[show abstract][hide abstract] ABSTRACT: We have synthesized a family of rhein-huprine hybrids to hit several key targets for Alzheimer's disease. Biological screening performed in vitro and in Escherichia coli cells has shown that these hybrids exhibit potent inhibitory activities against human acetylcholinesterase butyrylcholinesterase, and BACE-1, dual Aβ42 and tau anti-aggregating activity, and brain permeability. Ex vivo studies with the leads (+)- and (-)-7e in brain slices of C57bl6 mice have revealed that they efficiently protect against the Aβ-induced synaptic dysfunction , preventing the loss of synaptic proteins and/or have a positive effect on the induction of long term potentiation. In vivo studies in APP-PS1 transgenic mice treated i.p. for 4 weeks with (+)- and (-)-7e have shown a central soluble Aβ lowering effect, accompanied by an increase in the levels of mature amyloid precursor protein (APP). Thus, (+)- and (-)-7e emerge as very promising disease-modifying anti-Alzheimer drug candidates.
Journal of Medicinal Chemistry 02/2014; · 5.61 Impact Factor
[show abstract][hide abstract] ABSTRACT: A series of 1,2,3,4-tetrahydrobenzo[h][1,6]naphthyridines differently substituted at positions 1, 5, and 9 have been designed from the pyrano[3,2-c]quinoline derivative 1, a weak inhibitor of acetylcholinesterase (AChE) with predicted ability to bind to the AChE peripheral anionic site (PAS), at the entrance of the catalytic gorge. Fourteen novel benzonaphthyridines have been synthesized through synthetic sequences involving as the key step a multicomponent Povarov reaction between an aldehyde, an aniline and an enamine or an enamide as the activated alkene. The novel compounds have been tested against Electrophorus electricus AChE (EeAChE), human recombinant AChE (hAChE), and human serum butyrylcholinesterase (hBChE), and their brain penetration has been assessed using the PAMPA-BBB assay. Also, the mechanism of AChE inhibition of the most potent compounds has been thoroughly studied by kinetic studies, a propidium displacement assay, and molecular modelling. We have found that a seemingly small structural change such as a double O → NH bioisosteric replacement from the hit 1 to 16a results in a dramatic increase of EeAChE and hAChE inhibitory activities (>217- and >154-fold, respectively), and in a notable increase in hBChE inhibitory activity (>11-fold), as well. An optimized binding at the PAS besides additional interactions with AChE midgorge residues seem to account for the high hAChE inhibitory potency of 16a (IC50 = 65 nM), which emerges as an interesting anti-Alzheimer lead compound with potent dual AChE and BChE inhibitory activities.
European journal of medicinal chemistry 12/2013; 73C:141-152. · 3.27 Impact Factor
[show abstract][hide abstract] ABSTRACT: We present here an exhaustive characterization of the structure and properties of 6-selenoguanine, an isoster of guanine, and the impact of its introduction in DNA. This study reports the results of state-of-the-art quantum mechanical calculations and atomistic molecular dynamics simulations carried out to shed light on the impact of the replacement of guanine (G) by 6-selenoguanine (SeG) in different forms of DNA. The results point out that the G → SeG substitution leads to stable DNA duplex, antiparallel triplex and G-quadruplex structures, though local distortions are also found. These structural changes affect the thermodynamic stability of the mutation leading to a clear destabilization for all studied systems. Interestingly, the lowest effect has been found when the mutation was placed in the triplex-forming oligonucleotide strand in a reverse Hoogsteen orientation, which favours the antiparallel triplex formation regarding the G-tetraplex formation. Detailed QM studies strongly suggest that SeG impacts the HOMO-LUMO gap and accordingly the transfer properties of DNA, opening the way to modulate the conductivity properties of non-natural DNAs.
Physical Chemistry Chemical Physics 11/2013; · 3.83 Impact Factor
[show abstract][hide abstract] ABSTRACT: Since cyanide potentiates the inhibitory activity of several monoamine oxidase (MAO) inhibitors, a series of carbonitrile-containing aminoheterocycles was examined to explore the role of nitriles in determining the inhibitory activity against MAO. Dicarbonitrile aminofurans were found to be potent, selective inhibitors against MAO A. The origin of the MAO A selectivity was identified by combining spectroscopic and computational methods. Spectroscopic changes induced in MAO A by mono- and dicarbonitrile inhibitors were different, providing experimental evidence for distinct binding modes to the enzyme. Similar differences were also found between the binding of dicarbonitrile compounds to MAO A and to MAO B. Stabilization of the flavin anionic semiquinone by monocarbonitrile compounds, but destabilization by dicarbonitriles, provided further support to the distinct binding modes of these compounds and their interaction with the flavin ring. Molecular modelling studies supported the role played by the nitrile and amino groups in anchoring the inhibitor to the binding cavity. In particular, the results highlight the role of Asn181 and Ile335 in assisting the interaction of the nitrile-containing aminofuran ring. The network of interactions afforded by the specific attachment of these functional groups provides useful guidelines for the design of selective, reversible MAO A inhibitors.
Biochimica et Biophysica Acta 11/2013; · 4.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: The evolution of a ternary molecular system (imine, diene and nitrile) is analyzed to disclose the pathways leading to a divergent synthetic outcome. The Lewis acid catalyzed reaction between cyclohexadiene, 2-phenyl-indol-3-one and acetonitrile yields the imino-Diels-Alder adduct as the major product together with minor amounts of the Mannich-Ritter-amidine product. The experimental and computational data show that the relative orientation of the initial reactants dictates the synthetic outcome. The exo approach between imine and diene leads to the Diels-Alder adduct in a concerted process, whereas the endo mode leads to a polarized intermediate, which is trapped by acetonitrile to yield the multicomponent adduct.
[show abstract][hide abstract] ABSTRACT: We address the issue of whether chemical alterations of nucleobases are an effective tool to modulate charge transfer through DNA molecules. Our investigation uses a multi-level computational approach based on classical molecular dynamics and quantum chemistry. We find yet another evidence that structural fluctuations are a key factor to determine the electronic structure of double-stranded DNA. We argue that the electronic structure and charge transfer ability of flexible polymers is the result of a complex intertwining of various structural, dynamical and chemical factors. Chemical intuition may be used to design molecular wires, but this is not the sole component in the complex charge transfer mechanism through DNA.
[show abstract][hide abstract] ABSTRACT: The aim of this work was to perform a detailed study of the alkaloid content of Narcissus triandrus, as well as a complete analysis of the alkaloid profile of 18 wild populations, comprising all the taxa of the section Ganymedes. Through the application of a combination of spectroscopic and chromatographic methods, the isolation and structural elucidation of 3 compounds are reported for the first time from a natural source (2-oxomesembrenone, 7,7a-dehydromesembrenone and 2-oxoepimesembranol), together with the identification of 5 major common mesembrane alkaloids. Additionally, the GC-MS analysis of the alkaloid profile demonstrated the regular presence of mesembranes in all the studied plants, showing mesembrenone as the predominant compound without any typical Amaryllidaceae alkaloid being detected.
[show abstract][hide abstract] ABSTRACT: The presence of cavities and tunnels in the interior of proteins, in conjunction with the structural plasticity arising from the coupling to the thermal fluctuations of the protein scaffold, has profound consequences on the pathways followed by ligands moving through the protein matrix. In this perspective we discuss how quantitative analysis of experimental rebinding kinetics from laser flash photolysis, trapping of unstable conformational states by embedding proteins within the nanopores of silica gels, and molecular simulations can synergistically converge to gain insight into the migration mechanism of ligands. We show how the evaluation of the free energy landscape for ligand diffusion based on the outcome of computational techniques can assist the definition of sound reaction schemes, leading to a comprehensive understanding of the broad range of chemical events and time scales that encompass the transport of small ligands in hemeproteins.
Physical Chemistry Chemical Physics 06/2013; · 3.83 Impact Factor
[show abstract][hide abstract] ABSTRACT: We present a refinement of the backbone torsion parameters ε and ζ of the Cornell et al. AMBER force field for DNA simulations. The new parameters, denoted as εζOL1, were derived from quantum-mechanical calculations with inclusion of conformation-dependent solvation effects according to the recently reported methodology (J. Chem. Theory Comput. 2012, 7(9), 2886-2902). The performance of the refined parameters was analyzed by means of extended molecular dynamics (MD) simulations for several representative systems. The results showed that the εζOL1 refinement improves the backbone description of B-DNA double helices and G-DNA stem. In B-DNA simulations, we observed an average increase of the helical twist and narrowing of the major groove, thus achieving better agreement with X-ray and solution NMR data. The balance between populations of BI and BII backbone substates was shifted towards the BII state, in better agreement with ensemble-refined solution experimental results. Furthermore, the refined parameters decreased the backbone RMS deviations in B-DNA MD simulations. In the antiparallel guanine quadruplex (G-DNA) the εζOL1 modification improved the description of non-canonical α/γ backbone substates, which were shown to be coupled to the ε/ζ torsion potential. Thus, the refinement is suggested as a possible alternative to the current ε/ζ torsion potential, which may enable more accurate modeling of nucleic acids. However, long-term testing is recommended before its routine application in DNA simulations.
Journal of Chemical Theory and Computation 05/2013; 9(5):2339-2354. · 5.39 Impact Factor
[show abstract][hide abstract] ABSTRACT: Nitrophorins (NPs) are nitric oxide (NO)-carrying heme proteins found in the saliva of the blood-sucking insect Rhodnius prolixus. Though NP7 exhibits a large sequence resemblance with other NPs, two major differential features are the ability to interact with negatively charged cell surfaces and the presence of a specific N-terminus composed of three extra residues (Leu1-Pro2-Gly3). The aim of this study is to examine the influence of the N-terminus on the ligand binding, and the topological features of inner cavities in closed and open states of NP7, which can be associated to the protein structure at low and high pH, respectively. Laser flash photolysis measurements of the CO rebinding kinetics to NP7 and its variant NP7(Δ1-3), which lacks the three extra residues at the N-terminus, exhibit a similar pattern and support the existence of a common kinetic mechanism for ligand migration and binding. This is supported by the existence of a common topology of inner cavities, which consists of two docking sites in the heme pocket and a secondary site at the back of the protein. The ligand exchange between these cavities is facilitated by an additional site, which can be transiently occupied by the ligand in NP7, although it is absent in NP4. These features provide a basis to explain the enhanced internal gas hosting capacity found experimentally in NP7 and the absence of ligand rebinding from secondary sites in NP4. The current data allow us to speculate that the processes of docking to cell surfaces and NO release may be interconnected in NP7, thereby efficiently releasing NO into a target cell.
Biochimica et Biophysica Acta 04/2013; · 4.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: The accuracy and performance of implicit solvent methods for solvation free energy calculations were assessed on a set of 20 neutral drug molecules. Molecular dynamics (MD) provided ensembles of conformations in water and water-saturated octanol. The solvation free energies were calculated by popular implicit solvent models based on quantum mechanical (QM) electronic densities (COSMO-RS, MST, SMD) as well as on molecular mechanical (MM) point-charge models (GB, PB). The performance of the implicit models was tested by a comparison with experimental water-octanol transfer free energies (ΔGow) by using single- and multi-conformation approaches. MD simulations revealed difficulties in a priori estimation of the flexibility features of the solutes from simple structural descriptors, such as the number of rotatable bonds. An increasing accuracy of the calculated ΔGow was observed in the following order: GB1 ~ PB < GB7 < MST < SMD ~ COSMO-RS with a clear distinction identified between MM- and QM-based models, although for the set excluding three largest molecules, the differences between COSMO-RS, MST and SMD were negligible. It was shown that the single-conformation approach applied to crystal geometries provides a rather accurate estimate of ΔGow for rigid molecules yet fails completely for the flexible ones. The multi-conformation approaches improved the performance, but only when the deformation contribution was ignored. It was revealed that for large-scale calculations on small molecules a recent GB model, GB7, provided a reasonable accuracy/speed ratio. In conclusion, the study contributes to the understanding of solvation free energy calculations for physical and medicinal chemistry applications.
The Journal of Physical Chemistry B 04/2013; · 3.61 Impact Factor
[show abstract][hide abstract] ABSTRACT: This study reports a comparative analysis of the topological properties of inner cavities and the intrinsic dynamics of non-symbiotic hemoglobins AHb1 and AHb2 from Arabidopsis thaliana. The two proteins belong to the 3/3 globin fold and have a sequence identity of about 60%. However, it is widely assumed that they have distinct physiological roles. In order to investigate the structure-function relationships in these proteins, we have examined the bis-histidyl and ligand-bound hexacoordinated states by atomistic simulations using in silico structural models. The results allow us to identify two main pathways to the distal cavity in the bis-histidyl hexacoordinated proteins. Nevertheless, a larger accessibility to small gaseous molecules is found in AHb2. This effect can be attributed to three factors: the mutation Leu35(AHb1)→Phe32(AHb2), the enhanced flexibility of helix B, and the more favorable energetic profile for ligand migration to the distal cavity. The net effect of these factors would be to facilitate the access of ligands, thus compensating the preference for the fully hexacoordination of AHb2, in contrast to the equilibrium between hexa- and pentacoordinated species in AHb1. On the other hand, binding of the exogenous ligand introduces distinct structural changes in the two proteins. A well-defined tunnel is formed in AHb1, which might be relevant to accomplish the proposed NO detoxification reaction. In contrast, no similar tunnel is found in AHb2, which can be ascribed to the reduced flexibility of helix E imposed by the larger number of salt bridges compared to AHb1. This feature would thus support the storage and transport functions proposed for AHb2. This article is part of a Special Issue entitled: Oxygen Binding and Sensing Proteins.
Biochimica et Biophysica Acta 04/2013; · 4.66 Impact Factor
[show abstract][hide abstract] ABSTRACT: Bicyclic α-iminophosphonates were prepared via the first diastereoselective silver catalyzed [3 + 2] cycloaddition reaction of diethyl isocyanomethylphosphonate and diversely N-substituted maleimides. The reduction of the resulting imine by catalytic hydrogenation led to cyclic α-aminophosphonates, which are α-aminoester surrogates. The relative stereochemistry of the adducts was confirmed by X-ray crystallographic analysis of . The diastereoselectivity of the cycloaddition reaction was rationalised by theoretical studies.
[show abstract][hide abstract] ABSTRACT: Cytoglobin (Cygb) was recently discovered in the human genome and localized in different tissues. It was suggested to play tissue-specific protective roles, spanning from scavenging of reactive oxygen species in neurons to supplying oxygen to enzymes in fibroblasts. To shed light on the functioning of such versatile machinery, we have studied the processes supporting transport of gaseous heme ligands in Cygb. Carbon monoxide rebinding shows a complex kinetic pattern with several distinct reaction intermediates, reflecting rebinding from temporary docking sites, second order recombination, and formation (and dissociation) of a bis-histidyl heme hexacoordinated reaction intermediate. Ligand exit to the solvent occurs through distinct pathways, some of which exploit temporary docking sites. The remarkable change in energetic barriers, linked to heme bis-histidyl hexacoordination by HisE7, may be responsible for active regulation of the flux of reactants and products to and from the reaction site on the distal side of the heme. A substantial change in both protein dynamics and inner cavities is observed upon transition from the CO-liganded to the pentacoordinated and bis-histidyl hexacoordinated species, which could be exploited as a signalling state. These findings are consistent with the expected versatility of the molecular activity of this protein.
PLoS ONE 01/2013; 8(1):e49770. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: Glycogen synthase kinase 3β (GSK-3β) is widely recognised as a relevant player in the pathogenesis of several highly prevalent disorders such as Alzheimer's disease, mood disorders, diabetes and cancer. Therefore, this enzyme constitutes a highly attractive therapeutic target for the development of selective inhibitors as new promising drugs for the treatment of these pathologies. We describe here the isolation and biochemical characterization of the marine natural sesquiterpene palinurin as a GSK-3β inhibitor. Experimental studies performed for characterizing the inhibitory mechanism indicate that GSK-3β inhibition by palinurin cannot be competed out by ATP nor peptide substrate. Molecular modelling techniques have enabled us to propose an unconventional binding mode to GSK-3β. Moreover, molecular dynamics simulations have identified an allosteric mechanism by which binding of palinurin leads to GSK-3β inhibition. The inhibitory activities determined for a series of structurally related analogues support the proposed binding mode of palinurin, which is the first compound described to target this allosteric site. The results offer new opportunities for designing and developing selective inhibitors with novel mechanisms of action.
European journal of medicinal chemistry 12/2012; 60C:479-489. · 3.27 Impact Factor
[show abstract][hide abstract] ABSTRACT: When a door closes, a window opens! The use of geometrically or electronically restricted imines for Povarov-type processes does not afford the anti-Bredt tetrahydroquinolines, but leads instead to highly functionalized structures through novel reaction pathways (see picture; LA=Lewis acid). The exploration of "forbidden" routes constitutes a valuable approach in the search for new multicomponent reactions.
Angewandte Chemie International Edition 05/2012; 51(28):6874-7. · 13.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: The truncated hemoglobin N, HbN, of Mycobacterium tuberculosis is endowed with a potent nitric oxide dioxygenase (NOD) activity that allows it to relieve nitrosative stress and enhance in vivo survival of its host. Despite its small size, the protein matrix of HbN hosts a two-branched tunnel, consisting of orthogonal short and long channels, that connects the heme active site to the protein surface. A novel dual-path mechanism has been suggested to drive migration of O(2) and NO to the distal heme cavity. While oxygen migrates mainly by the short path, a ligand-induced conformational change regulates opening of the long tunnel branch for NO, via a phenylalanine (PheE15) residue that acts as a gate. Site-directed mutagenesis and molecular simulations have been used to examine the gating role played by PheE15 in modulating the NOD function of HbN. Mutants carrying replacement of PheE15 with alanine, isoleucine, tyrosine and tryptophan have similar O(2)/CO association kinetics, but display significant reduction in their NOD function. Molecular simulations substantiated that mutation at the PheE15 gate confers significant changes in the long tunnel, and therefore may affect the migration of ligands. These results support the pivotal role of PheE15 gate in modulating the diffusion of NO via the long tunnel branch in the oxygenated protein, and hence the NOD function of HbN.
PLoS ONE 01/2012; 7(11):e49291. · 3.73 Impact Factor
[show abstract][hide abstract] ABSTRACT: A new family of multitarget molecules able to interact with acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE), as well as with monoamino oxidase (MAO) A and B, has been synthesized. Novel compounds (3-9) have been designed using a conjunctive approach that combines the benzylpiperidine moiety of the AChE inhibitor donepezil (1) and the indolyl propargylamino moiety of the MAO inhibitor N-[(5-benzyloxy-1-methyl-1H-indol-2-yl)methyl]-N-methylprop-2-yn-1-amine (2), connected through an oligomethylene linker. The most promising hybrid (5) is a potent inhibitor of both MAO-A (IC50=5.2±1.1 nM) and MAO-B (IC50=43±8.0 nM) and is a moderately potent inhibitor of AChE (IC50=0.35±0.01 μM) and BuChE (IC50=0.46±0.06 μM). Moreover, molecular modeling and kinetic studies support the dual binding site to AChE, which explains the inhibitory effect exerted on Aβ aggregation. Overall, the results suggest that the new compounds are promising multitarget drug candidates with potential impact for Alzheimer's disease therapy.
Journal of Medicinal Chemistry 12/2011; 54(24):8251-70. · 5.61 Impact Factor