The Proteolytically Stable Peptidomimetic Pam-(Lys-ßNSpe)₆-NH₂ Selectively Inhibits Human Neutrophil Activation via Formyl Peptide Receptor 2

Immunomodulatory host defense peptides (HDPs) are considered to be lead compounds for novel anti-sepsis and anti-inflammatory agents. However, development of drugs based on HDPs has been hampered by problems with toxicity and low bioavailability due to in vivo proteolysis. Here, a subclass of proteolytically stable HDP mimics consisting of lipidated α-peptide/β-peptoid oligomers was investigated for their effect on neutrophil function. The most promising compound, Pam-(Lys-βNSpe)₆-NH₂, was shown to inhibit formyl peptide receptor 2 (FPR2) agonist-induced neutrophil granule mobilization and release of reactive oxygen species. The potency of Pam-(Lys-βNSpe)₆-NH₂ was comparable to that of PBP10, the most potent FPR2-selective inhibitor known. The immunomodulatory effects of structural analogues of Pam-(Lys-βNSpe)₆-NH₂ emphasized the importance of both the lipid and peptidomimetic parts. By using imaging flow cytometry in primary neutrophils and FPR-transfected cell lines we found that a fluorescently labelled analogue of Pam-(Lys-βNSpe)₆-NH₂ interacted selectively with FPR2. Furthermore the interaction between Pam-(Lys-βNSpe)₆-NH₂ and FPR2 was found to prevent binding of the FPR2-specific activating peptide agonist Cy5-WKYMWM, while the binding of a FPR1-selective agonist was not inhibited. To our knowledge, Pam-(Lys-βNSpe)₆-NH₂ is the first HDP mimic found to inhibit activation of human neutrophils via direct interaction with FPR2. Hence, we consider Pam-(Lys-βNSpe)₆-NH₂ to be a convenient tool in the further dissection of the role of FPR2 in inflammation and homeostasis as well as for investigation of the importance of neutrophil stimulation in anti-infective therapy involving HDPs.