title: Synthese, Charakterisierung und Anwendung neuer Inhibitoren der Proproteinkonvertase Furin
title: Synthesis, characterization and application of novel furin inhibitors
Creators: name: Hardes, Kornelia
Publisher: name: Philipps-Universität Marburg
Year: 2015
Subjects: Medizin, Gesundheit
proprotein convertase
serin protease
furin
Furin
Proproteinkonvertase
inhibitor
Inhibitor
Serinprotease

description: Die Typ-I Transmembranprotease
      Furin gehört zur Familie der Proproteinconvertasen und ist
      ubiquitär im menschlichen Körper verbreitet. Furin aktiviert
      verschiedenste Proproteine, die sowohl in normalen
      physiologischen aber auch in zahlreichen pathophysiologischen
      Prozessen involviert sind. Beispielsweise aktiviert Furin
      Wachstumsfaktoren, deren Rezeptoren aber auch
      Matrixmetalloproteasen, die beim Tumorwachstum und der
      Metastasierung beteiligt sind. Zusätzlich prozessiert Furin
      verschiedene virale Glykoproteine und bakterielle Toxine.
      Diese Spaltung ist essentiell für die Vermehrung zahlreicher
      Viren oder der toxischen Wirkung humanpathogener Bakterien.
      Daher ist Furin ein potentielles Target der
      Wirkstoffentwicklung.
description: The type-I transmembrane protein
      furin belongs to the family of proprotein convertases (PCs)
      and contains a Ca2+-dependent subtilisin-like serine protease
      domain. Furin is ubiquitously distributed in human tissues
      and catalyzes the maturation of inactive precursor proteins
      in physiological and pathogenic processes. It is involved in
      various viral and bacterial infections, tumorigenesis,
      neurodegenerative disorders, diabetes, and atherosclerosis.
      Furin has a strong preference to process its substrates at a
      multibasic cleavage site, like Arg-Xaa-Arg/Lys-Arg-Xaa. Based
      on this sequence highly potent substrate analogue inhibitors
      containing decarboxylated arginine mimetics in P1 position
      have been previously developed. Among a series of inhibitors
      containing various proteinogenic P3 amino acids strong
      differences in their inhibitory potencies have been observed,
      whereby the strongest inhibition was found for the Val and
      Ile analogues. Further optimization of the lead structure
      Phac-Arg-Xaa-Arg-4-amidinobenzylamide by incorporation of
      unnatural hydrophobic P3-residues provided new furin
      inhibitors with improved potency. The most potent analogues
      contain tert.leucine (Ki = 0,17 nM) or penicillamine (Ki =
      0,25 nM) as P3 residue. Combination of P3 tert.leucine with
      basic substituted phenylacetyl residues in P5 position
      resulted in picomolar furin inhibitors. The most potent
      derivative MI-1148 (4-GMe-Phac-Arg-Tle-Arg-4-Amba) was
      characterized as tight-binding inhibitor of furin possessing
      a Ki value of 5.5 pM. Selected derivatives were used in
      various cell culture assays to demonstrate their potency as
      anti-infective drugs. Low micromolar concentrations of
      inhibitors MI-1148 and of its closely related analogue
      MI-1150 effectively inhibit the cleavage of the hemagglutinin
      (HA) from highly pathogenic influenza strains (H5N1, H7N1)
      and their subsequent virus propagation in MDCK cells. Further
      decreased inhibitor concentrations reduce the propagation of
      canine distemper virus in VeroSLAMdogtag cells. Moreover,
      nanomolar concentrations of MI-1148 and its closely related
      analogue MI-1150 were sufficient to protected macrophages
      against anthrax and diphtheria toxin. The protective effect
      against the bacterial toxins, as wells as their antiviral
      activity suggest that inhibitors of furin could represent
      promising lead structures for the development of new
      broad-spectrum anti-infective drugs. The analogs of this
      series also inhibit the related PC1/3 with similar potency,
      whereas PC2 is less affected. It was possible to solve a
      crystal structure of human furin in complex with MI-1148,
      which revealed numerous key interactions explaining the
      excellent low picomolar potency of this analogue. In an
      additional approach the N-terminal segment of the lead
      structure was modified. Many furin substrates possess
      additional basic residues in the P5 – P8 region. For this
      reason, a constant Arg-Val-Arg-4-Amba segment was maintained
      in P4-P1 positions and its N-terminus has been elongated with
      a different number of arginine residues in D- and/or
      L-configuration. These N-terminally unprotected derivatives
      inhibit furin in the subnanomolar range between 30 – 90 pM,
      whereby a perfect fit of the enzyme kinetic data to the
      equation for tight-binding inhibition was not possible,
      although determination coefficients &gt;0.97 have been
      calculated. Compared to the most potent inhibitors these
      compounds showed a reduced inhibition of influenza virus
      propagation in MDCK cells at a concentration of 50 µM.
      Furthermore, the lead structure was extended with L-arginine
      residues in P5- and P6 position. The best derivative of this
      series MI-1103 (Phac-Arg-Arg-Arg-Val-Arg-4-Amba) inhibits
      furin with an inhibition constant of 13,8 pM. To decrease the
      molecular weight of this inhibitor type, several compounds
      without P5-moiety have been prepared. These compounds possess
      reduced inhibitory potency in enzyme kinetic studies.
      Nevertheless the best compounds of this series
      (5-guanidino)-valeryl-Val-Arg-4-Amba (MI-1146) and its P3-Tle
      analogue MI-1152 inhibit furin with Ki-values of 2.5 and 1.3,
      respectively. Both derivatives also showed a concentration
      dependent protection of macrophages against anthrax toxin,
      although their efficacy was relatively low.