ABSTRACT
Two new potential impurities of antiarrhythmic drug substance Dronedarone Hydrochloride together with debutyldronedarone were detected by LC-MS analysis during process development. A successful synthetic strategy for the synthesis of these potential impurities was developed facilitating the access to new impurity reference standards. Their synthesis and characterization are discussed in detail. The availability of these impurity standards allowed cost reduction through the increase of process control.
Subject(s)
Amiodarone/analogs & derivatives , Anti-Arrhythmia Agents/analysis , Anti-Arrhythmia Agents/chemical synthesis , Amiodarone/analysis , Amiodarone/chemical synthesis , Chemistry Techniques, Analytical/methods , Chromatography, Liquid/methods , Chromatography, Thin Layer , Dronedarone , Drug Contamination , Drug Design , Hydrogen/chemistry , Magnetic Resonance Spectroscopy/methods , Mass Spectrometry/methods , Spectrometry, Mass, Electrospray Ionization , Spectroscopy, Fourier Transform InfraredABSTRACT
Bacterial enoyl-acyl carrier protein reductase (ENR) catalyzes an essential step in fatty acid biosynthesis. ENR is an attractive target for narrow-spectrum antibacterial drug discovery because of its essential role in metabolism and its sequence conservation across many bacterial species. In addition, the bacterial ENR sequence and structural organization are distinctly different from those of mammalian fatty acid biosynthesis enzymes. High-throughput screening to identify inhibitors of Escherichia coli ENR yielded four structurally distinct classes of hits. Several members of one of these, the 2-(alkylthio)-4,6-diphenylpyridine-3-carbonitriles ("thiopyridines"), inhibited both purified ENR (50% inhibitory concentration [IC(50)] = 3 to 25 micro M) and the growth of Staphylococcus aureus and Bacillus subtilis (MIC = 1 to 64 micro g/ml). The effect on cell growth is due in part to inhibition of fatty acid biosynthesis as judged by inhibition of incorporation of [(14)C]acetate into fatty acids and by the increased sensitivity of cells that underexpress an ENR-encoding gene (four- to eightfold MIC shift). Synthesis of a variety of compounds in this chemical series revealed a correlation between IC(50) and MIC, and the results provided initial structure-activity relationships. Preliminary structure-activity relationships, potency on purified ENR, and activity on bacterial cells indicate that members of the thiopyridine chemical series are effective fatty acid biosynthesis inhibitors suitable for further antibacterial development.
Subject(s)
Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Escherichia coli/enzymology , Oxidoreductases/antagonists & inhibitors , Pyridines/chemical synthesis , Pyridines/pharmacology , Bacillus subtilis/drug effects , Bacteria/drug effects , Cloning, Molecular , Drug Evaluation, Preclinical , Enoyl-(Acyl-Carrier-Protein) Reductase (NADH) , Escherichia coli/drug effects , Fatty Acids/biosynthesis , Gene Expression Regulation, Bacterial/drug effects , Kinetics , Lac Operon/genetics , Microbial Sensitivity Tests , Oxidoreductases/biosynthesis , Oxidoreductases/genetics , Reverse Transcriptase Polymerase Chain Reaction , Staphylococcus/drug effects , Staphylococcus/genetics , Staphylococcus/metabolism , Structure-Activity RelationshipABSTRACT
A general, modular strategy for the first completely stereoselective synthesis of defined heparin oligosaccharides is described. Six monosaccharide building blocks (four differentially protected glucosamines, one glucuronic and one iduronic acid) were utilized to prepare di- and trisaccharide modules in a fully selective fashion. Installation of the alpha-glucosamine linkage was controlled by placing a conformational constraint on the uronic acid glycosyl acceptors thereby establishing a new concept for stereochemical control. Combination of disaccharide modules to form trans-uronic acid linkages was completely selective by virtue of C2 participating groups. Coupling reactions between disaccharide modules exhibited sequence dependence. While the union of many glucosamine uronic acid disaccharide modules did not meet any problems, certain sequences proved not accessible. Elaboration of glucosamine uronic acid disaccharide building blocks to trisaccharide modules by addition of either one additional glucosamine or uronic acid allowed for stereoselective access to oligosaccharides as demonstrated on the example of a hexasaccharide resembling the ATIII-binding sequence. Final deprotection and sulfation yielded the fully synthetic heparin oligosaccharides.
