Synthesis, antibacterial activities and molecular docking studies of Schiff bases derived from N-(2/4-benzaldehyde-amino) phenyl-N'-phenyl-thiourea

A series of novel Schiff base derivatives have been designed and synthesized, and their biological activities were also evaluated as potential inhibitors of FabH. These compounds were assayed for antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. Compounds with potent antibacterial activities were tested for their E. coli FabH inhibitory activity. Compound 3v showed the most potent antibacterial activity with MIC of 1.56-6.25 μg/mL against the tested bacterial strains and exhibited the most potent E. coli FabH inhibitory activity with IC(50) of 4.3 μM. Docking simulation was performed to position compound 3v into the E. coli FabH active site to determine the probable binding conformation.     

Synthesis, molecular modeling and biological evaluation of β-ketoacyl-acyl carrier protein synthase III (FabH) as novel antibacterial agents

A series of novel cinnamic acid secnidazole ester derivatives have been designed and synthesized, and their biological activities were also evaluated as potential inhibitors of FabH. These compounds were assayed for antibacterial activity against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. Compounds with potent antibacterial activities were tested for their E. coli FabH inhibitory activity. Compound 3n showed the most potent antibacterial activity with MIC of 1.56-6.25 μg/mL against the tested bacterial strains and exhibited the most potent E. coli FabH inhibitory activity with IC?? of 2.5 μM. Docking simulation was performed to position compound 3n into the E. coli FabH active site to determine the probable binding conformation.     

Discovery and development of substituted thiadiazoles as inhibitors of Staphylococcus aureus Sortase A

High-throughput screening of small-molecule libraries has led to the identification of thiadiazoles as a new class of inhibitors against Staphylococcus aureus sortase A (SrtA). N-(5-((4-nitrobenzyl)thio)-1,3,4-thiadiazol-2-yl)nicotinamide (IC₅₀ = 3.8 µM) was identified as a potent inhibitor of SrtA after synthetic modification of hit compounds. Additional ligands developed in this study displayed affinities in the low micromolar range without affecting bacterial growth in vitro. The study also suggest a new mode of action through covalent binding to the active site cysteine.     

Discovery of potent small molecule inhibitors of DYRK1A by structure-based virtual screening and bioassay

In this study, six novel dual-specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A) inhibitors with IC(50) values ranging from 1.51 to 88.13 μM were successfully identified through virtual screening and in vitro plus cell based bioassay. Compound 5 with IC(50) value of 1.51 μM is the most potent hit against DYRK1A in vitro, while compound 3 exhibited the most potent activity in cultured cells. The inhibition mechanism was explored by molecular docking approach. This study may provide a start point for further mechanism based study as well as discovery of drug candidate against Down syndrome (DS).     

Monofunctional transglycosylases are not essential for Staphylococcus aureus cell wall synthesis

The polymerization of peptidoglycan is the result of two types of enzymatic activities: transglycosylation, the formation of linear glycan chains, and transpeptidation, the formation of peptide cross-bridges between the glycan strands. Staphylococcus aureus has four penicillin binding proteins (PBP1 to PBP4) with transpeptidation activity, one of which, PBP2, is a bifunctional enzyme that is also capable of catalyzing transglycosylation reactions. Additionally, two monofunctional transglycosylases have been reported in S. aureus: MGT, which has been shown to have in vitro transglycosylase activity, and a second putative transglycosylase, SgtA, identified only by sequence analysis. We have now shown that purified SgtA has in vitro transglycosylase activity and that both MGT and SgtA are not essential in S. aureus. However, in the absence of PBP2 transglycosylase activity, MGT but not SgtA becomes essential for cell viability. This indicates that S. aureus cells require one transglycosylase for survival, either PBP2 or MGT, both of which can act as the sole synthetic transglycosylase for cell wall synthesis. We have also shown that both MGT and SgtA interact with PBP2 and other enzymes involved in cell wall synthesis in a bacterial two-hybrid assay, suggesting that these enzymes may work in collaboration as part of a larger, as-yet-uncharacterized cell wall-synthetic complex.     

A new class of small molecule RNA polymerase inhibitors with activity against rifampicin-resistant Staphylococcus aureus

The RNA polymerase holoenzyme is a proven target for antibacterial agents. A high-throughput screening program based on this enzyme from Staphylococcus aureus had previously identified a 2-ureidothiophene-3-carboxylate as a low micromolar inhibitor. An investigation of the relationships between the structures of this class of compounds and their inhibitory- and antibacterial activities is described here, leading to a set of potent RNA polymerase inhibitors with antibacterial activity. Characterization of this bioactivity, including studies of the mechanism of action, is provided, highlighting the power of the reverse chemical genetics approach in providing tools to inhibit the bacterial RNA polymerase.     

Discovery of MEK/PI3K dual inhibitor via structure-based virtual screening

Mitogen/extracellular signal-regulated kinase (MEK) and phosphoinositide 3-kinase (PI3Kα) are considered to be promising targets for the development of anticancer therapeutics. We report the first example of the successful application of structure-based virtual screening to identify novel inhibitors of MEK with IC(50) values ranging from 1 to 25 μM. One of the four newly identified MEK inhibitors was found to be also a potent inhibitor of PI3Kα with submicromolar inhibitory activity (IC(50)=0.3 μM). Because this dual inhibitor was screened for having desirable physicochemical properties as a drug candidate as well as the high inhibitory activities against MEK and PI3Kα, it warrants further development through structure-activity relationship (SAR) studies to optimize the inhibitory and anticancer activities. Structural features relevant to the stabilization of the dual inhibitor in the ATP-binding sites of MEK1 and PI3Kα are addressed in detail.     

Design, synthesis and antimicrobial activities of nitroimidazole derivatives containing 1,3,4-oxadiazole scaffold as FabH inhibitors

Nitroimidazoles and their derivatives have drawn continuing interest over the years because of their varied biological activities, recently found application in drug development for antimicrobial chemotherapeutics and antiangiogenic hypoxic cell radiosensitizers. In order to search for novel antibacterial agents, we designed and synthesized a series of secnidazole analogs based on oxadiazole scaffold (4-21). Among these compounds, 4 and 7-21 were reported for the first time. These compounds were tested for antibacterial activities against Escherichia coli, Pseudomonas aeruginosa, Bacillus subtilis and Staphylococcus aureus. This new nitroimidazole derivatives class demonstrated strong antibacterial activities. Escherichia coli β-ketoacyl-acyl carrier protein synthase III (FabH) inhibitory assay and docking simulation indicated that the compounds 2-(2-methoxyphenyl)-5-((2-methyl-5-nitro-1H-imidazol-1-yl)methyl)-1,3,4-oxadiazole (11) with MIC of 1.56-3.13 μg/mL against the tested bacterial strains and 2-((2-methyl-5-nitro-1H-imidazol-1-yl)methyl)-5-(2-methylbenzyl)-1,3,4-oxadiazole (12) with MIC of 1.56-6.25 μg/mL were most potent inhibitors of Escherichia coli FabH.     

Determination of selectivity and efficacy of fatty acid synthesis inhibitors

Type II fatty acid synthesis (FASII) is essential to bacterial cell viability and is a promising target for the development of novel antibiotics. In the past decade, a few inhibitors have been identified for this pathway, but none of them lend themselves to drug development. To find better inhibitors that are potential drug candidates, we developed a high throughput assay that identifies inhibitors simultaneously against multiple targets within the FASII pathway of most bacterial pathogens. We demonstrated that the inverse t(1/2) value of the FASII enzyme-catalyzed reaction gives a measure of FASII activity. The Km values of octanoyl-CoA and lauroyl-CoA were determined to be 1.1 +/- 0.3 and 10 +/- 2.7 microM in Staphylococcus aureus and Bacillus subtilis, respectively. The effects of free metals and reducing agents on enzyme activity showed an inhibition hierarchy of Zn2+ > Ca2+ > Mn2+ > Mg2+; no inhibition was found with beta-mercaptoethanol or dithiothreitol. We used this assay to screen the natural product libraries and isolated an inhibitor, bischloroanthrabenzoxocinone (BABX) with a new structure. BABX showed IC50 values of 11.4 and 35.3 microg/ml in the S. aureus and Escherichia coli FASII assays, respectively, and good antibacterial activities against S. aureus and permeable E. coli strains with minimum inhibitory concentrations ranging from 0.2 to 0.4 microg/ml. Furthermore, the effectiveness, selectivity, and the in vitro and in vivo correlations of BABX as well as other fatty acid inhibitors were elucidated, which will aid in future drug discovery.     

Mannich reaction derivatives of novobiocin with modulated physiochemical properties and their antibacterial activities

Synthetic derivatives of the natural product antibiotic novobiocin were synthesized in order to improve their physiochemical properties. A Mannich reaction was used to introduce new side chains at a solvent-exposed position of the molecule, and a diverse panel of functional groups was evaluated at this position. Novobiocin and the new derivatives were tested for their binding to gyrase B and their antibacterial activities against Staphylococcus aureus, Mycobacterium tuberculosis, Francisella tularensis and Escherichia coli. While the new derivatives still bound the gyrase B protein potently (0.07-1.8 μM, IC(50)), they had significantly less antibacterial activity. Two compounds were identified with increased antibacterial activity against M. tuberculosis, with a minimum inhibitory concentration of 2.5 μg/ml.     

Proteome analyses of cellular proteins in methicillin-resistant Staphylococcus aureus treated with rhodomyrtone, a novel antibiotic candidate

The ethanolic extract from Rhodomyrtus tomentosa leaf exhibited good antibacterial activities against both methicillin-resistant Staphylococcus aureus (MRSA) and S. aureus ATCC 29213. Its minimal inhibitory concentration (MIC) values ranged from 31.25-62.5 μg/ml, and the minimal bactericidal concentration (MBC) was 250 μg/ml. Rhodomyrtone, an acylphloroglucinol derivative, was 62.5-125 times more potent at inhibiting the bacteria than the ethanolic extract, the MIC and MBC values were 0.5 μg/ml and 2 μg/ml, respectively. To provide insights into antibacterial mechanisms involved, the effects of rhodomyrtone on cellular protein expression of MRSA have been investigated using proteomic approaches. Proteome analyses revealed that rhodomyrtone at subinhibitory concentration (0.174 μg/ml) affected the expression of several major functional classes of whole cell proteins in MRSA. The identified proteins involve in cell wall biosynthesis and cell division, protein degradation, stress response and oxidative stress, cell surface antigen and virulence factor, and various metabolic pathways such as amino acid, carbohydrate, energy, lipid, and nucleotide metabolism. Transmission electron micrographs confirmed the effects of rhodomyrtone on morphological and ultrastructural alterations in the treated bacterial cells. Biological processes in cell wall biosynthesis and cell division were interrupted. Prominent changes including alterations in cell wall, abnormal septum formation, cellular disintegration, and cell lysis were observed. Unusual size and shape of staphylococcal cells were obviously noted in the treated MRSA. These pioneer findings on proteomic profiling and phenotypic features of rhodomyrtone-treated MRSA may resolve its antimicrobial mechanisms which could lead to the development of a new effective regimen for the treatment of MRSA infections.     

Aaptamines as sortase A inhibitors from the tropical sponge Aaptos aaptos

Four aaptamines (1-4), 1H-benzo[de][1,6]-naphthyridine alkaloids, were isolated from the marine sponge Aaptos aaptos and their inhibitory activities against sortase A (SrtA), an enzyme that plays a key role in cell wall protein anchoring and virulence in Staphylococcus aureus, were evaluated. Isoaaptamine (2) was a potent inhibitor of SrtA, with an IC(50) value of 3.7+/-0.2 microg/mL. The suppression of fibronectin-binding activity by isoaaptamine (2) highlights its potential for the treatment of S. aureus infections via inhibition of SrtA activity. Our studies have identified a series of SrtA inhibitors, providing the basis for further development of potent inhibitors.     

Design, synthesis and biological evaluation of benzo[1.3.2]dithiazolium ylide 1,1-dioxide derivatives as potential dual cyclooxygenase-2/5-lipoxygenase inhibitors

3-(4-Bromophenyl)-6-nitrobenzo[1.3.2]dithiazolium ylide 1,1-dioxide (5) was discovered as a new prototype for dual inhibitors of cyclooxygenase-2 (COX-2) and 5-lipoxygenase (5-LOX). Thus, the structure-activity relationships of benzo[1.3.2]dithiazolium ylide 1,1-dioxide skeleton were carried out. The 6-NO(2) group played an essential role in the inhibitory activity. In addition, moderate-sized lipophilic substituents at the para-position of the 3-aryl moiety were required for dual COX-2/5-LOX inhibitory activity. Among the identified potent dual inhibitors, 3-(4-tbutylphenyl) derivative 30c (IC(50) values of 0.27 μM and 0.30 μM against COX-2 and 5-LOX, respectively) and 3-(4-biphenyl) derivative 30f (IC(50) values of 0.50 μM and 0.15μM against COX-2 and 5-LOX, respectively) were the most potent dual COX-2/5-LOX inhibitors. Intraperitoneal administration of 30c at 100mg/kg demonstrated potent acute anti-inflammatory activity. As a result, benzo[1.3.2]dithiazolium ylide 1,1-dioxide represented a novel scaffold for the exploitation in developing dual COX-2/5-LOX inhibitors.     

Discovery and modification of sulfur-containing heterocyclic pyrazoline derivatives as potential novel class of β-ketoacyl-acyl carrier protein synthase III (FabH) inhibitors

A series of sulfur-containing heterocyclic pyrazoline derivatives (C1-C18; D1-D9) have been synthesized and purified (all are new except one) to be screened for FabH inhibitory activity. Compound C14 showed the most potent biological activity against Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis (MIC values: 1.56-3.13 μg/mL), being comparable with the positive control, while D6 performed the best in the thiazolidinone series (MIC values: 3.13-6.25 μg/mL). They also demonstrated strong broad-spectrum antimicrobial activity. Compounds C14 and D6 exhibited the most potent E. coli FabH inhibitory activity with IC(50) of 4.6 and 8.4 μM, respectively, comparable with the positive control DDCP (IC(50)=2.8 μM). Docking simulation was performed to position compound C14 and D6 into the E. coli FabH structure active site to determine the probable binding model. The structurally modification of previous compounds and the attempt in innovative target have brought a positive progress.     

Methicillin-resistant Staphylococcus aureus (MRSA) pyruvate kinase as a target for bis-indole alkaloids with antibacterial activities

Novel classes of antimicrobials are needed to address the emergence of multidrug-resistant bacteria such as methicillin-resistant Staphylococcus aureus (MRSA). We have recently identified pyruvate kinase (PK) as a potential novel drug target based upon it being an essential hub in the MRSA interactome (Cherkasov, A., Hsing, M., Zoraghi, R., Foster, L. J., See, R. H., Stoynov, N., Jiang, J., Kaur, S., Lian, T., Jackson, L., Gong, H., Swayze, R., Amandoron, E., Hormozdiari, F., Dao, P., Sahinalp, C., Santos-Filho, O., Axerio-Cilies, P., Byler, K., McMaster, W. R., Brunham, R. C., Finlay, B. B., and Reiner, N. E. (2011) J. Proteome Res. 10, 1139-1150; Zoraghi, R., See, R. H., Axerio-Cilies, P., Kumar, N. S., Gong, H., Moreau, A., Hsing, M., Kaur, S., Swayze, R. D., Worrall, L., Amandoron, E., Lian, T., Jackson, L., Jiang, J., Thorson, L., Labriere, C., Foster, L., Brunham, R. C., McMaster, W. R., Finlay, B. B., Strynadka, N. C., Cherkasov, A., Young, R. N., and Reiner, N. E. (2011) Antimicrob. Agents Chemother. 55, 2042-2053). Screening of an extract library of marine invertebrates against MRSA PK resulted in the identification of bis-indole alkaloids of the spongotine (A), topsentin (B, D), and hamacanthin (C) classes isolated from the Topsentia pachastrelloides as novel bacterial PK inhibitors. These compounds potently and selectively inhibited both MRSA PK enzymatic activity and S. aureus growth in vitro. The most active compounds, cis-3,4-dihyrohyrohamacanthin B (C) and bromodeoxytopsentin (D), were identified as highly potent MRSA PK inhibitors (IC(50) values of 16-60 nM) with at least 166-fold selectivity over human PK isoforms. These novel anti-PK natural compounds exhibited significant antibacterial activities against S. aureus, including MRSA (minimal inhibitory concentrations (MIC) of 12.5 and 6.25 μg/ml, respectively) with selectivity indices (CC(50)/MIC) >4. We also report the discrete structural features of the MRSA PK tetramer as determined by x-ray crystallography, which is suitable for selective targeting of the bacterial enzyme. The co-crystal structure of compound C with MRSA PK confirms that the latter is a target for bis-indole alkaloids. It elucidates the essential structural requirements for PK inhibitors in "small" interfaces that provide for tetramer rigidity and efficient catalytic activity. Our results identified a series of natural products as novel MRSA PK inhibitors, providing the basis for further development of potential novel antimicrobials.     

Identification of inhibitors against Mycobacterium tuberculosis thiamin phosphate synthase, an important target for the development of anti-TB drugs

Tuberculosis (TB) continues to pose a serious challenge to human health afflicting a large number of people throughout the world. In spite of the availability of drugs for the treatment of TB, the non-compliance to 6-9 months long chemotherapeutic regimens often results in the emergence of multidrug resistant strains of Mycobacterium tuberculosis adding to the precariousness of the situation. This has necessitated the development of more effective drugs. Thiamin biosynthesis, an important metabolic pathway of M. tuberculosis, is shown to be essential for the intracellular growth of this pathogen and hence, it is believed that inhibition of this pathway would severely affect the growth of M. tuberculosis. In this study, a comparative homology model of M. tuberculosis thiamin phosphate synthase (MtTPS) was generated and employed for virtual screening of NCI diversity set II to select potential inhibitors. The best 39 compounds based on the docking results were evaluated for their potential to inhibit the MtTPS activity. Seven compounds inhibited MtTPS activity with IC(50) values ranging from 20-100 μg/ml and two of these exhibited weak inhibition of M. tuberculosis growth with MIC(99) values being 125 μg/ml and 162.5 μg/ml while one compound was identified as a very potent inhibitor of M. tuberculosis growth with an MIC(99) value of 6 μg/ml. This study establishes MtTPS as a novel drug target against M. tuberculosis leading to the identification of new lead molecules for the development of antitubercular drugs. Further optimization of these lead compounds could result in more potent therapeutic molecules against Tuberculosis.     

Discovery of potent antimicrobial peptide analogs of Ixosin-B

Antimicrobial peptides (AMPs) represent the first defense line against infection when organisms are infected by pathogens. These peptides are generally good targets for the development of antimicrobial agents. Peptide amide analogs of Ixosin-B, an antimicrobial peptide with amino acid sequence of QLKVDLWGTRSGIQPEQHSSGKSDVRRWRSRY, were designed, synthesized and examined for antimicrobial activities against Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa. Within the peptides synthesized, we discovered an 11-mer peptide, KRLRRVWRRWR-amide, which exhibited potent antimicrobial activity while very little hemolytic activity in human erythrocytes was observed even at high dose level (100 μM). With further modifications, this peptide could be developed into a potent antimicrobial agent in the future.     

Cyclic tripeptide-based potent human SIRT7 inhibitors

In the current study, two cyclic tripeptides respectively harboring a thiourea-type and a carboxamide-type of catalytic mechanism-based sirtuin inhibitory warheads as the central residue were found to behave as potent (low μM level) inhibitors against the tRNA-activated human SIRT7 deacetylase activity. Despite exhibiting a potent pan-inhibition against the deacylase activities of the five tested human sirtuins (i.e. SIRT1/2/3/6/7), these two compounds represent the first examples of potent SIRT7 inhibitors ever identified thus far, and their identification could facilitate the future development of more potent and selective SIRT7 inhibitors.     

Antimicrobial and antiviral screening of novel indole carboxamide and propanamide derivatives

A few series of indole derivatives were screened for antimicrobial, antifungal and anti-HBV activities. The compounds were tested for their in vitro antibacterial activity against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and for their antifungal activity against Candida albicans using a disc diffusion method, which measures the diameter of the inhibition zone around a paper disc soaked in a solution of the test compounds. The antimicrobial activity results showed that all compounds are as a active as the standard compound ampicillin against Staphylococcus aureus. It was also found that indole carboxamide derivatives, substituted at 3-position with several benzyl groups, showed better inhibition of Bacillus subtilis than their congeners substituted at 2-position. Activity patterns of the compounds against Escherichia coli and Staphylococcus aureus were found slightly different by the same method. In this case, there was no correlation between structure and activity of the compounds. The antifungal activity of carboxamide derivatives was found higher compared to that of the propanamide derivatives. The minimum inhibitory concentration (MIC) values of some indole derivatives were also determined by the tube dilution technique. The MIC values of the compounds were found nearly 20- to 100-fold smaller compared to the standard compounds ciprofloxacin and ampicillin (1.56-3.13 microg/ml and 1.56-12.5 microg/ml, respectively) against Staphylococcus aureus, Bacillus subtilis and Escherichia coli. The MIC values of the tested compounds showed that these are better inhibitors for Candida albicans. Indole derivatives were screened by the anti-HBV susceptibility test. No compound showed good inhibition against the HBV virus.