Rivastigmine-Bambuterol Hybrids as Selective Butyrylcholinesterase Inhibitors.

Selective butyrylcholinesterase inhibitors are considered promising drug candidates for the treatment of Alzheimer's disease. In this work, one rivastigmine-bambuterol hybrid (MTR-1) and fourteen of its analogues were synthesized, purified, and characterized. In vitro cholinesterase assays showed that all the compounds were more potent inhibitors of BChE when compared to AChE.

Dual-Emission GFP Chromophore-Based Derivative for Imaging and Discriminating Aβ Oligomers and Aggregates.

β-Amyloid deposition is one of the main pathological features of Alzheimer's disease (AD). The development of fluorescent probes targeting specific β-amyloid species has recently become an attractive strategy to achieve the early diagnosis of AD. In this work, a dual-channel fluorescent protein chromophore derivative was rationally designed and synthesized for the detection and discrimination of Aβ aggregates and oligomers.

Efficient genome editing for Pseudomonas aeruginosa using CRISPR-Cas12a.

The CRISPR-Cas12a system has been demonstrated as an attractive tool for bacterial genome engineering. In particular, FnCas12a recognizes protospacer-adjacent motif (PAM) sites with medium or low GC content, which complements the Cas9-based systems. Here we explored Francisella novicida Cas12a (FnCas12a) for genome editing in Pseudomonas aeruginosa.

Spy chemistry-enabled protein directional immobilization and protein purification.

Site-directed protein immobilization allows the homogeneous orientation of proteins with high retention of activity, which is advantageous for many applications. Here, we report a facile, specific, and efficient strategy based on the SpyTag-SpyCatcher chemistry. Two SpyTag-fused model proteins, that is, the monomeric red fluorescent protein (RFP) and the oligomeric glutaryl-7-aminocephalosporanic acid acylase, were easily immobilized onto a SpyCatcher-modified resin directly from cell lysates, with activity recoveries in the range of 85-91%.

Design, synthesis and biological evaluation of novel carbamates as potential inhibitors of acetylcholinesterase and butyrylcholinesterase.

Rivastigmine, a dual inhibitor of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), has been approved by U.S. Food and Drug Administration to treat Alzheimer's disease (AD) and Parkinson's disease (PD) dementia.

Kinetically-controlled mechanism-based isolation of metabolic serine hydrolases in active form from complex proteomes: butyrylcholinesterase as a case study.

In this work an activity-based probe containing a carbamate group was designed to isolate human butyrylcholinesterase (hBChE), a metabolic serine hydrolase (mSH), from complex proteomes. The method took advantage of the native interaction mechanism of mSHs with carbamate pseudo-substrates for temporarily capturing the enzyme on a resin functionalized with the carbamate probe and releasing the enzyme in active form after removal of the contaminating proteins. The isolation relied on the possibility of manipulating the carbamylation and decarbamylation kinetics favoring the former during the capture and wash steps and the latter in the release step.

New trends in aggregating tags for therapeutic protein purification.

The rapid growth of the therapeutic protein market calls for more efficient purification methods. Various aggregating tags have recently emerged as simple, fast, cost-effective and column-free technologies for protein (and peptide) purification. In general, these column-free protein purification technologies involve the use of aggregating tags that induce the target protein into insoluble aggregates.

HybridSim-VS: a web server for large-scale ligand-based virtual screening using hybrid similarity recognition techniques.

Summary: Molecular-similarity searches based on two-dimensional (2D) fingerprint and three-dimensional (3D) shape represent two widely used ligand-based virtual screening (VS) methods in computer-aided drug design. 2D fingerprint-based VS utilizes the binary fragment information on a known ligand, whereas 3D shape-based VS takes advantage of geometric information for predefined features from a 3D conformation. Given their different advantages, it would be desirable to hybridize 2D fingerprint and 3D shape molecular-similarity approaches in drug discovery.

Alterations in NO/ROS ratio and expression of Trx1 and Prdx2 in isoproterenol-induced cardiac hypertrophy.

The development of cardiac hypertrophy is a complicated process, which undergoes a transition from compensatory hypertrophy to heart failure, and the identification of new biomarkers and targets for this disease is greatly needed. Here we investigated the development of isoproterenol (ISO)-induced cardiac hypertrophy in an in vitro experimental model. After the induction of hypertrophy with ISO treatment in H9c2 cells, cell surface area, cell viability, cellular reactive oxygen species (ROS), and nitric oxide (NO) levels were tested.

A new method to characterize the kinetics of cholinesterases inhibited by carbamates.

The inhibition of cholinesterases (ChEs) by carbamates includes a carbamylation (inhibition) step, in which the drug transfers its carbamate moiety to the active site of the enzyme and a decarbamylation (activity recovery) step, in which the carbamyl group is hydrolyzed from the enzyme. The carbamylation and decarbamylation kinetics decide the extent and the duration of the inhibition, thus the full characterization of candidate carbamate inhibitors requires the measurement of the kinetic constants describing both steps. Carbamylation and decarbamylation rate constants are traditionally measured by two separate set of experiments, thus making the full characterization of candidate inhibitors time-consuming.

Quantitative estimation of cholinesterase-specific drug metabolism of carbamate inhibitors provided by the analysis of the area under the inhibition-time curve.

Several molecules containing carbamate groups are metabolized by cholinesterases. This metabolism includes a time-dependent catalytic step which temporary inhibits the enzymes. In this paper we demonstrate that the analysis of the area under the inhibition versus time curve (AUIC) can be used to obtain a quantitative estimation of the amount of carbamate metabolized by the enzyme.

Design, synthesis and biological evaluation of bambuterol analogues as novel inhibitors of butyrylcholinesterase.

An increase activity of butyrylcholinesterase is believed to contribute to Alzheimer's disease. Bambuterol is a known potent inhibitor of butyrylcholinesterase, but it has undesired cardiac effects and less lipophilicity. Thirteen bambuterol analogues were synthesized using 1-(3, 5-dihydroxyphenyl) ethanone as a starting material.

Improved analysis of Monascus pigments based on their pH-sensitive UV-Vis absorption and reactivity properties.

Monascus pigments, a mixture of azaphilones mainly composed of red, orange and yellow pigments, are usually prepared in aqueous ethanol and analysed by ultraviolet-visible (UV-Vis) spectroscopy. The pH of aqueous ethanol used during sample preparation and analysis has never been considered a key parameter to control; however, this study shows that the UV-Vis spectra and colour characteristics of the six major pigments are strongly influenced by the pH of the solvent employed. In addition, the increase of solvent pH results in a remarkable increase of the amination reaction of orange pigments with amino compounds, and at higher pH (≥ 6.

Controlling composition and color characteristics of Monascus pigments by pH and nitrogen sources in submerged fermentation.

Submerged fermentations of Monascus anka were performed with different nitrogen sources at different pH in 3 L bioreactors. The results revealed that the Monascus pigments dominated by different color components (yellow pigments, orange pigments or red pigments) could be selectively produced through pH control and nitrogen source selection. A large amount of intracellular pigments dominated by orange pigments and a small amount of water-soluble extracellular yellow pigments were produced at low pH (pH 2.

Characterization of the species-dependent ketoprofen/albumin binding modes by induced CD spectroscopy and TD-DFT calculations.

The stereospecificity of high-affinity biorecognition phenomena at the basis of the activity of drugs is an important topic of active research in medicinal chemistry. The binding of drugs to their targets or to carrier proteins may lead to the onset of an induced circular dichroism (ICD) signal, which can be detected experimentally. Quantum mechanical (QM) calculations based on density functional theory (DFT) and its time-dependent formulation (TD-DFT) can be used to determine the theoretical chiroptical response of all the possible conformations of drugs bound to their hosts; by comparison with the experimental ICD spectra of drug-host complexes, this approach can lead to the identification of possible binding modes in the absence of X-ray crystallography or NMR data.

Stereoselective inhibition of human butyrylcholinesterase by the enantiomers of bambuterol and their intermediates.

This work describes the sequential hydrolysis of bambuterol enantiomers and their monocarbamate metabolites (MONO) catalyzed by human butyrylcholinesterase (BChE) as well as the enzyme inhibition resulting from this process. Particular emphasis is given to the contribution given by MONO to the enzyme inhibition because it was not fully characterized in previous works. Bambuterol and MONO enantiomers displayed the same time- and concentration-dependent mechanism of interaction with the enzyme.

Simultaneous determination of bambuterol and its two major metabolites in human plasma by hydrophilic interaction ultra-performance liquid chromatography-tandem mass spectrometry.

In this study, a rapid and sensitive hydrophilic interaction ultra-performance liquid chromatography-tandem mass spectrometry (HILIC-UPLC-MS/MS) method was developed for simultaneous determination of bambuterol and its two major metabolites monocarbamate bambuterol and terbutaline in human plasma. All samples were simply precipitated using acetonitrile and separated on a UPLC-HILIC column under gradient elution with a mobile phase consisting of acetonitrile and water with the addition of 10mm ammonium acetate and 0.1% formic acid at 0.

Species-dependent binding of tocainide analogues to albumin: affinity chromatography and circular dichroism study.

A series of novel tocainide analogues were characterized for their HSA and RSA binding, by using high-performance liquid affinity chromatography (HPLAC) and circular dichroism (CD). In this HPLAC study, HSA and RSA were covalently immobilized to the silica matrix of HPLC columns, with a procedure that maintained unaltered the binding properties of the proteins. The tocainide analogues were ranked for their affinity to HSA and RSA on the basis of their bound fractions measured by the two albumin-based columns.

Inoculation and alkali coeffect in volatile fatty acids production and microbial community shift in the anaerobic fermentation of waste activated sludge.

Batch fermentations of waste activated sludge (WAS) at alkaline pH with different inocula were performed. Paper mill anaerobic granular sludge (PAS) and dyeing mill anaerobic sludge (DAS) were used as inocula. At pH 10 the inoculation did not increase the volatile fatty acids (VFAs) production compared to the non-inoculated samples fermented in the same conditions, and the maximal VFAs yield of non-inoculated WAS was higher than inoculated WAS.

Amyloid β-peptide 25-35 self-assembly and its inhibition: a model undecapeptide system to gain atomistic and secondary structure details of the Alzheimer's disease process and treatment.

Combined results of theoretical molecular dynamic simulations and in vitro spectroscopic (circular dichroism and fluorescence) studies are presented, providing the atomistic and secondary structure details of the process by which a selected small molecule may destabilize the β-sheet ordered "amyloid" oligomers formed by the model undecapeptide of amyloid β-peptide 25-35 [Aβ(25-35)]. Aβ(25-35) was chosen because it is the shortest fragment capable of forming large β-sheet fibrils and retaining the toxicity of the full length Aβ(1-40/42) peptides. The conformational transition, that leads to the formation of β-sheet fibrils from soluble unordered structures, was found to depend on the environmental conditions, whereas the presence of myricetin destabilizes the self-assembly and antagonizes this conformational shift.

5-Imino-1,2-4-thiadiazoles and quinazolines derivatives as glycogen synthase kinase 3β (GSK-3β) and phosphodiesterase 7 (PDE7) inhibitors: determination of blood-brain barrier penetration and binding to human serum albumin.

5-Imino-1,2,4-thiadiazoles and quinazolines derivatives as glycogen synthase kinase 3β (GSK-3β) and phosphodiesterase 7 (PDE7) inhibitors were characterized for their ability to pass the blood-brain barrier (BBB) together with their human serum albumin (HSA) binding using high-performance liquid affinity chromatography (HPLAC) and circular dichroism (CD). To study the blood-brain barrier penetration, a parallel artificial membrane permeability assay (PAMPA) using a porcine brain lipid was employed. For the HPLAC investigation, HSA was previously covalently immobilized to the silica matrix of the HPLC column.

Stereochemical characterization of fluorinated 2-(phenanthren-1-yl)propionic acids by enantioselective high performance liquid chromatography analysis and electronic circular dichroism detection.

Enantioselective high performance liquid chromatography (HPLC) coupled with a detection system based on the simultaneous measurement of UV absorption and electronic circular dichroism (ECD) allows a complete stereochemical characterization of chiral compounds, once the relationship between sign of the chiroptical properties and absolute configuration is determined. In the present communication, the development of enantioselective HPLC methods for the resolution of a series of fluorinated 2-phenanthrenylpropionic acids (1-6) is reported. Different chiral stationary phases (CSPs) were tested: Chiralcel OJ, Chiralcel OD, Chiralpak AD, (S,S)-Whelk-O 1, Chirobiotic T and α(1)-acid glycoprotein (AGP).

Reversible human serum albumin binding of lipocrine: a circular dichroism study.

Lipocrine has been selected as an effective candidate for in vivo investigation because of its multiple biological properties, namely inhibition of AChE and BChE activities, inhibition of AChE-induced Aβ aggregation, and ability to protect cells against reactive oxygen species. To evaluate the possibility for lipocrine to become a lead and to be developed as a multipotent drug for the treatment of Alzheimer's disease, ADMET (absorption, distribution, metabolism, excretion, and toxicity) parameters need to be determined. Among ADMET parameters, distribution plays a key role in determining the lead drugability, and the drug binding to plasma proteins greatly influences the drug distribution.

Camptothecin and thiocamptothecin: the role of sulfur in shifting the hydrolysis equilibrium towards the closed lactone form.

Adverse effects have limited the clinical potential of 20-(S)-camptothecin (CPT) and led to a growing interest in the development of CPT analogues that exhibit less severe drawbacks, while maintaining their therapeutic activity. Recently, a thiopyridone isostere of CPT, 20-(S)-thiocamptothecin (TCPT), was developed that resulted more potent than the parent compound in H460, HT29 and IGROV-1 cell lines. The improved activity of TCPT over CPT might be due to the greater stability of the lactone ring.