Synthesis, in silico, in vitro evaluation of furanyl- and thiophenyl-3-phenyl-1H-indole-2-carbohydrazide derivatives as tubulin inhibitors and anticancer agents.

Twenty-one new indole derivatives comprising of seven furanyl-3-phenyl-1H-indole-carbohydrazide derivatives and fourteen thiophenyl-3-phenyl-1H-indole-carbohydrazide derivatives were synthesised and biologically evaluated for their microtubule-destabilising effects, and antiproliferative activities against the National Cancer Institute 60 (NCI60) human cancer cell line panel. Among the derivatives, 6i showed the best cytotoxic activity exhibiting selectivity for COLO 205 colon cancer (LC50 = 71 nM), SK-MEL-5 melanoma cells (LC50 = 75 nM), and MDA-MB-435 (LC50 = 259 nM). Derivative 6j showed the strongest microtubule-destabilising effect. Both 6i and 6j were able to induce G2/M cell cycle arrest and apoptosis in MDA-MB-231 triple-negative breast cancer cells. Molecular docking simulation results suggested that these derivatives inhibit tubulin by binding at the colchicine site. The calculated molecular descriptors showed that the most potent derivatives have acceptable pharmacokinetic profiles and are favourable for oral drug administration.

Metal- and photocatalyst-free approach to visible-light-induced acylation of quinoxalinones.

A transition-metal- and photocatalyst-free photochemical reaction was successfully developed for the direct acylation of quinoxalin-2(1H)-ones, which was enabled by the formation of electron donor-acceptor (EDA) complexes. The use of 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU) as the electron donor allows efficient and operationally simple access to a series of C3-aroylated and acylated quinoxalin-2(1H)-ones with moderate to good yields.

Boesenbergia rotunda and Its Pinostrobin for Atopic Dermatitis: Dual 5-Lipoxygenase and Cyclooxygenase-2 Inhibitor and Its Mechanistic Study through Steady-State Kinetics and Molecular Modeling.

Human 5-lipoxygenase (5-LOX) and cyclooxygenase-2 (COX-2) are potential targets for suppressing pruritic skin inflammation in atopic dermatitis (AD). In addition, Staphylococcus aureus colonization and oxidative stress worsen AD skin conditions. We aimed to investigate anti-inflammatory activity, using 5-LOX and COX-2 inhibitions, and the anti-staphylococcal, and antioxidant potentials of several medicinal plants bio-prospected from traditional medicine related to AD pathogenesis. Essential oils and hexane fractions were prepared and analyzed using gas chromatography-mass spectrometry. Boesenbergia rotunda hexane extract displayed anti-Staphylococcus aureus (MIC = 10 µg/mL) and antioxidant activities (IC50 = 557.97 and 2651.67 µg/mL against DPPH and NO radicals, respectively). A major flavonoid, pinostrobin, was further nonchromatographically isolated. Pinostrobin was shown to be a potent 5-LOX inhibitor (IC50 = 0.499 µM) compared to nordihydroguaiaretic acid (NDGA; IC50 = 5.020 µM) and betamethasone dipropionate (BD; IC50 = 2.077 µM) as the first-line of AD treatment. Additionally, pinostrobin inhibited COX-2 (IC50 = 285.67 µM), which was as effective as diclofenac sodium (IC50 = 290.35 µM) and BD (IC50 = 240.09 µM). This kinetic study and molecular modeling showed the mixed-type inhibition of NDGA and pinostrobin against 5-LOX. This study suggests that B. rotunda and its bioactive pinostrobin have promising properties for AD therapy.

Virtual screening, identification and in vitro validation of small molecule GDP-mannose dehydrogenase inhibitors.

Upon undergoing mucoid conversion within the lungs of cystic fibrosis patients, the pathogenic bacterium Pseudomonas aeruginosa synthesises copious quantities of the virulence factor and exopolysaccharide alginate. The enzyme guanosine diphosphate mannose dehydrogenase (GMD) catalyses the rate-limiting step and irreversible formation of the alginate sugar nucleotide building block, guanosine diphosphate mannuronic acid. Since there is no corresponding enzyme in humans, strategies that could prevent its mechanism of action could open a pathway for new and selective inhibitors to disrupt bacterial alginate production. Using virtual screening, a library of 1447 compounds within the Known Drug Space parameters were evaluated against the GMD active site using the Glide, FRED and GOLD algorithms. Compound hit evaluation with recombinant GMD refined the panel of 40 potential hits to 6 compounds which reduced NADH production in a time-dependent manner; of which, an usnic acid derivative demonstrated inhibition six-fold stronger than a previously established sugar nucleotide inhibitor, with an IC50 value of 17 µM. Further analysis by covalent docking and mass spectrometry confirm a single site of GMD alkylation.

Assessing the Potential of Gallic Acid and Methyl Gallate to Enhance the Efficacy of ß-Lactam Antibiotics against Methicillin-Resistant Staphylococcus aureus by Targeting ß-Lactamase: In Silico and In Vitro Studies.

Methicillin-resistant Staphylococcus aureus (MRSA), a global health concern, has prompted research into antibiotic adjuvants as a potential solution. Although our group previously reported the enhancing effects of gallic acid (GA) and methyl gallate (MG) on penicillin G activity against MRSA, the synergistic potential with other ß-lactam antibiotics and the underlying mechanism have not been fully explored. Therefore, this study primarily aimed to investigate the antibacterial synergism with ß-lactam antibiotics through disc diffusion, checkerboard, and time-kill assays. The ß-lactamase inhibition was also examined through both molecular modeling and in vitro experiments. Additionally, bacterial morphology changes were studied using a scanning electron microscopy (SEM). The results revealed that both GA and MG exhibited anti-MRSA activity and showed indifferent effects when combined with ß-lactam antibiotics against methicillin susceptible S. aureus (MSSA). Interestingly, MG demonstrated synergism with only the ß-lactamase-unstable antibiotics against MRSA with the lowest fractional inhibitory concentration (FIC) indexes of ≤3.75. However, GA and MG exhibited weak ß-lactamase inhibition. Furthermore, GA, MG, and the combination with ampicillin induced the morphological changes in MRSA, suggesting a possible mechanism affecting the cell membrane. These findings suggest that MG could potentially serve as an adjunct to ß-lactam antibiotics to combat MRSA infections.

The redox-sensing mechanism of Agrobacterium tumefaciens NieR as a thiol-based oxidation sensor for hypochlorite stress.

NieR is a TetR family transcriptional repressor previously shown to regulate the NaOCl-inducible efflux pump NieAB in Agrobacterium tumefaciens. NieR is an ortholog of Escherichia coli NemR that specifically senses hypochlorite through the redox switch of a reversible sulfenamide bond between C106 and K175. The amino acid sequence of NieR contains only one cysteine. NieR has C104 and R166, which correspond to C106 and K175 of NemR, respectively. The aim of this study was to investigate the redox-sensing mechanism of NieR under NaOCl stress. C104 and R166 were subjected to mutagenesis to determine their roles. Although the substitution of R166 by alanine slightly reduced its DNA-binding activity, NieR retained its repressor function. By contrast, the DNA-binding and repression activities of NieR were completely lost when C104 was replaced by alanine. C104 substitution with serine only partially impaired the repressor function. Mass spectrometry analysis revealed an intermolecular disulfide bond between the C104 residues of NieR monomers. This study demonstrates the engagement of C104 in the mechanism of NaOCl sensing. C104 oxidation induced the formation of a disulfide-linked dimer that was likely to alter conformation, thus abolishing the DNA-binding ability of NieR and derepressing the target genes.

Phosphatidylcholine-Specific Phospholipase C as a Promising Drug Target.

Phosphatidylcholine-specific phospholipase C (PC-PLC) is an enzyme that catalyzes the formation of the important secondary messengers phosphocholine and diacylglycerol (DAG) from phosphatidylcholine. Although PC-PLC has been linked to the progression of many pathological conditions, including cancer, atherosclerosis, inflammation and neuronal cell death, studies of PC-PLC on the protein level have been somewhat neglected with relatively scarce data. To date, the human gene expressing PC-PLC has not yet been found, and the only protein structure of PC-PLC that has been solved was from Bacillus cereus (PC-PLCBc). Nonetheless, there is evidence for PC-PLC activity as a human functional equivalent of its prokaryotic counterpart. Additionally, inhibitors of PC-PLCBc have been developed as potential therapeutic agents. The most notable classes include 2-aminohydroxamic acids, xanthates, N,N'-hydroxyureas, phospholipid analogues, 1,4-oxazepines, pyrido[3,4-b]indoles, morpholinobenzoic acids and univalent ions. However, many medicinal chemistry studies lack evidence for their cellular and in vivo effects, which hampers the progression of the inhibitors towards the clinic. This review outlines the pathological implications of PC-PLC and highlights current progress and future challenges in the development of PC-PLC inhibitors from the literature.

A structure-based virtual high-throughput screening, molecular docking, molecular dynamics and MM/PBSA study identified novel putative drug-like dual inhibitors of trypanosomal cruzain and rhodesain cysteine proteases.

Virtual screening a collection of ~ 25,000 ChemBridge molecule collection identified two nitrogenous heterocyclic molecules, 12 and 15, with potential dual inhibitory properties against trypanosomal cruzain and rhodesain cysteine proteases. Similarity search in DrugBank found the two virtual hits with novel chemical structures with unreported anti-trypanosomal activities. Investigations into the binding mechanism by molecular dynamics simulations for 100 ns revealed the molecules were able to occupy the binding sites and stabilise the protease complexes. Binding affinities calculated using the MM/PBSA method for the last 20 ns showed that the virtual hits have comparable binding affinities to other known inhibitors from literature suggesting both molecules as promising scaffolds with dual cruzain and rhodesain inhibition properties, i.e. 12 has predicted ΔGbind values of - 38.1 and - 38.2 kcal/mol to cruzain and rhodesain, respectively, and 15 has predicted ΔGbind values of - 34.4 and - 25.8 kcal/mol to rhodesain. Per residue binding free energy decomposition studies and visual inspection at 100 ns snapshots revealed hydrogen bonding and non-polar attractions with important amino acid residues that contributed to the ΔGbind values. The interactions are similar to those previously reported in the literature. The overall ADMET predictions for the two molecules were favourable for drug development with acceptable pharmacokinetic profiles and adequate oral bioavailability.

Phlegcarines A-C, three Lycopodium alkaloids from Phlegmariurus carinatus (Desv. ex Poir.) Ching.

Three undescribed Lycopodium alkaloids, phlegcarines A-C, along with nine known alkaloids, were isolated from the aerial parts of a gardening clubmoss Phlegmariurus carinatus (Desv. ex Poir.) Ching. Phlegcarine A is an undescribed Lycopodium alkaloid possessing an unprecedented 5/9/6/6 fused-tetracyclic ring system consisting of an oxa-cyclononanone, a piperidine, a methylcyclohaxane and an oxazolidine. Phlegcarine B is the first N-chloromethyl piperidinium Lycopodium alkaloid of (+)-lycoflexine. The semi-synthesis of phlegcarine B was investigated from (+)-fawcettimine. Phlegcarine C, an undescribed epimer of 12-epilycodoline, is a rare lycopodine-type alkaloid with ß-oriented H-4. Transformation of phlegcarine C and lycodoline to 12-epilycopodine N-oxide via keto-enol tautomerization was investigated using m-CPBA. The structural assignments were established through comprehensive spectroscopic techniques and chemical correlations. Phlegcarines A-C were assayed for their anti-acetylcholinesterase activity, but none of them exhibited biological activity more potent than that of huperzine A.

Siamenflavones A-C, three undescribed biflavonoids from Selaginella siamensis Hieron. and biflavonoids from spike mosses as EGFR inhibitor.

Three undescribed biflavonoids (BFVs), siamenflavones A-C along with twelve BFVs were isolated from Selaginella siamensis Hieron. and Selaginella bryopteris (L.) Baker (Selaginellaceae). The chemical structures of undescribed compounds were established through comprehensive spectroscopic techniques, chemical correlations, and X-ray crystallography. The ten isolated BFVs, siamenflavones A-C, delicaflavone, chrysocauflavone, robustaflavone, robustaflavone-4-methylether, amentoflavone, tetrahydro-amentoflavone, and sciadopitysin were evaluated for the antiproliferative effects against four human cancer cell lines A549, H1975, HepG2 and T47D. Delicaflavone and robustaflavone 4'-methylether exerted strong effects on the four human cancer cell lines. Siamenflavone B, delicaflavone and robustaflavone 4'-methylether showed potent inhibitory activities against wild-type EGFR. The inhibition of the compounds was further supported by molecular docking and predictive intermolecular interactions. Molecular dynamics simulation studies of siamenflavone B and robustaflavone-4'-methylether complexed to EGFR-TK further supported inhibition of the compounds to the ATP binding site. Finally, analysis of pharmacokinetic and electronic properties using density-functional theory and known drug index calculations suggest that the compounds are pharmaceutically compatible for drug administration.

Characterisation of the triclosan efflux pump TriABC and its regulator TriR in Agrobacterium tumefaciens C58.

TriR serves as a repressor for a resistance-nodulation-cell division (RND) efflux pump TriABC involved in triclosan (TCS) resistance in Agrobacterium tumefaciens. The triR gene is transcribed divergently from the triABC operon. TriR specifically bound to the triR-triA intergenic region, at an imperfect 10 bp inverted repeat, 5'-TTGACTAttC-GgtTAGTCAA-3' (TriR box), that was revealed by DNase I footprinting and electrophoretic mobility shift assay. TCS treatment appeared to up-regulate triR and triABC expression, via preventing TriR binding to the triR-triA intergenic region. Promoter-lacZ fusions and ß-galactosidase activity assay further demonstrated TriR-mediated repression of triABC and triR autoregulation. Site-directed mutagenesis confirmed the identified TriR box is essential for TriR repression. A. tumefaciens mutant strains disrupting either triR or triA were constructed to determine their biological functions. The triA mutant showed hypersensitivity to TCS and sodium dodecyl sulfate (SDS), whereas the triR mutant was hyper-resistant, compared to wild-type. In addition to TCS and SDS, overproduction of TriABC from a multi-copy plasmid conferred enhanced resistance to a quaternary ammonium compound, benzalkonium chloride. Molecular modelling was able to predict the model of TriR and docking simulations were able to anticipate plausible binding interactions between TriR and TCS ligand.

Discovery of potent antiproliferative agents from selected oxygen heterocycles as EGFR tyrosine kinase inhibitors from the U.S. National Cancer Institute database by in silico screening and bioactivity evaluation.

A similarity search was conducted on the U.S. Enhanced National Cancer Institute Database Browser 2.2 to find structures related to 1,5-dihydroxy-9H-xanthen-9-one, a previously established EGFR-TK inhibitor. Compounds were virtually screened and selected for bioactivity testing revealed 5 candidates, mostly displayed stronger antiproliferative activities than erlotinib with IC50 values between 0.95 and 17.71 µM against overexpressed EGFR-TK cancer cell lines: A431 and HeLa. NSC107228 displayed the strongest antiproliferative effects with IC50 values of 2.84 and 0.95 µM against A431 and HeLa cancer cell lines, respectively. Three compounds, NSC81111, NSC381467 and NSC114126 inhibited EGFR-TK with IC50 values between 0.15 and 30.18 nM. NSC81111 was the best inhibitor with IC50 = 0.15 nM. Molecular docking analysis of the 3 compounds predicted hydrogen bonding and hydrophobic interactions with key residues were important for the bioactivities observed. Furthermore, calculations of the physicochemical properties suggest the compounds are drug-like and are potentially active orally.

Two new nor-lignans, siamensinols A and B, from Selaginella siamensis Hieron. and their biological activities.

Two new nor-lignans siamensinols A-B (1-2) and seven known compounds agatharesinol (3), syringaresinol-glucoside (4), noreugenin (5), 8-methyleugenitol (6), melachromone (7), uncinoside A (8) and daucosterol (9) were isolated from Selaginella siamensis Hieron. The structures of the new compounds were elucidated on the basis of comprehensive spectroscopic methods, including 1 D, 2 D-NMR, HR-ESI-MS and CD spectrometry. Compounds 1-2 showed moderate inhibitory effect on MOLT-3 cells while 8-methyleugenitol (6) exhibited moderate inhibitory effect on three tumor cells (HepG2, A549 and HuCCA-1). Compounds 2-3 showed the potent cancer chemoprevention in DPPH, XXO, IXO and AIA assays.

Impact of the ferrocenyl group on cytotoxicity and KSP inhibitory activity of ferrocenyl monastrol conjugates.

The incorporation of the ferrocenyl moiety into a bioactive molecule may significantly alter the activity of the resulting conjugate. By applying this strategy, we designed ferrocenyl analogs of monastrol - the first low molecular weight kinesin spindle protein (KSP) inhibitor. The obtained compounds showed low micromolar antiproliferative activity towards a panel of sensitive and ABC-overexpressing cancer cells. Most cytotoxic compounds exhibited also higher KSP modulatory activity and ability for ROS generation compared to monastrol. The increased bioactivity of the studied compounds can be attributed to the presence of the ferrocenyl group.

Visible-light-mediated decarboxylative alkylation of 2-pyridone derivatives via a C3-selective C-H functionalization.

A direct C-H functionalization approach to access C3-alkylated 2-pyridone derivatives is reported. This study utilizes N-hydroxyphthalimide (NHPI) esters of various carboxylic acids as sources of alkyl radicals by reductive cleavage under photocatalytic reaction conditions. The carbon-carbon bond formation occurred site-selectively at C3 of 2-pyridone to give the desired products in moderate to good yields. This method enables a faster access to C3-alkylated pyridone compounds which can be applied to the synthesis of small molecule drugs.

Diterpenoids with Aromatase Inhibitory Activity from the Rhizomes of Kaempferia elegans.

Investigation of bioactive compounds from the rhizomes of Kaempferia elegans led to the isolation and characterization of ten new diterpenoids, namely, five 12,13-seco-diterpenoids named elegansins A-E (1-5) and five new abietanes, elegansols A-E (6-10), together with seven known diterpenoids (11-17). The structure elucidation of the new compounds was achieved by HRESIMS, NMR, and ECD spectroscopic analysis. Compounds (1-5) are the first examples of 12,13-seco-diterpenoid-type compounds representing a decalin fused dihydropyran skeleton. Plausible biosynthetic pathways for compounds 1-5 are proposed. Aromatase inhibitory activities of all compounds were evaluated, and abieta-8,11,13-trien-11-ol (16) was found to be the most potent aromatase inhibitor with an IC50 value of 3.7 µM.

Identification of new 3-phenyl-1H-indole-2-carbohydrazide derivatives and their structure-activity relationships as potent tubulin inhibitors and anticancer agents: A combined in silico, in vitro and synthetic study.

Virtual screening of commercially available molecular entities by using CDRUG, structure-based virtual screening, and similarity identified eight new derivatives of 3-phenyl-1H-indole-2-carbohydrazide with anti-proliferative activities. The molecules were tested experimentally for inhibition of tubulin polymerisation, which revealed furan-3-ylmethylene-3-phenyl-1H-indole-2-carbohydrazide (27a) as the most potent candidate. Molecule 27a was able to induce G2/M phase arrest in A549 cell line, similar to other tubulin inhibitors. Synthetic modifications of 27a were focussed on small substitutions on the furan ring, halogenation at R1 position and alteration of furyl connectivity. Derivatives 27b, 27d and 27i exhibited the strongest tubulin inhibition activities and were comparable to 27a. Bromine substitution at R1 position showed most prominent anticancer activities; derivatives 27b-27d displayed the strongest activities against HuCCA-1 cell line and were more potent than doxorubicin and the parent molecule 27a with IC50 values <0.5 µM. Notably, 27b with a 5-methoxy substitution on furan displayed the strongest activity against HepG2 cell line (IC50 = 0.34 µM), while 27d displayed stronger activity against A549 cell line (IC50 = 0.43 µM) compared to doxorubicin and 27a. Fluorine substitutions at the R1 position tended to show more modest anti-tubulin and anticancer activities, and change of 2-furyl to 3-furyl was tolerable. The new derivatives, thiophenyl 26, displayed the strongest activity against A549 cell line (IC50 = 0.19 µM), while 1-phenylethylidene 21b and 21c exhibited more modest anticancer activities with unclear mechanisms of action; 26 and 21c demonstrated G2/M phase arrest, but showed weak tubulin inhibitory properties. Molecular docking suggests the series inhibit tubulin at the colchicine site, in agreement with the experimental findings. The calculated molecular descriptors indicated that the molecules obey Lipinski's rule which suggests the molecules are drug-like structures.

The 4-(Phenylsulfanyl) butan-2-one Improves Impaired Fear Memory Retrieval and Reduces Excessive Inflammatory Response in Triple Transgenic Alzheimer's Disease Mice.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by an excessive inflammatory response and impaired memory retrieval, including spatial memory, recognition memory, and emotional memory. Acquisition and retrieval of fear memory help one avoid dangers and natural threats. Thus, it is crucial for survival. AD patients with impaired retrieval of fear memory are vulnerable to dangerous conditions. Excessive expression of inflammatory markers is known to impede synaptic transmission and reduce the efficiency of memory retrieval. In wild-type mice, reducing inflammation response can improve fear memory retrieval; however, this effect of this approach is not yet investigated in 3xTg-AD model mice. To date, no satisfactory drug or treatment can attenuate the symptoms of AD despite numerous efforts. In the past few years, the direction of therapeutic drug development for AD has been shifted to natural compounds with anti-inflammatory effect. In the present study, we demonstrate that the compound 4-(phenylsulfanyl) butan-2-one (4-PSB-2) is effective in enhancing fear memory retrieval of wild-type and 3xTg-AD mice by reducing the expression of TNF-α, COX-2, and iNOS. We also found that 4-PSB-2 helps increase dendritic spine density, postsynaptic density protein-95 (PSD-95) expression, and long-term potentiation (LTP) in the hippocampus of 3xTg-AD mice. Our study indicates that 4-PSB-2 may be developed as a promising therapeutic compound for treating fear memory impairment of AD patients.

An optimised MALDI-TOF assay for phosphatidylcholine-specific phospholipase C.

The Bacillus cereus phosphatidylcholine-specific phospholipase C (PC-PLCBc) is an enzyme that catalyses the hydrolysis of phosphatidylcholines into phosphocholine and 1,2-diacylglycerols. PC-PLCBc has found applications in both the food industry and in medicinal chemistry. Herein, we report our work in the development and optimisation of a matrix assisted laser desorption ionisation time-of-flight (MALDI-TOF) mass spectrometry-based assay to monitor PC-PLCBc activity. The use of one-phase and two-phase reaction systems to assess the inhibition of PC-PLCBc with different structural classes of inhibitors was compared. We also highlighted the advantage of our assay over the commonly used commercially available Amplex Red assay. This method will also be applicable to work on the activity and inhibition of other phospholipases.

The discovery of novel antitrypanosomal 4-phenyl-6-(pyridin-3-yl)pyrimidines.

Human African trypanosomiasis, or sleeping sickness, is a neglected tropical disease caused by Trypanosoma brucei rhodesiense and Trypanosoma brucei gambiense which seriously affects human health in Africa. Current therapies present limitations in their application, parasite resistance, or require further clinical investigation for wider use. Our work herein describes the design and syntheses of novel antitrypanosomal 4-phenyl-6-(pyridin-3-yl)pyrimidines, with compound 13, the 4-(2-methoxyphenyl)-6-(pyridine-3-yl)pyrimidin-2-amine demonstrating an IC50 value of 0.38 µM and a promising off-target ADME-Tox profile in vitro. In silico molecular target investigations showed rhodesain to be a putative candidate, supported by STD and WaterLOGSY NMR experiments, however, in vitro evaluation of compound 13 against rhodesain exhibited low experimental inhibition. Therefore, our reported library of drug-like pyrimidines present promising scaffolds for further antikinetoplastid drug development for both phenotypic and target-based drug discovery.