Peter Norman

BAY u9773, a novel antagonist of cysteinyl-leukotrienes with activity against two receptor subtypes

The effects of BAY u9773 (6(R)-(4-carboxyphenylthio)-5(S)-hydroxy-7(E),9(E), 11(Z),14(Z)-eicosatetraenoic acid), a cysteinyl-leukotriene analogue, were investigated on a variety of smooth muscle preparations in order to determine its profile as a cysteinyl-leukotriene receptor antagonist. The tissues were contracted with leukotriene C4 or leukotriene D4 and their receptor characteristics defined as either typical or atypical according to the activity or inactivity, respectively, of the selective antagonists ICI 198615, MK 571 and SKF 104353. BAY u9773 antagonised typical cysteinyl-leukotriene receptors with pA2 (or pKB) values in the range 6.8-7.4 and also antagonised atypical receptors with pA2 values in the range 6.8-7.7. However, BAY u9773 had no effect at 10(-6) M against a selection of non-leukotriene stimuli in the same preparations. BAY u9773 competitively displaced [3H]leukotriene D4 binding to guinea-pig lung homogenate, with a pKi of 7.0 +/- 0.1. In the guinea-pig lung strip, BAY u9773 was found to be inactive at 10(-6)M against leukotriene C4- and leukotriene D4-induced contractions, which may suggest the existence of a third type of cysteinyl-leukotriene receptor. These data demonstrate that BAY u9773 is a selective cysteinyl-leukotriene receptor antagonist with comparable activity at both typical and atypical receptors and as such represents a valuable tool for the study of cysteinyl-leukotriene receptors.

BAY u3405, a potent and selective thromboxane A2 receptor antagonist on airway smooth muscle in vitro

1 BAY u3405 (3(R)‐[[(4‐fluorophenyl) sulphonyl]amino]‐1,2,3,4‐tetrahydro‐9H‐carbazole‐9‐propanoic acid) has been evaluated on airway smooth muscle, from a number of species including man, for its thromboxane A2 (TXA2) antagonist activity. 2 BAY u3405 was a potent, and competitive, antagonist of the TXA2‐mimetic U46619‐induced contractions of human, guinea‐pig, rat and ferret airway smooth muscle with pA2 values between 8.0 and 8.9 and with no inherent contractile activity (10−9–10−4 m). 3 The TXA2 antagonist activity of BAY u3405 was stereoselective. Its (S)‐enantiomer, BAY u3406, was approximately 50 fold less effective against U46619 on guinea‐pig and human airway smooth muscle. 4 BAY u3405 also competitively antagonized contractions of guinea‐pig airway smooth muscle induced by prostaglandin D2 (PGD2) or its metabolite 9α, 11β‐PGF2. On human and ferret airway smooth muscle it abolished contractions induced by PGD2, PGF2α and 16, 16‐dimethyl‐PGE2. 5 A high concentration (10−6 m) of BAY u3405 had no effect on the contraction, or relaxation, of airway smooth muscle induced by a range of other agonists, nor did BAY u3405 have any effect on other prostanoid receptor types (DP, EP1, EP2, FP or IP). 6 BAY u3405, in contrast to some other TXA2 antagonists, is a potent and selective antagonist on a wide range of airways including human. This high affinity, and the oral activity of the compound described elsewhere, suggest it may be an appropriate tool to investigate the role of prostanoids in airway diseases such as asthma.

The inhibition of [3H]leukotriene D4 binding to guinea-pig lung membranes. The correlation of binding affinity with activity on the guinea-pig ileum

Guinea-pig lung membranes contain high affinity (KD = 0.8 nM) binding sites for [3H]leukotriene D4 [( 3H]LTD4). The binding is inhibited by leukotriene antagonists, such as ICI 198615 and SK&F 104353, in a manner consistent with the Law of Mass Action at a single site. It is also inhibited by a range of leukotriene analogues in a dose-related manner. Inhibition by some of these e.g. LTC4 suggests that the [3H]LTD4 binding sites are heterogeneous. The binding affinity of the leukotriene analogues is significantly correlated (P less than 0.001) to their spasmogenic activity on guinea-pig ileum but not on guinea-pig lung strip. The binding affinity of the leukotriene antagonists is also correlated to their antagonist activity on guinea-pig ileum (P less than 0.05) but not on guinea-pig lung. These results indicate that the [3H]LTD4 binding site in guinea-pig lung is similar to the leukotriene receptor activated by LTD4 and LTE4 on guinea-pig ileum. The contractile response of guinea-pig lung strips to leukotrienes, in the presence of indomethacin, is mediated by a distinct type of leukotriene receptor.

Characterisation of the peptido-leukotriene receptor PL2 on the ferret spleen strip.

The peptido-leukotriene receptor(s) (PL) on the ferret isolated spleen strip have been characterised by functional studies using the naturally occurring leukotrienes (LTs), a range of structurally distinct PL antagonists, and by ligand binding studies. LTB4 (0.01-10 microM) was inactive on ferret spleen whereas LTC4, LTD4 and LTE4 produced concentration-related contractions with maximal responses, relative to noradrenaline, of 57% (EC50 0.28 microM), 60% (EC50 0.5 microM) and 7% respectively. The leukotriene responses were unaltered by L-serine borate, L-cysteine, indomethacin, phentolamine, propranolol, mepyramine, methysergide or atropine, suggesting that the peptido-leukotrienes were acting through distinct PL receptors. The PL1 antagonists, FPL 55712 (0.01-10 microM), ICI 198615 (10 microM), SK&F 104353 (10 microM) and MK541 (10 microM) were all inactive against LTC4- or LTD4-induced contractile responses. LTE4 was a partial agonist with respect to LTC4 and LTD4 with pKB values of 5.8 and 5.5 respectively. Nifedipine (0.1 microM) produced a rightward shift of the concentration-response curves to both LTC4 and LTD4 and depressed their maximal responses. An unacceptably high level of non-specific binding of [3H]LTD4 to membrane preparations of ferret spleen prevented characterisation of this receptor by ligand binding. These results suggest that the ferret spleen has a homogeneous population of a PL receptor type which is insensitive to existing PL1 receptor antagonists. The functional characteristics of this PL receptor type are similar to those of the PL2 receptor on other tissues. The absence of PL1 receptors on this tissue makes it particularly useful in identifying new and selective drug tools for the PL2 receptor.

The thromboxane receptors of rat and guinea-pig lung.

The thromboxane (TXA2) receptors on rat and guinea-pig lung strips were compared using TXA2 agonists and TXA2 receptor antagonists. On rat lung strip several TXA2 mimetics were full agonists whilst the primary prostanoids lacked agonist activity. On guinea-pig lung strip the same agonists displayed markedly different efficacies. Both preparations contained homogeneous populations of TXA2 receptors as evidenced by BAY u3405 giving comparable pA2 values against four TXA2 mimetics. However, the observed pA2s of nine different TXA2 receptor antagonists, determined against U46619, did not correlate between the two preparations. These results point to the existence of TXA2 receptor subtypes.

A new structural analogue antagonist of peptido-leukotrienes. The discovery of bay x7195

Abstract Systematic modification of LTD4 has led to the discovery of BAY x7195, a potent, selective, orally-active peptido-leukotriene antagonist.

Characterisation of leukotriene receptors on rat lung strip

The leukotriene receptor(s) present on rat lung strip have been characterised using the natural agonists, a selective mimetic, and potent (cysLT1) selective leukotriene receptor antagonists. Leukotriene C4 and leukotriene D4 displayed comparable contractile potencies whilst leukotriene E4 was less potent. However, both leukotriene D4 and leukotriene E4 were found to be partial agonists relative to leukotriene C4. Responses to all three leukotrienes were competitively antagonised by ICI 198615 (1-((2-methoxy-(4-phenylsulfonyl)-aminocarbonyl)-phenyl) methyl)-1H-indazol-6-yl) carbonic acid cyclopentyl ester), SK&F 104353 (2-(R)-hydroxy-3(S)-(2-carboxyethylthio)-3-(2-[8-phenyloctyl ]- phenyl)propanoic acid, and MK571 (+/-(E)-3-[3-[2-(7-chloro-2-quinolin-yl)ethenyl]-phenyl)- ([3-(dimethylamino)-3-oxopropyl]thio]methyl]thio]thio]propanoic acid) with comparable affinities irrespective of the agonist used. This indicates that rat lung contains a homogeneous population of leukotriene receptors and that they are of the CysLT1 type.

Antagonism of leukotriene responses in human airways by BAY X7195

Contractions induced by leukotriene and anti-IgE (sheep antiserum to human IgE) were antagonized by pretreatment of human airways with the cysteinyl leukotriene receptor antagonist BAY x7195 ((4S)-[4-carboxyphenylthio]-7-[4-(4-phenoxybutoxy)-phenyl]-h ept-5-(z)- enoic acid). However, this receptor antagonist did not inhibit either leukotriene D4- or leukotriene C4-induced contractions in human pulmonary veins. The pA2 value for BAY x7195 in human airways against leukotriene D4 was 7.83 +/- 0.16 with a slope of 1.07 +/- 0.15 (means +/- S.E.M; n = 11). The IC50 value for BAY x7195 in human airways contracted with anti-IgE was 0.31 +/- 0.08 microM (n = 11). These results were comparable to those obtained with ICI 204,219 (4-(5-cyclopentyl-oxycarbonylamino-1-methylindol-3-ylmeth yl)-3-methoxy-N-otolyl - sulfonylbenzamide). These data demonstrate that BAY x7195 is a potent selective leukotriene receptor antagonist which may block allergic reactions in the lung.