Research paper
Design, synthesis, and evaluation of amphiphilic sofalcone derivatives as potent Gram-positive antibacterial agents

https://doi.org/10.1016/j.ejmech.2020.112596Get rights and content

Highlights

  • Discovery of amphiphilic chalcone derivatives as small molecular membrane-active antimicrobial peptidomimetics.

  • Compound 14 exhibited potent in vitro antibacterial activity against Gram-positive bacteria.

  • Compound 14 exhibited low hemolytic activity, high membrane selectivity, and rapid bactericidal activity.

  • Compound 14 demonstrated excellent efficacy in a murine bacterial keratitis model induced by Gram-positive bacteria.

Abstract

New antimicrobial agents are urgently needed to overcome drug-resistant bacterial infections. Here we describe the design, synthesis and evaluation of a new class of amphiphilic sofalcone compounds as antimicrobial peptidomimetics. The most promising compound 14, bearing two arginine residues, showed poor hemolytic activity, low cytotoxicity, and excellent antimicrobial activity against Gram-positive bacteria, including MRSA. Compound 14, had good stability in various salt conditions, killed bacteria rapidly by directly disrupting bacterial cell membranes and was slow at developing bacterial resistance. Additionally, compound 14 exhibited effective in vivo efficacy in the murine model of bacterial keratitis caused by Staphylococcus aureus ATCC29213. Our studies suggested that compound 14 possessed promising potential to be used as a novel antimicrobial agent to combat drug-resistant Gram-positive bacteria.

Introduction

Infections by drug-resistant bacterial infection are a serious threat to human health, which has become a critical global healthcare problem [1,2]. The rapidly developing bacterial resistance is widely associated with clinical treatment failure, additional mortality, and high healthcare costs [[3], [4], [5]]. The efficacy of commercial antibiotics is being greatly reduced or even eliminated due to the emergence of bacterial resistance [6]. For instance, approximate 50% of the Staphylococcus aureus (S. aureus) strains isolated from US hospitals are resistant to methicillin, and some of them are even resistant to the last-resort antibiotics vancomycin and daptomycin [7,8]. The approaches that bacteria employ to develop drug resistance include (1) inactivation of antibiotics via enzymatic degradation or inactivation, (2) the formation of bacterial biofilm, (3) antibiotic target mutation, (4) modification of membranes permeability and (5) expression of efflux pumps [[9], [10], [11]]. The currently available antimicrobial agents including β-lactams, aminoglycosides, macrolides, oxazolidinones, glycopeptides, rifampicins, tetracyclines and quinolones, inhibit bacteria by targeting the bacterial intracellular components via a biochemical pathway [12]. However, these targets are susceptible to bacterial resistance [12]. Therefore, it is an urgent need for developing new classes of antimicrobial agents with new molecular entities (NME)or new modes of action [13].

Antimicrobial peptides (AMPs), also termed as host defense peptides, were considered to be the promising choice for antimicrobial agents [[14], [15], [16]]. As a key component of the host immune system, AMPs have been widely found in animals, plants, and bacteria [15,17]. AMPs generally contain cationic and hydrophobic residues, forming a highly amphiphilic topological structure, which plays a vital role in the membrane selectivity and antibacterial activity [16,18,19]. Additionally, AMPs have critical advantages over traditional antibiotics, such as a rapid bacterial killing, low toxicity to human cells, and a low probability of developing bacterial resistance [15,20,21]. However, AMPs have several obvious disadvantages including high costs, poor stability and systemic toxicity, seriously hindering their clinical applications [22,23].

Small-molecule based antimicrobial peptidomimetics which mimic the molecular structure and antimicrobial mechanism of AMPs, are expected to overcome the inherent disadvantages of AMPs and become a new generation of antimicrobial therapeutics [[24], [25], [26]]. Several peptidomimetics were developed using a non-peptide skeleton, such as m-phenylene ethynylenes [27], cationic steroid antibiotics [28], acrylamide oligomer [29], and norbornane-based antimicrobial peptidomimetics [30]. These peptidomimetics displayed potent broad-spectrum antibacterial activity and a low tendency of developing bacterial resistance. Moreover, they display great stability under physiological conditions and are not susceptible to proteolytic degradation, so they can remain potent in vivo antimicrobial activity [[27], [28], [29], [30], [31]]. Previously, our group have focused on the design and development of xanthone-based peptidomimetics and successfully prepared several highly effective and low toxicity antimicrobial candidate drugs [32,33].

Considering these issues, we designed and synthesized a series of amphiphilic sofalcone (1) derivatives as membrane-targeting antimicrobial peptidomimetics. Sofalcone (Scheme 1), an isoprenyl chalcone derivative, is prepared from soforadine isolated from the root of the plant Sophora subprostrata [34]. Sofalcone is mainly used in the treatment of gastric ulcer and gastritis [35]. To our knowledge, this is the first report on the discovery of sofalcone-based antimicrobials. In addition, natural or synthetic chalcone derivatives have been reported to have various pharmacological properties, including antibacterial and antifungal activities [[36], [37], [38]]. Our design hypothesis is that the incorporation of cationic groups to sofalcone can greatly increase the interaction between the sofalcone derivative and the negatively charged bacterial membrane through electrostatic interaction. Meanwhile, the hydrophobic moieties including the chalcone core and two isoprenyl groups, would enhance the interaction between sofalcone derivatives and the phospholipid bilayer in the bacterial membrane, thereby promoting cationic amphiphilic sofalcone derivatives to penetrate into the bacterial lipid membrane. By contrast, the interaction between the sofalcone derivatives and the electrically neutral mammal cell membrane should be very weak. Therefore, cationic sofalcone derivatives can selectively act on bacteria. Additionally, sofalcone can be applied as a special type of non-natural hydrophobic amino acid residue to design antimicrobial peptidomimetics. Since sofalcone has good pharmacokinetic properties, high safety, and excellent drugability [39], it could serve as a privileged structure for the discovery and development of antibacterial drugs. In this study, the antibacterial and hemolytic activity of prepared amphiphilic sofalcone derivatives were screened by testing their minimum inhibitory concentrations (MICs) and HC50 values (concentration of compound required to lyse 50% rabbit red blood cells). Finally, time-kill kinetics, drug resistance development, cytotoxicity toward mammalian cells, mode of action, and in vivo efficacy of the selected sofalcone compound were studied.

Section snippets

Chemistry

The general synthetic routes used for the synthesis of cationic amphiphilic sofalcone derivatives are shown in Scheme 1, Scheme 2. The starting material sofalcone was reacted to the corresponding basic amino acid or aliphatic amine in the presence of HATU or HBPyU to yield compounds 211 and 1314. The key intermediate 12 was prepared by treating compound 9 with LiOH. The acid 12 was then coupled with 3-(dimethylamino)-1-propylamine, or 1,3-propanediamine using HBPyU to produce compounds 15 and

Conclusions

In summary, a new series of amphiphilic sofalcone compounds as small-molecule based antimicrobial peptidomimetics has been designed, synthesized, and evaluated. The incorporation of different types of amine moieties can dramatically enhance the antibacterial activity of sofalcone derivatives. The most promising compound (14) displayed potent activity against Gram-positive bacteria (MICs = 1.56 μg/mL), including MRSA, comparable to that was observed with the commercial antibacterial drug

Experimental section

Sofalcone (purity of 97%) was purchased from Shanghai Bidepharm (Shanghai, China). Vancomycin, Triton X-100, DMSO, sodium chloride, potassium chloride, calcium chloride, and magnesium chloride were purchased from Shanghai Aladdin (Shanghai, China). Mueller-Hinton agar (MHA) and MHB were purchased from Qingdao Hopebio (Shandong, China). Chloramphenicol was purchased from Shanghai Macklin (Shanghai, China). PBS was purchased from Beijing HyCone (Beijing, China). RPMI Medium 1640 basic were

Declaration of competing interest

The authors declare no conflict of interest.

Acknowledgements

This work was supported by the Talent Fund for High-Level University Construction of Guangzhou (B195002009029 to Liu, S.P., B195002009030 to Lin, S.M.) and the National Natural Science Foundation of China (21907019to Lin, S.M.).

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