Comparative in vitro study on cytotoxicity of recombinant β-hairpin peptides.

Natural antimicrobial peptides (AMPs) are important components of the innate immune system with a wide spectrum of biological activity. In this study, we investigated the cytotoxic effect of three recombinant β-hairpin cationic AMPs: arenicin-1 from the polychaeta Arenicola marina, tachyplesin I from the horseshoe crab Tachypleus tridentatus, and gomesin from the spider Acanthoscurria gomesiana. All the three β-hairpin AMPs were overexpressed in Escherichia coli. Different cell lines were incubated with various concentrations of the investigated AMPs in order to evaluate their cytotoxic activity. Double staining with subsequent flow cytometric analysis was used to determine the predominant way of cell death mediated by each AMP. Hemolytic activity of the peptides was tested against fresh human red blood cells. Our results indicated that all the three AMPs exhibited significant cytotoxic effect against cancer cells that varied depending on the cell line type and, in most cases, on the presence of serum components. Flow cytometric analysis implicitly indicated that tachyplesin I mostly promoted late apoptosis/necrosis, while arenicin-1 and gomesin induced early apoptosis under the same conditions. Tachyplesin I proved to be the most promising therapeutic candidate as it displayed the highest specific cytotoxicity against cancer cell lines, independent of the serum presence.