N-benzylpiperidine derivatives as α7 nicotinic receptor antagonists

This document is the accepted manuscript version of a Published Work that appeared in final form in ACS Chemical Neuroscience 7.8, copyright © American Chemical Society after peer review and technical editing by the publisher. To access the final edited and published work see DOI: 10.1021/acschemneuro.6b00122.A series of multitarget directed propargylamines, as well as other differently susbstituted piperidines have been screened as potential modulators of neuronal nicotinic acetylcholine receptors (nAChRs). Most of them showed antagonist actions on α7 nAChRs. Especially, compounds 13, 26, and 38 displayed submicromolar IC50 values on homomeric α7 nAChRs, whereas they were less effective on heteromeric α3β4 and α4β2 nAChRs (up to 20-fold higher IC50 values in the case of 13). Antagonism was concentration dependent and noncompetitive, suggesting that these compounds behave as negative allosteric modulators of nAChRs. Upon the study of a series of less complex derivatives, the N-benzylpiperidine motif, common to these compounds, was found to be the main pharmacophoric group. Thus, 2-(1-benzylpiperidin-4-yl)-ethylamine (48) showed an inhibitory potency comparable to the one of the previous compounds and also a clear preference for α7 nAChRs. In a neuroblastoma cell line, representative compounds 13 and 48 also inhibited, in a concentration-dependent manner, cytosolic Ca2+ signals mediated by nAChRs. Finally, compounds 38 and 13 inhibited 5-HT3A serotonin receptors whereas they had no effect on α1 glycine receptors. Given the multifactorial nature of many pathologies in which nAChRs are involved, these piperidine antagonists could have a therapeutic potential in cases where cholinergic activity has to be negatively modulated.This work was supported by grants SAF2011-22802 to S.S., SAF2012-33304 to J.M.-C., CSD2008-00005 (the Spanish Ion Channel Initiative-CONSOLIDER INGENIO 2010) to M.C. from the Spanish Ministry of Science and Innovation (Ministerio de Economía y Competitividad)

Preclinical Characterization of Antioxidant Quinolyl Nitrone QN23 as a New Candidate for the Treatment of Ischemic Stroke

Nitrones are encouraging drug candidates for the treatment of oxidative stress-driven diseases such as acute ischemic stroke (AIS). In a previous study, we found a promising quinolylnitrone, QN23, which exerted a neuroprotective effect in neuronal cell cultures subjected to oxygen–glucose deprivation and in experimental models of cerebral ischemia. In this paper, we update the biological and pharmacological characterization of QN23. We describe the suitability of intravenous administration of QN23 to induce neuroprotection in transitory four-vessel occlusion (4VO) and middle cerebral artery occlusion (tMCAO) experimental models of brain ischemia by assessing neuronal death, apoptosis induction, and infarct area, as well as neurofunctional outcomes. QN23 significantly decreased the neuronal death and apoptosis induced by the ischemic episode in a dose-dependent manner and showed a therapeutic effect when administered up to 3 h after post-ischemic reperfusion onset, effects that remained 11 weeks after the ischemic episode. In addition, QN23 significantly reduced infarct volume, thus recovering the motor function in a tMCAO model. Remarkably, we assessed the antioxidant activity of QN23 in vivo using dihydroethidium as a molecular probe for radical species. Finally, we describe QN23 pharmacokinetic parameters. All these results pointing to QN23 as an interesting and promising preclinical candidate for the treatment of AIS.This work was supported by the Instituto de Salud Carlos III and co-financed by the European Development Regional Fund (FEDER) through grants PI18/00255, RD16/0019/0006, and RD21/0006/0019 to J.M. and A.A., and RD16/0019/0008 and RD21/0006/0014 to J.B.S.; the MINECO grant SAF2015-65586-R to J.M.-C.; and the Comunidad de Madrid Neurocentro project B2017/BMD-3760 to D.G.-N

Acetylcholinesterase Inhibition of Diversely Functionalized Quinolinones for Alzheimer's Disease Therapy

In this communication, wereport the synthesis and cholinesterase (ChE)/monoamine oxidase (MAO) inhibition of 19 quinolinones (QN1-19) and 13 dihydroquinolinones (DQN1-13) designed as potential multitarget small molecules (MSM) for Alzheimer¿s disease therapy. Contrary to our expectations, none of them showed significant human recombinant MAO inhibition, but compounds QN8, QN9, and DQN7 displayed promising human recombinant acetylcholinesterase (hrAChE) and butyrylcholinesterase (hrBuChE) inhibition. In particular, molecule QN8 was found to be a potent and quite selective non-competitive inhibitor of hrAChE (IC50 = 0.29 M), with Ki value in nanomolar range (79 nM). Pertinent docking analysis confirmed this result, suggesting that this ligand is an interesting hit for further investigation.R.A., M.S., P.B., and K.M. were supported by European Regional Development Fund/European Social Fund (ERDF/ESF, project PharmaBrain, no. CZ.02.1.01/0.0/0.0/16_025/0007444), University of Hradec Kralove (no. SV2113-2019, VT2019-2021), and EU COST action CA15135 MuTaLig. J.M.C. thanks Ministerio de Economía (MINECO, SAF2015-65586-R) and Universidad Camilo José Cela (UCJC, grants UCJC 2020-03, and UCJC 2020-33) for support

In vitro and in silico ADME-Tox profiling and safety significance of multifunctional monoamine oxidase inhibitors targeting neurodegenerative deseases

Herein we report in vitro metabolic stability in human liver microsomes (HLMs), interactions with cytochrome P450 isoenzymes (CYP3A4, CYP2D6, and CYP2C9), and cytotoxicity analyses on HEK-293, HepG2, Huh7, and WTIIB cell lines of our most recent multitarget directed ligands PF9601N, ASS234, and contilisant. Based on these results, we conclude that (1) PF9601N and contilisant are metabolically stable in the HLM assay, in contrast to the very unstable ASS234; (2) CYP3A4 activity was decreased by PF9601N at all the tested concentrations and by ASS234 and contilisant only at the highest concentration; CYP2D6 activity was reduced by ASS234 at 1, 10, and 25 μM and by PF9601N at 10 and 25 μM, whereas contilisant increased its activity at the same concentrations; CYP2C9 was inhibited by the three compounds; (3) contilisant did not affect cell viability in the widest range of concentrations: up to 10 μM on HEK-293 cells, up to 30 μM on Huh7 cells, up to 50 μM on HepG2 cells, and up to 30 or 100 μM on WTIIB cells. Based on these results, we selected contilisant as a metabolically stable and nontoxic lead compound for further studies in Alzheimer's disease therapy.This study received financial support from the National Science Centre Poland (Grant No. 2016/23/D/NZ7/01328). J.M.-C. thanks AEI (Government of Spain) for grants PDI- 2019-105813RB-C21 and SAF2015-65586-R. J.M.-C. and F.L.- M. thank UCJC (Grants UCJC 2020-33 UCJC 2020-03) for support

Synthesis and biological evaluation of benzochromenopyrimidinones as cholinesterase inhibitors and potent antioxidant, non-hepatotoxic agents for Alzheimer’s disease

We report herein the straightforward two-step synthesis and biological assessment of novel racemic benzochromenopyrimidinones as non-hepatotoxic, acetylcholinesterase inhibitors with antioxidative properties. Among them, compound 3Bb displayed a mixed-type inhibition of human acetylcholinesterase (IC50 = 1.28 ± 0.03 μM), good antioxidant activity, and also proved to be non-hepatotoxic on human HepG2 cell line.JMC thanks Government of Spain for support (SAF2016-65586-R), JJ and OS thank MH CZ- DRO (UHHK 00179906).We acknowledge support by the CSIC Open Access Publication Initiative through its Unit of Information Resources for Research (URICI)

Synthesis, antioxidant properties and neuroprotection of α-phenyl-tert-butylnitrone derived HomoBisNitrones in in vitro and in vivo ischemia models

We herein report the synthesis, antioxidant power and neuroprotective properties of nine homo-bis-nitrones HBNs1–9 as alpha-phenyl-N-tert-butylnitrone (PBN) analogues for stroke therapy. In vitro neuroprotection studies of HBNs1–9 against Oligomycin A/Rotenone and in an oxygen-glucose-deprivation model of ischemia in human neuroblastoma cell cultures, indicate that (1Z,1′Z)-1,1′-(1,3-phenylene)bis(N-benzylmethanimine oxide) (HBN6) is a potent neuroprotective agent that prevents the decrease in neuronal metabolic activity (EC = 1.24 ± 0.39 μM) as well as necrotic and apoptotic cell death. HBN6 shows strong hydroxyl radical scavenger power (81%), and capacity to decrease superoxide production in human neuroblastoma cell cultures (maximal activity = 95.8 ± 3.6%), values significantly superior to the neuroprotective and antioxidant properties of the parent PBN. The higher neuroprotective ability of HBN6 has been rationalized by means of Density Functional Theory calculations. Calculated physicochemical and ADME properties confirmed HBN6 as a hit-agent showing suitable drug-like properties. Finally, the contribution of HBN6 to brain damage prevention was confirmed in a permanent MCAO setting by assessing infarct volume outcome 48 h after stroke in drug administered experimental animals, which provides evidence of a significant reduction of the brain lesion size and strongly suggests that HBN6 is a potential neuroprotective agent against stroke.We would like to thank Soledad Martinez Montero for the excellent technical assistance. This work was supported by grants from the Spanish Ministry of Economy and Competitiveness (SAF2015-65586-R to JMC; CTQ2016- 78205-P and CTQ2016-81797-REDC to IF, and NEUROCENTRO-CM S2017/BMD3760 to RMM and DNG), and Camilo José Cela University (UCJC-2018-04) to MJOG. DDI thanks the University of Alcalá and Spanish Ministry of Science, Innovation and Universities for pre-doctoral FPU grants. BCG thanks the Spanish Ministr

Synthesis and Pharmacological Evaluation of New N-Sulfonylureas as NLRP3 Inflammasome Inhibitors: Identification of a Hit Compound to Treat Gout

NLRP3 is involved in the pathophysiology of several inflammatory diseases. Therefore, there is high current interest in the clinical development of new NLRP3 inflammasome small inhibitors to treat these diseases. Novel N-sulfonylureas were obtained by the replacement of the hexahydroindacene moiety of the previously described NLRP3 inhibitor MCC950. These new derivatives show moderate to high potency in inhibiting IL-1β release in vitro. The greatest effect was observed for compound 4b, which was similar to MCC950. Moreover, compound 4b was able to reduce caspase-1 activation, oligomerization of ASC, and therefore, IL-1β processing. Additional in silico predictions confirmed the safety profile of compound 4b, and in vitro studies in AML12 hepatic cells confirmed the absence of toxicological effects. Finally, we evaluated in vivo anti-inflammatory properties of compound 4b, which showed a significant anti-inflammatory effect and reduced mechanical hyperalgesia at 3 and 10 mg/kg (i.p.) in an in vivo mouse model of gout.J.E. thanks Fondo de Investigaciones Sanitarias (ISCIII/ FEDER) (Programa Miguel Servet: CP19/00005 and PI19/ 00082) and Fundación Mutua Madrileñ a. D.D.-I. thanks the Spanish Ministry of Science, Innovation, and Universities for predoctoral FPU grant

Multipotent cholinesterase/monoamine oxidase inhibitors for the treatment of Alzheimer's disease: design, synthesis, biochemical evaluation, ADMET, molecular modeling, and QSAR analysis of novel donepezil-pyridyl hybrids

The design, synthesis, and biochemical evaluation of donepezil-pyridyl hybrids (DPHs) as multipotent cholinesterase (ChE) and monoamine oxidase (MAO) inhibitors for the potential treatment of Alzheimer's disease (AD) is reported. The 3D-quantitative structure-activity relationship study was used to define 3D-pharmacophores for inhibition of MAO A/B, acetylcholinesterase (AChE), and butyrylcholinesterase (BuChE) enzymes and to design DPHs as novel multi-target drug candidates with potential impact in the therapy of AD. DPH14 (Electrophorus electricus AChE [EeAChE]: half maximal inhibitory concentration [IC50] = 1.1 +/- 0.3 nM; equine butyrylcholinesterase [eqBuChE]: IC50 = 600 +/- 80 nM) was 318-fold more potent for the inhibition of AChE, and 1.3-fold less potent for the inhibition of BuChE than the reference compound ASS234. DPH14 is a potent human recombinant BuChE (hBuChE) inhibitor, in the same range as DPH12 or DPH16, but 13.1-fold less potent than DPH15 for the inhibition of human recombinant AChE (hAChE). Compared with donepezil, DPH14 is almost equipotent for the inhibition of hAChE, and 8.8-fold more potent for hBuChE. Concerning human monoamine oxidase (hMAO) A inhibition, only DPH9 and 5 proved active, compound DPH9 being the most potent (IC50 [MAO A] = 5,700 +/- 2,100 nM). For hMAO B, only DPHs 13 and 14 were moderate inhibitors, and compound DPH14 was the most potent (IC50 [MAO B] = 3,950 +/- 94 nM). Molecular modeling of inhibitor DPH14 within EeAChE showed a binding mode with an extended conformation, interacting simultaneously with both catalytic and peripheral sites of EeAChE thanks to a linker of appropriate length. Absortion, distribution, metabolism, excretion and toxicity analysis showed that structures lacking phenyl-substituent show better druglikeness profiles; in particular, DPHs13-15 showed the most suitable absortion, distribution, metabolism, excretion and toxicity properties. Novel donepezil-pyridyl hybrid DPH14 is a potent, moderately selective hAChE and selective irreversible hMAO B inhibitor which might be considered as a promising compound for further development for the treatment of AD