Synthesis and antibacterial activities of novel pleuromutilin derivatives bearing an aminothiophenol moiety

We synthesized a series of novel thioether pleuromutilin derivatives incorporating 2‐aminothiophenol moieties into the C14 side chain via acylation reactions under mild conditions. We evaluated the in‐vitro antibacterial activities of the derivatives against methicillin‐resistant Staphylococcus aureus (MRSA, ATCC 43300), Staphylococcus aureus (ATCC 29213) and Escherichia coli (ATCC 25922). The majority of the synthesized derivatives possessed moderate antibacterial activities. Compound 8 was found to be the most active antibacterial derivative against MRSA. We conducted docking experiments to understand the possible mode of interactions between compounds 8 , 9b , 11a and 50S ribosomal subunit. The docking results proved that there is a reasonable correlation between the binding free energy and the antibacterial activity. Compound 8 was evaluated for its in‐vivo antibacterial activity and showed higher efficacy than tiamulin against MRSA in mouse infection model.     

From Zn to Mn: the study of novel manganese-binding groups in the search for new drugs against tuberculosis

In most eubacteria, apicomplexans, and most plants, including the causal agents for diseases such as malaria, leprosy, and tuberculosis, the methylerythritol phosphate pathway is the route for the biosynthesis of the C(5) precursors to the essential isoprenoid class of compounds. Owing to their absence in humans, the enzymes of the methylerythritol phosphate pathway have become attractive targets for drug discovery. This work investigates a new class of inhibitors against the second enzyme of the pathway, 1-deoxy-D-xylulose 5-phosphate reductoisomerase. Inhibition of this enzyme may involve the chelation of a crucial active site Mn ion, and the metal-chelating moieties studied here have previously been shown to be successful in application to the zinc-dependent metalloproteinases. Quantum mechanics and docking calculations presented in this work suggest the transferability of these metal-chelating compounds to Mn-containing 1-deoxy-D-xylulose 5-phosphate reductoisomerase enzyme, as a promising starting point to the development of potent inhibitors.     

Three-dimensional quantitative structure-activity relationship modeling of gamma-secretase inhibitors using molecular field analysis

Three-dimensional quantitative structure-activity relationship analysis of a set of 79 analogs of gamma-secretase inhibitors was performed by molecular field analysis with genetic partial least squares method to investigate the substitutional requirements to derive a predictive model and for the favorable receptor-drug interaction that may be used for the designing of a novel gamma-secretase inhibitors. The developed molecular field analysis model has a good fit, with r2 value of 0.952 and cross-validated coefficient, r2(cv), value of 0.931. Predictive ability of the developed model was further assessed using test set of 19 compounds and r2(pred) was found to be 0.665.     

Novel Glycoconjugate of 8‐Fluoro Norfloxacin Derivatives as Gentamicin‐resistant Staphylococcus aureus Inhibitors: Synthesis and Molecular Modelling Studies

Antibiotic resistance has been the subject of interest in clinical practice due to high prevalence of antibiotic‐resistant pathogenic organisms. In view of the prevalence of lesser resistance in antibiotics belonging to aminoglycoside class of compounds viz. Food and Drug Administration‐approved gentamicin for the treatment of Staphylococcus infections, which also has instances of resistance in the clinical isolates of Staphylococcus aureus, a series of novel glycoconjugates of 8‐fluoro norfloxacin analogues with high regio‐selectivity by employing copper (I)‐catalyzed 1, 3‐dipolar cycloaddition of 1‐O‐propargyl monosaccharides has been synthesized and evaluated for the antibacterial activity against gentamicin resistance Staphylococcus aureus. Among these compounds, the compound 10g showed better antibacterial activity (MIC = 3.12  μ g/ml) than gentamicin (Escherichia coli (12.5  μ g/ml), Staphylococcus aureus (6.25  μ g/ml) and Klebsiella pneumonia (6.25  μ g/ml), including gentamicin resistant (>50  μ g/ml) strain in vitro). The docking studies suggest DNA gyrase of Staphylococcus aureus as a probable target for the antibacterial action of compound 10g .     

Isonicotinohydrazones as inhibitors of alkaline phosphatase and ecto‐5′‐nucleotidase

A series of isonicotinohydrazide derivatives was synthesized and tested against recombinant human and rat ecto‐5′‐nucleotidases (h‐e5′NT and r‐e5′NT) and alkaline phosphatase isozymes including both bovine tissue‐non‐specific alkaline phosphatase (b‐TNAP) and tissue‐specific calf intestinal alkaline phosphatase (c‐IAP). These enzymes are implicated in vascular calcifications, hypophosphatasia, solid tumors, and cancers, such as colon, lung, breast, pancreas, and ovary. All tested compounds were active against both enzymes. The most potent inhibitor of h‐e5′NT was derivative (E)‐N′‐(1‐(3‐(4‐fluorophenyl)‐5‐phenyl‐4,5‐dihydro‐1H‐pyrazol‐1‐yl)ethylidene)isonicotinohydrazide ( 3j ), whereas derivative (E)‐N′‐(4‐hydroxy‐3‐methoxybenzylidene)isonicotinohydrazide ( 3g ) exhibited significant inhibitory activity against r‐e5′NT . In addition, the derivative (E)‐N′‐(4′‐chlorobenzylidene)isonicotinohydrazide ( 3a ) was most potent inhibitor against calf intestinal alkaline phosphatase and the derivative (E)‐N′‐(4‐hydroxy‐3‐methoxybenzylidene)isonicotinohydrazide ( 3g ) was found to be most potent inhibitor of bovine tissue‐non‐specific alkaline phosphatase. Furthermore, putative binding modes of potent compounds against e5′NT (human and rat e5′NT) and AP (including b‐TNAP and c‐IAP) were determined computationally.     

Computer-Aided Drug Discovery Approach Finds Calcium Sensitizer of Cardiac Troponin

In the fight against heart failure, therapeutics that have the ability to increase the contractile power of the heart are urgently needed. One possible route of action to improve heart contractile power is increasing the calcium sensitivity of the thin filament. From a pharmaceutical standpoint, calcium sensitizers have the distinct advantage of not altering cardiomyocyte calcium levels and thus have lower potential for side-effects. Small chemical molecules have been shown to bind to the interface between cTnC and the cTnI switch peptide and exhibit calcium-sensitizing properties, possibly by stabilizing cTnC in an open conformation. Building on existing structural data of a known calcium sensitizer bound to cardiac troponin, we combined computational structure-based virtual screening drug discovery methods and solution NMR titration assays to identify a novel calcium sensitizer 4-(4-(2,5-dimethylphenyl)-1-piperazinyl)-3-pyridinamine (NSC147866) which binds to cTnC and the cTnC-cTnI_147–163 complex. Its presence increases the affinity of switch peptide to cTnC by approximately a factor of two. This action is comparable to that of known levosimendan analogues.     

Molecular Mechanisms of Action and In Vivo Validation of an M4 Muscarinic Acetylcholine Receptor Allosteric Modulator with Potential Antipsychotic Properties

We recently identified LY2033298 as a novel allosteric potentiator of acetylcholine (ACh) at the M₄ muscarinic acetylcholine receptor (mAChR). This study characterized the molecular mode of action of this modulator in both recombinant and native systems. Radioligand-binding studies revealed that LY2033298 displayed a preference for the active state of the M₄ mAChR, manifested as a potentiation in the binding affinity of ACh (but not antagonists) and an increase in the proportion of high-affinity agonist–receptor complexes. This property accounted for the robust allosteric agonism displayed by the modulator in recombinant cells in assays of [³⁵S]GTPγS binding, extracellular regulated kinase 1/2 phosphorylation, glycogen synthase kinase 3β phosphorylation, and receptor internalization. We also found that the extent of modulation by LY2033298 differed depending on the signaling pathway, indicating that LY2033298 engenders functional selectivity in the actions of ACh. This property was retained in NG108-15 cells, which natively express rodent M₄ mAChRs. Functional interaction studies between LY2033298 and various orthosteric and allosteric ligands revealed that its site of action overlaps with the allosteric site used by prototypical mAChR modulators. Importantly, LY2033298 reduced [³H]ACh release from rat striatal slices, indicating retention of its ability to allosterically potentiate endogenous ACh in situ. Moreover, its ability to potentiate oxotremorine-mediated inhibition of condition avoidance responding in rodents was significantly attenuated in M₄ mAChR knockout mice, validating the M₄ mAChR as a key target of action of this novel allosteric ligand.     

Sequential virtual screening approach to the identification of small organic molecules as potential BACE-1 inhibitors

In this letter, we report on the sequential application of two different in silico screening approaches combined with bioassays aimed at the identification of small organic molecules as potential BACE-1 inhibitors. Two hits endowed of micromolar inhibitory potency were selected, and the binding mode of the most potent compound was further characterized through docking simulations.     

The Unique Binding Mode of Laulimalide to Two Tubulin Protofilaments

Laulimalide, a cancer chemotherapeutic in preclinical development, has a unique binding site located on two adjacent β‐tubulin units between tubulin protofilaments of a microtubule. Our extended protein model more accurately mimics the microtubule environment, and together with a 135 ns molecular dynamics simulation, identifies a new binding mode for laulimalide, which differs from the modes presented in work using smaller protein models. The new laulimalide–residue interactions that are computationally revealed explain the contacts observed via independent mass shift perturbation experiments. The inclusion of explicit solvent shows that many laulimalide–tubulin interactions are water mediated. The new contacts between the drug and the microtubule structure not only improve our understanding of laulimalide binding but also will be essential for efficient derivatization and optimization of this prospective cancer chemotherapy agent. Observed changes in secondary protein structure implicate the S7–H9 loop (M–loop) and H1′–S2 loop in the mechanism by which laulimalide stabilizes microtubules to exert its cytotoxic effects.     

CXCR4, inhibitors and mechanisms of action

In this review, the author discusses recent advances in anti-HIV inhibitors, targeting CXCR4, including natural and modified chemokines, peptides and organic compounds, their mechanisms of action, and the molecular process of virus invasion of immune cells. Peptides with strong anti-HIV activity exhibit several common features, such as electrostatic charges, cyclization, beta-turns and dimerization induced by a sulphide bond. Organic compounds, such as cyclams, display a unique metal-mediated mechanism in the binding process to its target CXCR4. Understanding of their mechanisms of action may be useful for the design of more effective drugs. Consecutive interactions of viral glycoprotein gp120 with CD4 and the co-receptor, CXCR4 or another co-receptor CCR5 on the cell surface leads to virus invasion into host cells. The molecular details of the binding between HIV glycoproteins and the co-receptors also provide a basis for anti-HIV therapy.     

Discovery and Development of Toll-Like Receptor 4 (TLR4) Antagonists: A New Paradigm for Treating Sepsis and Other Diseases

Sepsis remains the most common cause of death in intensive care units in the USA, with a current estimate of at least 750,000 cases per year, and 215,000 deaths annually. Despite extensive research still we do not quite understand the cellular and molecular mechanisms that are involved in triggering and propagation of septic injury. Endotoxin (lipopolysaccharide from Gram-negative bacteria, or LPS) has been implicated as a major cause of this syndrome. Inflammatory shock as a consequence of LPS release remains a serious clinical concern. In humans, inflammatory responses to LPS result in the release of cytokines and other cell mediators from monocytes and macrophages, which can cause fever, shock, organ failure and death. A number of different approaches have been investigated to try to treat and/or prevent the septic shock associated with infections caused by Gram-negative bacteria, including blockage of one or more of the cytokines induced by LPS. Recently several novel amphipathic compounds have been developed as direct LPS antagonists at the LPS receptor, TLR4. This review article will outline the current knowledge on the TLR4-LPS synthesis and discuss the signaling, in vitro pre-clinical and in vivo clinical evaluation of TLR4 antagonists and their potential use in sepsis and a variety of diseases such as atherosclerosis as well as hepatic and renal malfunction.     

Suitability of MMGBSA for the selection of correct ligand binding modes from docking results

The estimation of the correct binding mode and affinity of a ligand into a target protein using computational methods is challenging. However, docking can introduce poses from which the correct binding mode could be identified using other methods. Here, we analyzed the reliability of binding energy estimation using the molecular mechanics‐generalized Born surface area (MMGBSA) method without and with energy minimization to identify the likely ligand binding modes within docking results. MMGBSA workflow (a) outperformed docking in recognizing the correct binding modes of androgen receptor ligands and (b) improved the correlation coefficient of computational and experimental results of rescored docking poses to phosphodiesterase 4B. Combined with stability and atomic distance analysis, MMGBSA helped to (c) identify the binding modes and sites of metabolism of cytochrome P450 2A6 substrates. The standard deviation of estimated binding energy within one simulation was lowered by minimization in all three example cases. Minimization improved the identification of the correct binding modes of androgen receptor ligands. Although only three case studies are shown, the results are analogous and indicate that these behaviors could be generalized. Such identified binding modes could be further used, for example, with free energy perturbation methods to understand binding energetics more accurately.     

pso@autodock: a fast flexible molecular docking program based on Swarm intelligence

On the quest of novel therapeutics, molecular docking methods have proven to be valuable tools for screening large libraries of compounds determining the interactions of potential drugs with the target proteins. A widely used docking approach is the simulation of the docking process guided by a binding energy function. On the basis of the molecular docking program autodock, we present pso@autodock as a tool for fast flexible molecular docking. Our novel Particle Swarm Optimization (PSO) algorithms varCPSO and varCPSO-ls are suited for rapid docking of highly flexible ligands. Thus, a ligand with 23 rotatable bonds was successfully docked within as few as 100 000 computing steps (rmsd = 0.87 A), which corresponds to only 10% of the computing time demanded by autodock. In comparison to other docking techniques as gold 3.0, dock 6.0, flexx 2.2.0, autodock 3.05, and sodock, pso@autodock provides the smallest rmsd values for 12 in 37 protein-ligand complexes. The average rmsd value of 1.4 A is significantly lower then those obtained with the other docking programs, which are all above 2.0 A. Thus, pso@autodock is suggested as a highly efficient docking program in terms of speed and quality for flexible peptide-protein docking and virtual screening studies.     

Emerging Drugs: The Prospect for Improved Medicines (Vol. 3)

edited by W. C. Bowman, J. D. Fitzgerald and J. B. Taylor, Ashley Publications, 1998. £395.00 (397 pages) ISSN 1361-9195     

The Future of Combinatorial Chemistry as a Drug Discovery Paradigm

Pharmaceutical Research -     

Combined pharmacophore and 3D-QSAR study on a series of Staphylococcus aureus Sortase A inhibitors

Methicillin resistant Staphylococcus aureus has become a major health concern and it requires new therapeutic agents. Staphylococcus aureus Sortase A enzyme contributes in adherence of bacteria with the cell wall of host cell; consequently, inhibition of S. aureus Sortase A by small molecules could be employed as potential antibacterial agents against methicillin resistant S. aureus. Current study focused on the identification of 3D pharmacophoric features within a series of rhodanine, pyridazinone, and pyrazolethione analogs as S. aureus Sortase A inhibitors. Pharmacophore model was constructed employing representative molecules using Genetic Algorithm with Linear Assignment of Hypermolecular Alignment of Database. The identified pharmacophoric points were then utilized to create alignment hypothesis for three-dimensional quantitative structure-activity relationships. Outcome of comparative molecular field analysis and comparative molecular similarity indices analysis experiments were in good agreement (comparative molecular field analysis: q(2) = 0.562 and r(2) = 0.995, comparative molecular similarity indices analysis: q(2) = 0.549 and r(2) = 0.978) and capable of explaining the variance in biological activities coherently with respect to the structural features of compounds. The results were also found in concurrence with the outcome of pharmacophoric features.     

Design,synthesis, and structure–activity relationship studies of novel pleuromutilin derivatives having a piperazine ring

A series of novel pleuromutilin derivatives possessing piperazine moieties were synthesized under mild conditions. The in vitro antibacterial activities of these derivatives against Staphylococcus aureus and Escherichia coli were tested by the agar dilution method. Structure–activity relationship studies resulted in compounds 11b , 13b , and 14a with the most potent in vitro antibacterial activity among the series (minimal inhibitory concentration = 0.0625–0.125 μg/mL). The binding of compounds 11b , 13b , and 14a to the E. coli ribosome was investigated by molecular modeling, and it was found that there is a reasonable correlation between the binding free energy and the antibacterial activity.     

The chemical tuning of a weak zinc binding motif for histone deacetylase using electronic effects

The hydroxamic acid moiety is an effective metal-binding warhead for a variety of metalloenzyme targets of interest in drug-discovery. For the zinc-containing histone deacetylase enzymes in particular, this chemical group has been widely incorporated and studied in the clinic. An alternative chemical functionality for binding zinc is the α-aminocarbonyl motif, which has been shown to bind to histone deacetylase enzymes. The current article explores the minimal binding site theoretical approach combined with structural knowledge to explore the ideal chemical substitution pattern of the α-aminocarbonyl motif within HDAC8. The metal-binding strength of the group is predicted to be highly tunable to chemical substitution at the carbonyl and the α-amino carbon. A fixed receptor model approach with a dispersion-corrected density functional, clearly discerned the effect of different substituents at both these positions using either a flexible or partially fixed ligand optimized in the presence of a fixed receptor model of the HDAC8 binding site. An electron donating substituent such as methyl at the C(α) in combination with NMe(2) substitution at the carbonyl position, similar to observed crystal structures, result in the optimal energetic profile for binding the zinc atom in the HDAC8 enzyme.     

Applications of In Silico Solvent Screening and an Interactive Web-Based Portal for Pharmaceutical Crystallization Process Development

In an effort to reduce development time and costs associated with active pharmaceutical ingredient process solvent selection and crystallization design, a tiered approach to crystallization solvent selection was developed that leverages different solubility modeling tools selected on the basis of available data and the intended use of the prediction. To facilitate easy access to routine solubility modeling functionality with a high level of automation and parallelization, a web-based in silico solvent-screening tool was also developed as well as a user interface to visualize and interpret the large number of predicted results. Examples are presented to illustrate the utility of the workflow and solvent-screening tool at various stages of development for a diverse range of crystallization processes. Implementation of the in silico solvent selection workflow has led to a ∼10× reduction in active pharmaceutical ingredient usage and 20% reduction in full-time employee time per project based on average after the first year.