Research paper
Optimization of biaryloxazolidinone as promising antibacterial agents against antibiotic-susceptible and antibiotic-resistant gram-positive bacteria

https://doi.org/10.1016/j.ejmech.2019.111781Get rights and content

Highlights

  • Novel biaryloxazolidinone analogues were designed to improve the metabolic stability.

  • Compound 14a-7 exhibited a MIC value of 0.125 μg/mL against S.aureus.

  • Compound 14a-7 was stable in human liver microsome.

  • Compound 14a-7 exhibited lower inhibitory activity against human MAO-A compared to linezolid.

Abstract

We previously discovered a series of novel biaryloxazolidinone analogues bearing a hydrazone moiety with potent antibacterial activity. However, the most potent compound OB-104 exhibited undesirable chemical and metabolic instability. Herein, novel biaryloxazolidinone analogues were designed and synthesized to improve the chemical and metabolic stability. Compounds 6a-1, 6a-3, 14a-1, 14a-3 and 14a-7 showed significant antibacterial activity against the tested Gram-positive bacteria as compared to radezolid and linezolid. Further studies indicated that most of them exhibited improved water solubility and chemical stability. Compound 14a-7 had MIC values of 0.125–0.25 μg/mL against all tested Gram-positive bacteria, and showed excellent antibacterial activity against clinical isolates of antibiotic-susceptible and antibiotic-resistant bacteria. Moreover, it was stable in human liver microsome. From a safety viewpoint, it showed non-cytotoxic activity against hepatic cell and exhibited lower inhibitory activity against human MAO-A compared to linezolid. The potent antibacterial activity and all these improved drug-likeness properties and safety profile suggested that compound 14a-7 might be a promising drug candidate for further investigation.

Introduction

The emergence and outbreak of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant Enterococci (VRE) and penicillin-resistant Streptococcus pneumoniae (PRSP) made antibiotic-resistant infections a critical healthcare concern worldwide [[1], [2], [3]]. Antibiotic-resistant infections resulted in increasing patient mortality and social economic burden. In the United States, there were approximately 19000 people dying each year from MRSA, which is known as the first superbug [4,5]. Finding effective ways to resolve antibiotic-resistant crisis is an urgent task. Karen B et al. summarized the approaches for resolving this global health crisis, of which, finding antibiotics with new structural classes or mechanisms was the main concern for drug chemists [6]. Due to developing antibiotics with new mechanisms is a long-term work, we focus our efforts on modifications of old antibiotics with known mechanisms to overcome drug resistance.

The ribosomes synthesize proteins and the bacterial ribosome is one of the main targets of antibiotics [7]. Many natural, semi-synthetic and synthetic antibiotics bind to bacterial ribosome to inhibit bacterial protein synthesis. In recent years, important progresses on the crystal structures of antibiotics binding to ribosome have been reported, which give drug chemists insight into the mechanism of action of ribosome-targeting antibiotics. And these progresses are crucial for drug chemists to improve the potency and overcome bacterial resistance by further drug optimization [8,9]. Linezolid (Figure 1), the first oxazolidinone drug, was approved in 2000 by FDA for the treatment of antibiotic-resistant Gram-positive infections, including MRSA, PRSP, and VRE [10,11]. It binds to 23S RNA of the 50S ribosomal subunit to inhibit bacterial protein biosynthesis, which belongs to a ribosome-targeting antibiotic [12]. However, bacterial resistance to linezolid has been discovered due to the overuse of linezolid. Resistance to linezolid was mediated by a number of mechanisms, such as alteration of microbial permeability and target mutation [7,13].

The C-ring of linezolid and many other ribosome-targeting antibiotics, such as chloramphenicol and clindamycin, share an overlapping region on the binding sites of ribosomal RNA, which inspired many research groups to develop oxazolidinone derivatives with C-ring and D-ring to overcome bacterial resistance and to improve the potency [7,14,15]. Radezolid was developed based on the crystal structures of linezolid binding to ribosomal RNA and it exhibited more potent activity against antibiotic-resistant Gram-positive bacteria than linezolid [16,17]. As reported, oxazolidinone derivatives containing biaryl scaffold showed increased antibacterial potency against Gram-positive bacteria, especially antibiotic-resistant Gram-positive bacteria compared with linezolid [[18], [19], [20], [21], [22]]. These compounds revealed outstanding antibacterial activity by forming π–π stacking or hydrogen bond interactions with the 50S subunit .

In our previous work, we discovered compound OB-104, a biaryloxazolidinone derivative bearing a hydrazone moiety, showed excellent antibacterial activity against five tested Gram-positive bacteria in vitro, which were a 15- to 30-fold increase compared to linezolid in activity against linezolid-resistant Enterococcus faecalis [23]. However, compound OB-104 was unstable under simulated gastric acid condition (pH = 1.2) and showed short half-life time (72.85 min) in human liver microsome. Further pharmacokinetic study in rats by intragastric administration revealed that the half-life time (T1/2) of OB-104 was 2.7 h. As shown in Fig. 2, the metabolites of OB-104 in rats were further studied by UPLC-QTOF/MS method. Four main metabolites were found in plasma of rats given intragastric administration of OB-104. The main metabolite M-1 was confirmed by MS spectra and the reference substance, which was formed by the hydroxylation of C-ring at C-3 position. At the same time, nitrogen demethylation occurred on the piperazine ring to form metabolite M-2. The Cdouble bondN bond of the hydrazone moiety of OB-104 was hydrolyzed to the aldehyde metabolite M-3, which was further oxidized to the carboxyl metabolite M-4. Metabolite M-1 with hydroxyl group at C-ring attracted our interest, and it was synthesized and its antibacterial activity was evaluated. Remarkably, M-1 showed significant antibacterial activity against the tested Gram-positive bacteria (data are shown in Table 1). As we know, the electronic or sterically hindered effects of substituents in C-ring may have a large effect on the stability of hydrazone bond. Basing on the structure of M-1 and other reported biaryloxazolidinone analogues [24,25], we focused on the structural optimization of compound OB-104 in following strategies, which are shown in Fig. 3: (I) introduction of a sterically hindered substituent or hydrogen bond donor (OH)/hydrogen bond receptor (F) at the ortho-position of the hydrazone bond on the C-ring (compounds 6a-6e and 16). (Ⅱ) replacement of C-ring with nitrogen-containing heterocyclic ring (compounds 14a-14b and 15a), of which, the electron-withdrawing effect of nitrogen atom was expected to improve the chemical stability of hydrazone bond [21] .

Section snippets

Chemistry

The synthesis of target compounds 6 is depicted in Scheme 1. 4-Bromo-3-fluoroaniline was acylated with methyl chloroformate to give 2 with a high yield. Intermediate 3 was obtained by cyclization reaction of intermediate 2 with (S)-1-((acetylamino)methyl)-2-chloroethyl acetate. Intermediate 4 was prepared by Miyaura-Ishiyama-Hartwig borylation reaction of intermediate 3 with bis(pinacolato)diboron, which was subjected to substituted p-bromobenzaldehydes to give the corresponding

Evaluation of in vitro antibacterial activity

All the target biaryloxazolidinone compounds were screened for their in vitro antibacterial activity against Gram-positive, antibiotic-susceptible and antibiotic-resistant bacteria strains (S.aureus ATCC29213, MRSA, MSSA, LREF and VRE), using linezolid and radezolid as positive control. The results of minimum inhibitory concentrations (MICs, μg/mL) determined by the broth liquid microdilution method were summarized in Table 1, Table 2.

As shown in Table 1, we firstly synthesized the putative

Conclusions

In summary, we described the structural optimization of biaryloxazolidinone analogue OB-104, which was reported in our previous work. Compound OB-104 was unstable in simulated gastric acid conditions (pH = 1.2) and human liver microsome. However, the metabolite M-1 (6a-1) of OB-104 showed significant antibacterial activity, which prompted us to conduct structural optimization strategies to improve the stability of hydrazone moiety while maintaining or enhancing antibacterial activity.

Chemistry

All materials were obtained from commercial suppliers and were used without further purification unless stated otherwise. Reactions’ time of the compounds were monitored by TLC (silica gel GF254). Column chromatography was run on silica gel (200–300 mesh) from Qingdao Ocean Chemicals (Qingdao, Shandong, China). All melting points were measured with a Büchi Melting Point B-540 apparatus (Büchi Labortechnik, Flawil, Switzerland) and were uncorrected. Mass spectra (MS) were taken in ESI mode on

Acknowledge

Supported by Basic Research Projects of Higher Education Institutions in Liaoning Province (2017LZD06), and LiaoNing Revitalization Talents Program (XLYC1805014).

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    Yachuang Wu and Xiudong Ding contributed equally to this work.

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