In vitro antibacterial activity of acyl-lysyl oligomers against Helicobacter pylori.

The gastric pathogen Helicobacter pylori has developed resistance to virtually all current antibiotics; thus, there is a pressing need to develop new anti-H. pylori therapies. The goal of this work was to evaluate the antibacterial effect of oligo-acyl-lysyl (OAK) antimicrobial peptidomimetics to determine if they might represent alternatives to conventional antibiotic treatment of H. pylori infection. A total of five OAK sequences were screened for growth-inhibitory and/or bactericidal effects against H. pylori strain G27; four of these sequences had growth-inhibitory and bactericidal effects. The peptide with the highest efficacy against strain G27, C12K-2beta12, was selected for further characterization against five additional H. pylori strains (26695, J99, 7.13, SS1, and HPAG1). C12K-2beta12 displayed MIC and minimum bactericidal concentration (MBC) ranges of 6.5 to 26 microM and 14.5 to 90 microM, respectively, across the six strains after 24 h of exposure. G27 was the most sensitive H. pylori strain (MIC = 6.5 to 7 microM; MBC = 15 to 20 microM), whereas 26695 was the least susceptible strain (MIC = 25 to 26 microM; MBC = 70 to 90 microM). H. pylori was completely killed after 6 to 8 h of incubation in liquid cultures containing two times the MBC of C12K-2beta12. The OAK demonstrated strong in vitro stability, since efficacy was maintained after incubation at extreme temperatures (4 degrees C, 37 degrees C, 42 degrees C, 50 degrees C, 55 degrees C, 60 degrees C, and 95 degrees C) and at low pH, although reduced killing kinetics were observed at pH 4.5. Additionally, upon transient exposure to the bacteria, C12K-2beta12 showed irreversible and significant antibacterial effects and was also nonhemolytic. Our results show a significant in vitro effect of C12K-2beta12 against H. pylori and suggest that OAKs may be a valuable resource for the treatment of H. pylori infection.

Design and characterization of a broad -spectrum bactericidal acyl-lysyl oligomer.

Previously characterized chemical mimics of host defense peptides belonging to the oligo-acyl-lysyl (OAK) family have so far failed to demonstrate broad-spectrum antibacterial potency combined with selectivity toward host cells. Here, we investigated OAK sequences and characterized a promising representative, designated C(12)K-3beta(10), with broad-spectrum activity (MIC(90) = 6.2 microM) and low hemotoxicity (LC(50) > 100 microM). Whereas C(12)K-3beta(10) exerted an essentially bactericidal effect, E. coli bacteria were killed faster than S. aureus (minutes versus hours). Mechanistic studies addressing this difference revealed that unlike E. coli, S. aureus bacteria undergo a transient rapid bactericidal stage that over time converts to a bacteriostatic effect. This behavior was dictated by interactions with cell wall-specific components. Preliminary efficacy studies in mice using the thigh infection model demonstrated the OAK's ability to significantly affect bacterial viability upon single-dose systemic treatment (2 mg/kg).

Structure-activity relationships of antibacterial acyl-lysine oligomers.

We describe structure-activity relationships that emerged from biophysical data obtained with a library of antimicrobial peptide mimetics composed of 103 oligoacyllysines (OAKs) designed to pin down the importance of hydrophobicity (H) and charge (Q). Based on results obtained with OAKs displaying minimal inhibitory concentration < or = 3 microM, the data indicate that potent inhibitory activity of the gram-negative Escherichia coli and the gram-positive Staphylococcus aureus required a relatively narrow yet distinct window of HQ values where the acyl length played multiple and critical roles, both in molecular organization and in selective activity. Thus, incorporation of long-but not short-acyl chains within a peptide backbone is shown to lead to rigid supramolecular organization responsible for poor antibacterial activity and enhanced hemolytic activity. However, sequence manipulations, including introduction of a tandem lysine motif into the oligomer backbone, enabled disassembly of aggregated OAKs and subsequently revealed tiny, nonhemolytic, yet potent antibacterial derivatives.