Dermaseptin AA-3-1

Over the last 20 years, natural peptides playing a key role in defense mechanisms and innate immunity have been isolated from unicellular organisms. Amphibian skin secretes dermaseptins, 24-34 amino acids in length that have a wide antimicrobial spectrum incorporating yeast, fungi, protozoa, bacteria and enveloped viruses. The anti-rabies virus (RABV) activity of dermaseptins S3 (30aa) and S4 (28aa) from Phyllomedusa sauvagei has been investigated, and further dissected its molecular basis by comparing punctual mutation or deletion of S4 analogues. The results showed that: (1) S4 is more active than S3 against RABV infection, 89% versus 38% inhibition at 7.5 μM; (2) the 5 NH2-aa of S4 are crucial for its inhibitory potential (S46-28 lost any inhibition) but the COOH terminus stabilizes the inhibitory potential (S41-16 showed only 23% inhibition at 7.5 μM); (3) there is a correlation between viral inhibition and dermaseptin cytotoxicity, which remains however moderated for BSR cells (≤12% at 10 μM). A single mutation in position 4 (S4M4K) slightly reduced cytotoxicity while keeping its antiviral activity, 97% at 7.5 μM. S4 and S4M4K showed an antiviral activity in vitro when provided 1 h after infection. In vivo experiments in mice by intramuscular injection of non-toxic doses of dermaseptin S4M4K 1 h post-infection by a lethal dose of RABV at the same site allowed more than 50% improvement in mice survival. This study highlights the potential interest of dermaseptins as non-expansive alternatives to rabies immunoglobulins for the treatment of rabies that continues to claim about 60,000 human lives per year worldwide, almost exclusively in developing countries.

Antimicrobial resistance has been an increasing public health threat in recent years. Antimicrobial peptides are considered as potential drugs against drug-resistant bacteria because they are mainly broad-spectrum and are unlikely to cause resistance. In this study, a novel peptide was obtained from the skin secretion of Agalychnis callidryas using the "shotgun" cloning method. The amino acid sequence, molecular weight, and secondary structure of Dermaseptin-AC were determined. The in vitro antimicrobial activity, hemolysis, and cytotoxicity of Dermaseptin-AC were evaluated. MICs and minimum bactericidal concentrations (MBCs) of Dermaseptin-AC against seven different bacterial strains ranged between 2 ~ 4 μM and 2 ~ 8 μM. The HC50 (50% maximum hemolysis concentration) of Dermaseptin-AC against horse erythrocytes was 76.55 μM. The in vivo anti-MRSA effect was tested on immune-suppressed MRSA pneumonia in mice. Dermaseptin-AC showed anti-MRSA effects similar to the same dose of vancomycin (10 mg/kg body weight). Short-term (7 days of intraperitoneal injection, 10 mg/kg body weight) in vivo safety evaluation of Dermaseptin-AC was tested on mice. The survival rate during the 7-day injection was 80%. Dermaseptin-AC showed no obvious effect on the liver, heart, spleen, kidney, and blood, but did induce slight pulmonary congestion. The skin safety of Dermaseptin-AC was evaluated on wounds on the back skin of a rat, and no irritation was observed. IMPORTANCE In this study, we discovered a new antimicrobial peptide, Dermaseptin-AC, and studied its in vitro and in vivo antimicrobial activity. These studies provide some data for finding new antimicrobial peptides for overcoming antimicrobial resistance. Dermaseptin-AC showed strong broad-spectrum antibacterial activity and relatively low hemolysis, and was more cytotoxic to cancer cells than to normal cells. Dermaseptin-AC was active in vivo, and its anti-MRSA effect was similar to that of vancomycin when administered by intraperitoneal injection. Safety studies found that continuous injection of Dermaseptin-AC may cause mild pulmonary congestion, while there was no obvious irritation when it was applied to skin wounds. Chronic wounds are often accompanied by high bacterial burdens and, at the same time, antimicrobial resistance is more likely to occur during repeated infections and treatments. Therefore, developing Dermaseptin-AC to treat chronic wound infection may be an attractive choice.

Dermaseptin, a 34-amino acid residue cationic peptide, was recently shown to inhibit the growth of pathogenic fungi responsible for severe opportunistic infections accompanying immunodeficiency syndrome and the use of immunosuppressive agents. To improve our understanding of the mechanism by which dermaseptin exerts its potent antimicrobial action, a series of either NH2- or COOH-terminally truncated analogs was synthesized. These analogs were evaluated for their ability to inhibit the growth of various pathogenic agents in culture medium. Dermaseptin exerted a lytic action upon bacteria, protozoa, yeasts, and filamentous fungi at micromolar concentrations. No inhibition of proliferation was observed with human KB cells, and dermaseptin did not lyse guinea pig lymphocytes or rabbit erythrocytes at doses up to 200 micrograms/ml. Shortening the peptide chain of dermaseptin to dermaseptin-(3-34) slightly reduced the activity of the peptide, while further reduction of the chain length to residues 14-34, 16-34, 20-34, and 28-34 yielded peptide derivatives devoid of antimicrobial activity. On the other hand, lengthening the peptide chain starting from residues 1-4 to residues 1-8 and 1-18 led to a progressive recovery of the activity of the parent molecule. Whereas the central core of dermaseptin (residues 10-19) was virtually inactive, alteration of the COOH-terminal carboxylic group of dermaseptin-(1-18) to a carboxamide yielded a peptide exhibiting enhanced antimicrobial potency, yet displaying even less in vitro toxicity compared with dermaseptin. Overall, the data indicate that molecular elements responsible for the exceptional antimicrobial potency of dermaseptin are to be traced to the NH2-terminal alpha-helical amphipathic segment spanning residues 1-18 of the molecule. Dermaseptin-(1-18)-NH2 may therefore be considered as a useful and highly tractable tool for identifying key features responsible for membrane permeabilization and as a starting point for the design of new therapeutic agents.