Matthew Joseph Fisher

p38 mitogen-activated protein kinase phosphorylates cytosolic phospholipase A2 (cPLA2) in thrombin-stimulated platelets. Evidence that proline-directed phosphorylation is not required for mobilization of arachidonic acid by cPLA2

The Ca2+-sensitive 85-kDa cytosolic phospholipase A2 (cPLA2) is responsible for thrombin-stimulated mobilization of arachidonic acid for the synthesis of thromboxane A2 in human platelets. We have previously shown that thrombin activates p38 kinase, a recently discovered new member of the mitogen-activated protein kinase family (Kramer, R. M., Roberts, E. F., Strifler, B. A., and Johnstone, E. M. (1995) J. Biol. Chem. 270, 27395-27398) and also induces phosphorylation of cPLA2, thereby increasing its intrinsic catalytic activity. In the present study we have examined the role of p38 kinase in the phosphorylation and activation of cPLA2 in stimulated platelets. We have observed that activation of p38 kinase accompanies receptor-mediated events in platelets and coincides with cPLA2 phosphorylation. Furthermore, in the presence of inhibitors of p38 kinase, the proline-directed phosphorylation of cPLA2 was completely blocked in platelets stimulated with the thrombin receptor agonist peptide SFLLRN and was suppressed during the early (up to 2 min) phase of platelet stimulation caused by thrombin. Unexpectedly, we found that prevention of proline-directed phosphorylation of cPLA2 in stimulated platelets did not attenuate its ability to release arachidonic acid from platelet phospholipids. We conclude that: 1) cPLA2 is a physiological target of p38 kinase; 2) p38 kinase is involved in the early phosphorylation of cPLA2 in stimulated platelets; and 3) proline-directed phosphorylation of cPLA2 is not required for its receptor-mediated activation.

Small-molecule direct thrombin inhibitors

Because of its central role in the control of thrombus formation, the trypsin-like serine protease thrombin is an important target for therapeutic intervention in thrombotic disease. The patent literature from the past several years illustrates that the design of low molecular weight, active-site thrombin inhibitors has followed several major themes. These include tripeptides and peptide mimimics of the cleavage site of fibrinogen, analogues of tertiary arginine amides, and non-peptide inhibitors derived from screening and rational design. This report reviews the concurrent development of these approaches.

Melanocortin - 4 Receptor Agonists for the Treatment of Obesity

The melanocortin family of receptors (MC 1-5R) and their endogenous peptide ligands (alpha, beta, gamma- MSH and ACTH) have been implicated in the control of a wide variety of behavioral and physiological functions including the homeostatic control of food intake and body weight. In rodent models, melanocortin agonists including the nonselective peptide MTII have been shown to decrease food intake and body weight while antagonists such as SHU9119 and AGRP have been shown to stimulate food intake and increase body weight. Deletion of either the MC3R or MC4R in mice was found to be associated with obesity although hyperphagia was only observed in the MC4R deficient mice. Similarly in humans, inactivating mutations of the MC4R have been found in as many as six percent of obese individuals. The suggestion from these findings that activation of MC4Rs would have an anorectic effect in humans has resulted in efforts to produce selective agonists for the treatment of obesity. Over the past decade, efforts to develop MC4R selective small molecule and peptide agonists have been met with fractional success. Many small molecule agonists have been identified; however, few have been shown to have activity in vivo. While their use as therapeutics may have limitations, selective and potent peptide agonists have been shown by several investigators to decrease food intake and body weight in rodent models. The subject of the current review is to examine the progress made to date on producing both small molecule and peptide MC4R agonists as potential therapeutics for obesity.

1,2-Disubstituted indole, azaindole and benzimidazole derivatives possessing amine moiety : A novel series of thrombin inhibitors

A novel series of 1,2-disubstituted indole, azaindole and benzimidazole derivatives possessing an amine moiety was identified as thrombin inhibitors. An indole with basic diamine moieties (12a) was the most potent thrombin inhibitor in the series with Kass= 197 x 10(6) L/mol.

Identification and Characterization of Novel Microsomal Prostaglandin E Synthase-1 Inhibitors for Analgesia

Prostaglandin (PG) E2 plays a critical role in eliciting inflammation. Nonsteroidal anti-inflammatory drugs and selective inhibitors of cyclooxygenase, which block PGE2 production, have been used as key agents in treating inflammation and pain associated with arthritis and other conditions. However, these agents have significant side effects such as gastrointestinal bleeding and myocardial infarction, since they also block the production of prostanoids that are critical for other normal physiologic functions. Microsomal prostaglandin E2 synthase-1 is a membrane-bound terminal enzyme in the prostanoid pathway, which acts downstream of cyclooxygenase 2 and is responsible for PGE2 production during inflammation. Thus, inhibition of this enzyme would be expected to block PGE2 production without inhibiting other prostanoids and would provide analgesic efficacy without the side effects. In this report, we describe novel microsomal prostaglandin E2 synthase-1 inhibitors that are potent in blocking PGE2 production and are efficacious in a guinea pig monoiodoacetate model of arthralgia. These molecules may be useful in treating the signs and symptoms associated with arthritis.

3-Phenyl-5-isothiazole carboxamides with potent mGluR1 antagonist activity

The disclosed 3-phenyl-5-isothiazole carboxamides are potent allosteric antagonists of mGluR1 with generally good selectivity relative to the related group 1 receptor mGluR5. Pharmacokinetic properties of a member of this series (1R,2R)-N-(3-(4-methoxyphenyl)-4-methylisothiazol-5-yl)-2-methylcyclopropanecarboxamide (14) are good, showing acceptable plasma and brain exposure after oral dosing. Oral administration of isothiazole 14 gave robust activity in the formalin model of persistent pain which correlated with CNS receptor occupancy.

Fused bicyclic Gly-Asp β-turn mimics with potent affinity for GPIIb-IIIa. Exploration of the arginine isostere

6-[4-Amidinobenzoyl]amino]-tetralone-2-acetic acid is a potent antagonist of GPIIb-IIIa. Substitution in the meta position of the benzamidine, or replacement with a heteroaryl amidine was tolerated in this series. Use of an acyl-linked 4-alkyl piperidine as an arginine isostere also provided active compounds. Compounds from this series provided substantial systemic exposure in the rat following oral administration.

Dihydroisoquinolone RGD mimics. Exploration of the aspartate isostere

Abstract Disubstituted 3,4-dihydroisoquinolones that contain an ether-linked benzamidine at C6 and a β-substituted aspartate mimic at C2 offer enhanced affinity for GPIIb–IIIa relative to the non-substituted isoquinolone propionate. Alkyl substituents afforded a 10-fold increase in intrinsic activity while aryl substituents yielded a 40-fold improvement.

An acyliminium ion approach towards the synthesis of β-substituted 3,4-dihydroisoquinolone propionates

A variety of α-methoxy amides were prepared either by selective reduction of the exocyclic carbonyl of a 2-acyl-3,4-dihydroisoquinolone and subsequent trapping of the resultant α-hydroxy amide with acidic methanol or by reacting the sodium salt of 3,4-dihydroisoquinolone with a functionalized α-chloro ether. These intermediates were reacted with 1-tert-butoxy-1-tert-butyldimethylsiloxy ethene in the presence of BF3·Et2O providing access to β-substituted isoquinolone propionates in good yield.

Characterization of 3,3-dimethyl substituted N -aryl piperidines as potent microsomal prostaglandin E synthase-1 inhibitors

Here we report on novel, potent 3,3-dimethyl substituted N-aryl piperidine inhibitors of microsomal prostaglandin E synthases-1(mPGES-1). Example 14 potently inhibited PGE2 synthesis in an ex vivo human whole blood (HWB) assay with an IC50 of 7nM. In addition, 14 had no activity in human COX-1 or COX-2 assays at 30μM, and failed to inhibit human mPGES-2 at 62.5μM in a microsomal prep assay. These data are consistent with selective mPGES-1-mediated reduction of PGE2. In dog, 14 had oral bioavailability (74%), clearance (3.62mL/(min*kg)) and volume of distribution (Vd,ss=1.6L/kg) values within our target ranges. For these reasons, 14 was selected for further study.