Oligomeric formylpeptide activity on human neutrophils.

A series of oligomeric formylpeptides were synthesized by cross-linking the prototype fMLP using a Lys residue. They were then investigated for their ability to stimulate chemotaxis, superoxide anion production, and lytic enzyme release in human neutrophils. Although active in stimulating the different receptor isoforms, leading to the different biological responses, these analogues showed a lesser potency and affinity than the standard peptide. On the basis of the results reported here, we can hypothesise that: (i) the increased bulk of these molecules seems to hinder their correct positioning into the receptor pocket, thereby hindering favourable receptor interaction; and that: (ii) fMLP space positions do not seem to allow the ligand to increase biological responses.

New chemotactic dimeric peptides show high affinity and potency at the human formylpeptide receptor.

A number of analogues of the prototypical peptide for-Met-Leu-Phe-OMe (fMLP-OMe) have been studied in order to evaluate their ability to interact with formylpeptide receptors and to induce specific biological responses in human neutrophils. In vitro assays were carried out and receptor binding, chemotaxis, superoxide anion release and secretagogue activity were evaluated. The fMLP-OMe analogues synthesized, with the general formula for-Met-Leu-Phe-Xaa-Lys(OMe)-Phe-Leu-Met-for (Xaa=Gly, beta-Ala, gamma-aminobutyric acid, 5-aminovaleric acid, and 6-aminocaproic acid), were constituted by two fMLP units linked by a Lys residue, with an amino acid spacer between them. Competition binding experiments revealed that the new compounds have much more affinity for formylpeptide receptors than the reference ligand, with good correlation between receptor affinity and length of spacer. The EC(50) values for the killing mechanisms of each analogue were similar to each other, the affinity and potency, once again, being strictly dependent on the chain length. Furthermore the analogues proved to be more potent full agonists than the prototype fMLP-OMe in these functions, while chemotaxis was poorly induced. The dimeric fMLP-OMe analogues are one of the few examples of formylpeptides which exhibit a receptor affinity greater than the parent fMLP-OMe thereby rendering them suitable to be used as carriers for various drugs.

Structure-activity relationship of for-L-Met L-Leu-L-Phe-OMe analogues in human neutrophils.

Neutrophils constitute the first line of defence against bacterial invasion. They migrate to infected tissues along a concentration gradient of chemoattractant molecules, the most important of which is for-Met-Leu-Phe-OH (fMLP). Different responses arise from formylpeptides binding to different isoforms of the specific receptor. The aim of the studies reported herein was to clarify (i) the role of fMLP-OMe amide bonds in receptor-ligand cross-linking, (ii) the nature of the group occupying the N- and C-terminal positions, (iii) the features peculiar to the Met, Leu, and Phe receptor pockets, and (iv) the features which determine the specific neutrophil response.

Biological activity of for-Met-Leu-Phe-OMe analogs: relevant substitutions specifically trigger killing mechanisms in human neutrophils.

Two analogs of the prototypical peptide for-Met-Leu-Phe-OMe (fMLP-OMe), for-Gln-Tyr-Phe-OMe (1) and for-Gln-Tyr-Tyr-OMe (2), carrying unusual hydrophilic residues, were synthesized in order to investigate whether they provoked specific biological responses, as well as intracellular calcium mobilization, in human neutrophils. Whereas neither compound stimulates chemotaxis, both are able to elicit lysosomal enzyme production. However compound 1 is able to trigger copious superoxide anion production while compound 2 only elicits minor superoxide anion production. In binding experiments on formylpeptide receptors, the newly synthesized compounds for-Gln-Tyr-Phe-OMe (1) and for-Gln-Tyr-Tyr-OMe (2) showed affinity values in the micromolar range. These derivatives demonstrate inability to find a positive contribute from single substitutions. A very important result of this research is the evidence of the ability of the formyl group alone to trigger the primary target of the human neutrophil activity, i.e. killing mechanisms, by activating the specific receptor conformation.