A Systematic Study of the Stability, Safety, and Efficacy of the de novo Designed Antimicrobial Peptide PepD2 and Its Modified Derivatives Against Acinetobacter baumannii.

Sung-Pang Chen,Tsai-Chen Yang,Chun-Hung Lin,Pin-Shin Kuo,Ming-Yen Hsieh,Eric H-L Chen,Kung-Ta Lee,Hau-Ming Jan,Rita P-Y Chen,Sheng-Yung Yang

doi:10.3389/fmicb.2021.678330

Abstract

Searching for new antimicrobials is a pressing issue to conquer the emergence of multidrug-resistant (MDR) bacteria and fungi. Antimicrobial peptides (AMPs) usually have antimicrobial mechanisms different from those of traditional antibiotics and bring new hope in the discovery of new antimicrobials. In addition to antimicrobial activity, stability and target selectivity are important concerns to decide whether an antimicrobial peptide can be applied in vivo. Here, we used a simple de novo designed peptide, pepD2, which contains only three kinds of amino acid residues (W, K, L), as an example to evaluate how the residues and modifications affect the antimicrobial activity against Acinetobacter baumannii, stability in plasma, and toxicity to human HEK293 cells. We found that pepI2 with a Leu→Ile substitution can decrease the minimum bactericidal concentrations (MBC) against A. baumannii by one half (4 μg/mL). A D-form peptide, pepdD2, in which the D-enantiomers replaced the L-enantiomers of the Lys(K) and Leu(L) residues, extended the peptide half-life in plasma by more than 12-fold. PepD3 is 3-residue shorter than pepD2. Decreasing peptide length did not affect antimicrobial activity but increased the IC50 to HEK293 cells, thus increased the selectivity index (SI) between A. baumannii and HEK293 cells from 4.7 to 8.5. The chain length increase of the N-terminal acyl group and the Lys→Arg substitution greatly enhanced the hemolytic activity, hence those modifications are not good for clinical application. Unlike colistin, the action mechanism of our peptides relies on negatively charged lipids rather than lipopolysaccharides. Therefore, not only gram-negative bacteria but also gram-positive bacteria can be killed by our peptides.

Highlights

Over the last three decades, many microbial isolates resistant to penicillins, cephalosporins, carbapenems, fluoroquinolones, and aminoglycosides have been reported
The culture after peptide treatment was spotted on an Müller–Hinton agar (MHA) plate, and the peptide concentration that could kill all the bacteria was defined as the minimum bactericidal concentrations (MBC)
Antimicrobial peptides (AMPs) have the advantages of broad-spectrum antimicrobial activity and do not induce microbial resistance

Summary

Introduction

Over the last three decades, many microbial isolates resistant to penicillins, cephalosporins, carbapenems, fluoroquinolones, and aminoglycosides have been reported. The emergence of multidrug-resistant (MDR) bacteria and fungi is a severe threat to global human health. A peptide antibiotic against Gram-negative bacteria, was discovered in the 1940s. Due to the emergence of MDR bacteria, polymyxins (colistin and its derivatives) are clinically used as the last-line therapy for drug-resistant bacterial infection despite the possibility of colistin-induced acute kidney injury (Evans et al, 1999; Falagas et al, 2005; Falagas and Kasiakou, 2006; Zavascki et al, 2007; Lim et al, 2010; Gai et al, 2019; Yousfi et al, 2019). Colistin targets lipopolysaccharide (LPS) in the outer membrane of Gram-negative bacteria, displaces divalent cations (Ca2+ and Mg2+), changes membrane permeability, and disrupts the bacterial membrane. Colistin resistance rapidly emerged by a change in the lipid composition in the bacterial membrane to reduce its binding affinity with colistin (Bialvaei and Samadi Kafil, 2015)

Methods

Results

Conclusion