Modification of Triclosan Scaffold in Search of Improved Inhibitors for Enoyl‐Acyl Carrier Protein (ACP) Reductase in Toxoplasma gondii

Through our focused effort to discover new and effective agents against toxoplasmosis, a structure‐based drug design approach was used to develop a series of potent inhibitors of the enoyl‐acyl carrier protein (ACP) reductase (ENR) enzyme in Toxoplasma gondii (TgENR). Modifications to positions 5 and 4′ of the well‐known ENR inhibitor triclosan afforded a series of 29 new analogues. Among the resulting compounds, many showed high potency and improved physicochemical properties in comparison with the lead. The most potent compounds 16 a and 16 c have IC₅₀ values of 250 nM against Toxoplasma gondii tachyzoites without apparent toxicity to the host cells. Their IC₅₀ values against recombinant TgENR were found to be 43 and 26 nM , respectively. Additionally, 11 other analogues in this series had IC₅₀ values ranging from 17 to 130 nM in the enzyme‐based assay. With respect to their excellent in vitro activity as well as improved drug‐like properties, the lead compounds 16 a and 16 c are deemed to be excellent starting points for the development of new medicines to effectively treat Toxoplasma gondii infections.     

Modification of Biphenolic Anti-Bacterial to Achieve Broad-Spectrum Activity

The Gram-positive bacteria, methicillin-resistant Staphylococcus aureus (MRSA) and Gram-negative bacteria, Acinetobacter baumannii, are pathogens responsible for millions of nosocomial infections worldwide. Due to the threat of bacteria evolving resistance to antibiotics, scientists are constantly looking for new classes of compounds to treat infectious diseases. The biphenolic analogs of honokiol that were most potent against oral bacteria had similar bioactivity against MRSA. However, all the compounds proved ineffective against A. baumannii. The inability to inhibit A. baumannii is due to the difficult-to-penetrate lipopolysaccharide-coated outer membrane that makes it challenging for antibiotics to enter Gram-negative bacteria. The C 2 scaffold was optimized from the inhibition of Gram-positive bacteria to broad-spectrum antibacterial compounds that inhibit the dangerous Gram-negative pathogen A. baumannii.     

Modifications to a Biphenolic Antibacterial Compound: Activity against ESKAPE Pathogens

Forty-four analogs of honokiol, a compound with known antibacterial activity, especially with respect to oral bacteria, were synthesized to explore the structure-activity relationships against the ESKAPE pathogens. Compounds with high therapeutic indices (hemolysis₂₀/MIC) were identified. In particular, ester-linked compounds that would be less than environmentally durable than biaryl ether antibacterials such as the broadly used triclosan were found to be active. MRSA mutants could be generated against some, but not all, of the highly active compounds. Based on gene sequencing results, membrane permeability, intracellular sodium, and intracellular pH assays revealed overlapping mechanisms of action.     

Molecular Basis of Selectivity and Activity for the Antimicrobial Peptide Lynronne-1 Informs Rational Design of Peptide with Improved Activity

Antibiotic resistance is a significant threat to human health, with natural products remaining the best source for new antimicrobial compounds. Antimicrobial peptides (AMPs) are natural products with great potential for clinical use as they are small, amenable to customization, and show broad-spectrum activities. Lynronne-1 is a promising AMP identified in the rumen microbiome that shows broad-spectrum activity against pathogens such as methicillin-resistant Staphylococcus aureus and Acinetobacter baumannii. Here we investigated the structure of Lynronne-1 using solution NMR spectroscopy and identified a 13-residue amphipathic helix containing all six cationic residues. We used biophysical approaches to observe folding, membrane partitioning and membrane lysis selective to the presence of anionic lipids. We translated our understanding of Lynronne-1 structure to design peptides which varied in the size of their hydrophobic helical face. These peptides displayed the predicted continuum of membrane-lysis activities in vitro and in vivo, and yielded a new AMP with 4-fold improved activity against A. baumannii and 32-fold improved activity against S. aureus.     

Antibacterial agents and cystic fibrosis: synthesis and antimicrobial evaluation of a series of N-thiomethylazetidinones

The increasing emergence of multidrug-resistant microorganisms is one of the greatest challenges in the clinical management of infectious disease. New antimicrobial agents are therefore urgently required, particularly in the treatment of chronic and recurrent infections often associated with antibiotic-resistant pathogens, as in the case of cystic fibrosis (CF) patients. This study reports the antibacterial activity of a series of monocyclic β-lactams with an alkylidenecarboxyl chain or electron-withdrawing groups such as 4-OAc, 4-SAc, and 4-SO(2)Ph at the C4 position of the ring. N-Unsubstituted and N-thiomethyl derivatives were compared. A total of 33 azetidinones were tested for their activity against Gram-positive and Gram-negative bacterial clinical isolates. The combination of an N-thiomethyl group and a benzyl ester on the 4-alkylidene side chain were found to increase the potency against Gram-positive bacteria. The N-thiomethyl group clearly elevated the activity of 4-acetoxyazetidinones relative to the corresponding NH derivatives. The most active compounds showed minimum inhibitory concentration (MIC) values of 4 and 8 mg L(-1) against methicillin-resistant Staphylococcus aureus isolated from pediatric patients with CF.     

Towards Catalytic Antibiotics: Redesign of Aminoglycosides To Catalytically Disable Bacterial Ribosomes

The emergence of multidrug-resistant pathogens that are resistant to the majority of currently available antibiotics is a significant clinical problem. The development of new antibacterial agents and novel approaches is therefore extremely important. We set out to explore the potential of catalytic antibiotics as a new paradigm in antibiotics research. Herein, we describe our pilot study on the design, synthesis, and biological testing of a series of new derivatives of the natural aminoglycoside antibiotic neomycin B for their potential action as catalytic antibiotics. The new derivatives showed significant antibacterial activity against wild-type bacteria and were especially potent against resistant and pathogenic strains including Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus. Selected compounds displayed RNase activity even though the activity was not as high and specific as we would have expected. On the basis of the observed chemical and biochemical data, along with the comparative molecular dynamics simulations of the prokaryotic rRNA decoding site, we postulate that the rational design of catalytic antibiotics should involve not only their structure but also a comprehensive analysis of the rRNA A-site dynamics.     

Evaluation of 4,5-disubstituted-2-aminoimidazole-triazole conjugates for antibiofilm/antibiotic resensitization activity against MRSA and Acinetobacter baumannii

A library of 4,5-disubstituted-2-aminoimidazole-triazole conjugates (2-AITs) was synthesized, and the antibiofilm activity was investigated. This class of small molecules was found to inhibit biofilm formation by methicillin-resistant Staphylococcus aureus (MRSA) at low-micromolar concentrations; 4,5-disubstituted-2-AITs were also able to inhibit and disperse Acinetobacter baumannii biofilms. The activities of the lead compounds were compared against the naturally occurring biofilm dispersant cis-2-decenoic acid and were revealed to be more potent. The ability of selected compounds to resensitize MRSA to traditional antibiotics (resensitization activity) was also determined. Lead compounds were observed to resensitize MRSA to oxacillin by 2-4-fold.     

中氮茚并喹啉二酮衍生物的合成、表征及抗菌活性

以自制的6,7-二氯喹啉-5,8-二酮为原料,经一步反应合成了目标化合物9-氯-5,12-二酮-5,12-二氢中氮茚并[2,3-g]喹啉-6-甲酸乙酯,并通过1HNMR 、HRMS以及X-射线单晶衍射对其结构进行表征.活性测试结果表明,该化合物对革兰氏阳性菌具有选择性抑制作用,对耐甲氧西林金黄色葡萄球菌(MRSA)的最小抑菌浓度(MIC)为1μg/mL,优于阳性对照药物万古霉素.通过化合物与DNA促旋酶的分子对接实验进一步说明了化合物可能的作用机理.     

Expanding the Spectrum of Antibiotics Capable of Killing Multidrug-Resistant Staphylococcus aureus and Pseudomonas aeruginosa

Infections from antibiotic-resistant Staphylococcus aureus and Pseudomonas aeruginosa are a serious threat because reduced antibiotic efficacy complicates treatment decisions and prolongs the disease state in many patients. To expand the arsenal of treatments against antimicrobial-resistant (AMR) pathogens, 600-Da branched polyethylenimine (BPEI) can overcome antibiotic resistance mechanisms and potentiate β-lactam antibiotics against Gram-positive bacteria. BPEI binds cell-wall teichoic acids and disables resistance factors from penicillin binding proteins PBP2a and PBP4. This study describes a new mechanism of action for BPEI potentiation of antibiotics generally regarded as agents effective against Gram-positive pathogens but not Gram-negative bacteria. 600-Da BPEI is able to reduce the barriers to drug influx and facilitate the uptake of a non-β-lactam co-drug, erythromycin, which targets the intracellular machinery. Also, BPEI can suppress production of the cytokine interleukin IL-8 by human epithelial keratinocytes. This enables BPEI to function as a broad-spectrum antibiotic potentiator, and expands the opportunities to improve drug design, antibiotic development, and therapeutic approaches against pathogenic bacteria, especially for wound care.     

Novel Phenyl-Based Bis-quaternary Ammonium Compounds as Broad-Spectrum Biocides

Herein we report the synthesis and microbiological evaluation of novel phenyl based bis-quaternary ammonium compounds (bis-QACs). Using a simple 2-step synthetic route from dibromo- and dihydroxybenzenes, we obtained a structurally diverse broad panel of bis-QACs with topologically distinct bridging connections between pyridinium heads. Selected analogs possessed potent broad-spectrum biocidal activity against both bacterial and fungal pathogens: methicillin-resistant Staphylococcus aureus (ATCC 43300); Escherichia coli (ATCC 25922), Klebsiella pneumonia (ATCC 700603), Acinetobacter baumannii (ATCC 19606), Pseudomonas aeruginosa (ATCC 27853), Candida albicans (ATCC 90028), Cryptococcus neoformans var. grubii (ATCC 208821). Promising compounds displayed minimum inhibitory concentrations (MIC) values ≤0.25 μg/mL alongside improved cytotoxicity and hemolytic profiles compared to modern antiseptics. Thus, synthesized bis-QACs represent a promising class of biocides with the potential to replace existing household sanitizers.     

Search for Shorter Portions of the Proline-Rich Antimicrobial Peptide Fragment Bac5(1–25) That Retain Antimicrobial Activity by Blocking Protein Synthesis

The spread of antibiotic-resistant pathogens has boosted the search for new antimicrobial drugs. Proline-rich antimicrobial peptides are promising lead compounds for the development of next-generation antibiotics, given their very low cytotoxicity and their good antimicrobial activity targeting the bacterial ribosome. Bac5(1–25) is an N-terminal fragment of the bovine proline-rich antimicrobial peptide Bac5, whose mode of action has been recently described. In this work we tested a number of Bac5(1–25) fragments, and we characterized their antimicrobial activity against Escherichia coli, Acinetobacter baumannii, Klebsiella pneumoniae, Staphylococcus aureus, Salmonella enterica, and Pseudomonas aeruginosa. We evaluated their cytotoxicity toward human cells and their efficacy in inhibiting bacterial protein synthesis. This allowed us to identify some shorter fragments of Bac5(1–25) with a good balance between antibacterial efficacy, protein synthesis inhibition, and ease/cost-effectiveness of synthesis, suitable as lead compounds to develop new antibacterials.     

Rheological and curing properties of reactive blending products of epoxidised natural rubber and cassava starch

Abstract

Epoxidised natural rubber (ENR) has been prepared and used as a blending ingredient together with a compatibiliser for blending of natural rubber (air dry sheet, ADS) and cassava starch. Mooney viscosities of the blends were quantified at 100°C and rheological properties in terms of shear stress and shear viscosity were plotted against shear rates. The results showed that pure ENR gave a lower Mooney viscosity, shear stress, and shear viscosity than blends with cassava starch. Mooney viscosity, shear stress, and shear viscosity for the blends increased with increasing levels of starch. At the same level of cassava starch blended, the highest values of these quantities were observed for the blends with ENR. The blend of ADS with ENR as a compatibiliser showed lower values than those of ENR itself, but higher than those of ADS with the starch. The results are described in terms of the level of chemical interaction between polar groups in ENR and in cassava starch. Curing behaviour for compounds of ENR, ADS, and ADS with ENR as a compatibiliser were studied. The results found that ENR exhibited a long delay (～ 10 min) before the vulcanisation took place compared with 1 min for ADS compounds. In the curing curve for ENR, an equilibrium value at maximum torque was not found indicating that the stiffness of the ENR compounds still increased with increasing testing time until 60 min. Stiffness of the ENR compounds also increased with increasing levels of cassava starch.     

Ligand- and Structure-Based Approaches of Escherichia coli FabI Inhibition by Triclosan Derivatives: From Chemical Similarity to Protein Dynamics Influence

Enoyl-acyl carrier protein reductase (FabI) is the limiting step to complete the elongation cycle in type II fatty acid synthase (FAS) systems and is a relevant target for antibacterial drugs. E. coli FabI has been employed as a model to develop new inhibitors against FAS, especially triclosan and diphenyl ether derivatives. Chemical similarity models (CSM) were used to understand which features were relevant for FabI inhibition. Exhaustive screening of different CSM parameter combinations featured chemical groups, such as the hydroxy group, as relevant to distinguish between active/decoy compounds. Those chemical features can interact with the catalytic Tyr156. Further molecular dynamics simulation of FabI revealed the ionization state as a relevant for ligand stability. Also, our models point the balance between potency and the occupancy of the hydrophobic pocket. This work discusses the strengths and weak points of each technique, highlighting the importance of complementarity among approaches to elucidate EcFabI inhibitor's binding mode and offers insights for future drug discovery.     

Lipidation of Temporin-1CEb Derivatives as a Tool for Activity Improvement,Pros and Cons of the Approach

The alarming raise of multi-drug resistance among human microbial pathogens makes the development of novel therapeutics a priority task. In contrast to conventional antibiotics, antimicrobial peptides (AMPs), besides evoking a broad spectrum of activity against microorganisms, could offer additional benefits, such as the ability to neutralize toxins, modulate inflammatory response, eradicate bacterial and fungal biofilms or prevent their development. The latter properties are of special interest, as most antibiotics available on the market have limited ability to diffuse through rigid structures of biofilms. Lipidation of AMPs is considered as an effective approach for enhancement of their antimicrobial potential and in vivo stability; however, it could also have undesired impact on selectivity, solubility or the aggregation state of the modified peptides. In the present work, we describe the results of structural modifications of compounds designed based on cationic antimicrobial peptides DK5 and CAR-PEG-DK5, derivatized at their N-terminal part with fatty acids with different lengths of carbon chain. The proposed modifications substantially improved antimicrobial properties of the final compounds and their effectiveness in inhibition of biofilm development as well as eradication of pre-formed 24 h old biofilms of Candida albicans and Staphylococcus aureus. The most active compounds (C5-DK5, C12-DK5 and C12-CAR-PEG-DK5) were also potent against multi-drug resistant Staphylococcus aureus USA300 strain and clinical isolates of Pseudomonas aeruginosa. Both experimental and in silico methods revealed strong correlation between the length of fatty acid attached to the peptides and their final membranolytic properties, tendency to self-assemble and cytotoxicity.     

Effect of Secondary Structure and Side Chain Length of Hydrophobic Amino Acid Residues on the Antimicrobial Activity and Toxicity of 14-Residue-Long de novo AMPs

Herein we report the efficacy and toxicity of three de novo designed cationic antimicrobial peptides (AMPs) LL-14, VV-14 and ββ-14, where side chains of the hydrophobic amino acids were reduced gradually. The AMPs showed broad-spectrum antimicrobial activity against three pathogens from the ESKAPE group and two fungal strains. This study showed that side chains which are either too long or too short increase toxicity and lower antimicrobial activity, respectively. VV-14 was found to be non-cytotoxic and highly potent under physiological salt concentrations against several pathogens, especially Salmonella typhi TY2. These AMPs acted via membrane deformation, depolarization, and lysis. The activity of the AMPs is related to their ability to take on amphipathic helical conformations in the presence of microbial membrane mimics. Among AMPs with the same charge, hydrophobic interactions between the side chains of the residues with cell membrane lipids determine their antimicrobial potency and cytotoxicity. Strikingly, an optimum hydrophobic interaction is the crux of generating highly potent non-cytotoxic AMPs.     

Emerging Options for the Diagnosis of Bacterial Infections and the Characterization of Antimicrobial Resistance

Precise and rapid identification and characterization of pathogens and antimicrobial resistance patterns are critical for the adequate treatment of infections, which represent an increasing problem in intensive care medicine. The current situation remains far from satisfactory in terms of turnaround times and overall efficacy. Application of an ineffective antimicrobial agent or the unnecessary use of broad-spectrum antibiotics worsens the patient prognosis and further accelerates the generation of resistant mutants. Here, we provide an overview that includes an evaluation and comparison of existing tools used to diagnose bacterial infections, together with a consideration of the underlying molecular principles and technologies. Special emphasis is placed on emerging developments that may lead to significant improvements in point of care detection and diagnosis of multi-resistant pathogens, and new directions that may be used to guide antibiotic therapy.     

Thermal and mechanical properties of epoxidized natural rubber modified epoxy matrices

Two sets (A and B) of bisphenol A–diglycidyl ether (DGEBA) based epoxy resin formulations were modified with epoxidized natural rubber (ENR 50) and its liquid version (LENR 50), and cured with amino propoxylate initiator/accelerator at ambient temperatures. The ENR 50 loading range was 1.6–5.9 wt%. Both sets could be loaded up to 12 wt% with LENR 50. Significant improvements in tensile toughness and impact toughness could only be observed for set A formulations. At the maximum LENR 50 loading achieved, the improvement in tensile toughness is 250% in comparison with that of the neat formulation; that for impact toughness is 125%. Differential scanning calorimetry reveals multiple transitions, characteristic of these systems. Scanning electron micrographs of fractured surfaces show uniform rubber dispersions in the submicrometre size range. At the loading levels used, LENR 50 particle dispersions fall within the range of 0.33–0.47 µm in size; those of ENR 50 are 0.48–0.67 µm in average size. Improvements in toughness are similar for both versions of epoxidized natural rubber. For both LENR 50 and ENR 50 modified epoxy systems, the extremes of 0.80 (set A) and 1.95 (set B) in glycidyl ether/reactive hydrogen molar ratios considered show distinct failure mechanisms, that of ductile failure with yielding in the former and brittle failure in the latter, irrespective of reactive diluent content. © 1999 Society of Chemical Industry     

Biological Evaluation of Potent Triclosan‐Derived Inhibitors of the Enoyl–Acyl Carrier Protein Reductase InhA in Drug‐Sensitive and Drug‐Resistant Strains of Mycobacterium tuberculosis

New triclosan (TRC) analogues were evaluated for their activity against the enoyl–acyl carrier protein reductase InhA in Mycobacterium tuberculosis (Mtb). TRC is a well‐known inhibitor of InhA, and specific modifications to its positions 5 and 4′ afforded 27 derivatives; of these compounds, seven derivatives showed improved potency over that of TRC. These analogues were active against both drug‐susceptible and drug‐resistant Mtb strains. The most active compound in this series, 4‐(n‐butyl)‐1,2,3‐triazolyl TRC derivative 3 , had an MIC value of 0.6 μg mL^?1 (1.5 μM ) against wild‐type Mtb. At a concentration equal to its MIC, this compound inhibited purified InhA by 98 %, and showed an IC₅₀ value of 90 nM . Compound 3 and the 5‐methylisoxazole‐modified TRC 14 were able to inhibit the biosynthesis of mycolic acids. Furthermore, mc²4914, an Mtb strain overexpressing inhA, was found to be less susceptible to compounds 3 and 14 , supporting the notion that InhA is the likely molecular target of the TRC derivatives presented herein.     

Selectively guanidinylated aminoglycosides as antibiotics

The emergence of virulent, drug-resistant bacterial strains coupled with a minimal output of new pharmaceutical agents to combat them makes this a critical time for antibacterial research. Aminoglycosides are a well-studied, highly potent class of naturally occurring antibiotics with scaffolds amenable to modification, and therefore, they provide an excellent starting point for the development of semisynthetic, next-generation compounds. To explore the potential of this approach, we synthesized a small library of aminoglycoside derivatives selectively and minimally modified at one or two positions with a guanidine group replacing the corresponding amine or hydroxy functionality. Most guanidino-aminoglycosides showed increased affinity for the ribosomal decoding rRNA site, the cognate biological target of the natural products, when compared with their parent antibiotics, as measured by an in vitro fluorescence resonance energy transfer (FRET) A-site binding assay. Additionally, certain analogues showed improved minimum inhibitory concentration (MIC) values against resistant bacterial strains, including methicillin-resistant Staphylococcus aureus (MRSA). An amikacin derivative holds particular promise with activity greater than or equal to the parent antibiotic in the majority of bacterial strains tested.     

Eradicating Biofilms of Carbapenem-Resistant Enterobacteriaceae by Simultaneously Dispersing the Biomass and Killing Planktonic Bacteria with PEGylated Branched Polyethyleneimine

Carbapenem-resistant Enterobacteriaceae (CRE) are emerging pathogens that cause variety of severe infections. CRE evade antibiotic treatments because these bacteria produce enzymes that degrade a wide range of antibiotics including carbapenems and β-lactams. The formation of biofilms aggravates CRE infections, especially in a wound environment. These difficulties lead to persistent infection and non-healing wounds. This creates the need for new compounds to overcome CRE antimicrobial resistance and disrupt biofilms. Recent studies in our lab show that 600 Da branched polyethyleneimine (BPEI) and its derivative PEG350-BPEI can overcome antimicrobial resistance and eradicate biofilms in methicillin-resistant S. aureus, methicillin-resistant S. epidermidis, P. aeruginosa, and E. coli. In this study, the ability of 600 Da BPEI and PEG350-BPEI to eradicate carbapenem-resistant Enterobacteriaceae bacteria and their biofilms is demonstrated. We show that both BPEI and PEG350-BPEI have anti-biofilm efficacy against CRE strains expressing Klebsiella pneumoniae carbapenemases (KPCs) and metallo-β-lactamases (MBLs), such as New Delhi MBL (NDM-1). Furthermore, our results illustrate that BPEI affects planktonic CRE bacteria by increasing bacterial length and width from the inability to proceed with normal cell division processes. These data demonstrate the multi-functional properties of 600 Da BPEI and PEG350-BPEI to reduce biofilm formation and mitigate virulence in carbapenem-resistant Enterobacteriaceae.