The increasing emergence of medically important pathogenic bacteria (Staphylococci, Streptococci, Enterococci, Pseudomonas aeruginosa, Mycobacterium tuberculosis) with high resistance to numerous antibacterial agents constitutes a serious public threat in the treatment of infectious diseases. Therefore, extensive research in the development of novel antibacterial agents has been carried out in the past few years. One of the most important and validated targets for antibacterial drug discovery research is bacterial peptidoglycan, a heteropolymer composed of alternating glycan chains cross-linked by short peptides. As an essential component of bacterial cell wall, peptidoglycan is important for the maintenance of a defined cell shape and withstanding the internal osmotic pressure. Interference with its synthesis or structure leads to loss of cellular integrity and ultimately to bacterial cell death. Biosynthesis of peptidoglycan involves numerous enzymes. Amongst them are ATP-dependant ligases such as Mur ligases and D-alanyl-D-alanine ligase (Ddl), which are essential for the synthesis of UDP-N-acetylmuramyl-pentapetide, a crucial intracellular precursor of peptidoglycan.MurC to MurF ligases catalyze stepwise addition of L-alanine, D-glutamic acid, L-lysine or meso-diaminopimelic acid and dipeptide D-alanyl-D-alanine (prepared by Ddl from two Dalanines) to UDP-N-acetylmuramic acid. In addition to Ddl, the members of D-alanine:D-X ligases superfamily are also the Van enzymes, which are crucial for vancomycin resistance in
Enterococci. Phosphinate, phosphonate and sulfonamide transition-state analogs are wellknown inhibitors of Mur ligases and Ddl. Therefore, we decided to design, synthesize and evaluate the hydroxyethylamines as new potential inhibitors of Mur ligases, VanA and Ddl. To design a new class of inhibitors we used »de novo« ligand design software SPROUT and ligand docking tool eHiTS. In the following step we developed and optimized an efficient synthetic approach to hydroxyethylamines via oxidation of the corresponding allylamine derivatives and subsequent opening of epoxide ring with N-nucleophyles in the presence of calcium trifluoromethanesulphonate as a catalyst. The synthesized compounds were evaluated for their inhibitory activities against DdlB, MurC, MurD and MurF from Echerichia coli, MurE from Staphylococcus aureus and VanA from Enterococcus faecium. The compounds composed of hydroxyethylamine moiety and L-alanine or D-glutamic acid did not show any inhibitory activity on MurC or MurD. Hydroxyethylamine derivatives of pyroglutamic acid substituted with trifluoromethylphenyl substituent inhibited the activity of DdlB and VanA in the concentrations above 500 μM. The strongest inhibitors were obtained when a phosphate group was attached to hydroxyl group of hydroxyethylamine moiety. Hydroxyethylamine derivative 1-(4-methoxy-phenylsulfonamido)-3-orpholinopropan-2-yl dihydrogen phosphate was found as an inhibitor of DdlB (IC50 = 110 μM) and VanA (IC50 =224 μM). Since VanA is responsible for vancomycin resistance the aformentioned inhibitor has the potential to be developed into drugs that would reverse bacterial resistance to vancomycin. This compound also showed promising inhibitory activity against MurE with IC50 value in low micromolar range (IC50 = 6 μM). If the metoxy group was replaced by a trifluoromethyl group the inhibitor of MurC and MurF ligases was obtained. The hydroxyethylamine derivatives of 1-morpholino-3-(phenylsulfonamido)propan-2-yl dihydrogen phosphate are therefore an important starting point for development of multiple inhibitors of peptidoglycan biosynthesis with broad-spectrum antibacterial activity. Moreover, due to inhibition of MurE ligase from Staphylococcus aureus this compounds could be developed into antibacterial drugs against MRSA (Methicillin-resistant S. aureus).
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