Bacterial membrane disrupting dodecapeptide SC4 improves survival of mice challenged with Pseudomonas aeruginosa

Abstract

Background: Dodecapeptide SC4 is a broad-spectrum bactericidal agent that functions by disintegrating bacterial membranes and neutralizing endotoxins. For insight into which SC4 amino acids are functionally important, we assessed Gram-negative bactericidal effects in structure-activity relationship experiments. Subsequently, SC4 was tested in a murine bacteremia model to combine and compare the efficacy with Zosyn, a first-line antibiotic against Pseudomonas aeruginosa (P. aeruginosa).

Methods: SC4 alanine-scanning analogs and their activities on were tested on P. aeruginosa. Survival studies in P. aeruginosa challenged mice were executed to monitor overall efficacy of SC4 and Zosyn, as a single modality and also as combination treatment. ELISAs were used to measure blood serum levels of selected inflammatory cytokines during treatment.

Results: Cationic residues were found to play a crucial role in terms of bactericidal activity against P. aeruginosa. In vivo, while only 9% (3/34) of control animals survived to day two and beyond, 44% (12/27) to 41% (14/34) of animals treated with SC4 or Zosyn, respectively, survived beyond one week. Combination treatment of SC4 and Zosyn demonstrated improved survival, i.e. 60% (12/20). The TNFα, IL-1, and IL-6 serum levels were attenuated in each treatment group compared to the control group.

Conclusions: These data show that combination treatment of SC4 and Zosyn is most effective at killing P. aeruginosa and attenuating inflammatory cytokine levels in vivo.

General significance: Combination treatment of SC4 and Zosyn may be useful in the clinic as a more effective antibiotic therapy against Gram-negative infectious diseases.

Fig. 1

Examples of dose-response curves SC4 and alanine-scanning peptide analogues. Bactericidal activities of SC4 alanine-scanning peptide analogues against Pseudomonas aeruginosa (P. a.). Lines are sigmoidal curve fits used to determine LD₅₀ values.

Fig. 2

Combination of SC4 and Zosyn show enhanced protection against bacteria. SC4 and Zosyn were used in mouse bacteria model to assess in vivo efficacy against P.a.. The compounds were administered for 3 days by osmotic mini-pumps starting one day prior to i.p. injection of P.a. into C57/BL6 mice (n=20–34/group). Each mouse received a lethal dose of P.a. (300 µl of Klett 70). Survival of the mice were monitored for 14 days, however since no change in survival was noted after 120 hours, up to day 5 is plotted. *p < 0.03 (treatment group vs. control).

Fig. 3

Serum cytokine levels upon treatment with SC4, Zosyn and combination after P.a. bacteria challenge. SC4 (10 mg/kg) and Zosyn (400 mg/kg) were used in mouse bacteria model to assess cytokine serum levels of TNF-α (A), IL-1 (B) and IL-6 (C) after P.a. challenge. The compounds were administered by osmotic mini-pumps one day prior to i.p. injection of P.a. into C57/BL6 mice. Cytokines levels were assessed by ELISA (R&D systems, Minneapolis, MN). Data points shown are means of 2 independent experiments (n=2–4) ± SEM. *p < 0.03 (combination vs. control).