[Show abstract][Hide abstract] ABSTRACT: Polythiazole amino acids clasp linear peptides to generate cyclic derivatives, however, the resulting species are not merely stapled peptides but bear a complex heterocyclic moiety displaying its intrinsic set of interactions. As a proof of concept, a bisthiazole moiety has been grafted onto an RGD sequence to deliver a new cilengitide analogue with improved integrin selectivity and remarkable in vivo antiangiogenic activity.
[Show abstract][Hide abstract] ABSTRACT: Optimization of an essentially inactive 3,4-dihydro-2H-pyrano[3,2-c]quinoline carboxylic ester derivative as acetylcholinesterase (AChE) peripheral anionic site (PAS)-binding motif by double O → NH bioisosteric replacement, combined with molecular hybridization with the AChE catalytic anionic site (CAS) inhibitor 6-chlorotacrine and molecular dynamics-driven optimization of the length of the linker has resulted in the development of the trimethylene-linked 1,2,3,4-tetrahydrobenzo[h][1,6]naphthyridine-6-chlorotacrine hybrid 5a as a picomolar inhibitor of human AChE (hAChE). The tetra-, penta-, and octamethylene-linked homologues 5b-d have been also synthesized for comparison purposes, and found to retain the nanomolar hAChE inhibitory potency of the parent 6-chlorotacrine. Further biological profiling of hybrids 5a-d has shown that they are also potent inhibitors of human butyrylcholinesterase and moderately potent Aβ42 and tau anti-aggregating agents, with IC50 values in the submicromolar and low micromolar range, respectively. Also, in vitro studies using an artificial membrane model have predicted a good brain permeability for hybrids 5a-d, and hence, their ability to reach their targets in the central nervous system. The multitarget profile of the novel hybrids makes them promising leads for developing anti-Alzheimer drug candidates with more balanced biological activities.
European journal of medicinal chemistry. 07/2014; 84C:107-117.
[Show abstract][Hide abstract] ABSTRACT: The α4β2 nicotinic acetylcholine receptor (nAChR) is a molecular target of 3,4-methylenedioxymethamphetamine (MDMA), a synthetic drug also known as ecstasy, and it modulates the MDMA-mediated reinforcing properties. However, the enantioselective preference of the α4β2 nAChR subtype still remains unknown. Since the two enantiomers exhibit different pharmacological profiles and stereoselective metabolism, the aim of this study is to assess a possible difference in the interaction of the MDMA enantiomers with this nAChR subtype. To this end, we report a novel simple, yet highly efficient enantioselective synthesis of the MDMA enantiomers, in which the key step is the diastereoselective reduction of imides derived from optically pure tert-butylsulfinamide. The enantioselective binding to the receptor is examined using [(3)H]epibatidine in a radioligand assay. Even though the two enantiomers induced a concentration-dependent binding displacement, (S)-MDMA has an inhibition constant 13-fold higher than (R)-MDMA, which shows a Hill's coefficient not significantly different from unity, implying a competitive interaction. Furthermore, when NGF-differentiated PC12 cells were pretreated with the compounds, a significant increase in binding of [(3)H]epibatidine was found for (R)-MDMA, indicating up-regulation of heteromeric nAChR in the cell surface. Finally, docking and molecular dynamics studies have been used to identify the binding mode of the two enantiomers, which provides a structural basis to justify the differences in affinity from the differential interactions played by the substituents at the stereogenic centre of MDMA. The results provide a basis to explore the distinct psychostimulant profiles of the MDMA enantiomers mediated by the α4β2 nAChR subtype.
European journal of medicinal chemistry. 06/2014; 81:35-46.
[Show abstract][Hide abstract] ABSTRACT: Amantadine inhibits the M2 proton channel of influenza A virus, yet most of the currently circulating strains of the virus carry mutations in the M2 protein that render the virus amantadine-resistant. While most of the research on novel amantadine analogs has revolved around the synthesis of novel adamantane derivatives, we have recently found that other polycyclic scaffolds effectively block the M2 proton channel, including amantadine-resistant mutant channels. In this paper, we have synthesized and characterized a series of pyrrolidine derivatives designed as analogs of amantadine. Inhibition of the wild-type M2 channel and the A/M2-S31N, A/M2-V27A and A/M2-L26F mutant forms of the channel were measured in Xenopus oocytes using two-electrode voltage clamp assays. Most of the novel compounds inhibited the wild type ion channel in the low micromolar range. Of note, two of the compounds inhibited the amantadine-resistant A/M2-V27A and A/M2-L26F mutant ion channels with submicromolar and low micromolar IC50, respectively. None of the compounds was found to inhibit the S31N mutant ion channel.
[Show abstract][Hide abstract] ABSTRACT: Many pathogenic microorganisms have evolved hemoglobin mediated nitric-oxide (NO) detoxification mechanisms, where a globin domain in conjunction with a partner reductase catalyzes the conversion of toxic NO to innocuous nitrate. The truncated hemoglobin HbN of Mycobacterium tuberculosis displays a potent NO-dioxygenase activity despite lacking a reductase domain. The mechanism by which HbN recycles itself during NO-dioxygenation and the reductase that participates in this process are currently unknown. This study demonstrates that the NADH-ferredoxin/flavodoxin system is a fairly efficient partner for electron transfer to HbN with an observed reduction rate of 6.2 micromole/min-1, which is nearly 3- and 5-fold faster than the ones reported for Vitreoscilla hemoglobin and myoglobin, respectively. Structural docking of the HbN with E. coli NADH-flavodoxin reductase (FdR) together with site-directed mutagenesis revealed that the CD loop of the HbN forms contacts with the reductase, and that Gly48 may have a vital role. The electron coupling parameters calculated using the semiempirical pathway method amounts to an average of ca. 6.4 10-5 eV, which is lower than the value obtained for E. coli flavoHb (8.0 10-4 eV), but still supports the feasibility of an efficient electron transfer. The deletion of Pre-A abrogated the heme iron reduction by FdR in the HbN, thus signifying its involvement during intermolecular interactions of the HbN and the FdR. The present study, thus, unravels novel role of the CD loop and Pre-A motif in assisting the interactions of the HbN with the reductase and the electron-cycling, which may be vital for its NO-scavenging function.
[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: Multitarget compounds are increasingly being pursued for the effective treatment of complex diseases. Herein, we describe the design and synthesis of a novel class of shogaol-huprine hybrids, purported to hit several key targets involved in Alzheimer's disease. The hybrids have been tested in vitro for their inhibitory activity against human acetylcholinesterase and butyrylcholinesterase and antioxidant activity (ABTS(+), DPPH and Folin-Ciocalteu assays), and in intact Escherichia coli cells for their Aβ42 and tau anti-aggregating activity. Also, their brain penetration has been assessed (PAMPA-BBB assay). Even though the hybrids are not as potent AChE inhibitors or antioxidant agents as the parent huprine Y and -shogaol, respectively, they still exhibit very potent anticholinesterase and antioxidant activities and are much more potent Aβ42 and tau anti-aggregating agents than the parent compounds. Overall, the shogaol-huprine hybrids emerge as interesting brain permeable multitarget anti-Alzheimer leads.
[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