The design and synthesis of second generation leukotriene B4 (LTB4) receptor antagonists related to SC-41930

SC-41930, 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid, is a selective, orally active, LTB₄ receptor antagonist currently in clinical trials for psoriasis and ulcerative colitis. Exhaustive SAR studies found a potential backup compound, SC-50605, which was 7–16 times more potent than SC-41930 in LTB₄ receptor binding, chemotaxis and degranulation assays. SC-50605 also inhibited LTB₄-induced intradermal chemotaxis in cavine skin at an oral dose of 0.10 mg/kg and displayed good activity in animal models of colitis and epidermal inflammation both orally and topically.     

The effect of leukotriene-B4 receptor antagonist,SC-41930, on acetic acid-induced colonic inflammation

SC-41930, 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)-propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid, is a potentin vitro leukotriene-B₄ (LTB₄) receptor antagonist. LTB₄ levels are elevated in colonic tissue of inflammatory bowel disease (IBD) patients which may account for the high degree of neutrophil (PMN) infiltration. The guinea pig acetic acid-induced colonic inflammation model has characteristics of IBD including PMN infiltration, edema, ulceration and necrosis. The model was used to evaluate the effect of SC-41930. SC-41930 was given orally, 30 min before and after intrarectal administration of 3% acetic acid. The PMN marker enzyme, myeloperoxidase, was measured along with histological evaluation to assess inflammation. Both parameters showed significantly less inflammation in SC-41930 treated animals with an oral ED₅₀ of 20 mg/kg. These study results with an LTB₄ receptor antagonist indicate a role for LTB₄ in colonic inflammation and that an LTB₄ receptor antagonist may be beneficial for treatment of IBD.     

Dermal inflammation in primates,mice, and guinea pigs: Attenuation by second-generation leukotriene B4 receptor antagonist,SC-53228

Granulocyte infiltration is a prominent feature of human psoriasis. Psoriatic lesional skin contains abnormally high amounts of immunoreactive leukotriene B₄ (LTB₄), a potent granulocyte chemotaxin in vivo and in vitro. SC-53228 [(+)-(S)-7-(3-{2-(cyclopropylmethyl)-3-methoxy-4-[(methylamino)carbonyl]phenoxy}propoxy)-3, 4-dihydro-8-propyl-2H-1-benzopyran-2-propanoic acid], a second-generation LTB₄ receptor antagonist, was tested topically and orally in phorbol ester-induced dermal inflammation in three species. Skin inflammation was induced by topical application of phorbol-12-myristate-13-acetate-(PMA/TPA) and assessed by ear thickness, levels of the neutrophil marker enzyme myeloperoxidase (MPO) and histological examination. In mice, SC-53228 inhibited inflammation with a topical ED₅₀ value of 200 ± 18 g. When applied to guinea pigs, SC-53228 (100 g) inhibited the MPO increase by 86%, while 1000 g abrogated inflammation in rhesus macaques with no plasma accumulation of the drug. A 1 % gel formulation was also efficacious in guinea pig PMA-induced epidermal inflammation. Furthermore, single oral dose administration to mice was efficacious (ED₅₀ < 2.5 mg/kg) as was multidose administration to rhesus macaques. PMA-induced skin inflammation possesses some of the attributes of human psoriasis and an agent such as SC-53228 may have utility in the medical management of this condition.     

Effects of SC-41930 on leukocyte adherence and emigration in rat mesenteric venules

This study demonstrates that SC-41930, 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)-propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid, an orally active LTB₄ receptor antagonist, reduces LTB₄-induced leukocyte adhesion and emigration in rat mesenteric venules. The mesentery of Sprague-Dawley rats was prepared for intravital microscopic examination and venules of 25–35 m were chosen for evaluation. In control animals, LTB₄ (20nM) was superfused over the mesentery for 30 min. In the treatment group SC-41930 (5 M) was superfused for 30 min, followed by a 30 min superfusion with SC-41930 and LTB₄. The LTB₄-induced increase in leukocyte adherence and emigration in postcapillary venules was significantly attenuated by pretreatment with SC-41930. Other experiments demonstrated that platelet-activating-factor-induced leukocyte adherence was not affected by SC-41930. These results indicate that SC-41930 is a potent inhibitor of LTB₄-induced leukocyte-endothelial cell adhesive interactions in postcapillary venules.     

SC-41930, a leukotriene B4 receptor antagonist,inhibits 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE) binding to epidermal cells

SC-41930, 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)-propoxyl]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid, a potent leukotriene-B₄ (LTB₄) receptor antagonist, inhibitsin vivo 12-hydroxyei-cosatetraenoic acid (12-HETE)-induced neutrophil infiltration, suggesting a potential 12-HETE receptor antagonist effect, as well. Since 12-HETE is assumed to have a pathophysiological role in inflammatory skin diseases, and epidermal cells possess high affinity binding sites for 12(S)-HETE, we studied the effect of SC-41930 on 12(S)-HETE binding to the human epidermal cell line, SCL-II. SC-41930 antagonized the 12(S)-HETE binding to SCL-II cells with a Ki of 480 nM. This Ki value is similar to that obtained for the inhibition of LTB₄ binding to human neutrophils. Our results show that SC-41930, in addition to its LTB₄ receptor antagonist effect, exhibits 12-HETE receptor antagonist effect as well, and therefore may be of benefit in skin diseases with elevated 12-HETE levels.Dr. Kemény is on leave from the Department of Dermatology, Albert Szent-Györgyi Medical University, Szeged, Hungary, as a recipient of the Humboldt Fellowship, and his work was supported by the Alexander von Humboldt Foundation.     

Leukotriene B4-induced granulocyte trafficking in guinea pig dermis

Leukotriene B₄ (LTB₄) is a proinflammatory product of arachidonic acid metabolism that has teen implicated as a mediator in a number of inflammatory diseases. When injected intradermally into the guinea pig, LTB₄ elicits a dose-dependent migration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-1 -benzopyran-2-carboxylic acid, a first-generation LTB₄ receptor antagonist inhibitedthe chemotactic actions of LTB₄ when coadministered into the dermal site and when given orally with ED₅₀ values of 340 ng and 1.7 mg/kg, respectively. The secondgeneration LTB₄ receptor antagonists SC-50605 7-[3-[2(cyclopropylmethyl)-3-methoxy-4-(4-thiazolyl)phenoxy] propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid and SC-51146 7-[3-[2(cyclopropylmethyl)-3-methoxy-4-[(methylamino)carbonyl]phenoxy]propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran2-propanoic acid inhibited LTB₄-induced chemotaxis when coadministered with ED₅₀ values of 70 ng and 32 ng, respectively, and when given intragastrically with ED₅₀ values of 0.10 and 0.09 mg/kg, respectively. SC-41930, SC-50605, and SC-51146 had oral durations of action of 5.5, 15, and 21 h, respectively. These potent, LTB₄ receptor antagonists may well have application in the medical management of disease states such as asthma, rheumatoid arthritis, inflammatory bowel disease, contact dermatitis, and psoriasis, where LTB₄ is implicated as an inflammatory mediator.     

Comparative evaluation of arachidonic acid (AA)- and tetradecanoylphorbol acetate (TPA)-induced dermal inflammation

The effects of topical application of arachidonic acid (AA) or phorbol ester, tetradecanoylphorbol 13-acetate (TPA), on edema response, vascular permeability, MPO, NAG, and generation of eicosanoids were studied in two murine models of cutaneous inflammation. AA produced a short-lived edema response with a rapid onset that was associated with marked increases in levels of prostaglandins (PGE₂, 6-keto-PGF₁, PGF₂), thromboxane B₂ (TxB₂) and leukotriene B₄ (LTB₄), with smaller increases in levels of LTC₄. TPA produced a longer-lasting edema that was associated with marked influx of neutrophils and predominant formation of LTB₄ along with significant changes in levels of TxB₂. Circulating T lymphocytes have no apparent role in the acute inflammatory responses induced by either agent. Arachidonic acid-induced vascular permeability preceded the edema response and neutrophil influx, whereas TPA-induced vascular permeability paralleled the edema response and influx of neutrophils. Mast cells appear to be important in the complete expression of inflammatory response, i.e., edema, cellular influx, and vascular permeability induced by either AA or TPA, as these responses were blunted in mast cell-deficient mice. Inhibitors of CO or 5-LO attenuated inflammatory responses in both models. The LTB₄ receptor antagonist, SC-41930, inhibited the inflammatory response to TPA but had little effect on that initiated by AA. This suggests that LTB₄ is an important mediator in the phorbol ester-induced inflammatory response, whereas peptidoleukotrienes and prostaglandins regulate vascular permeability responses in the arachidonate model.     

A23187-induced pulmonary gas trapping and inflammation in the guinea pig

A brief A23187 aerosol exposure produced prolonged airway obstruction with granulocyte accumulation in conscious guinea pigs. Aminophylline, atropine, pyrilamine, salbutamol, SC-41930 (a leukotriene B₄ antagonist) and WEB 2086 (a platelet activating factor antogonist) were administered intravenously (i.v.) to evaluate their ability to prevent these changes. Inhaled salbutamol was also assessed. Aminophylline, atropine, and salbutamol (i.v. and aerosol) inhibited A23187-induced gas trapping (p<0.01). However, pyrilamine, SC-41930 and WEB 2086 did not influence this airway obstructive effect. Only atropine, inhaled salbutamol and SC-41930 inhibited the cell influx (p<0.01), while pyrilamine potentiated the inflammation (p<0.05). We conclude that A23187 produces a sustained bronchospasm and an intense granulocyte accumulation. The treatment agents tested differ considerably in their ability to alter A23187-induced airway obstruction and inflammation.     

Inflammatory mediator changes in cotton-top tamarins (CTT) after SC-41930 anti-colitic therapy

Use of the CTT model provides insight into the inflammatory mediator contribution in the pathogenesis of idiopathic colitis. To evaluate anti-colitic efficacy, the leukotriene B₄ receptor antagonist and anti-inflammatory agent, SC-41930, was administered (10 mg/kg BW by gavage BID) for 8 weeks to CTTs with histologically confirmed persistent and defined active colitis. The inflammatory mediators LTB₄, PGE₂, TXB₂, and PAF were assayed in colonic dialysate that was collected after 11/2 h from four CTTs pre-, mid-, and post-treatment, frozen at −70°C, and analyzed by RIA after HPLC purification. LTB₄ levels were lower at mid- and post-treatment and had little inter-animal variation post-treatment. PGE₂ and PAF levels were elevated during SC-41930 treatment, but there was a trend towards lower thromboxane B₂ levels. Reduced LTB₄ (PMN degranulation and chemotaxis) and increased PGE₂ (mucosal-protective effect) may, in part, explain the observed efficacy of SC-41930 in active tamarin colitis.

     

Antiinflammatory effects of second-generation leukotriene B4 receptor antagonist,SC-53228: Impact upon Leukotriene B4- and 12(R)-HETE- mediated events

Leukotriene B₄ (LTB₄) and 12(R)-hydroxyeicosatetraenoic acid [12(R)-HETE] are proinflammatory products of arachidonic acid metabolism that have been implicated as mediators in a number of inflammatory diseases. When injected intradermally into the guinea pig, LTB₄ and 12(R)-HETE elicit a dose-dependent migration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 {7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxyl]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid}, a first-generation LTB₄ receptor antagonist, inhibited the chemotactic actions of LTB₄ when given orally with an ED₅₀ value of 1.7 mg/kg. The second-generation LTB₄ receptor antagonist, SC-53228 [(+)-(S)-7-(3-{2-(cyclopropylmethyl)-3-methoxy-4-[(methylamino)carbonyl]phenoxy} propoxy)-3,4-dihydro-8-propyl-2H-1-benzopyran-2-propanoic acid], inhibited LTB₄-induced chemotaxis when given intragastrically with an ED₅₀ value of 0.07 mg/kg. Furthermore, SC-53228 inhibited 12(R)-HETE-induced granulocyte chemotaxis with an oral ED₅₀ value of 5.8 mg/kg. When dosed orally over a range of 0.03–100 mg/kg, SC-53228 gaveC _max plasma concentrations of 0.015–41.1g/ml. SC-53228 inhibited LTB₄-primed membrane depolarization of human neutrophils with an IC₅₀ value of 34 nM. As a potent LTB₄ receptor antagonist, SC-53228 may well have application in the medical management of disease states such as asthma, rheumatoid arthritis, inflammatory bowel disease, contact dermatitis, and psoriasis, in which LTB₄ and/or 12(R)-HETE are implicated as inflammatory mediators.     

Effect of a leukotriene B4 receptor antagonist on leukotriene B4-induced neutrophil chemotaxis in cavine dermis

Leukotriene B₄ (LTB₄) is a proinflammatory product of arachidonic acid metabolism that has been implicated as a mediator in a number of inflammatory diseases. When injected intradermally into the cavine, LTB₄ elicits a dose-dependent immigration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 {7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxy]-3,4-dihydro-8-propyl-2H-l-benzopyran-2-carboxylic acid, a potent LTB₄ receptor antagonist inhibited the chemotactic actions of LTB₄ when coadministered into the dermal site and when given intravenously or orally with ED₅₀ values of 200 ng, 0.5 mg/kg, and 0.6 mg/ kg respectively. This compound may well have application in disease states, such as inflammatory bowel disease and psoriasis, where LTB₄ is implicated as a proinflammatory mediator.     

Anti-inflammatory activity of the leukotriene B4 receptor antagonist,SC-41930, in colitic cotton-top tamarins

Cotton-top tamarins (CTTs) with histologically confirmed persistent and active colitis were given the leukotriene B₄ (LTB₄) receptor antagonist, SC-41930, (10 mg/kg BW, by gavage b.i.d.) for eight weeks. Anti-inflammatory activity was evaluated by colonic biopsy, stool consistency and the level of the lipid mediators LTB₄ and prostaglandin E₂ (PGE₂) in rectal dialysates. Stool consistency did not improve with treatment but did not worsen. Blood chemistry (ALT, AST, LDH) and hematological parameters neither showed any untoward effects of SC-41930 treatment nor was there any effect on body weight. In rectal dialysate LTB₄ levels were significantly reduced from pretreatment level of 4.87±1.46 ng/ml to 1.07±0.67 and 2.45±0.13 ng/ml at 4 and 8 weeks, respectively, and higher prostaglandin E₂ (PGE₂) over time. Histologically, 5/7 improved, 1/7 remained the same and 1/7 worsened.

Oral SC-41930 treatment was safe and associated with an anti-colitic effect. The reduced LTB₄ levels (affecting granulocyte degranulation and recruitment into tissues) and increased PGE₂ (perhaps exerting a mucosal protective effect) may, in part explain the observed efficacy of this compound in active tamarin colitis. Use of the CTT model could provide insight into the inflammatory mediator contribution to idiopathic colitis and serve as a useful bridge between preclinical pharmacology and the assessment of these compounds in the medical management of human inflammatory bowel disease.

     

Effect of the leukotriene B4 receptor antagonist,SC-41930, on experimental allergic encephalomyelitis (EAE) in the guinea pig

The accepted model for the human demyelinating disease, multiple sclerosis (MS), is experimental allergic encephalomyelitis (EAE). We assessed the ability of SC-41930(7-[3(4-acetyl-3-methoxy-2-propylphenoxy)-propoxy]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxyl acid), to modulate the symptoms of acute EAE generated in guinea pigs. Animals were pretreated with SC-41930 (20 mg/kg, i.p.) for two days followed by thrice-weekly maintenance. At day 52, a significant number of the SC-41930-treated animals were alive as compared to EAE alone. Control animals had an increase in body weight while EAE animals lost over 20% (p<0.5) of their body weight by day 18. SC-41930-treatment significantly reduced, but did not completely inhibit the cachectic response. The results indirectly implicate LTB₄ in the pathogenesis of EAE. Agents that modify this model be useful in the treatment of human MS.     

Are leukotrienes or PAF involved in hyperbaric oxygen toxicity?

Several very selective leukotriene inhibitors, and a PAF inhibitor, suitable forin vivo use, have been tested for their effects on hyperbaric oxygen toxicity.The leukotriene D₄ inhibitor, L660 771, and the 5-lipoxygenase pathway inhibitor L663 563, failed to affect convulsions or lung damage induced by hyperbaric oxygen (pressure range 515–615 kPa) in either rats or mice. The specific PAF antagonist L659 989 showed marginal protection against hyperoxic convulsions and did not alter pulmonary damage. The specific LTB₄ antagonist SC-41930 was very effective in inhibiting hyperbaric oxygen-induced convulsions in both rats and mice. SC-41930 also very significantly protected rats against pulmonary oxygen toxicity, but had only marginally significant effects on pulmonary protection in mice.     

Effect of Topically Applied Cyclooxygenase-2-Selective Inhibitors on Arachidonic Acid- and Tetradecanoylphorbol Acetate-Induced Dermal Inflammation in the Mouse

The topical effects of cyclooxygenase-2 (COX-2)-selective inhibitors, flosulide (CGP 28238), L-745,337 and SC-57,666 were examined in AA- and TPA-induced ear dermal inflammation in the mouse. The doses that caused 50% inhibition in AA edema (ED₅₀) were 2.4, 0.45 and 0.35 mg/ear for flosulide, L-745,337 and SC-57,666, respectively. The respective ED_50s in TPA-edema were 1, 0.45 and 0.14. Indomethacin and zileuton showed higher activity than the COX-2-selective inhibitors in both models. Flosulide and L-745,337 inhibited the AA-induced increase in 6-keto-PGF₁, while SC-57,666 was inactive. 80% inhibition was seen with indomethacin while zileuton had no effect. COX-2 selective inhibitors and indomethacin had no effect on LTB₄ levels, while zileuton produced a 50% inhibition. The TPA-induced increase in 6-keto-PGF₁ was greatly inhibited by all COX-2 inhibitors while LTB₄was potentiated by both flosulide and L-745,337. Indomethacin inhibited 6-keto-PGF₁ and zileuton reduced 6-keto-PGF and strongly reduced LTB₄. The neutrophil influx induced by AA was lower than that of TPA. Myeloperoxidase (MPO) levels were lowered by flosulide and L-745,337 but not by SC-57,666. TPA-induced MPO increase was decreased by all COX-2 inhibitors. Indomethacin and zileuton had similar effect on AA and TPA-induced increase in MPO. The results indicate that COX-2-selective inhibitors showed lower topical anti-inflammatory activity than indomethacin or zileuton.     

Arachidonic Acid (AA) and Tetradecanoylphorbol Acetate (TPA) Exert Systemic Effects When Applied Topically in the Mouse

We studied the systemic actions of topically applied arachidonic acid (AA) and tetradecanoylphorbol acetate (TPA) in the mouse. AA or TPA-induced ear edema, increases in vascular permeability, eicosanoid levels, neutrophil and mononuclear influx were determined in both phlogogen-treated and contralateral vehicle-treated ear of each mouse and were compared with vehicle-treated ears from control mice. Edema and vascular permeability increases appeared only in AA- or TPA-applied ears. Moreover, in contralateral ears from AA-treated mice an increase in 6-keto-PGF₁ and LTB₄ was found. Only LTB₄ increased in the contralateral ear after TPA. Contralateral ears from AA- or TPA-treated mice also showed a significant increase in MPO levels. The increased levels of 6-keto-PGF₁ but not those of LTB₄ in contralateral ears were reduced by indomethacin applied simultaneously with the phlogogen. AA also increased plasma and serum levels of LTB₄, but not those of 6-keto-PGF₁. In contrast, TPA increased plasma and serum levels of 6-keto-PGF₁ and LTB₄. The results show that both AA and TPA applied topically exert systemic effects.     

Induction of colitis in rats by 2-2′-azobis(2-amidinopropane) dihydrochloride

Reactive oxygen metabolites (ROM) may play a role in the pathophysiology of inflammatory bowel disease (IBD) and ischemia-reperfusion-induced intestinal injury. Although there are many reports of intestinal mucosal injury associated with neutrophil-derived ROM, free radicals themselves have not been reported to induce intestinal mucosal injury. We administered intrarectally 2,2-azobis(2-amidinopropane) dihydrochloride (AAPH) to rats, an azo compound that generates free radicals in vitro. Acute mucosal injury was assessed histologically by light microscopy and biochemically by myeloperoxidase (MPO) activity. Intrarectal administration of AAPH (60, 90, 150 mg/kg) caused erythema, edema, and histologically verifiable mucosal inflammation. MPO activity was increased 9- to 18-fold above the control level. The levels of thiobarbituric acid (TBA) reactants and sulfhydryls (SH) were significanty (P<0.01) increased and decreased, respectively, by 90 mg/kg AAPH. Sulfasalazine, 5-aminosalicylic acid, the LTB₄ receptor antagonist SC-41930, and the antioxidant glutathione prevented the inflammation. This model of mucosal inflammation may be useful in evaluating new therapeutic agents for the treatment of IBD.     

Leukotriene B4 and inflammatory disease

Polymorphonuclear leukocytes (PMNL) are prominent at sites of acute inflammation. Their infiltration is stimulated under pathological conditions by a variety of agents which include bacteria, immune complexes and complement derived chemotactic peptides. Recently attention was focussed on the 5-lipoxygenase product leukotriene B₄ (LTB₄) which has been demonstrated to induce the key features associated with an acute inflammatory reaction. However, evidence supporting a pro-inflammatory role for LTB₄, and therefore the anti-inflammatory efficacy of 5-lipoxygenase inhibitors, is largely circumstantial. Moreover, there are concerns that other chemotactic factors, notably C₅a, may compensate for the absence of LTB₄. Here we challenge this view and, on the basis of recent experimental and clinical data suggest that LTB₄ does not simply duplicate the activity of C₅a. Instead we propose that their predominant site(s) of action differ in such a way that they may synergise in mediating PMNL recruitment.     

Modulation of mouse ear edema by cyclooxygenase and lipoxygenase inhibitors and other pharmacologic agents

Inhibitors of arachidonic acid (AA) metabolism and other pharmacologic agents were evaluated against ear edema produced in mice by tetradecanoylphorbol acetate (TPA) or AA. Drugs were administered orally and topically either 30 min prior to AA or 30 min after TPA, except for steroids which were administered 2.5–3 hr prior to AA. Several cyclooxygenase (CO) inhibitors including indomethacin, aspirin, piroxicam and timegadine were without effect when administered orally against either irritant; the same drugs inhibited TPA edema when they were administered topically. Mixed CO/lipoxygenase (LO) inhibitors, phenidone and BW755C, were active orally against AA edema (ED₅₀s of 84 and 65 mg/kg, respectively) and against TPA edema (ED₅₀s of 235 and 88 mg/kg, respectively). Phenidone was more active topically against AA edema (ED₅₀, 0.2 mg/ear) than (ED₅₀, 2.8 mg/ear); however, BW755C was more active topically against TPA edema (ED₅₀, 0.2 mg/ear) than phenidone (ED₅₀, 0.6 mg/ear). Methylprednisolone was very effective in the AA (oral ED₅₀, 17 mg/kg; topical ED₅₀,>1 mg/ear) and TPA models (oral ED₅₀, 4.3 mg/kg; topical ED₅₀, 0.03 mg/ear. MK-447 was topically and orally effective only in the TPA model. Not surprisingly, drugs were more effective models were somewhat selective for CO and CO/LO inhibitors; however, dapsone was orally effective in the ear models, and a number of mediator antagonists and CNS drugs, especially anti-psychotics, were topically active primarily against TPA edema. These models may be useful for the detection ofin vivo activity of CO/LO or 5-LO inhibitors.     

Function and stimulus-specific effects of phorbol 12-myristate 13-acetate on human polymorphonuclear neutrophils: Autoregulatory role for protein kinase C in signal transduction

Exposure of polymorphonuclear neutrophils (PMNs) to phorbol 12-myristate 13-acetate (PMA) resulted in a concentration-dependent (1–10 ng/ml) inhibition of granule exocytosis induced with the receptor-specific ligands,N-formylmethionyl-leucyl-phenylalanine (FMLP), pepstatin A, 5(S),12(R)-dihydroxy-6, 14-cis-8, 10-trans-eicosatetraenoic acid (LTB₄), and acetyl-sn-glyceryl-3-phosphorylcholine (AGEPC), PMA exerted a marginal inhibitory effect on calcium ionophore A23187-induced PMN degranulation, and the PMA analog, 4-phorbol 12, 13-didecanoate (4-PDD), was inactive. However, PMA potentiated AGEPC, pepstatin A, FMLP, LTB₄, and A23187-stimulated Superoxide anion (O ₂ ^– ) production. The mobilization of intracellular sequestered calcium (Ca²⁺) by the receptor-specific ligands, as reflected by a rise in the cytosolic-free Ca²⁺ concentration ([Ca²⁺]_i) in PMNs loaded with the CA²⁺-sensitive dye, Fura-2, was suppressed by PMA. A protein kinase C (PKC) inhibitor, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7) reversed the PMA-mediated inhibition of PMN degranulation and intracellular CA²⁺ mobilization. However, another, but less potent PKC inhibitor,N-(2-guanidinoethyl)-5-isoquinolinesulfonamide (HA 1004), had no effect on the inhibition of PMN activation by PMA.