The Bitter Taste Receptor TAS2R14 as a Drug Target

G protein-coupled receptors (GPCRs) mediate most of our physiological responses to hormones, neurotransmitters and environmental stimulants. Besides human senses like vision and olfaction, taste perception is mostly mediated by GPCRs. Hence, the bitter taste receptor family TAS2R comprises 25 distinct receptors and plays a key role in food acceptance and drug compliance. The TAS2R14 subtype is the most broadly tuned bitter taste receptor, recognizing a range of chemically highly diverse agonists. Besides other tissues, it is expressed in human airway smooth muscle and may represent a novel drug target for airway diseases. Several natural products as well as marketed drugs including flufenamic acid have been identified to activate TAS2R14, but higher potency ligands are needed to investigate the ligand-controlled physiological function and to facilitate the targeted modulate for potential future clinical applications. A combination of structure-based molecular modeling with chemical synthesis and in vitro profiling recently resulted in new flufenamic acid agonists with improved TAS2R14 potency and provided a validated and refined structural model of ligand–TAS2R14 interactions, which can be applied for future drug design projects

Structure-based development of caged dopamine D2/D3 receptor antagonists

Dopamine is a neurotransmitter of great physiological relevance. Disorders in dopaminergic signal transduction are associated with psychiatric and neurological pathologies such as Parkinson's disease, schizophrenia and substance abuse. Therefore, a detailed understanding of dopaminergic neurotransmission may provide access to novel therapeutic strategies for the treatment of these diseases. Caged compounds with photoremovable groups represent molecular tools to investigate a biological target with high spatiotemporal resolution. Based on the crystal structure of the D-3 receptor in complex with eticlopride, we have developed caged D-2/D-3 receptor ligands by rational design. We initially found that eticlopride, a widely used D-2/D-3 receptor antagonist, was photolabile and therefore is not suitable for caging. Subtle structural modification of the pharmacophore led us to the photostable antagonist dechloroeticlopride, which was chemically transformed into caged ligands. Among those, the 2-nitrobenzyl derivative 4 (MG307) showed excellent photochemical stability, pharmacological behavior and decaging properties when interacting with dopamine receptor-expressing cells

FAUC 213, a highly selective dopamine D4 receptor full antagonist, exhibits atypical antipsychotic properties in behavioural and neurochemical models of schizophrenia

Rationale: 2-[4-(4-Chlorophenyl)piperazin-1-ylmethyl]pyrazolo[1,5-a]pyridine (FAUC 213) is a highly selective antagonist at the dopamine D4 receptor subtype. It was designed as a derivative of two partial antagonists and has been proven to be a complete antagonist in mitogenesis assay. Objectives: In the present study, FAUC 213 was examined for antipsychotic properties in animal models of behavioural neurobiology and neurochemistry. Methods: Different concentrations of FAUC 213 were screened for effects on spontaneous, as well as amphetamine-induced, locomotor activity and apomorphine-induced prepulse disruption. The liability of causing extrapyramidal side effects was investigated in models of catalepsy and by high-performance liquid chromatography (HPLC) detection of dopamine turnover in several brain regions. The application schedule was validated, and the bioavailability of the compound determined, by means of a HPLC-pharmacokinetic study. Results: A significant effect in both the reduction of amphetamine-induced locomotor hyperactivity and the restoration of apomorphine-disrupted prepulse inhibition was found at 30mg/kg. This dose proved not to be high enough to induce catalepsy or to increase dopamine turnover in the dorsal striatum, nucleus accumbens and medial prefrontal cortex. The selective D4 antagonist FAUC 213, therefore, is not believed to mediate the above-mentioned effects via D2 receptor antagonism, but a partial involvement of 5-HT2- and α1-receptors cannot be ruled out at present. Conclusions: We have gathered evidence that FAUC 213 exhibits atypical antipsychotic characteristic

Structure-based discovery of opioid analgesics with reduced side effects

Morphine is an alkaloid from the opium poppy used to treat pain. The potentially lethal side effects of morphine and related opioids—which include fatal respiratory depression—are thought to be mediated by μ-opioid-receptor (μOR) signalling through the β-arrestin pathway or by actions at other receptors. Conversely, G-protein μOR signalling is thought to confer analgesia. Here we computationally dock over 3 million molecules against the μOR structure and identify new scaffolds unrelated to known opioids. Structure-based optimization yields PZM21—a potent Gi activator with exceptional selectivity for μOR and minimal β-arrestin-2 recruitment. Unlike morphine, PZM21 is more efficacious for the affective component of analgesia versus the reflexive component and is devoid of both respiratory depression and morphine-like reinforcing activity in mice at equi-analgesic doses. PZM21 thus serves as both a probe to disentangle μOR signalling and a therapeutic lead that is devoid of many of the side effects of current opioids

Quinpramine Ameliorates Rat Experimental Autoimmune Neuritis and Redistributes MHC Class II Molecules

Activation of inflammatory cells is central to the pathogenesis of autoimmune demyelinating diseases of the peripheral nervous system. The novel chimeric compound quinpramine—generated from imipramine and quinacrine—redistributes cholesterol rich membrane domains to intracellular compartments. We studied the immunological and clinical effects of quinpramine in myelin homogenate induced Lewis rat experimental autoimmune neuritis (EAN), a model system for acute human inflammatory neuropathies, such as the Guillain-Barré syndrome. EAN animals develop paresis of all limbs due to autoimmune inflammation of peripheral nerves. Quinpramine treatment ameliorated clinical disease severity of EAN and infiltration of macrophages into peripheral nerves. It reduced expression of MHC class II molecules on antigen presenting cells and antigen specific T cell proliferation both in vitro and in vivo. Quinpramine exerted its anti-proliferatory effect on antigen presenting cells, but not on responder T cells. Our data suggest that quinpramine represents a candidate pharmaceutical for inflammatory neuropathies

Genomic attributes of airway commensal bacteria and mucosa

Microbial communities at the airway mucosal barrier are conserved and highly ordered, in likelihood reflecting co-evolution with human host factors. Freed of selection to digest nutrients, the airway microbiome underpins cognate management of mucosal immunity and pathogen resistance. We show here the initial results of systematic culture and whole-genome sequencing of the thoracic airway bacteria, identifying 52 novel species amongst 126 organisms that constitute 75% of commensals typically present in heathy individuals. Clinically relevant genes encode antimicrobial synthesis, adhesion and biofilm formation, immune modulation, iron utilisation, nitrous oxide (NO) metabolism and sphingolipid signalling. Using whole-genome content we identify dysbiotic features that may influence asthma and chronic obstructive pulmonary disease. We match isolate gene content to transcripts and metabolites expressed late in airway epithelial differentiation, identifying pathways to sustain host interactions with microbiota. Our results provide a systematic basis for decrypting interactions between commensals, pathogens, and mucosa in lung diseases of global significance

Self-association of 8-anilino-1-naphthalene-sulfonate Molecules: Spectroscopic Characterization and Application to the Investigation of Protein Folding

It was suggested long ago that the reason for the considerable increase of 8-anilino-1-naphthalene-sulfonate (8-ANS) fluorescence intensity upon the transition from aqueous to organic solvents is the dissociation of 8-ANS associates. To clarify this point the dependence of spectral properties of the dye on concentration and solvent composition was investigated by means of steady-state and time-resolved fluorescence spectroscopy. It was shown that the increase of 8-ANS concentration leads to the changes in the shape of absorption and fluorescence spectra of the dye, accompanied by the decrease in its fluorescence decay time values. Such changes were observed in aqueous and organic solvents for Mg2+- and NH+4-salts of 8-anilino-1-naphthalene-sulfonateic acid and reflect the existence of self-association of the dye molecules in both media. However, the decrease in fluorescence intensity induced by the self-association of the probe molecules is too small to explain weak fluorescence of 8-ANS in water. At the same time, it expounds the difference between the decay times of protein-embedded 8-ANS molecules upon interaction of this probe with native and molten globule proteins

Inhibition of human ether à go-go potassium channels by Ca(2+)/calmodulin

Intracellular Ca(2+) inhibits voltage-gated potassium channels of the ether à go-go (EAG) family. To identify the underlying molecular mechanism, we expressed the human version hEAG1 in Xenopus oocytes. The channels lost Ca(2+) sensitivity when measured in cell-free membrane patches. However, Ca(2+) sensitivity could be restored by application of recombinant calmodulin (CaM). In the presence of CaM, half inhibition of hEAG1 channels was obtained in 100 nM Ca(2+). Overlay assays using labelled CaM and glutathione S-transferase (GST) fusion fragments of hEAG1 demonstrated direct binding of CaM to a C-terminal domain (hEAG1 amino acids 673–770). Point mutations within this section revealed a novel CaM-binding domain putatively forming an amphipathic helix with both sides being important for binding. The binding of CaM to hEAG1 is, in contrast to Ca(2+)-activated potassium channels, Ca(2+) dependent, with an apparent K(D) of 480 nM. Co-expression experiments of wild-type and mutant channels revealed that the binding of one CaM molecule per channel complex is sufficient for channel inhibition

Use of Fluorescence Decay Times of 8-ans-protein Complexes to Study the Conformational Transitions in Proteins Which Unfold Through the Molten Globule State

The conformational transitions starting with the native protein, passing the molten globule state and finally approaching the unfolded state of proteins was investigated for bovine carbonic anhydrase B (BCAB) and human α-lactalbumin (α-HLA) by means of fluorescence decay time measurements of the dye 8-anilinonaphthalene-1-sulphonic acid (8-ANS). Stepwise denaturation was realized by using the denaturant guanidinium chloride (GdmCl). It was shown that 8-ANS bound with protein yields a double-exponential fluorescence decay, where both decay times considerably exceed the decay time of free 8-ANS in water. This finding reflects the hydrophobic environment of the dye molecules attached to the proteins. The fluorescence lifetime of the short-time component is affected by protein association and can be effectively quenched by acrylamide, indicating that 8-ANS molecules preferentially bind at the protein surface. The fluorescence lifetime of the long-time component is independent of the protein and acrylamide concentration and may be related to protein-embedded dye molecules. Changes of the long lifetime component upon GdmCl-induced denaturation and unfolding of BCAB and α-HLA correlate well with overall changes of the protein conformation. The transition from native protein to the molten globule state is accompanied by an increase of the number of protein-embedded 8-ANS molecules, while the number of dye molecules located at the protein surface decreases. For the transition from the molten globule to the unfolded state was the opposite behaviour observed

Alkoholabhängigkeit : ein Leitfaden zur Gruppentherapie

Dieser Leitfaden beinhaltet eine umfassende, übersichtliche und in sich abgeschlossene Therapieeinheit für die strukturierte Behandlung von Patienten mit Alkoholabhängigkeit. Er wurde für ambulante und stationäre Gruppentherapien entwickelt und evaluiert, bietet aber auch wertvolle Anregungen für die einzeltherapeutische Arbeit mit alkoholabhängigen Patienten. Neu ist die Integration aktueller Befunde aus der Neurobiologie sowie der sich daraus ableitenden Folgerungen für die psychotherapeutische Behandlung. Die Beschreibung der einzelnen Therapiemodule mit einfacher Verfügbarkeit der Arbeitsmaterialien als Download ermöglicht eine schnelle und praxisnahe Integration des Konzeptes in den therapeutischen Alltag