equation with hydrophobic coefficient and electronic parameters has been formulated. Both hydrophobic and electrostatic interactions it was established to have a great significance for the inhibition of classical pathway of complement activation.
Effects of nafamostat mesilate and minimal-dose aprotinin on blood-foreign surface interactions in cardiopulmonary bypass
2004, Annals of Thoracic Surgery
Citation Excerpt :
Theoretically, the strong inhibition of XIIa and kallikrein by NM should shut down the release of elastase by neutrophils. It has been reported that NM inhibits the protease involved in the release of cytokines from cytokine-producing cells [23]. On the other hand, the effects of aprotinin on the inflammatory response have been thought to be limited because its inhibiting dose for kallikrein is about 100 times greater than that for plasmin [5].
The pharmacological inhibition of blood-foreign surface interactions is an attractive strategy for reducing the morbidity associated with cardiopulmonary bypass. We compared the inhibitory effects of nafamostat mesilate (a broad-spectrum synthetic protease inhibitor) and minimal-dose aprotinin on blood-surface interactions in clinical cardiopulmonary bypass.
Eighteen patients undergoing coronary surgery were divided into three groups: (1) the control group (heparin, 4 mg/kg; n = 6), (2) the nafamostat mesilate group (heparin plus nafamostat, 0.2 mg/kg bolus followed by 2.0 mg/kg/h during cardiopulmonary bypass; n = 6), and (3) the aprotinin group (heparin plus aprotinin, 2.0 × 10⁴ KIU/kg; n = 6). Platelet count, platelet aggregation, β-thromboglobulin, prothrombin fragment F1.2, thrombin-antithrombin complex, plasminogen activator inhibitor-1, α2-plasmin inhibitor-plasmin complex, D-dimer, neutrophil elastase, and interleukin-6 were measured before, during, and after bypass. Bleeding times and blood loss were recorded.
There were no significant differences between groups in platelet count, β-thromboglobulin, plasminogen activator inhibitor-1, interleukin-6, bleeding times, or blood loss. Platelet aggregation was better preserved at 12 hours after surgery in the nafamostat and aprotinin groups than in the control group. Prothrombin fragment F1.2, thrombin-antithrombin complex and neutrophil elastase levels were significantly reduced by aprotinin, but not by nafamostat as compared with the control group. The α2-plasmin inhibitor-plasmin complex and D-dimer were significantly lower with either of the drugs. Aprotinin showed better control of D-dimer than did nafamostat.
Nafamostat mesilate fails to reduce thrombin formation and neutrophil elastase release, whereas minimal-dose aprotinin inhibits both. Neither nafamostat nor aprotinin inhibits platelet activation. Nafamostat reduces fibrinolysis during cardiopulmonary bypass, although its effect is not as potent as aprotinin.

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International Journal of Immunopharmacology

Abstract

References (29)

Nafamstat mesilate

Drugs of the Future

Pharmacological studies of FUT-175, nafamstat mesilate I. inhibition of protease activity in in vitro and in vivo experiments

Jap. J. Pharmac.

Interleukin 1 acts on cultured human vascular endothelium to increase the adhesion of polymorphonuclear leukocytes, monocytes and related leukocyte cell lines

J. clin. Invest.

The arthus reaction: a model of neutrophil and complement-mediated injury.

Depletion of plasma complement in vivo by a protein of cobra venom: its effect on various immunologic reactions

J. Immun.

Acute inflammation induced by immune complexes in the microcirculation

Exp. molec. Path.

Chemotactic response to human C3_a and C5_a anaphylatoxins. 1. Evaluation of C3_a and C5_a leukotaxis in vitro and under simulated in vivo conditions

J. Immun.

New synthetic inhibitors of C1r, C1 esterase, thrombin, plasmin, kallikrein and trypsin

Biochim. biophys. Acta

Stimulation of the adherence of neutrophils to umbilical vein endothelium by human recombinant tumor necrosis factor

Tissue injury in inflammation

J. clin. Invest.

The structural basis for anaphylatoxin and chemotactic functions of C3_a, C4_a and C5_a

CRC Critic. Rev. Immun.

New synthetic inhibitor to the alternative complement pathway

Immunology

Effect of FUT-175, a new synthetic protease inhibitor, on the development of lupus nephritis in (NZBxNZW) F₁, mice

Immunology

The effect of nonsteroidal anti-inflammatory agents on E. coli-induced inflammation

Immunopharmacology

Cited by (37)

Nafamostat mesilate, a serine protease inhibitor, suppresses interferon-gamma-induced up-regulation of programmed cell death ligand 1 in human cancer cells

Effect of synthetic protease inhibitor gabexate mesilate on attenuation of coagulant activity and cytokine release in a rat model of islet transplantation

Inhibiting the C5-C5a receptor axis

Nafamostat mesilate attenuates colonic inflammation and mast cell infiltration in the experimental colitis

Inhibition of classical pathway of complement activation with negative charged derivatives of bisphenol a and bisphenol disulphates

Effects of nafamostat mesilate and minimal-dose aprotinin on blood-foreign surface interactions in cardiopulmonary bypass

The effect of FUT-175 (nafamstat mesilate) on C3a, C4a and C5a generation in vitro and inflammatory reactions in vivo

Abstract

Nafamstat mesilate

Drugs of the Future

Pharmacological studies of FUT-175, nafamstat mesilate I. inhibition of protease activity in in vitro and in vivo experiments

Jap. J. Pharmac.

Interleukin 1 acts on cultured human vascular endothelium to increase the adhesion of polymorphonuclear leukocytes, monocytes and related leukocyte cell lines

J. clin. Invest.

The arthus reaction: a model of neutrophil and complement-mediated injury.

Depletion of plasma complement in vivo by a protein of cobra venom: its effect on various immunologic reactions

J. Immun.

Acute inflammation induced by immune complexes in the microcirculation

Exp. molec. Path.

Chemotactic response to human C3a and C5a anaphylatoxins. 1. Evaluation of C3a and C5a leukotaxis in vitro and under simulated in vivo conditions

J. Immun.

New synthetic inhibitors of C1r, C1 esterase, thrombin, plasmin, kallikrein and trypsin

Biochim. biophys. Acta

Stimulation of the adherence of neutrophils to umbilical vein endothelium by human recombinant tumor necrosis factor

Tissue injury in inflammation

J. clin. Invest.

The structural basis for anaphylatoxin and chemotactic functions of C3a, C4a and C5a

CRC Critic. Rev. Immun.

New synthetic inhibitor to the alternative complement pathway

Immunology

Effect of FUT-175, a new synthetic protease inhibitor, on the development of lupus nephritis in (NZBxNZW) F1, mice

Immunology

The effect of nonsteroidal anti-inflammatory agents on E. coli-induced inflammation

Immunopharmacology

The effect of FUT-175 (nafamstat mesilate) on C3_a, C4_a and C5_a generation in vitro and inflammatory reactions in vivo

Chemotactic response to human C3_a and C5_a anaphylatoxins. 1. Evaluation of C3_a and C5_a leukotaxis in vitro and under simulated in vivo conditions

The structural basis for anaphylatoxin and chemotactic functions of C3_a, C4_a and C5_a

Effect of FUT-175, a new synthetic protease inhibitor, on the development of lupus nephritis in (NZBxNZW) F₁, mice