Carlos R. Roman

ARL 17477, a selective nitric oxide synthase inhibitor, with neuroprotective effects in animal models of global and focal cerebral ischaemia

In the present studies, we have evaluated the effects of N-[4-(2-¿[(3-Chlorophenyl)methyl]amino¿ethyl)phenyl]-2-thiophenecarbo ximidamide dihydrochloride (ARL 17477) on recombinant human neuronal NOS (nNOS) and endothelial NOS (eNOS). We then carried out pharmacokinetic studies and measured cortical nitric oxide synthase (NOS) inhibition to determine that the compound crossed the blood brain barrier. Finally, the compound was evaluated in a model of global ischaemia in the gerbil and two models of transient focal ischaemia in the rat. The IC(50) values for ARL 17477 on human recombinant human nNOS and eNOS were 1 and 17 microM, respectively. ARL 17477 (50 mg/kg i.p.) produced a significant reduction in the ischaemia-induced hippocampal damage following global ischaemia when administered immediately post-occlusion, but failed to protect when administration was delayed until 30 min post-occlusion. In the endothelin-1 model of focal ischaemia, ARL 17477 (1 mg/kg i.v.) significantly attenuated the infarct volume when administered at either 0, 1 or 2 h post-endothelin-1 (P<0.05). In the intraluminal suture model, ARL 17477 at both 1 and 3 mg/kg i.v. failed to reduce the infarct volume measured at 1, 3 or 7 days post-occlusion. These results demonstrate that ARL 17477 protects against global ischaemia in gerbils and provides some reduction in infarct volume following transient middle cerebral artery occlusion in rats, indicating that nNOS inhibition may be a useful treatment of ischaemic conditions.

LY393615, a novel neuronal Ca2+ and Na+ channel blocker with neuroprotective effects in models of in vitro and in vivo cerebral ischemia

In the present studies we have examined the effects of a new calcium channel blocker, LY393615 ((N-Butyl-[5,5-bis-(4-fluorophenyl)tetrahydrofuran-2-yl]methylamine hydrochloride, NCC1048) in a model of hypoxia-hypoglycaemia in vitro and in a gerbil model of global and in two rat models of focal cerebral ischaemia in vivo. Results indicated that LY393615 protected against hypoxia-hypoglycaemic insults in brain slices and also provided significant protection against ischaemia-induced hippocampal damage in gerbil global cerebral ischaemia when dosed at 10, 12.5 (P<0.05) or 15 mg/kg i.p. (P<0.01) 30 min before and 2 h 30 min after occlusion. The compound penetrated the brain well after a 15 mg/kg i.p. dose and had a half-life of 2.5 h. In further studies LY393615 was protective 1 h post-occlusion when administered at 15 mg/kg i.p. followed by 2 doses of 5 mg/kg i.p. 2 and 3 h later. LY393615 dosed at 15 mg/kg i.p. followed by 2 further doses of 5 mg/kg i.p. (2 and 3 h later) also produced a significant reduction in the infarct volume following Endothelin-1 (Et-1) middle cerebral artery occlusion in the rat when administration was initiated immediately (P<0.01) or 1 h (P<0.05) after occlusion. The compound was also evaluated in the intraluminal monofilament model of focal ischaemia. The animals had the middle cerebral artery occluded for 2 h, and 15 min after reperfusion LY393615 was administered at 15 mg/kg i.p. followed by 2 mg/kg/h i.v. infusion for 6 h. There was no reduction in infarct volume using this dosing protocol. In conclusion, in the present studies we have reported that a novel calcium channel blocker, LY393615, with good bioavailability protects against neuronal damage caused by hypoxia-hypoglycaemia in vitro and both global and focal cerebral ischaemia in vivo. The compound is neuroprotective when administered post-occlusion and may therefore be a useful anti-ischaemic agent.

Pulmonary actions of LY255283, a leukotriene B4 receptor antagonist

The actions of LY255283, a leukotriene (LT) B4 receptor antagonist, were examined on guinea pig lung. LTB4 and LY255283 displaced [3H]LTB4 from its binding site on lung membranes with pKi values of 9.9 and 7.0, respectively. In the functional correlate of the binding studies, LY255283 competitively reduced contractile responses of lung parenchyma to LTB4 (pA2 = 7.2). LTB4 produced airway obstruction which was reduced by LY255283 administered i.v. (ED50 = 2.8 mg/kg) or orally (ED50 = 11.0 mg/kg). Contractile responses to histamine, LTD4 and the thromboxane mimetic, U46619, were not reduced by LY255283. The compound also did not inhibit cyclooxygenase or 5-lipoxygenase enzymes. We conclude that LY255283 selectively antagonized pharmacologic responses to LTB4 on lung tissue and appears to be a useful tool to investigate the role of LTB4 in pulmonary disease.

Differential relaxant responses of guinea-pig lung strips and bronchial rings to sodium nitroprusside : a mechanism independent of cGMP formation

The biochemical mechanism subserving smooth muscle relaxant effects of sodium nitroprusside was examined on U46619, 9, 11‐dideoxy‐9α, 11α‐methanoepoxy PGF2α, precontracted guinea‐pig lung strips and hilar bronchial rings.

Structure-activity relationships (SAR) of the 3-alkyl substituents among a series of hydroxyacetophenone leukotriene antagonists

LY171883 is an orally active antagonist of leukotriene (LT) D4 and LTE4. A series of related compounds varying the position and nature of the alkyl side chain were synthesized and evaluated for their ability to block LTD4-induced contraction of guinea pig ileum. Maximal activity was obtained with n-propyl, n-butyl, and n-pentyl substituents with slightly reduced activity for longer side chains. Polar groups on the side chain substantially reduced activity. Thus, it appears that the leukotriene receptor site requires a nonpolar alkyl group of moderate size at the 3-position on this type of receptor antagonist.

Contraction of guinea pig inferior vena cava by eicosanoids

SummaryGuinea pig inferior vena cava contracted in response to leukotriene (LT)C4, LTD4, LTE4 U46619, phenylephrine, histamine, and KCl. Although LTC4, LTD4, and U46619 were the most potent agonists, active tension generated by these eicosanoids was only about half that of histamine or KCl. LTE4 and phenylephrine were marginally active. Biochemical analysis showed vena cava able to convert about 23% LTC4 to LTD4 and LTE4 in 45 min. Pretreatment with acivicin prevented this by abrogating conversion of LTC4 to LTD4. A subthreshold concentration of LTE4 reduced responses to LTC4 and LTD4. LY171883 and WY-48252 competitively antagonized LTD4-induced contractions of vena cava. In contrast, these antagonists blocked contractions to LTC4 in a biphasic manner. Lower segments of the LTC4 concentration-response curves were less affected than the upper portion suggesting the possibility of 2 LTC4 receptor subtypes. Our results indicate that LTE4 is a weak or partial agonist in this tissue and furthermore they suggest a lack of high affinity receptors for LTE4 favoring LTC4 and LTD4. Indomethacin did not influence contractions to the leukotrienes or histamine. However, the response to U46619 was greatly enhanced suggesting release of a vasodilator prostaglandin as part of the overall response of the vena cava to the thromboxane A2 mimetic.

QUINOLINE-SUBSTITUTED DIHYDROINDOLES AS CYSLT1 (LTD4 RECEPTOR) ANTAGONISTS

Abstract A series of quinoline-substituted dihydroindoles has been synthesized and evaluated as antagonists of the cysLT 1 receptor. This series, exemplified by 2 (LY302905; pKi = 8.3 for inhibition of binding of 3 H-LTD 4 to guinea pig lung membranes), represents reduced analogues of the corresponding indoles that were previously shown to be potent, orally active cysLT 1 receptor antagonists. These dihydroindole compounds generally displayed increased in vitro and in vivo (oral) activity.

Leukotriene (LT) receptor antagonists. Heterocycle-linked tetrazoles and carboxylic acids. LY203647

LY171883, 1-[2-hydroxy-3-propyl-4-[4-(1H-tetrazol-5-yl)butoxy]phenyl] ethanone, is an orally active antagonist of LTD4- and LTE4-induced responses in a variety of test systems. We prepared a new series of LT antagonists based on a proposed model of LY171883 binding to the LTE4 receptor in which then-propyl and tetrazole moieties of LY171883 occupy those parts of the receptor to which the C1-C5 chain and the cysteinyl carboxyl of LTE4 bind, respectively. The new compounds have an acidic function corresponding to the glycine carboxyl of LTD4 linked through a heterocyclic group which is proposed to bind to the LTD4 receptor where the cysteinyl glycine amide bond of LTD4 binds. LY203647, 1-[2-hydroxy-3-propyl-4-[4-[2-[4-(1H-tetrazol-5-yl) butyl]-2H-tetrazol-5-yl]butoxy]phenyl] ethanone, showed good LTD4 antagonist activity with a suitable pharmacologic and toxicologic profile and has been chosen for clinical evaluation.

2-alkyl-4-ethyl-5-[6-methyl-6-(2H-tetrazol-5-yl)heptyloxy]phenol leukotriene B4 receptor antagonists

A series of 2-n-alkyl-4-ethyl-5-[6-methyl-6-(2H-tetrazol-5-yl)heptyloxy]phenols were prepared and shown to be potent leukotriene B4 (LTB4) receptor antagonists. They bound to the human neutrophil and guinea pig lung LTB4 receptors with high affinity. Each compound was also shown to be effective at antagonizing the effects of LTB4-induced integrin up-regulation on human neutrophils and on LTB4-mediated contraction of guinea pig lung parenchyma.

Leukotriene Receptor Antagonism and Augmentation of β‐Receptor‐mediated Events by LY1718831

Abstract— LY171883, (1‐[2‐hydroxy‐3‐propyl‐4‐((4(1H‐tetrazol‐5‐yl)butoxy)phenyl]ethanone), a leukotriene (LT) D4/E4 receptor antagonist, was assessed in comparison with two well known phosphodiesterase inhibitors, isobutylmethyl‐xanthine (IBMX) and theophylline, for its ability to augment β‐receptormediated responses. Relaxation of carbachol‐contracted guinea‐pig trachea by isoprenaline was enhanced by the three agents in a dose‐dependent manner. A two‐fold enhancement of isoprenaline‐induced smooth muscle relaxation was produced by 2.5 μM IBMX, 28 μM LY171883, or 140 μM theophylline. Similar concentrations of IBMX or theophylline did not antagonize LTE4‐induced tracheal contractions; LY171883 totally inhibited the response and had significant LTE4 receptor antagonist activity even at 10‐fold lower concentrations. Antigen‐induced release of histamine and LTC4 from guinea‐pig lung was reduced by isoprenaline. Prior treatment with LY171883, IBMX, or theophylline did not enhance this action. Isoprenaline reduced histamine‐induced bronchospasm in anaesthetized guinea‐pigs. LY171883, 30 mg kg−1, or IBMX, 1 mg kg−1, did not affect the isoprenaline‐induced decrease in the histamine response. IBMX, 3 mg kg−1, and theophylline, 30 mg kg−1, augmented the isoprenaline‐induced bronchodilation. LTE4‐induced bronchoconstriction was not affected by IBMX or theophylline whereas LY171883 antagonized this response at doses as low as 3 mg kg−1. Therefore, in both in‐vitro and in‐vivo test systems, LY171883 functioned primarily as a leukotriene receptor antagonist with minimal pharmacological activity attributable to its ability to potentiate isoprenaline.