Gordon L. Bundy

Prostaglandin d2 as a potential antithrombotic agent.

Prostaglandin D2 was found to be a potent inhibitor of platelet aggregation. Aggregation of human platelets by ADP, collagen and prostaglandin G2 was inhibited more strongly by PGD2 than by PGE1. Although ADP-induced aggregation of rabbit platelets was inhibited more strongly by PGE1 than by PGD2 the latter prostaglandin gave a more long-lasting inhibitory effect on platelet aggregation following intravenous or oral administration. These results coupled with the finding that PGD2 has less hypotensive effects on the cardiovascular system than PGE1 suggest the possible use of PGD2 as an antithrombotic agent.

NOVEL PROSTAGLANDIN SYNTHESES

Syntheses of 2 different classes of prostaglandin (PG) analogs are described that of the 3-oxa-PGs and that of the 15-methyl PGs the former of which inhibit beta-oxidative degradation and the latter of which prevent degradation by the C-15 dehydrogenase. 7 sterochemical figures accompany the descriptions of the synthesis procedures. The analogs though inert to 2 of the recognized modes of enzymatic catabolism maintain appreciable smooth muscle activity. Furthermore as an alternative to totally synthetic routes a partially synthetic route by which the 15-methyl analogs of PGE and PGF types as well as the partially synthetic route by which the types as well as the parent PGs may be derived from the non-mammalian marine source Plexaura Homomall is described.

Novel membrane localized iron chelators as inhibitors of iron-dependent lipid peroxidation.

Attachment of various iron chelating moieties to hydrophobic steroids greatly enhanced their abilities to inhibit iron-dependent lipid peroxidation. Using whole rat brain homogenates, lipid peroxidation initiated by the addition of 200 microM Fe2+ was assessed by the formation of thiobarbituric acid reactive products (TBAR). Under these conditions, 50% inhibitory concentrations of Fe3+ chelators such as desferrioxamine or N1,N8-bis(2,3-dihydroxybenzoyl) spermidine hydrobromide (compound II) were around 170 and 50 microM respectively. Coupling desferrioxamine or compound II to a steroid at the D ring increased their potency in lipid peroxidation assays by 5- to 10-fold. Evidence that inhibition of lipid peroxidation by the steroid-chelator adducts was due to iron chelation was suggested by the fact that methylation of the catechol oxygens of compound II, which are essential for chelation, completely eliminated activity of the steroid adduct. A series of 21-aminosteroids which complex Fe2+ iron and potently inhibit iron-dependent lipid peroxidation has also been synthesized. Coupling Fe2+ chelators to hydrophobic steroids increased their inhibitory potencies by as much as 10- to 100-fold. Some steroid-based Fe2+ chelators stimulated lipid peroxidation at low concentrations in the presence of Fe3+. The degree of stimulation was related to the affinity of a compound for Fe2+ with the stronger chelators causing greater stimulation. The most potent inhibitors of lipid peroxidation in the 21-aminosteroid series were found to be those compounds forming the weakest Fe2+ complexes. The findings suggest that it is iron at or near the membrane that is responsible for the catalysis of lipid peroxidation. The compounds described should provide useful tools for studies of the involvement of iron in the lipid peroxidation process.

Discovery and Characterization of QPT-1, the Progenitor of a New Class of Bacterial Topoisomerase Inhibitors

ABSTRACT QPT-1 was discovered in a compound library by high-throughput screening and triage for substances with whole-cell antibacterial activity. This totally synthetic compound is an unusual barbituric acid derivative whose activity resides in the (−)-enantiomer. QPT-1 had activity against a broad spectrum of pathogenic, antibiotic-resistant bacteria, was nontoxic to eukaryotic cells, and showed oral efficacy in a murine infection model, all before any medicinal chemistry optimization. Biochemical and genetic characterization showed that the QPT-1 targets the β subunit of bacterial type II topoisomerases via a mechanism of inhibition distinct from the mechanisms of fluoroquinolones and novobiocin. Given these attributes, this compound represents a promising new class of antibacterial agents. The success of this reverse genomics effort demonstrates the utility of exploring strategies that are alternatives to target-based screens in antibacterial drug discovery.

Evidence for 15-HETE synthesis by human umbilical vein endothelial cells.

Incubation of cultured human umbilical vein endothelial cells with [1-14C]-arachidonic acid, followed by RP-HPLC analysis, resulted in the appearance of two principal radioactive products besides 6-keto-PGF1 alpha. The first peak was HHT, a hydrolysis product of the prostaglandin endoperoxide. The second peak was esterified, converted to the trimethylsilyl ether derivative, and analyzed by GC/MS and was shown to be the lipoxygenase product 15-HETE. Stimulation of endothelial cells with thrombin enhanced 15-HETE synthesis from arachidonate. Subsequent experiments showed that 5-HETE and 12-HETE were also synthesized by endothelial cells, but no evidence of leukotriene synthesis was found. Incubation of the 15-HETE precursor 15-HPETE with endothelial cells resulted in the formation of four distinct ultraviolet light-absorbing peaks. Ultraviolet and GC/MS analysis showed these peaks to be 8,15-diHETEs that differed only in their hydroxyl configuration and cis-trans double-bond geometry. Formation of 8,15-diHETE molecules suggests the prior formation of the unstable epoxide molecule 14,15-LTA4 or an attack at C-10 of 15-HPETE by an enzyme with mechanistic features in common with a 12-lipoxygenase. The observation that endothelial cells can synthesize both 15-HETE and 8,15-diHETE molecules suggest that this cell type contains both a 15-lipoxygenase and a system that can synthesize 14,15-LTA4.

Synthesis and biological properties of 9-deoxo-16,16-dimethyl-9-methylene-PGE2.

The in vivo monkey uterine stimulating potency of 9-deoxy-16,16-dimethyl-9-methylene-PGE2 is similar to that of 16,16-dimethyl-PGE2 and approximately 15 times that of PGE2. Low doses of this compound stimulated uterine contractions when administered vaginally. Pregnancy was terminated prematurely following subcutaneous, intramuscular or vaginal suppository treatment. Estimates of potential for gastrointestinal side effects using the rat enteropooling assay and in vivo monkey effects indicate that diarrhea will be substantially reduced with retention of uterine stimulating potency.

The synthesis of 2,3-dinorprostacyclin metabolites—a new approach to spirolactone hemiacetals

Abstract The major human urinary metabolites of prostacyclin and 6-keto-PGF 1 α have been synthesized by a direct route, involving three-carbon homologation of bicyclic lactone intermediates and spontaneous spirolactonization of the products. The fact that these 2,3-dinor-6-oxo metabolites exist almost exclusively as spirolactone hemiacetals under acidic conditions (pH 5 and below) may explain the reported difficulties in derivatizing samples of biological origin. Several 19,19,20,20-d 4 metabolites have also been synthesized.

Arachidonic acid and 15(S)-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid modulate human polymorphonuclear neutrophil activation by monocyte derived neutrophil activating factor

Exposure of human polymorphonuclear neutrophils (PMN) to human monocyte derived neutrophil activating factor(s) (NAF) resulted in a concentration-dependent extracellular release of granule constituents. NAF also induced the generation of 5(S),12(R)-dihydroxy-6,14-cis-8,10-trans-eicosatetraenoic acid [Leukotriene B4 (LTB4)] by PMNs which was enhanced in the presence of exogenous arachidonic acid (AA). In contrast to its enhancing effect on LTB4 production, AA inhibited NAF-stimulated PMN degranulation. 15(S)-hydroxy-5,8,11-cis-13-trans-eicosatetraenoic acid (15-HETE), a product of the 15-lipoxy-genation of AA in PMNS, caused a concentration-dependent suppression of degranulation and LTB4 generation by PMNs in contact with NAF. 15-HETE also inhibited the rise in cytosolic-free calcium [( Ca2+]i) observed in NAF activated PMNs. These data suggest that AA and a 15-lipoxygenase product modulate the NAF-associated activation pathway in human PMNs.

Prevention of cecitis in hamsters by certain prostaglandins

Acute inflammation of the colon (cecitis) was produced in hamsters by daily subcutaneous administration of an antibiotic for 3 days. The following prostaglandins completely prevented the cecitis: 16,16-dimethyl-PGE2, 15(R)-15-methyl-PGE2, and 2-acetyl-2-decarboxy-15(S)-15-methyl-PGF2 alpha. PGF2 beta was less active. The synthesis of 2-acetyl-2-decarboxy-15(S)-methyl-PGF2 alpha is described. Castor oil also prevented the cecitis and peanut oil exerted partial protection. Since these oils contain linoleic acid, a precursor of PGE1, protection may have been due to endogenous formation of that prostaglandin. A partial block of the protective effect of castor oil by treatment with indomethacin supports such mechanism. The tissue level of endogenous prostaglandins seems to exert protection since administration of cyclooxygenase inhibitors, indomethacin and aspirin, markedly increased the incidence of cecitis. Magnesium sulfate given orally and sodium salicylate given subcutaneously reduced the incidence of cecitis only partially. The following agents were inactive: loperamide, an antidiarrheic agent; carbachol, a cholinergic and diarrheogenic agent, atropine, an anticholinergic agent; and acetazolamide, a carbonic anhydrase inhibitor. These results, show that certain prostaglandins, which have been shown earlier to be cytoprotective for the stomach and the small intestine, are cytoprotective for the large intestine as well.

Inhibition of human platelet thromboxane synthetase by 11a-carbathromboxane A2 analogs

Summary Three 11a-carbathromboxane A 2 analogs were synthesized and evaluated for “thromboxane A 2 -like” biological activity in human platelets and rat aortic strips. All three analogs were potent inhibitors of either prostaglandin H 2 or arachidonic acid-induced human platelet aggregation, and proved to be both receptor level antagonists of thromboxane A 2 and thromboxane synthetase inhibitors. The three 11a-carba analogs were devoid of either agonist or antagonist activity on rat aortic strips, suggesting that the thromboxane A 2 receptor in platelets exhibits different binding characteristics than the thromboxane receptor in vascular smooth muscle.