Easily Accessible Substituted Heterocyclic Hemithioindigos as Bistable Molecular Photoswitches.

Thioaurone chromophores, part of the indigoid family and commonly named hemithioindigos, have recently gained attention due to their interesting photoswitching properties. The study focuses on heterocyclic hemithioindigos (Het-HTIs) and investigates their synthesis using electron-rich and electron-poor heterocycles and modifications to the thioindigo moiety. Furthermore, it aims to evaluate the photoswitching performances of these synthesised compounds, with a particular emphasis on the influence of the heterocycles on the photoisomerization capabilities, which was found to be more prominent than the modifications made to the thioindigo moiety. Among the 44 Het-HTIs tested, several exhibited highly efficient photoswitchable properties, demonstrating Z-to-E photoisomerization in the blue region, and E-to-Z photoisomerization in the green or the red regions. Additionally, the metastable E-isomer displayed an impressive half-life of up to 54 days in a polar solvent (DMSO). These results suggest that heterocyclic hemithioindigos hold great promise as photoswitches for researchers interested in light-controlled molecular mechanisms.

Potent 3-Hydroxy-2-Pyridine Aldoxime Reactivators of Organophosphate-Inhibited Cholinesterases with Predicted Blood-Brain Barrier Penetration.

A new series of 3-hydroxy-2-pyridine aldoxime compounds have been designed, synthesised and tested in vitro, in silico, and ex vivo as reactivators of human acetylcholinesterase (hAChE) and butyrylcholinesterase (hBChE) inhibited by organophosphates (OPs), for example, VX, sarin, cyclosarin, tabun, and paraoxon. The reactivation rates of three oximes (16-18) were determined to be greater than that of 2-PAM and comparable to that of HI-6, two pyridinium aldoximes currently used by the armies of several countries. The interactions important for a productive orientation of the oxime group within the OP-inhibited enzyme have been clarified by molecular-modelling studies, and by the resolution of the crystal structure of the complex of oxime 17 with Torpedo californica AChE. Blood-brain barrier penetration was predicted for oximes 15-18 based on their physicochemical properties and an in vitro brain membrane permeation assay. Among the evaluated compounds, two morpholine-3-hydroxypyridine aldoxime conjugates proved to be promising reactivators of OP-inhibited cholinesterases. Moreover, efficient ex vivo reactivation of phosphylated native cholinesterases by selected oximes enabled significant hydrolysis of VX, sarin, paraoxon, and cyclosarin in whole human blood, which indicates that the oximes have scavenging potential.

Use of an Air-Stable Cu(I)-NHC Catalyst for the Synthesis of Peptidotriazoles.

We report the use of air-stable Cu(I)-NHC complex 4a as a catalyst for the efficient microwave-assisted synthesis of peptidotriazoles on solid phase. Compared with the usual conditions (CuI or CuSO4/NaAsc), catalyst 4a allowed the preparation of a series of peptidomimetic compounds containing a 1,2,3-triazole ring in their backbone without the oxidation of common side-chains. Overall, the peptidotriazoles were obtained in good yields (61-87%), in excellent purity (higher than 94%) and with low copper contamination.

Increasing Polarity in Tacrine and Huprine Derivatives: Potent Anticholinesterase Agents for the Treatment of Myasthenia Gravis.

Symptomatic treatment of myasthenia gravis is based on the use of peripherally-acting acetylcholinesterase (AChE) inhibitors that, in some cases, must be discontinued due to the occurrence of a number of side-effects. Thus, new AChE inhibitors are being developed and investigated for their potential use against this disease. Here, we have explored two alternative approaches to get access to peripherally-acting AChE inhibitors as new agents against myasthenia gravis, by structural modification of the brain permeable anti-Alzheimer AChE inhibitors tacrine, 6-chlorotacrine, and huprine Y. Both quaternization upon methylation of the quinoline nitrogen atom, and tethering of a triazole ring, with, in some cases, the additional incorporation of a polyphenol-like moiety, result in more polar compounds with higher inhibitory activity against human AChE (up to 190-fold) and butyrylcholinesterase (up to 40-fold) than pyridostigmine, the standard drug for symptomatic treatment of myasthenia gravis. The novel compounds are furthermore devoid of brain permeability, thereby emerging as interesting leads against myasthenia gravis.

Neisseria meningitidis serogroup W, Burkina Faso, 2012.

In 2010, Burkina Faso became the first country to introduce meningococcal serogroup A conjugate vaccine (PsA-TT). During 2012, Burkina Faso reported increases in Neisseria meningitidis serogroup W, raising questions about whether these cases were a natural increase in disease or resulted from serogroup replacement after PsA-TT introduction. We analyzed national surveillance data to describe the epidemiology of serogroup W and genotyped 61 serogroup W isolates. In 2012, a total of 5,807 meningitis cases were reported through enhanced surveillance, of which 2,353 (41%) were laboratory confirmed. The predominant organism identified was N. meningitidis serogroup W (62%), and all serogroup W isolates characterized belonged to clonal complex 11. Although additional years of data are needed before we can understand the epidemiology of serogroup W after PsA-TT introduction, these data suggest that serogroup W will remain a major cause of sporadic disease and has epidemic potential, underscoring the need to maintain high-quality case-based meningitis surveillance after PsA-TT introduction.

Reaction site-driven regioselective synthesis of AChE inhibitors.

The enzyme-directed synthesis is an emerging fragment-based lead discovery approach in which the biological target is able to assemble its own multidentate ligands from a pool of building blocks. Here, we report for the first time the use of the human acetylcholinesterase (AChE) as an enzyme for the design and synthesis of new potent heterodimeric huprine-based inhibitors. Both the specific click chemistry site within the protein and the regioselectivity of the Huisgen cycloaddition observed suggest promising alternatives in the design of efficient mono- and dimeric ligands of AChE. Finally, a detailed computational modelling of the click reaction was conducted to further understand the origin of this TGS selectivity.

Synthesis of fluorinated agonist of sphingosine-1-phosphate receptor 1.

The bioactive metabolite sphingosine-1-phosphate (S1P), a product of sphingosine kinases (SphKs), mediates diverse biological processes such as cell differentiation, proliferation, survival and angiogenesis. A fluorinated analogue of S1P receptor agonist has been synthesized by utilizing a ring opening reaction of oxacycles by a lithiated difluoromethylphosphonate anion as the key reaction. In vitro activity of this S1P analogue is also reported.

Crystal structures of human cholinesterases in complex with huprine W and tacrine: elements of specificity for anti-Alzheimer's drugs targeting acetyl- and butyryl-cholinesterase.

The multifunctional nature of Alzheimer's disease calls for MTDLs (multitarget-directed ligands) to act on different components of the pathology, like the cholinergic dysfunction and amyloid aggregation. Such MTDLs are usually on the basis of cholinesterase inhibitors (e.g. tacrine or huprine) coupled with another active molecule aimed at a different target. To aid in the design of these MTDLs, we report the crystal structures of hAChE (human acetylcholinesterase) in complex with FAS-2 (fasciculin 2) and a hydroxylated derivative of huprine (huprine W), and of hBChE (human butyrylcholinesterase) in complex with tacrine. Huprine W in hAChE and tacrine in hBChE reside in strikingly similar positions highlighting the conservation of key interactions, namely, π-π/cation-π interactions with Trp86 (Trp82), and hydrogen bonding with the main chain carbonyl of the catalytic histidine residue. Huprine W forms additional interactions with hAChE, which explains its superior affinity: the isoquinoline moiety is associated with a group of aromatic residues (Tyr337, Phe338 and Phe295 not present in hBChE) in addition to Trp86; the hydroxyl group is hydrogen bonded to both the catalytic serine residue and residues in the oxyanion hole; and the chlorine substituent is nested in a hydrophobic pocket interacting strongly with Trp439. There is no pocket in hBChE that is able to accommodate the chlorine substituent.

Cholesterol Oxime Olesoxime Assessed as a Potential Ligand of Human Cholinesterases.

Olesoxime, a cholesterol derivative with an oxime group, possesses the ability to cross the blood-brain barrier, and has demonstrated excellent safety and tolerability properties in clinical research. These characteristics indicate it may serve as a centrally active ligand of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE), whose disruption of activity with organophosphate compounds (OP) leads to uncontrolled excitation and potentially life-threatening symptoms. To evaluate olesoxime as a binding ligand and reactivator of human AChE and BChE, we conducted in vitro kinetic studies with the active metabolite of insecticide parathion, paraoxon, and the warfare nerve agents sarin, cyclosarin, tabun, and VX. Our results showed that both enzymes possessed a binding affinity for olesoxime in the mid-micromolar range, higher than the antidotes in use (i.e., 2-PAM, HI-6, etc.). While olesoxime showed a weak ability to reactivate AChE, cyclosarin-inhibited BChE was reactivated with an overall reactivation rate constant comparable to that of standard oxime HI-6. Moreover, in combination with the oxime 2-PAM, the reactivation maximum increased by 10-30% for cyclosarin- and sarin-inhibited BChE. Molecular modeling revealed productive interactions between olesoxime and BChE, highlighting olesoxime as a potentially BChE-targeted therapy. Moreover, it might be added to OP poisoning treatment to increase the efficacy of BChE reactivation, and its cholesterol scaffold could provide a basis for the development of novel oxime antidotes.

Reactivators of acetylcholinesterase inhibited by organophosphorus nerve agents.

Since the September 11, 2001, terrorist attacks in the United States, the specter of a chemical threat against civilian populations has renewed research interest in chemical warfare agents, their mechanisms of action, and treatments that reverse their effects. In this Account, we focus specifically on organophosphorus nerve agents (OPNAs). Although some OPNAs are used as pest control, the most toxic chemicals in this class are used as chemical warfare agents in armed conflicts. The acute toxicity of OPNAs results from the irreversible inhibition of acetylcholinesterase (AChE, EC 3.1.1.7) via the formation of a covalent P-O bond at the serine hydroxyl group in the enzyme active site. AChE breaks down the neurotransmitter acetylcholine at neuronal synapses and neuromuscular junctions. The irreversible inhibition of AChE causes the neurotransmitter to accumulate in the synaptic cleft, leading to overstimulation of cholinergic receptors, seizures, respiratory arrest, and death. The current treatment for OPNA poisoning combines an antimuscarinic drug (e.g., atropine), an anticonvulsant drug (e.g., diazepam), and an AChE reactivator of the pyridinium aldoxime family (pralidoxime, trimedoxime, obidoxime, HI-6, HLö-7). Because of their high nucleophilicity, oximes can displace the phosphyl group from the catalytic serine, thus restoring the enzyme's catalytic activity. During 50 years of research in the reactivator field, researchers have synthesized and tested numerous structural modifications of monopyridinium oximes and bispyridinium oximes. In the past decade, medicinal chemists have focused their research on the more efficient bispyridinium reactivators, but all known reactivators have several drawbacks. First, due to their permanent positive charge, they do not cross the blood-brain barrier (BBB) efficiently and do not readily reactivate AChE in the central nervous system. Second, no single oxime is efficient against a wide variety of OPNAs. Third, oximes cannot reactivate "aged" AChE. This Account summarizes recent strategies for the development of AChE reactivators capable of crossing the BBB. The use of nanoparticulate transport and inhibition of P-glycoprotein efflux pumps improves BBB transport of these AChE reactivators. Chemical modifications that increased the lipophilicity of the pyridinium aldoximes, the addition of a fluorine atom and the replacement of a pyridyl ring with a dihydropyridyl moiety, enhances BBB permeability. The glycosylation of pyridine aldoximes facilitates increased BBB penetration via the GLUT-1 transport system. The development of novel uncharged reactivators that can move efficiently across the BBB represents one of the most promising of these new strategies.

Influence of Different Sterilization Methods on the Surface Chemistry and Electrochemical Behavior of Biomedical Alloys.

Sterilization is a prerequisite for biomedical devices before contacting the human body. It guarantees the lack of infection by eliminating microorganisms (i.e., bacteria, spores and fungi). It constitutes the last fabrication process of a biomedical device. The aim of this paper is to understand the effect of different sterilization methods (ethanol-EtOH, autoclave-AC, autoclave + ultraviolet radiation-ACUV and gamma irradiation-G) on the surface chemistry and electrochemical reactivity (with special attention on the kinetics of the oxygen reduction reaction) of CoCrMo and titanium biomedical alloys used as prosthetic materials. To do that, electrochemical measurements (open circuit potential, polarization resistance, cathodic potentiodynamic polarization and electrochemical impedance spectroscopy) and surface analyses (Auger Electron Spectroscopy) of the sterilized surfaces were carried out. The obtained results show that the effect of sterilization on the corrosion behavior of biomedical alloys is material-dependent: for CoCrMo alloys, autoclave treatment increases the thickness and the chromium content of the passive film increasing its corrosion resistance compared to simple sterilization in EtOH, while in titanium and its alloys, autoclave and UV-light accelerates its corrosion rate by accelerating the kinetics of oxygen reduction.

Biological response and cell death signaling pathways modulated by tetrahydroisoquinoline-based aldoximes in human cells.

The uncharged 3-hydroxy-2-pyridine aldoximes with protonatable tertiary amines are studied as antidotes in toxic organophosphates (OP) poisoning. Due to some of their specific structural features, we hypothesize that these compounds could exert diverse biological activity beyond their main scope of application. To examine this further, we performed an extensive cell-based assessment to determine their effects on human cells (SH-SY5Y, HEK293, HepG2, HK-2, myoblasts and myotubes) and possible mechanism of action. As our results indicated, aldoxime having a piperidine moiety did not induce significant toxicity up to 300 µM within 24 h, while those with a tetrahydroisoquinoline moiety, in the same concentration range, showed time-dependent effects and stimulated mitochondria-mediated activation of the intrinsic apoptosis pathway through ERK1/2 and p38-MAPK signaling and subsequent activation of initiator caspase 9 and executive caspase 3 accompanied with DNA damage as observed already after 4 h exposure. Mitochondria and fatty acid metabolism were also likely targets of 3-hydroxy-2-pyridine aldoximes with tetrahydroisoquinoline moiety, due to increased phosphorylation of acetyl-CoA carboxylase. In silico analysis predicted kinases as their most probable target class, while pharmacophores modeling additionally predicted the inhibition of a cytochrome P450cam. Overall, if the absence of significant toxicity for piperidine bearing aldoxime highlights the potential of its further studies in medical counter-measures, the observed biological activity of aldoximes with tetrahydroisoquinoline moiety could be indicative for future design of compounds either in a negative context in OP antidotes design, or in a positive one for design of compounds for the treatment of other phenomena like cell proliferating malignancies.

A New Class of Bi- and Trifunctional Sugar Oximes as Antidotes against Organophosphorus Poisoning.

Recent events demonstrated that organophosphorus nerve agents are a serious threat for civilian and military populations. The current therapy includes a pyridinium aldoxime reactivator to restore the enzymatic activity of acetylcholinesterase located in the central nervous system and neuro-muscular junctions. One major drawback of these charged acetylcholinesterase reactivators is their poor ability to cross the blood-brain barrier. In this study, we propose to evaluate glucoconjugated oximes devoid of permanent charge as potential central nervous system reactivators. We determined their in vitro reactivation efficacy on inhibited human acetylcholinesterase, the crystal structure of two compounds in complex with the enzyme, their protective index on intoxicated mice, and their pharmacokinetics. We then evaluated their endothelial permeability coefficients with a human in vitro model. This study shed light on the structural restrains of new sugar oximes designed to reach the central nervous system through the glucose transporter located at the blood-brain barrier.

Synthesis and luminescence properties of new red-shifted absorption lanthanide(III) chelates suitable for peptide and protein labelling.

The synthesis and photo-physical properties of an original bis-pyridinylpyrazine chromophore efficiently sensitising europium(III) and samarium(III) are described. The corresponding lanthanide(III) complexes display in aqueous solutions a maximum excitation wavelength which is significantly red-shifted compared to the usual terpyridine-based chelates, and a valuable luminescence brightness above 2,000 dm(3) mol(-1) cm(-1) at 345 nm was obtained with a europium(III) derivative. Further functionalisation with three different bioconjugatable handles was also investigated and their ability to efficiently label a model hexapeptide was evaluated and compared. Finally, the best bioconjugatable europium(III) chelate was used in representative labelling experiments involving monoclonal antibodies and the luminescence features of the corresponding bioconjugates remained satisfactory.

Discovery of a Potent Dual Inhibitor of Acetylcholinesterase and Butyrylcholinesterase with Antioxidant Activity that Alleviates Alzheimer-like Pathology in Old APP/PS1 Mice.

The combination of the scaffolds of the cholinesterase inhibitor huprine Y and the antioxidant capsaicin results in compounds with nanomolar potencies toward human acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) that retain or improve the antioxidant properties of capsaicin. Crystal structures of their complexes with AChE and BChE revealed the molecular basis for their high potency. Brain penetration was confirmed by biodistribution studies in C57BL6 mice, with one compound (5i) displaying better brain/plasma ratio than donepezil. Chronic treatment of 10 month-old APP/PS1 mice with 5i (2 mg/kg, i.p., 3 times per week, 4 weeks) rescued learning and memory impairments, as measured by three different behavioral tests, delayed the Alzheimer-like pathology progression, as suggested by a significantly reduced Aß42/Aß40 ratio in the hippocampus, improved basal synaptic efficacy, and significantly reduced hippocampal oxidative stress and neuroinflammation. Compound 5i emerges as an interesting anti-Alzheimer lead with beneficial effects on cognitive symptoms and on some underlying disease mechanisms.

Functional characterization of multifunctional ligands targeting acetylcholinesterase and alpha 7 nicotinic acetylcholine receptor.

Alzheimer's disease (AD) is a neurodegenerative disorder associated with cholinergic dysfunction, provoking memory loss and cognitive dysfunction in elderly patients. The cholinergic hypothesis provided over the years with molecular targets for developing palliative treatments for AD, acting on the cholinergic system, namely, acetylcholinesterase and α7 nicotinic acetylcholine receptor (α7 nAChR). In our synthetic work, we used "click-chemistry" to synthesize two Multi Target Directed Ligands (MTDLs) MB105 and MB118 carrying tacrine and quinuclidine scaffolds which are known for their anticholinesterase and α7 nAChR agonist activities, respectively. Both, MB105 and MB118, inhibit human acetylcholinesterase and human butyrylcholinesterase in the nanomolar range. Electrophysiological recordings on Xenopus laevis oocytes expressing human α7 nAChR showed that MB105 and MB118 acted as partial agonists of the referred nicotinic receptor, albeit, with different potencies despite their similar structure. The different substitution at C-3 on the 2,3-disubstituted quinuclidine scaffold may account for the significantly lower potency of MB118 compared to MB105. Electrophysiological recordings also showed that the tacrine precursor MB320 behaved as a competitive antagonist of human α7 nAChR, in the micromolar range, while the quinuclidine synthetic precursor MB099 acted as a partial agonist. Taken all together, MB105 behaved as a partial agonist of α7 nAChR at concentrations where it completely inhibited human acetylcholinesterase activity paving the way for the design of novel MTDLs for palliative treatment of AD.

Pharmacokinetic Evaluation of Brain Penetrating Morpholine-3-hydroxy-2-pyridine Oxime as an Antidote for Nerve Agent Poisoning.

Nerve agents, the deadliest chemical warfare agents, are potent inhibitors of acetylcholinesterase (AChE) and cause rapid cholinergic crisis with serious symptoms of poisoning. Oxime reactivators of AChE are used in medical practice in the treatment of nerve agent poisoning, but the search for novel improved reactivators with central activity is an ongoing pursuit. For numerous oximes synthesized, in vitro reactivation is a standard approach in biological evaluation with little attention given to the pharmacokinetic properties of the compounds. This study reports a comprehensive physicochemical, pharmacokinetic, and safety profiling of five lipophilic 3-hydroxy-2-pyridine aldoximes, which were recently shown to be potent AChE reactivators with a potential to be centrally active. The oxime JR595 was singled out as highly metabolically stable in human liver microsomes, noncytotoxic oxime for SH-SY5Y neuroblastoma and 1321N1 astrocytoma cell lines, and its pharmacokinetic profile was determined after intramuscular administration in mice. JR595 was rapidly absorbed into blood after 15 min with simultaneous distribution to the brain at up to about 40% of its blood concentration; however, it was eliminated from both the brain and blood within an hour. In addition, the MDCKII-MDR1 cell line assay showed that oxime JR595 was not a P-glycoprotein efflux pump substrate. Finally, the preliminary antidotal study against multiple LD50 doses of VX and sarin in mice showed the potential of JR595 to provide desirable therapeutic outcomes with future improvements in its circulation time.

Preparation, modification, and evaluation of cruentaren A and analogues.

An expeditious total synthesis of the highly cytotoxic F-ATPase inhibitor cruentaren A (1) is described based on a ring-closing alkyne metathesis (RCAM) reaction for the formation of the macrocylic ring. Other key transformations comprise a C-acylation of the benzyl lithium reagent derived from orsellinic acid ester 9 with Weinreb amide 7, a CBS reduction of the resulting ketone 10, and a Soderquist propargylation of aldehyde 21 with allenylborane (S)-27 to set the C-15 chiral center of the required alcohol fragment 25. The RCAM precursor 33 was assembled by acylation of 25 with acid fluoride 32, since more conventional methods for ester bond formation were unproductive. Moreover, the choice of the protecting groups, in particular for the secondary alcohol at C-9, which is prone to engage in translactonization, turned out to be critical; a relatively stable TBDPS ether had to be chosen for this site, which was removed in the final step of the synthesis with aqueous HF since other fluoride sources met with failure. The successful synthetic route was then expanded beyond the natural product, bringing a series of analogues into reach that feature incremental but deep-seated structural modifications. Three of these fully synthetic compounds turned out to be as or even more cytotoxic than cruentaren A itself against L-929 mouse fibroblast cells, reaching IC(50) values as low as 0.7 ng mL(-1).

Self-cleavable chemiluminescent probes suitable for protease sensing.

A new generation of dioxetane-based chemiluminescent substrates suitable for detecting protease activities is described. Our strategy involves the use of a self-cleavable spacer as the key molecular component of these protease-sensitive chemiluminescent probes. Among the assayed strategies, the PABA (para-aminobenzylic alcohol) linker associated with an ether linkage enables the release of the light-emitting phenolic 1,2-dioxetane moiety through an enzyme-initiated domino reaction. To validate this strategy, two proteolytic enzymes were chosen: penicillin amidase and caspase-3, and the corresponding self-cleavable chemiluminescent substrates were synthesised. Their evaluation using an in vitro assay has enabled us to prove the decomposition of the linker under physiological conditions and the selectivity for the targeted enzyme.

Synthesis and structure-activity relationship of Huprine derivatives as human acetylcholinesterase inhibitors.

New series of Huprine (12-amino-6,7,10,11-tetrahydro-7,11-methanocycloocta[b]quinolines) derivatives have been synthesized and their inhibiting activities toward recombinant human acetylcholinesterase (rh-AChE) are reported. We have synthesized two series of Huprine analogues; in the first one, the benzene ring of the quinoline moiety has been replaced by different heterocycles or electron-withdrawing or electron-donating substituted phenyl group. The second one has been designed in order to evaluate the influence of modification at position 12 where different short linkers have been introduced on the Huprine X, Y skeletons. All these molecules have been prepared from ethyl- or methyl-bicyclo[3.3.1]non-6-en-3-one via Friedländer reaction involving selected o-aminocyano aromatic compounds. The synthesis of two heterodimers based on these Huprines has been also reported. Activities from moderate to same range than the most active Huprines X and Y taken as references have been obtained, the most potent analogue being about three times less active than parent Huprines X and Y. Topologic data have been inferred from molecular dockings and variations of activity between the different linkers suggest future structural modifications for activity improvement.