1,721,149 research outputs found
Design, synthesis and pharmacological properties of alkylbisammonio bitopic ligands of muscarinic acetylcholine receptors
No full textOver the past two decades, novel opportunities for drug discovery have risen from a greater understanding of the complexity of GPCR signaling. A striking example of this is the appreciation that GPCRs possess functional allosteric binding sites.[1] The five muscarinic acetylcholine receptor (mAChR) subtypes bind their physiological transmitter in the highly conserved orthosteric site within the transmembrane domain of the receptors. Orthosteric muscarinic agonists have negligible binding selectivity and poor signaling specificity. A less conserved allosteric site has been also characterized at the extracellular entrance of the binding pocket of mAChRs. Our interest in the research field on mAChR ligands has been recently extended to the study of derivatives designed to simultaneously interact with the orthosteric and the allosteric site (bitopic ligands) of these receptors, thus exploiting the complementary characteristics of the two different binding sites by a single ligand molecule.[2,3] This approach is a variation of the more traditional bivalent ligand concept and shares some of the same challenges, including the choice and role of the linker between the two pharmacophores and the validation of mechanism of action.[4] The most interesting bitopic compounds which emerged from our investigation (represented by the general molecular skeleton A in Figure 1) are alkylbisammonio derivatives incorporating a) iperoxo, an oxotremorine-related unselective muscarinic superagonist, b) a polymethylene spacer chain, and c) a heteroaromatic fragment targeting the allosteric site.
The synthetic approach of target compounds and the most relevant results of their pharmacological investigation will be presented. In particular, the analgesic effects observed for a group of the compounds under study will be illustrated in detail.
References
1. Wang, L.; Martin, B.; Brenneman, R.; Luttrell, L. M.; Maudsley, S. Allosteric Modulators of G Protein-Coupled Receptors: Future Therapeutics for Complex Physiological Disorders. J. Pharmacol. Exp. Ther. 2009, 331, 340-348
2. Antony, J.; Kellershohn, K.; Disingrini, T.; Dallanoce, C.; Tränkle, C.; Christopoulos, A.; Barocelli, E.; De Amici, M.; Holzgrabe, U.; Mohr, K. et al. Dualsteric GPCR targeting: a novel route to binding and signaling pathway selectivity. FASEB J. 2009, 23, 442-450
3. Bock, A.; Merten, N.; Schrage, R.; Dallanoce, C.; Matera, C.; Hoffmann, C.; De Amici, M.; Holzgrabe, U.; Kostenis, E.; Mohr, K. et al. The allosteric vestibule of a seven transmembrane helical receptor controls G-protein coupling. Nat. Commun. 2012, 3, 1044
4. Valant, C.; Lane, J. R.; Sexton, P. M.; Christopoulos, A. The Best of Both Worlds? Bitopic Orthosteric/Allosteric Ligands of G Protein-Coupled Receptors. Annu. Rev. Pharmacol. Toxicol. 2012, 52, 153-17
Photoswitchable Molecular Tools: Applications to Enzymes, GPCRs and Ion Channels
No full textPhotocontrolled molecular tools provide powerful means to manipulate and interrogate biological functions with high spatiotemporal precision and low invasiveness. Our research efforts in the field have focused on the design of reversible photoswitchable compounds to photocontrol enzymes, GPCRs, and ion channels. Among others, we have developed phototrexate, the first photoswitchable inhibitor of the human dihydrofolate reductase with demonstrated cytotoxicity in vitro and in zebrafish larvae,1 and PAI, a light-controlled dualsteric agonist of muscarinic M2 receptors that enabled photomodulation of cardiac function in tadpoles and brain states in mice.2 More recently, we have designed a fast photoswitchable tethered ligand of ionotropic glutamate receptors to enable activation of the auditory neurons with light. This compound, named TCPfast, induced photocurrents in untransfected neurons upon covalently tethering to endogenous glutamate receptors and activating them reversibly with blue light. We applied it to the ultrafast synapses of cochlear auditory neurons that encode sound and provide auditory input to the brain. TCPfast functions as a molecular prosthesis that bypasses the neurotransmitter-encoded signal with a photonic signal. Photosensitization of cochlear spiral ganglion neurons (SGNs) by locally administered TCPfast enabled temporally precise light-evoked firing up to a rate of approximately 1 kHz, matching the fastest optogenetic SGN stimulation.3 Hence, TCPfast shows that photopharmacology might serve as an interesting alternative for the development of optical cochlear implants for hearing restoration. The main results of all these studies will be presented and discussed.
References
1. Matera C et al. Journal of the American Chemical Society 2018, 140 (46), 15764–15773.
2. Riefolo F, Matera C et al. Journal of the American Chemical Society 2019, 141 (18), 7628–7636.
3. Garrido-Charles A, Huet A, Matera C et al., Journal of the American Chemical Society 2022, 144 (21), 9229–9239
Design, synthesis and pharmacological investigation of novel bifunctional ligands targeting nicotine and dopamine receptor subtypes
No full textBackground - An increasing number of receptor interactions depends on their physical association, and receptor hetero(di)mers usually show pharmacological and functional properties different from those of their constituent partners, thus behaving as new receptor entities.1 In particular, striatal dopaminergic transmission appears to be under the control of receptor heteromers containing D2 autoreceptors and non-alpha7 nicotinic acetylcholine receptors (nAChRs).2 This evidence stimulated our interest in investigating the biochemical and functional interactions between dopaminergic autoreceptors and nAChRs containing the beta2 subunit.
Aims - The deliberate, rational design of bifunctional (multiple) ligands has gained a relevant interest in the medicinal chemistry research. These compounds may serve as suitable molecular probes to study in depth receptor architectures and, in some instances, represent innovative approaches for therapeutic intervention.3,4 The results of the present research project are directed to explore new potential pharmacological approaches to the therapy of nicotinic addiction on one hand and of Parkinson’s disease on the other.
Methods - A common strategy was applied to the synthesis of two classes of designed bifunctional derivatives, that is we linked the chosen monovalent fragments with spacers of different length. Since the spacer should in principle not interfere with the ligand-receptor interactions involving the monomeric moieties, polymethylene chains were used to connect the pharmacophoric component parts. We synthesized bivalent derivatives formed by a) a D2/D3 agonist moiety and a nicotinic alpha4beta2 antagonist fragment and b) a D2/D3 antagonist moiety and a nicotinic alpha4beta2 agonist fragment. The structures of the parent ligands were selected after a critical inspection of the literature.
Results - We initially prepared bivalent derivatives characterized by a D2/D3 agonist moiety and a nicotinic alpha4beta2 antagonist fragment. The structural features of the chosen selective alpha4beta2 antagonists were those of N-n-alkylnicotinium analogues (NONI and NDNI), whereas we chose 2-(aminomethyl)chromane (2-AMC) was the molecular portion endowed with the required D2/D3 receptor agonist properties. Bitopic compounds with the above cited molecular features should ideally inhibit dopamine release, a condition that could be exploited in therapeutic protocols for nicotine addiction.
On the other hand, the synthesis of bivalent derivatives incorporating a D2/D3 antagonist moiety and a nicotinic alpha4beta2 agonist fragment was achieved using the D2/D3 antagonist Raclopride and the selective highly potent alpha4beta2 agonist A-84543 as building blocks. Bitopic compounds with such a molecular skeleton, able to recognize presynaptic heterodimeric receptor populations, should functionally enhance dopamine release, thus representing potential drug candidates for the treatment as Parkinson’s disease.
The synthetic routes to target compounds will be presented and commented.
Both classes of hybrid derivatives and their individual precursor ligands were assayed a) for their affinity and specificity at the nAChRs in striatal tissues and at the D2 receptor subtypes transfected into HEK cells, and b) for their functional activity, that is the capacity to favor or inhibit the release of [3H]dopamine (DA) from striatal slices stimulated by nicotinic agonists and/or the depolarizing agent potassium chloride. As an example, in the first group of derivatives, one of the bivalent compounds showed an interesting functional profile, since it totally inhibited the release of [3H]DA, being more potent then its parent nicotinic alpha4beta2antagonist.
Conclusion - The PhD experimental activity was devoted to the design, synthesis, and structure-activity studies of two distinct groups of bifunctional ligands targeting specific nicotinic and dopaminergic receptor subtypes. The preliminary results obtained for some of the studied derivatives encourage a further deepening of their functional profile (i.e., with electrophysiological experiments) coupled with the proof of concept of a truly bivalent mode of action.
References
1. Fiorentini, C.; Busi, C.; Gorruso, E.; Gotti, C.; Spano, P.; Missale, C. Reciprocal regulation of dopamine D1 and D3 receptor function and trafficking by heterodimerization. Mol. Pharmacol. 2008, 74, 59-69.
2. Quarta, D.; Ciruela, F.; Patkar, K.; Borycz, J.; Solinas, M.; Lluis, C.; Franco, R.; Wise, R. A.; Goldberg, S. R.; Hope, B. T.; Woods, A.; Ferré, S. Heteromeric nicotinic acetylcholine-dopamine autoreceptor complexes modulate striatal dopamine release. Neuropsychopharmacol. 2007, 32, 35-42.
3. Morphy, R.; Rankovic, Z. Designed multiple ligands. An emerging drug discovery paradigm. J. Med. Chem. 2005, 48, 6523-6543.
4. Rozenfeld, R.; Devi, L. A. Receptor heteromerization and drug discovery. Trends Pharmacol. Sci. 2010, 31, 124-130
Novel epibatidine-related analogues and epiboxidine enantiomers: synthesis and binding affinity at neuronal nicotinic acetylcholine receptor subtypes
No full textNeuronal nicotinic acetylcholine receptors (nAChRs) belong to a heterogeneous family of ligand-operated ion channels, differently expressed in the central and peripheral nervous systems, which are assembled in a
pentameric architecture to form a pore permeable to cations such as Na+, K+, or Ca++. Of the seventeen different
nAChR subunits which have been cloned, twelve (a2 to a10 and b2 to b4) were found in the neuronal receptors of vertebrates. Different subunit combinations characterize the nAChRs expressed within the diverse areas of the central nervous system (CNS), and affect their biophysical and functional properties, such as ion selectivity,
conductance, mean open-channel time, rate of desensitization as well as the sensitivity to neurotoxins.
Ongoing investigations on the pharmacology and neurobiology of CNS nAChRs allowed a better understanding of their involvement in various neuropsychiatric pathologies, such as Alzheimer’s and Parkinson’s diseases, mild cognitive impairment (MCI), Tourette’s syndrome, schizophrenia, depression,
anxiety, attention-deficit hyperactivity disorder (ADHD), and nicotine addiction. More than 90% of the heteromeric channels localized in the CNS contain the a4 and b2 subunits, whereas the most abundant
homomeric channel is the a7 pentamer. Thus, research efforts have mainly focused on the discovery of selective a4b2 or a7 ligands to ascertain the physiological and pathophysiological relevance of these receptor subtypes.
A significant impulse to the development of selective a4b2 nAChR ligands was given by the discovery of Epibatidine (−)-1, a natural toxin possessing an analgesic potency roughly one hundred times higher than that of morphine. Among the numerous Epibatidine-related analogues investigated so far, (±)-Epiboxidine 2, characterized by a 3-methylisoxazole ring, emerged as a potent a4b2 nicotinic receptor agonist, 10-fold less potent than Epibatidine as antinociceptive agent but 20-fold less toxic. Since we aimed at deepening the
pharmacological profile of Epiboxidine, we developed a novel synthetic sequence for the racemate and extended this approach to the preparation of the enantiomers (1R,2S,4S)-2 and (1S,2R,4R)-2 in enantiomeric
excess higher than 99%. The same approach allowed to prepare a group of Epibatidine-related unsaturated analogues represented by the general formula A (R=H, Me).
The synthesis of target derivatives took advantage of a palladium-catalyzed Stille reaction involving racemic or enantiopure enoltriflates and the suitable tributylisoxazolylstannane. The synthetic details, the affinity profile
of (+)-2 and (−)-2 and that of related analogues at a4b2 and a7 nAChRs will be outlined
In silico design of agonists targeting the alpha7 nicotinic acetylcholine receptors, their synthesis and preliminary pharmacological evaluation
No full textAs an extension of previous molecular modeling studies on nicotinic acetylcholine receptor (nAChR) ligands,[1] we now adopted the LIR (Linear Interaction Response) approach to reach a realistic compromise between accuracy and calculation rate, during the delta(G)bind value estimation of a training set of known agonists for the alpha7 nAChR subtype.
The same strategy was applied to a text set of potential alpha7 nAChR agonists, which allowed identification of general structures A and B as the most promising ligands in the series. Thus, some of these derivatives were synthesized along with procedures previously utilized by us,[2] and were then assayed for binding affinity at alpha7 and alpha4beta2 nAChR subtypes.
[1] Grazioso, G.; Cavalli, A.; De Amici, M.; Recanatini, M.; De Micheli, C. J. Comput. Chem. 2008, 29, 2593-2602. [2] Rizzi, L.; Dallanoce, C.; Matera, C.; Magrone, P.; Pucci, L.; Gotti, C.; Clementi, F.; De Amici, M. Bioorg. Med. Chem. Lett. 2008, 18, 4651-4654
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