Conjugate of Enkephalin and Temporin Peptides as a Novel Therapeutic Agent for Sepsis

Abstract

Antimicrobial peptides (AMPs) exhibit a wide spectrum of actions, ranging from a direct bactericidal effect to multifunctional activities as immune effector molecules. The aim of this study was to examine the anti-inflammatory properties of a DAL-PEG-DK5 conjugate composed of a lysine-rich derivative of amphibian temporin-1CEb (DK5) and dalargin (DAL), the synthetic Leu-enkephalin analogue. Detailed study of the endotoxin-neutralizing activity of the peptide revealed that DAL-PEG-DK5 interacts with LPS and the LPS binding protein (LBP). Moreover, DAL-PEG-DK5 prevented dimerization of TLR4 at the macrophage surface upon LPS stimulation. This inhibited activation of the NF-κB signaling pathway and markedly reduced pro-inflammatory cytokine production. Finally, we showed that aggregation of DAL-PEG-DK5 into amyloid-like structures induced by LPS neutralized the endotoxin proinflammatory activity. Consequently, DAL-PEG-DK5 reduced morbidity and mortality in vivo, in a mouse model of endotoxin-induced septic shock. Collectively, the data suggest that DAL-PEG-DK5 is a promising therapeutic compound for sepsis.

The Bactericidal Activity of Temporin Analogues Against Methicillin Resistant Staphylococcus aureus

“…Although we documented the direct bactericidal activity of DAL-PEG-DK5 and its efficient permeation of eukaryotic cells, we cannot rule out the possibility that an indirect mechanism contributes to the eradication of intracellular staphylococci from macrophages. Such a scenario is highly possible in light of the finding that DAL-PEG-DK5 exerts significant immunomodulatory effects on human macrophages [19].…”

“…In conclusion, we demonstrate that modification of temporin-1CEb to generate a peptide conjugate, DAL-PEG-DK5, makes the peptide an attractive candidate lead compound for the generation of a new agent to treat MRSA-related skin infections. Detailed in vivo studies are needed to confirm this hypothesis; however, it should be mentioned that we previously documented the stability and activity of this peptide in a murine model of sepsis [19]. The peptide inhibits planktonic growth of different clinical MRSA strains and kills them in the biofilm.…”

“…Here, we focused on temporin-1CEb, a naturally occurring 12-residue peptide derived from frog skin secretions that display antimicrobial activity against a broad spectrum of gram-positive and (albeit to a lesser extent) gram-negative bacteria [17,18]. Moreover, temporins, including temporin-1CEb, play a significant anti-inflammatory role both in vitro and in vivo [19]. The emergence of MRSA has resulted in an epidemic of SSTIs; thus, we created a library of compounds based on an analogue of temporin-1CEb (DK5) conjugated to peptides showing cytoprotective, pro-proliferative, and pro-migratory effects against human keratinocytes (cells that facilitate wound-healing) [20,21,22,23,24].…”

Application of the In Vitro HoxB8 Model System to Characterize the Contributions of Neutrophil–LPS Interaction to Periodontal Disease

“…Where indicated, cells were pre-treated with LPS inactivator polymyxin B (100 μg/mL, Invivogen, cat no. tlrlpmb) for 30 min prior to LPS addition, as published before [ 31 ].…”

Nanosystems as Vehicles for the Delivery of Antimicrobial Peptides (AMPs)

“…During 2017–2018, some review articles, including state-of-the-art polymer-based strategies to improve in vivo biocompatibility and delivery systems for AMPs, were published [10,46,47]. Furthermore, in the following paragraphs, we gather an update on the most recently published works reporting the use of polymer nanoparticles [48,49,50], nanofibers [51,52], multilayers [53,54], polymer-coated surfaces [55,56], and polymer conjugates [57,58,59,60,61,62,63] for AMP delivery.…”