title: Highly potent irreversible inhibitors of neutrophil elastase generated by selection from a randomized DNA-valine phosphonate library.
abstract: We incorporated a phosphonate irreversible inhibitor of neutrophil elastase into a randomized DNA library and, using the SELEX process, iteratively selected these assemblies for the most potent elastase inhibitors. The inhibitors were selected against purified elastase and against secreted elastase in the presence of activated neutrophils. Very active aptamer inhibitors were obtained by both methods, with second-order rate constants for inactivation of human neutrophil elastase ranging (1-3) x 10(8) M(-1) min(-1). These rates exceed those of any reported irreversible inhibitor of elastase and exceed the previous best phosphonate inhibitors by 80-fold. The selected inhibitors are also significantly more potent than alpha-1 proteinase inhibitor in blocking degradation of elastin by activated neutrophils. In contrast to a previous experiment [Smith et al. (1995) Chem. Biol. 2, 741-750], a single-enantiomer form of the valyl phosphonate was used rather than a racemic mixture. Our analysis shows that this use of a chirally resolved valyl phosphonate results in selection of much more potent inhibitors and that these inhibitors specifically potentiate a single enantiomeric form of the phosphonate.
authors.affiliation: NeXstar Pharmaceuticals, Inc., Boulder, Colorado 80301, USA.
authors.forename: J
authors.initials: J
authors.lastname: Charlton
authors.forename: G P
authors.initials: GP
authors.lastname: Kirschenheuter
authors.forename: D
authors.initials: D
authors.lastname: Smith
mesh_terms: D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors; D019793:Fluorescein; D005452:Fluoresceins / Q000378:metabolism; D005453:Fluorescence; D015723:Gene Library; D006801:Humans; D007700:Kinetics; D019272:Leukocyte Elastase / Q000037:antagonists & inhibitors*; D008956:Models, Chemical; D008968:Molecular Conformation; D009504:Neutrophils / Q000187:drug effects / Q000201:enzymology*; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry / Q000494:pharmacology*; D063065:Organophosphonates / Q000737:chemistry / Q000494:pharmacology*; D012492:Salts / Q000494:pharmacology; D017421:Sequence Analysis; D012697:Serine Endopeptidases; D015842:Serine Proteinase Inhibitors / Q000138:chemical synthesis* / Q000494:pharmacology*; D013379:Substrate Specificity; D014633:Valine
chemical_list: D005452:Fluoresceins; D009838:Oligodeoxyribonucleotides; D063065:Organophosphonates; D012492:Salts; D015842:Serine Proteinase Inhibitors; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D019272:Leukocyte Elastase; D014633:Valine; D019793:Fluorescein