A Unique Hybridization Route to Access Hydrazylnaphthalimidols as Novel Structural Scaffolds of Multitargeting Broad-Spectrum Antifungal Candidates.

This study developed a class of novel structural antifungal hydrazylnaphthalimidols (HNs) with multitargeting broad-spectrum potential via multicomponent hybridization to confront increasingly severe fungal invasion. Some prepared HNs exhibited considerable antifungal potency; especially nitrofuryl HN 4a (MIC = 0.001 mM) exhibited a potent antifungal activity against Candida albicans, which is 13-fold higher than that of fluconazole. Furthermore, nitrofuryl HN 4a displayed low cytotoxicity, hemolysis and resistance, as well as a rapid fungicidal efficacy. Preliminary mechanistic investigations revealed that nitrofuryl HN 4a could inhibit lactate dehydrogenase to decrease metabolic activity and promote the accumulation of reactive oxygen species, leading to oxidative stress. Moreover, nitrofuryl HN 4a did not exhibit membrane-targeting ability; it could embed into DNA to block DNA replication but could not cleave DNA. These findings implied that HNs are promising as novel structural scaffolds of potential multitargeting broad-spectrum antifungal candidates for treating fungal infection.

Biocompatible 7-nitro-2,1,3-benzoxadiazole-embedded naphthalimide for exploring endogenous H2S in living cells.

Hydrogen sulfide (H2S) is a central signaling and antioxidant biomolecule involved in various biological processes. As inappropriate levels of H2S in the human body are closely related to various diseases, including cancer, a tool capable of detecting H2S with high selectivity and sensitivity in living systems is urgently required. In this work, we intended to develop a biocompatible and activatable fluorescent molecular probe for detecting H2S generation in living cells. The 7-nitro-2,1,3-benzoxadiazole-imbedded naphthalimide (1) probe presented here responds specifically to H2S and produces readily detectable fluorescence at 530 nm. Interestingly, probe 1 exhibited significant fluorescence responses to changes in endogenous H2S levels as well as high biocompatibility and permeability in living HeLa cells. This allowed for the real-time monitoring of endogenous H2S generation as an antioxidant defense response in the oxidatively stressed cells.

A bio-friendly biotin-coupled and azide-functionalized naphthalimide for real-time endogenous hydrogen sulfide analysis in living cells.

Hydrogen sulfide (H2S) is involved in various biological processes. Thereby, abnormal levels of H2S are reported to be related to various human diseases including cancer. Currently, many fluorescent probes are pioneered to detect H2S by taking advantages of naphthalimides' unique internal charge transfer (ICT) property. However, most probes often require a high content of organic solvents or surfactants, and are limited to the analysis of exogenous H2S treated externally in live cell studies, and have difficulties in analyzing endogenous H2S, thus limiting their practical use. In this study, we developed a bio-friendly biotin-coupled and azide-functionalized naphthalimide (1) as a fluorescent probe enabling real-time analysis of H2S in living system. Probe was able to provide a fluorescence at 545 nm via H2S-mediated azide reduction selectively without interference by biologically abundant constituents and pH effects. In a biological study using A549 cells, probe readily penetrated living cells without cytotoxicity, and unreacted probes showed almost no fluorescence, enabling real-time detection of H2S in living cells without requiring separate washing process. More importantly, under stimulation with various H2S inducers and inhibitors, probe was able to provide an effective fluorescence response against fluctuations in endogenous H2S, a key requirement for H2S studies. Probe 1 can be applied as a useful chemical tool and enables the analysis of H2S and the study of H2S-related cell functions in a variety of environments.

An unanticipated discovery towards novel naphthalimide corbelled aminothiazoximes as potential anti-MRSA agents and allosteric modulators for PBP2a.

Available therapeutic strategies are urgently needed to conquer multidrug resistance of MRSA. A visible effort was guided towards the advancement of novel antibacterial framework of naphthalimide corbelled aminothiazoximes, and desired to assert some insight on the conjunction of individual pharmacophore with distinct biological activities and unique action mechanism. Preliminary assessment displayed that dimethylenediamine derivative 13d presented a wonderful inhibition on MRSA (MIC = 0.5 µg/mL), and showed excellent membrane selectivity (HC50 > 200 µg/mL) from an electrostatic distinction of the electronegative bacterial membranes and the electroneutral mammalian membranes. Moreover, 13d could effectually relieve the development of MRSA resistance. Investigations into explaining the mechanism of anti-MRSA disclosed that 13d displayed strong lipase affinity, which facilitated its permeation into cell membrane, causing membrane depolarization, leakage of cytoplasmic contents and lactate dehydrogenase (LDH) inhibition. Meanwhile, 13d could exert interaction with DNA to hinder biological function of DNA, and disrupt the antioxidant defense system of MRSA through up-regulation of ROS subjected the strain to oxidative stress. In particular, the unanticipated mechanism for naphthalimide corbelled aminothiazoximes that 13d could suppress the expression of PBP2a by inducing allosteric modulation of PBP2a and triggering the open of the active site, was discovered for the first time. These findings of naphthalimide corbelled aminothiazoximes as a small-molecule class of anti-MRSA agents held promise in strategies for treatment of MRSA infections.

Strategies of Detecting Bacteria Using Fluorescence-Based Dyes.

Identification of bacterial strains is critical for the theranostics of bacterial infections and the development of antibiotics. Many organic fluorescent probes have been developed to overcome the limitations of conventional detection methods. These probes can detect bacteria with "off-on" fluorescence change, which enables the real-time imaging and quantitative analysis of bacteria in vitro and in vivo. In this review, we outline recent advances in the development of fluorescence-based dyes capable of detecting bacteria. Detection strategies are described, including specific interactions with bacterial cell wall components, bacterial and intracellular enzyme reactions, and peptidoglycan synthesis reactions. These include theranostic probes that allow simultaneous bacterial detection and photodynamic antimicrobial effects. Some examples of other miscellaneous detections in bacteria have also been described. In addition, this review demonstrates the validation of these fluorescent probes using a variety of biological models such as gram-negative and -positive bacteria, antibiotic-resistant bacteria, infected cancer cells, tumor-bearing, and infected mice. Prospects for future research are outlined by presenting the importance of effective in vitro and in vivo detection of bacteria and development of antimicrobial agents.

Novel naphthalimide nitroimidazoles as multitargeting antibacterial agents against resistant Acinetobacter baumannii.

AIM: The increasing emergence of resistant bacteria imposed an urgent request to discover novel antibacterial agents. This work was to develop naphthalimide nitroimidazoles as potentially antibacterial agents. Results/methodology: Compound 9e showed the strong antibacterial activity (minimal inhibitory concentration = 0.013 µmol/ml) against resistant Acinetobacter baumannii (A. baumannii) with rapid killing effect and no obvious triggering of the development of resistance. Its combination use with chloromycin, norfloxacin or clinafloxacin improved the antibacterial potency. It could not only effectively permeate membrane of resistant A. baumannii bacteria, but also intercalate into resistant A. baumannii DNA to form 9e-DNA complex. The interaction with bacterial DNA gyrase B was driven by hydrogen bonds. CONCLUSION: Compound 9e should be a potentially multitargeting antibacterial agent against resistant A. baumannii.

Discovery of 2-aminothiazolyl berberine derivatives as effectively antibacterial agents toward clinically drug-resistant Gram-negative Acinetobacter baumanii.

Aminothiazolyl berberine derivatives as potentially antimicrobial agents were designed and synthesized in an effort to overcome drug resistance. The antimicrobial assay revealed that some target compounds exhibited significantly inhibitory efficiencies toward bacteria and fungi including drug-resistant pathogens, and the aminothiazole and Schiff base moieties were helpful structural fragments for aqueous solubility and antibacterial activity. Especially, aminothiazolyl 9-hexyl berberine 9c and 2,4-dichlorobenzyl derivative 18a exhibited good activities (MIC = 2 nmol/mL) against clinically drug-resistant Gram-negative Acinetobacter baumanii with low cytotoxicity to hepatocyte LO2 cells, rapidly bactericidal effects and quite slow development of bacterial resistance toward A. baumanii. Molecular modeling indicated that compounds 9c and 18a could bind with GLY-102, ARG-136 and/or ALA-100 residues of DNA gyrase through hydrogen bonds. It was found that compounds 9c and 18a were able to disturb the drug-resistant A. baumanii membrane effectively, and molecule 9c could not only intercalate but also cleave bacterial DNA isolated from resistant A. baumanii, which might be the preliminary antibacterial action mechanism of inhibiting the growth of A. baumanii strain. In particular, the combination use of compound 9c with norfloxacin could enhance the antibacterial activity, broaden antibacterial spectrum and overcome the drug resistance.

Novel Naphthalimide Aminothiazoles as Potential Multitargeting Antimicrobial Agents.

A series of novel naphthalimide aminothiazoles were developed for the first time and evaluated for their antimicrobial activity. Some prepared compounds possessed good inhibitory activity against the tested bacteria and fungi. Noticeably, the piperazine derivative 4d displayed superior antibacterial activity against MRSA and Escherichia coli with MIC values of 4 and 8 µg/mL, respectively, to reference drugs. The most active compound 4d showed low toxicity against mammalian cells with no obvious triggering of the development of bacterial resistance, and it also possessed rapid bactericidal efficacy and efficient membrane permeability. Preliminarily investigations revealed that compound 4d could not only bind with gyrase-DNA complex through hydrogen bonds but could effectively intercalate into MRSA DNA to form 4d-DNA supramolecular complex, which might be responsible for the powerful bioactivity. Further transportation behavior evaluation indicated that molecule 4d could be effectively stored and carried by human serum albumin, and the hydrophobic interactions and hydrogen bonds played important roles in the binding process.

Discovery of novel nitroimidazole enols as Pseudomonas aeruginosa DNA cleavage agents.

A series of nitroimidazole enols as new bacterial DNA-targeting agents were for the first time designed, synthesized and characterized by NMR, IR and HRMS spectra. The antimicrobial screening revealed that 2-methoxyphenyl nitroimidazole enol 3i possessed stronger anti-P. aeruginosa efficacy (MIC = 0.10 µmol/mL) than reference drugs Norfloxacin and Metronidazole. Time-kill kinetic assay manifested that the active molecule 3i could rapidly kill the tested strains. Molecular docking indicated that the interactions between compound 3i and topoisomerase II were driven by hydrogen bonds. Quantum chemical study was also performed on 3i to understand the structural features essential for activity. Further research found that compound 3i was not able to effectively intercalate into bacterial DNA but could cleave DNA isolated from the standard P. aeruginosa strain, which might block DNA replication to exert the efficient bioactivities, and this active molecule was also able to be stored and carried by human serum albumin via hydrophobic interactions and hydrogen bonds.

Novel potentially antifungal hybrids of 5-flucytosine and fluconazole: Design, synthesis and bioactive evaluation.

A series of novel potentially antifungal hybrids of 5-flucytosine and fluconazole were designed, synthesized and characterized by 1H NMR, 13C NMR, IR and HRMS spectra. Bioactive assay manifested that some prepared compounds showed moderate to good antifungal activities in comparison with fluconazole and 5-flucytosine. Remarkably, the 3,4-dichlorobenzyl hybrid 7h could inhibit the growth of C. albicans ATCC 90023 and clinical resistant strain C. albicans with MIC values of 0.008 and 0.02 mM, respectively. The active molecule 7h could not only rapidly kill C. albicans but also efficiently permeate membrane of C. albicans. Molecular docking study revealed that compound 7h could interact with the active site of CACYP51 through hydrogen bond. Quantum chemical studies were also performed to explain the high antifungal activity. Further preliminary mechanism research suggested that molecule 7h could intercalate into calf thymus DNA to form a steady supramolecular complex, which might block DNA replication to exert the powerful bioactivities.

Design, synthesis and antimicrobial evaluation of novel benzimidazole-incorporated sulfonamide analogues.

A novel series of benzimidazole-incorporated sulfonamide analogues were designed and synthesized with an effort to overcome the increasing antibiotic resistance. Compound 5c gave potent activities against Gram-positive bacteria and fungi, and 2,4-dichlorobenzyl derivative 5g showed good activities against Gram-negative bacteria. Both of these two active molecules 5c and 5g could effectively intercalate into calf thymus DNA to form compound-DNA complex respectively, which might block DNA replication to exert their powerful antimicrobial activity. Molecular docking experiments suggested that compounds 5c and 5g could insert into base-pairs of DNA hexamer duplex by the formation of hydrogen bonds with guanine of DNA. The transportation behavior of these highly active compounds by human serum albumin (HSA) demonstrated that the electrostatic interactions played major roles in the strong association of active compounds with HSA, and which was also confirmed by the full geometry calculation optimizations.

Novel benzimidazolyl tetrahydroprotoberberines: Design, synthesis, antimicrobial evaluation and multi-targeting exploration.

A series of novel benzimidazolyl tetrahydroprotoberberines were conveniently designed and efficiently synthesized from berberine via direct cyclization of tetrahydroprotoberberine aldehyde and o-phenylene diamines under metal-free aerobic oxidation. All the new compounds were characterized by IR, 1H NMR, 13C NMR and HRMS spectra. The antimicrobial evaluation revealed that the 5-fluorobenzimidazolyl derivative 5b was the most active antibacterial and antifungal molecule with broad spectrum in comparison to Berberine, Chloromycin, Norfloxacin and Fluconazole. It triggered almost no resistance development against MRSA even after 15 passages. Further studies demonstrated that compound 5b could not only effectively interact with Topo IA by hydrogen bonds, but also intercalate into calf thymus DNA and cleave pBR322 DNA, which might be responsible for its powerful bioactivities.

Design, synthesis and biological evaluation of novel Schiff base-bridged tetrahydroprotoberberine triazoles as a new type of potential antimicrobial agents.

A series of novel Schiff base-bridged tetrahydroprotoberberine (THPB) triazoles were designed, synthesized and characterized for the first time. Antimicrobial assay showed that some of the prepared compounds exerted stronger antibacterial and antifungal activities than the reference drugs. Especially, THPB triazole 7a gave low MIC values of 0.5, 1 and 2 µg mL-1 against B. yeast, M. luteus and MRSA, respectively. Further experiments indicated that the highly active molecule 7a was able to rapidly kill the MRSA strain and did not trigger the development of bacterial resistance even after 14 passages. The preliminary exploration for the antimicrobial mechanism revealed that compound 7a could effectively intercalate into calf thymus DNA to form a 7a-DNA supramolecular complex, and its Zn2+ complex had the ability to directly cleave pUC19 DNA, which suggested that compound 7a might be a potentially dual-targeting antibacterial molecule. It was also found that compound 7a could be efficiently stored and carried by human serum albumin (HSA), and the hydrophobic interactions and hydrogen bonds played important roles in the transportation of HSA to the active molecule 7a.