Chemotactic tripeptides incorporating at position 2 alpha-aminoacid residues with unsaturated side chains.

New N-For-Met-Leu-Phe-OMe (fMLF-OMe) analogues incorporating three different gamma-delta-didehydro-alpha-aminoacid residues (namely: Alg = (S)-Allylglycine; Dag = Diallylglycine; Cpg = 1-Aminocyclopent-3-ene-1-carboxylic acid) replacing the native (S)-Leucine have been synthesized and their activity towards human neutrophils has been evaluated in comparison with that shown by the reference tripeptide fMLF-OMe. Chemotaxis, lysozyme release and superoxide anion production have been measured. (1)H NMR titration experiments and NOESY spectrum of the Cpg containing model 10 have been discussed in order to ascertain the preferred solution conformations. A fully extended (C(5)) conformation at position 2 and a folded conformation with two consecutive gamma-turns (C(7) structure) have been proposed for the Dag and Cpg containing tripeptides, respectively.

Biscarbamate analogues of the chemotactic tripeptide fMLF-OMe.

Based on the sequence of the prototypical chemotactic tripeptide HCO-Met-Leu-Phe-OH (fMLF) and by taking into account the versatility shown by its N-terminal carbamate analogues, the new biscarbamates MeOCO-Met-Leu-gPhe-COOMe (2) and Boc-Met-Leu-gPhe-COOMe (4) were synthesized. These two new ligands are characterized by the presence of a gem-diamino residue (gPhe) replacing the C-terminal Phe and a carbamate functionality positioned at both the ends of the molecule. The activity of the two new compounds has been determined on human neutrophils and compared to that shown by the corresponding N-terminal monocarbamates MeOCO-Met-Leu-Phe-OMe (1) and Boc-Met-Leu-Phe-OMe (3).

Inhibition of adamalysin II and MMPs by phosphonate analogues of snake venom peptides.

Phosphonate analogues of the peptidomimetic N-(Furan-2-yl)carbonyl-Leu-Trp-OH were prepared with the goal of evaluating the effect of phosphonate for carboxylate replacement on binding with snake venom metalloproteinases and MMPs. N-(Furan-2-yl)carbonyl-Leu-L-Trp(P)-(OH)2 showed a 75-fold increase of the inhibiting activity against adamalysin II, a snake venom metalloproteinase structurally related to MMPs and TACE. Both the phosphonate and carboxylate peptidomimetics fit into the active site adopting a retrobinding mode and provide the structural base for a new class of metalloproteinases inhibitors.

2 angstrom X-ray structure of adamalysin II complexed with a peptide phosphonate inhibitor adopting a retro-binding mode.

The search of reprolysin inhibitors offers the possibility of intervention against both matrixins and ADAMs. Here we report the crystal structure of the complex between adamalysin II, a member of the reprolysin family, and a phosphonate inhibitor modeled on an endogenous venom tripeptide. The inhibitor occupies the primed region of the cleavage site adopting a retro-binding mode. The phosphonate group ligates the zinc ion in an asymmetric bidentate mode and the adjacent Trp indole system partly fills the primary specificity subsite S1'. An adamalysin-based model of tumor necrosis factor-alpha-converting enzyme (TACE) reveals a smaller S1' pocket for this enzyme.

The role of (Z)-2,3-didehydrophenylalanine and phenylalanine C-terminal residues in determining the chemotactic activity of formylpeptides.

Several formylpeptides, analogs of the chemotactic agent HCO-Met-Leu-Phe-OMe, having the HCO-Xaa-Leu-delta ZPhe-OMe and HCO-Xaa-Leu-delta ZPhe-Phe-OMe structures (delta ZPhe = (Z)-2,3-didehydrophenylalanine), have been synthesized. The biological activity of these ligands has been determined on human neutrophils and compared to that of the corresponding HCO-Xaa-Leu-Phe-OMe derivatives not containing the unsaturated residue. The replacement of the C-terminal Phe with delta ZPhe causes, in all the examined tripeptides, the loss of any biological activity. On the other hand, the introduction into the delta ZPhe containing models of an additional C-terminal Phe residue leads to the formyltetrapeptides HCO-Xaa-Leu-delta ZPhe-Phe-OMe which show a biological activity very similar to that exhibited by the corresponding HCO-Xaa-Leu-Phe-OMe analogues.

Chemotactic peptide analogues. Synthesis and chemotactic activity of N-formyl-Met-Leu-Phe analogues containing (S)-phenylalaninol derivatives.

The synthesis and the biological activity towards human neutrophils of some N-formyl-Met-Leu-Phe-OMe analogues containing (S)-phenylalaninol (Pheol) or its derivatives in place of the native phenylalanine are reported. While the analogue containing Pheol (4) was found to be devoid of significant biological activity, its esters 3 and 5, although inactive as chemoattractants, are able to strongly stimulate superoxide production and are active with a lower efficacy in the lysozyme release.

For-Met-Lys-Phe-For-Met-Lys-Phe-: a new cyclic analogue of the chemotactic formylpeptides.

As a continuation of the studies on chemotactic N-formylpeptides, we report here the synthesis and activity of a new cyclic analogue of the prototypical ligand For-Met-Leu-Phe-OMe. The new compound For-Met-Lys-Phe-For-Met-Lys-Phe- (4) contains a 20-membered cyclic moiety made up of a dimeric -Lys-Phe- sequence in which For-Met is attached to each Lys alpha-NH2 and hence remains outside the ring. The conformation in the crystal of the cyclic precursor of 4, namely Boc-Lys-Phe-Boc-Lys-Phe- (2) and the activity of the structurally related linear analogue For-Met-Lys(Z)-Phe-OBzl (6), have also been examined. The new analogues 4 and 6 are active as chemoattractants, secretagogues, and superoxide anion generating agents, when tested on human neutrophils. The structure-activity relationship is discussed and related to that of a previously studied cyclic model.

Two new formylated peptides able to activate chemotaxis and respiratory burst selectively as tools for studying human neutrophil responses.

Two new For-Met-Leu-Phe-OH (FMLP) methyl ester analogues, For-Thp-Leu-Ain-OMe [Thp1, Ain3] and For-Met-delta zLeu-Phe-OMe [delta zLeu2], able to activate selectively chemotaxis and superoxide anion (O2-) release, respectively modulate intracellular cyclic AMP (cAMP) levels in different ways. FMLP and [delta zLeu2] enhance human neutrophil cAMP levels per se, and this effect is potentiated by Ro 201724, a non-xanthinic phosphodiesterase (PDE) inhibitor, whereas it is counteracted by 3-isobutyl-1-methyl-xanthine (IBMX), a blocker of both phosphodiesterase and adenosine receptors. In contrast, [Thp1, Ain3] is ineffective. However, no formylated peptides influence cAMP phosphodiesterase activity. Neutrophil preincubation with Ro 201724 or IBMX drastically reduces chemotaxis and superoxide anion (O2-) production triggered by peptides. Our results suggest that: (1) peptide-induced cAMP increase is probably indirect, and due mainly to the action on adenosine-sensitive adenylate cyclase; (2) formylated peptide, endowed solely with chemotactic activity is unable to increase neutrophil cAMP concentration; (3) cAMP elevation may represent a feed-back mechanism to inhibit the physiological responses induced by formylated peptides.

Synthesis and chemotactic activity of the fMLP analog HCO-Hmb-Leu-Phe-OMe.

The new fMLP analog HCO-Hmb-Leu-Phe-OMe (1), containing (S)-2-hydroxy-4-(methylthio)butyric acid (Hmb) in place of L-methionine at the N-terminal position, has been synthesized and fully characterized. The peptide 1 has been designed in order to improve the understanding of the role exerted by the formamido group in the binding interaction with the formylpeptide chemotactic receptors. Chemotaxis, superoxide anion production, and lysozyme release have been measured for both 1 and its deformylated analog Hmb-Leu-Phe-OMe 2. Results indicate that a strong hydrogen bond of the OH....O = C type may complement a weak H-bonding interaction involving the formylic proton as H-bond donor.

Synthesis and properties of chemotactic peptide analogs. I. Crystal structure and molecular conformation of HCO-Met-leu-Ain-OMe.

HCO-Met-Leu-Ain-OMe (2), an analog of the chemotactic peptide HCO-Met-Leu-Phe-OH, containing the conformationally blocked residue of the 2-aminoindane-2-carboxylic acid (Ain) has been synthesized and its crystal and molecular conformation has been determined. Crystals of 2 are monoclinic, space group P2(1), with a = 15.059(7), b = 18.548(7), c = 9.600(4) A; beta = 85.04(3) degrees. The structure has been solved by direct methods and refined to R = 0.069 for 2813 independent reflections with I greater than 2.5 sigma (I). Two independent molecules A and B have been found in the asymmetric unit of the crystal of 2. Their conformation can be described as extended at the Met and Leu residues, but folded at the C-terminal Ain residue. The helical folding is left- and right-handed in the A and B molecule, respectively. The crystal packing is characterized by ribbons of intermolecular hydrogen bonded molecules extended along the c direction. The constrained analog 2 is highly active in the superoxide production, thus indicating that a stabilization of a helical folding at the C-terminal region of chemotactic tripeptides maintains the activity. The orientation of the aromatic ring, with respect to its adjacent backbone atoms, does not seem critical for the activity.