Lawrence Wayne Torley

Modulation of Eicosanoid Biosynthesis by Novel Pyridinylpyrimidines

A simple but effective strategy for drug development has been controlling those molecules that cause disease either through specific antagonists or synthesis inhibitors. Accordingly, structural elucidation of key mediators profoundly affects this approach by providing clear targets. The 1911 discovery of histamine is a classic example, eventually leading to a number of specific antagonists that remain therapeutically useful today. More recently, the discovery of the slow-reacting substance of anaphylaxis (SRS-A) as the sulfidopeptidyl leukotrienes (LT),’ and the rabbit aorta-constricting substance as a mixture of thromboxane Az (TXA2), prostaglandin Hz (PGHz), and PGGz,2 for example, have led to the synthesis of a number of specific inhibitors and antagonists with therapeutic potential. Although specific inhibition or antagonism is advantageous with respect to toxicity by minimally affecting other homeostatic processes, it is often insufficient due to redundancy of mediators of diverse origin and structure such as histamine and LTC. Because of this, an alternative to specific inhibitors and antagonists is the design of cell “stabilizing” agents or mediator release inhibitors. These agents prevent the release of both preformed mediators and the biosynthesis of other mediators such as eicosanoids. The demonstration that chromolyn sodium-a useful drug for the management of asthma-inhibits mediator release from antigen-stimulated mast cells’ greatly bolstered the mast cell-based search for mediator release inhibitors, despite clear evidence that this was chromolyn’s mechanism of action in asthma. A new generation of mediator release inhibitors containing a pyridinylpyrimidine structure is described below: but unlike chromolyn, the pyridinylpyrimidines also inhibit basophils and phagocytes. This series of compounds inhibits eicosanoid biosynthesis primarily at levels proximal to the phospholipase Az (PLA,), but to a lesser extent at distal sites such as the thromboxane synthetase. From in vivo and in vitro data, the pyridinylpyrimidines appear to be a promising new class of compounds for development as therapy for allergic and inflammatory diseases where eicosanoids and granule release are essential components of the etiology.

Studies of the effect of a platelet-activating factor antagonist, CL 184,005, in animal models of gram-negative bacterial sepsis.

The effect of CL 184,005, a potent and specific platelet-activating factor antagonist, has been examined in a variety of animal models relevant to gram-negative bacterial sepsis. Pretreatment of mice with CL 184,005 protected them from the lethal effects of platelet-activating factor. When rats or primates rendered hypotensive with endotoxin were treated with CL 184,005, blood pressure was normalized. Pretreatment of rats with CL 184,005 protected them from the gastrointestinal lesions induced by endotoxin. Pretreatment of rats and mice with CL 184,005 protected them from the lethal effects of endotoxin. Plasma tumor necrosis factor levels in endotoxin-treated mice were lower when the mice were pretreated with CL 184,005. These observations suggest that CL 184,005 may be potentially useful in the treatment of gram-negative bacterial sepsis, and the agent is undergoing clinical evaluation.

Effect of a series of 1-alkyl ether lipids on inhibition of phospholipase A2 activity and PAF responses.

Several 1-alkyl ether lipids were studied for their ability to inhibit PLA2 and antagonize PAF responses. Studies with synthetic micellar substrate (1-stearyl-2-arachidonyl phosphocholine), at concentrations ranging from 0.02 to 1000μM, demonstrate that CL 118326 inhibits porcine pancreatic PLA2 in vitro. As the substrate concentration increases, there is a dose-dependent increase in the IC50 value (IC50 ranges: 1.6–84.6μg/ml or 2.6–137μM). CL 118326 inhibits mammalian pancreatic PLA2, but not snake or bee venom PLA2. CL 118326 inhibits thrombin (IC50 =7.9μM), but not Na arachidonate- (IC50 > 100μM) induced platelet aggregation, indicative of inhibition of cellular PLA2. CL 118326 inhibits other PLA2-dependent processes such as antigen-induced leukotriene (LTC4) release (IC50=2.3μg/ml or 3.8μM) and histamine release (IC50=1.4μg/ml or 2.2μM) in basophil-enriched WBCs. Intradermal coinjection of CL 118326 (10μg) with PLA2 into guinea pig skin inhibits pancreatic PLA2-induced increase in vascular permeability and leakage, but not snake or bee venom PLA2-induced leakage. CL 118326 shows no PAF-like agonist activity in stimulating rabbit platelet-rich plasma. It inhibits PAF-induced aggregation (IC50=5.8μM), but not ADP-induced aggregation. CL 118326 has greater efficacy as a PLA2 inhibitor than as a PAF antagonist since the IC50-substrate concentration ratio for PLA2 inhibition is <- 1.0 at substrate concentrations of 10–1000μM while the IC50-agonist ratio for PAF antagonism is > 100. Results for four other compounds related to CL 118326 are also presented.

Pluronic F 127 liquid sensitizes mice to low doses of Escherichia coli lipopolysaccharide.

Background and MethodsIn murine models of endotoxemia, large amounts of lipopolysaccharide have to be administered to induce mortality. If mice are pretreated with D-galactosamine, the amount of lipopolysaccharide required to induce mortality is significantly lowered. Pluronic F 127 liquid is a relatively nontoxic copolymer that exhibits reverse gelation properties. Thus, it is a liquid at cold temperature and a gel at body temperature. The present studies were performed to ascertain whether the reverse gelation properties of Pluronic F 127 liquid could be used in devising a model of septic shock where a sustained delivery of lipopolysaccharide occurred. In evaluating this model, doseresponse studies were conducted with lipopolysaccharide when a) it was administered intraperitoneally in saline or in Pluronic F 127 liquid, and b) it was administered intravenously to mice that had been pretreated with saline or Pluronic F127 liquid. Mortality was followed for up to 72 hrs. ResultsVarious doses of Escherichia coli lipopolysaccharide dissolved in saline or in Pluronic F127 liquid were administered intraperitoneally to mice. The lethal dose of lipopolysaccharide required to kill 50% of the mice (LD50) administered in Pluronic F127 liquid was approximately ten- to 15-fold less than the values obtained for lipopolysaccharide administered in saline. This decrease in the LD50 of lipopolysaccharide was also observed if the mice were treated intraperitoneally with Pluronic F 127 liquid and challenged 6 hrs later with iv lipopolysaccharide. The concentrations of tumor necrosis factor and interleukin-6 in the plasma were significantly higher when a low dose of lipopolysaccharide was administered to mice that had been pretreated with Pluronic F 127 liquid. While there was no effect on the liver enzymes, Pluronic F 127 liquid caused an increase in the plasma triglycerides. ConclusionsThe data reported in this paper indicate that the LD50 of lipopolysaccharide is significantly decreased if it is administered in Pluronic F 127 liquid or administered to mice that have been pretreated with the Pluronic F 127 liquid. Thus, Pluronic F127 liquid appears to sensitize mice to low levels of lipopolysaccharide. Unlike the D-galactosamine model, lipopolysaccharide can be administered as late as 6 hrs after treatment with Pluronic F 127 liquid. While the mechanisms by which Pluronic F 127 liquid sensitizes mice is not known, plasma triglycerides were increased in mice treated with this agent, suggesting that tissues responsible for the synthesis and/or degradation of triglycerides play a role in this sensitization process.