Publications by authors named "Espargaró Alba"

44 Publications

Dual Inhibitors of Amyloid-β and Tau Aggregation with Amyloid-β Disaggregating Properties: Extended , , and Kinetic Studies of Multifunctional Anti-Alzheimer's Agents.

ACS Chem Neurosci 2021 06 21;12(11):2057-2068. Epub 2021 May 21.

Department of Physicochemical Drug Analysis, Faculty of Pharmacy, Jagiellonian University Medical College, Medyczna 9, 30-688 Krakow, Poland.

In Alzheimer's disease, neurons slowly degenerate due to the accumulation of misfolded amyloid β and tau proteins. In our research, we performed extended studies directed at amyloid β and tau aggregation inhibition using ( model of protein aggregation), , and kinetic studies. We tested our library of 1-benzylamino-2-hydroxyalkyl multifunctional anti-Alzheimer's agents and identified very potent dual aggregation inhibitors. Among the tested derivatives, we selected compound , which exhibited a unique profile of biological activity. This compound was the most potent and balanced dual aggregation inhibitor (Aβ inhibition (inh.) 80.0%, tau inh. 68.3% in 10 μM), with previously reported inhibitory activity against BuChE, BACE1, and Aβ (BuChE IC = 5.74 μM; BACE1 IC = 41.6 μM; Aβ aggregation (aggr.) inh. IC = 3.09 μM). In docking studies for both proteins, we tried to explain the different structural requirements for the inhibition of Aβ vs tau. Moreover, docking and kinetic studies showed that compound could inhibit the amyloid aggregation process at several steps and also displayed disaggregating properties. These results may help to design the next generations of dual or selective aggregation inhibitors.
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http://dx.doi.org/10.1021/acschemneuro.1c00235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8291496PMC
June 2021

Azobioisosteres of Curcumin with Pronounced Activity against Amyloid Aggregation, Intracellular Oxidative Stress, and Neuroinflammation.

Chemistry 2021 Apr 5;27(19):6015-6027. Epub 2021 Mar 5.

Pharmaceutical and Medicinal Chemistry, Institute of, Pharmacy and Food Chemistry, University of Würzburg, Am Hubland, 97074, Würzburg, Germany.

Many (poly-)phenolic natural products, for example, curcumin and taxifolin, have been studied for their activity against specific hallmarks of neurodegeneration, such as amyloid-β 42 (Aβ42) aggregation and neuroinflammation. Due to their drawbacks, arising from poor pharmacokinetics, rapid metabolism, and even instability in aqueous medium, the biological activity of azobenzene compounds carrying a pharmacophoric catechol group, which have been designed as bioisoteres of curcumin has been examined. Molecular simulations reveal the ability of these compounds to form a hydrophobic cluster with Aβ42, which adopts different folds, affecting the propensity to populate fibril-like conformations. Furthermore, the curcumin bioisosteres exceeded the parent compound in activity against Aβ42 aggregation inhibition, glutamate-induced intracellular oxidative stress in HT22 cells, and neuroinflammation in microglial BV-2 cells. The most active compound prevented apoptosis of HT22 cells at a concentration of 2.5 μm (83 % cell survival), whereas curcumin only showed very low protection at 10 μm (21 % cell survival).
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http://dx.doi.org/10.1002/chem.202005263DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8048673PMC
April 2021

Dual Effect of Prussian Blue Nanoparticles on Aβ40 Aggregation: β-Sheet Fibril Reduction and Copper Dyshomeostasis Regulation.

Biomacromolecules 2021 02 8;22(2):430-440. Epub 2021 Jan 8.

Department of Pharmacy and Pharmaceutical Technology and Physical-Chemistry, Faculty of Pharmacy and Food Sciences, University of Barcelona, Avda. Joan XXIII, 27-31 Barcelona, Catalonia, Spain.

Alzheimer's disease (AD), affecting almost 50 million individuals worldwide, is currently the first cause of dementia. Despite the tremendous research efforts in the last decade, only four supportive or palliative drugs, namely, acetylcholinesterase (AChE) inhibitors donepezil, galantamine, and rivastigmine and the glutamate NMDA receptor antagonist memantine, are currently available. New therapeutic strategies are becoming prominent, such as the direct inhibition of amyloid formation or the regulation of metal homeostasis. In the present report, the potential use of Prussian blue (PB), a drug that is in the World Health Organization Model List of Essential Medicines, in AD treatment is demonstrated. Both and studies indeed suggest that PB nanoparticles (PBNPs) are capable of reducing the formation of typical amyloid-β fibers (detected by thioflavin T fluorescence) and restoring the usual amyloid fibrillation pathway chelation/sequestration of copper, which is found in high concentrations in senile plaques.
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http://dx.doi.org/10.1021/acs.biomac.0c01290DOI Listing
February 2021

Pharmacophore Modeling and 3D-QSAR Study of Indole and Isatin Derivatives as Antiamyloidogenic Agents Targeting Alzheimer's Disease.

Molecules 2020 Dec 7;25(23). Epub 2020 Dec 7.

Department of Pharmacy-Pharmaceutical Sciences, University of Bari Aldo Moro, Via E. Orabona 4, 70125 Bari, Italy.

Thirty-six novel indole-containing compounds, mainly 3-(2-phenylhydrazono) isatins and structurally related 1-indole-3-carbaldehyde derivatives, were synthesized and assayed as inhibitors of beta amyloid (Aβ) aggregation, a hallmark of pathophysiology of Alzheimer's disease. The newly synthesized molecules spanned their IC values from sub- to two-digit micromolar range, bearing further information into structure-activity relationships. Some of the new compounds showed interesting multitarget activity, by inhibiting monoamine oxidases A and B. A cell-based assay in tau overexpressing bacterial cells disclosed a promising additional activity of some derivatives against tau aggregation. The accumulated data of either about ninety published and thirty-six newly synthesized molecules were used to generate a pharmacophore hypothesis of antiamyloidogenic activity exerted in a wide range of potencies, satisfactorily discriminating the 'active' compounds from the 'inactive' (poorly active) ones. An atom-based 3D-QSAR model was also derived for about 80% of 'active' compounds, i.e., those achieving finite IC values lower than 100 μM. The 3D-QSAR model (encompassing 4 PLS factors), featuring acceptable predictive statistics either in the training set ( = 45, q = 0.596) and in the external test set ( = 14, r = 0.695), usefully complemented the pharmacophore model by identifying the physicochemical features mainly correlated with the Aβ anti-aggregating potency of the indole and isatin derivatives studied herein.
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http://dx.doi.org/10.3390/molecules25235773DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7731220PMC
December 2020

Centrally Active Multitarget Anti-Alzheimer Agents Derived from the Antioxidant Lead CR-6.

J Med Chem 2020 09 11;63(17):9360-9390. Epub 2020 Aug 11.

Laboratory of Medicinal Chemistry (CSIC Associated Unit), Faculty of Pharmacy and Food Sciences, and Institute of Biomedicine (IBUB), University of Barcelona (UB), E-08028 Barcelona, Spain.

Oxidative stress is a major pathogenic factor in Alzheimer's disease, but it should not be tackled alone rather together with other key targets to derive effective treatments. The combination of the scaffold of the polar antioxidant lead 7-methoxy-2,2-dimethylchroman-6-ol (CR-6) with that of the lipophilic cholinesterase inhibitor 6-chlorotacrine results in compounds with favorable brain permeability and multiple activities in vitro (acetylcholinesterase, butyrylcholinesterase, β-site amyloid precursor protein (APP) cleaving enzyme-1 (BACE-1), and Aβ42 and tau aggregation inhibition). In in vivo studies on wild-type and APP/presenilin 1 (PS1) mice, two selected compounds were well tolerated and led to positive trends, albeit statistically nonsignificant in some cases, on memory performance, amyloid pathology (reduced amyloid burden and potentiated non-amyloidogenic APP processing), and oxidative stress (reduced cortical oxidized proteins and increased antioxidant enzymes superoxide dismutase 2 (SOD2), catalase, glutathione peroxidase 1 (GPX1), and heme oxygenase 1 (Hmox1) and transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2)). These compounds emerge as interesting brain-permeable multitarget compounds, with a potential as anti-Alzheimer agents beyond that of the original lead CR-6.
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http://dx.doi.org/10.1021/acs.jmedchem.0c00528DOI Listing
September 2020

Thiosemicarbazone Derivatives as Inhibitors of Amyloid-β Aggregation: Effect of Metal Coordination.

Inorg Chem 2020 May 5;59(10):6978-6987. Epub 2020 May 5.

nanoBIC, Departament de Química Inorgànica i Orgànica, Facultat de Química, Universitat de Barcelona, Martí i Franquès, 1-11, 08028 Barcelona, Spain.

Three thiosemicarbazone derivatives, namely 4-(dimethylamino)benzaldehyde 4,4-dimethylthiosemicarbazone (), 4-(dimethylamino)benzaldehyde thiosemicarbazone (), and 4-(dimethylamino)benzaldehyde 4-methylthiosemicarbazone (), have been synthesized and characterized. The three palladium(II) complexes - were prepared respectively from , , and . The crystal structures of two coordination compounds, namely Pd() () and Pd() (), were obtained, which showed the expected square-planar environment for the metal centers. The ligand and the Pd(II) complexes -, which are stable in buffered solutions containing up to 5% DMSO, exhibit remarkable inhibitory properties against the aggregation of amyloid-β, reducing the formation of fibrils. , , , and display IC values (i.e., the concentrations required to reduce Aβ fibrillation by 50%) below 1 μM, lower that of the reference compound catechin (IC = 2.8 μM). Finally, studies with cells revealed that the palladium(II) compounds are significantly more efficient than the free ligands in inhibiting Aβ aggregation inside bacterial inclusion bodies, thus illustrating a beneficial effect of metal coordination.
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http://dx.doi.org/10.1021/acs.inorgchem.0c00467DOI Listing
May 2020

On the Binding of Congo Red to Amyloid Fibrils.

Angew Chem Int Ed Engl 2020 05 19;59(21):8104-8107. Epub 2020 Feb 19.

Department of Pharmacy and Pharmaceutical Technology and Physical-Chemistry, School of Pharmacy and Food Sciences, University of Barcelona, Joan XXIII, 27-31, 08028, Barcelona, Spain.

Amyloids are characterized by their capacity to bind Congo red (CR), one of the most used amyloid-specific dyes. The structural features of CR binding were unknown for years, mainly because of the lack of amyloid structures solved at high resolution. In the last few years, solid-state NMR spectroscopy enabled the determination of the structural features of amyloids, such as the HET-s prion forming domain (HET-s PFD), which also has recently been used to determine the amyloid-CR interface at atomic resolution. Herein, we combine spectroscopic data with molecular docking, molecular dynamics, and excitonic quantum/molecular mechanics calculations to examine and rationalize CR binding to amyloids. In contrast to a previous assumption on the binding mode, our results suggest that CR binding to the HET-s PFD involves a cooperative process entailing the formation of a complex with 1:1 stoichiometry. This provides a molecular basis to explain the bathochromic shift in the maximal absorbance wavelength when CR is bound to amyloids.
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http://dx.doi.org/10.1002/anie.201916630DOI Listing
May 2020

A novel class of multitarget anti-Alzheimer benzohomoadamantane‒chlorotacrine hybrids modulating cholinesterases and glutamate NMDA receptors.

Eur J Med Chem 2019 Oct 17;180:613-626. Epub 2019 Jul 17.

Laboratory of Pharmaceutical Chemistry (CSIC Associated Unit), Faculty of Pharmacy and Food Sciences, Institute of Biomedicine (IBUB), University of Barcelona, Av. Joan XXIII 27-31, E-08028, Barcelona, Spain. Electronic address:

The development of multitarget compounds against multifactorial diseases, such as Alzheimer's disease, is an area of very intensive research, due to the expected superior therapeutic efficacy that should arise from the simultaneous modulation of several key targets of the complex pathological network. Here we describe the synthesis and multitarget biological profiling of a new class of compounds designed by molecular hybridization of an NMDA receptor antagonist fluorobenzohomoadamantanamine with the potent acetylcholinesterase (AChE) inhibitor 6-chlorotacrine, using two different linker lengths and linkage positions, to preserve or not the memantine-like polycyclic unsubstituted primary amine. The best hybrids exhibit greater potencies than parent compounds against AChE (IC 0.33 nM in the best case, 44-fold increased potency over 6-chlorotacrine), butyrylcholinesterase (IC 21 nM in the best case, 24-fold increased potency over 6-chlorotacrine), and NMDA receptors (IC 0.89 μM in the best case, 2-fold increased potency over the parent benzohomoadamantanamine and memantine), which suggests an additive effect of both pharmacophoric moieties in the interaction with the primary targets. Moreover, most of these compounds have been predicted to be brain permeable. This set of biological properties makes them promising leads for further anti-Alzheimer drug development.
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http://dx.doi.org/10.1016/j.ejmech.2019.07.051DOI Listing
October 2019

Bacterial Inclusion Bodies for Anti-Amyloid Drug Discovery: Current and Future Screening Methods.

Curr Protein Pept Sci 2019 ;20(6):563-576

Institute of Nanoscience and Nanotechnology (IN2UB), University of Barcelona, E-08028 Barcelona, Spain.

Amyloid aggregation is linked to an increasing number of human disorders from nonneurological pathologies such as type-2 diabetes to neurodegenerative ones such as Alzheimer or Parkinson's diseases. Thirty-six human proteins have shown the capacity to aggregate into pathological amyloid structures. To date, it is widely accepted that amyloid folding/aggregation is a universal process present in eukaryotic and prokaryotic cells. In the last decade, several studies have unequivocally demonstrated that bacterial inclusion bodies - insoluble protein aggregates usually formed during heterologous protein overexpression in bacteria - are mainly composed of overexpressed proteins in amyloid conformation. This fact shows that amyloid-prone proteins display a similar aggregation propensity in humans and bacteria, opening the possibility to use bacteria as simple models to study amyloid aggregation process and the potential effect of both anti-amyloid drugs and pro-aggregative compounds. Under these considerations, several in vitro and in cellulo methods, which exploit the amyloid properties of bacterial inclusion bodies, have been proposed in the last few years. Since these new methods are fast, simple, inexpensive, highly reproducible, and tunable, they have aroused great interest as preliminary screening tools in the search for anti-amyloid (beta-blocker) drugs for conformational diseases. The aim of this mini-review is to compile recently developed methods aimed at tracking amyloid aggregation in bacteria, discussing their advantages and limitations, and the future potential applications of inclusion bodies in anti-amyloid drug discovery.
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http://dx.doi.org/10.2174/1389203720666190329120007DOI Listing
August 2019

Amyloid Pan-inhibitors: One Family of Compounds To Cope with All Conformational Diseases.

ACS Chem Neurosci 2019 03 14;10(3):1311-1317. Epub 2018 Nov 14.

Department of Pharmacy and Pharmaceutical Technology and Physical-Chemistry, Faculty of Pharmacy and Food Sciences , University of Barcelona , E-08028 Barcelona , Spain.

Amyloids are ubiquitous protein aggregates sharing common internal structural features; they are present in all organisms, from prokaryotes to eukaryotes, where they play physiological or pathological roles. Importantly, amyloids, which are generated by aggregation of a range of distinct proteins, could be a key factor in a number of major human disorders, the so-called conformational diseases. Because all amyloids exhibit similar cross-β motifs, one may envisage that molecules capable of blocking the formation of β-sheet structures could abolish aggregation of all amyloid proteins, albeit with different efficacies. Herein, two different β-sheet blockers were tested against a selection of amyloidogenic proteins, encompassing all the major types of amyloid-based disorders. Analysis of their blocking efficiency, using a simple but contrasted cell-based screening procedure, unequivocally confirms that they indeed behave as aggregation pan-inhibitors. The significant inhibitory effects observed for these compounds against all tested amyloidogenic proteins could spur a broader biological evaluation of other known and new amyloid aggregation inhibitors to further determine the potential use of this class of compounds for the universal treatment of conformational diseases.
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http://dx.doi.org/10.1021/acschemneuro.8b00398DOI Listing
March 2019

Design, synthesis and multitarget biological profiling of second-generation anti-Alzheimer rhein-huprine hybrids.

Future Med Chem 2017 06 20;9(10):965-981. Epub 2017 Jun 20.

Laboratory of Pharmaceutical Chemistry (CSIC Associated Unit), Faculty of Pharmacy & Food Sciences, University of Barcelona, Av. Joan XXIII 27-31, E-08028 Barcelona, Spain.

Aim: Simultaneous modulation of several key targets of the pathological network of Alzheimer's disease (AD) is being increasingly pursued as a promising option to fill the critical gap of efficacious drugs against this condition.

Materials & Methods: A short series of compounds purported to hit multiple targets of relevance in AD has been designed, on the basis of their distinct basicities estimated from high-level quantum mechanical computations, synthesized, and subjected to assays of inhibition of cholinesterases, BACE-1, and Aβ42 and tau aggregation, of antioxidant activity, and of brain permeation.

Results: Using, as a template, a lead rhein-huprine hybrid with an interesting multitarget profile, we have developed second-generation compounds, designed by the modification of the huprine aromatic ring. Replacement by [1,8]-naphthyridine or thieno[3,2-e]pyridine systems resulted in decreased, although still potent, acetylcholinesterase or BACE-1 inhibitory activities, which are more balanced relative to their Aβ42 and tau antiaggregating and antioxidant activities.

Conclusion: Second-generation naphthyridine- and thienopyridine-based rhein-huprine hybrids emerge as interesting brain permeable compounds that hit several crucial pathogenic factors of AD.
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http://dx.doi.org/10.4155/fmc-2017-0049DOI Listing
June 2017

Evidence of Protein Adsorption in Pegylated Liposomes: Influence of Liposomal Decoration.

Nanomaterials (Basel) 2017 Feb 10;7(2). Epub 2017 Feb 10.

Department of Pharmacy, Pharmaceutical Technology and Physical Chemistry, Faculty of Pharmacy and Food Sciences, University of Barcelona, Avda. Joan XXIII, 27-31, 08028 Barcelona, Spain.

In order to contribute to a better knowledge of the events involved in the formation of the protein corona when nanoparticles (NPs) come in contact with proteins, we report a study about the changes on the physicochemical properties of pristine, PEGylated and Cyclic Arginine-Glycine-Aspartate peptide (RGD)-functionalized large unilamelar liposomes (LUVs) or magnetoliposomes (MLs) upon incubation with Bovine Serum Albumin (BSA). The main phospholipid component of both LUVs and MLs was l-α-phosphatydylcholine (PC) or 1,2-dimyristoyl--glycero-3-phosphocholine (DMPC) with 20% of cholesterol. The most obvious indication of the interaction of BSA-nanosystems is given by changes in the hydrodynamic diameter of the particles but other evidence is needed to corroborate the process. Our findings indicate that size modification is a process that is accomplished in few hours and that is strongly dependent not only on the surface decoration but also of the lipid composition of both LUVs and MLs. Fluorescence quenching experiments as well as cryogenic transmission electron microscopy (Cryo-TEM) images assessed these changes and confirmed that although each system has to be studied in a particular way, we can establish three distinctive features that turn into more reactive systems: (a) compositions containing PC compared with their DMPC counterparts; (b) the presence of PEG and/or RGD compared to the pristine counterparts; and (c) the presence of SPIONs: MLs show higher interaction than LUVs of the same lipid composition. Consequently, PEGylation (that is supposed to make stealth NPs) actually fails in preventing complete protein binding.
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http://dx.doi.org/10.3390/nano7020037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5333022PMC
February 2017

Combined in Vitro Cell-Based/in Silico Screening of Naturally Occurring Flavonoids and Phenolic Compounds as Potential Anti-Alzheimer Drugs.

J Nat Prod 2017 02 27;80(2):278-289. Epub 2017 Jan 27.

Department of Pharmacy and Pharmaceutical Technology and Physical-Chemistry, School of Pharmacy, and Institute of Nanoscience and Nanotechnology (IN2UB), University of Barcelona , E-08028, Barcelona, Spain.

Alzheimer's disease (AD) is the main cause of dementia in people over 65 years. One of the major culprits in AD is the self-aggregation of amyloid-β peptide (Aβ), which has stimulated the search for small molecules able to inhibit Aβ aggregation. In this context, we recently reported a simple, but effective in vitro cell-based assay to evaluate the potential antiaggregation activity of putative Aβ aggregation inhibitors. In this work this assay was used together with docking and molecular dynamics simulations to analyze the anti-Aβ aggregation activity of several naturally occurring flavonoids and phenolic compounds. The results showed that rosmarinic acid, melatonin, and o-vanillin displayed zero or low inhibitory capacity, curcumin was found to have an intermediate inhibitory potency, and apigenin and quercetin showed potent antiaggregation activity. Finally, the suitability of the combined in vitro cell-based/in silico approach to distinguish between active and inactive compounds was further assessed for an additional set of flavonols and dihydroflavonols.
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http://dx.doi.org/10.1021/acs.jnatprod.6b00643DOI Listing
February 2017

Key Points Concerning Amyloid Infectivity and Prion-Like Neuronal Invasion.

Front Mol Neurosci 2016 22;9:29. Epub 2016 Apr 22.

Faculty of Pharmacy, Laboratory of Conformational Diseases, Department of Physical Chemistry, Institute of Nanoscience and Nanotechnology, University of Barcelona Barcelona, Spain.

Amyloid aggregation has been related to an increasing number of human illnesses, from Alzheimer's and Parkinson's diseases (AD/PD) to Creutzfeldt-Jakob disease. Commonly, only prions have been considered as infectious agents with a high capacity of propagation. However, recent publications have shown that many amyloid proteins, including amyloid β-peptide, α-synuclein (α-syn) and tau protein, also propagate in a "prion-like" manner. Meanwhile, no link between propagation of pathological proteins and neurotoxicity has been demonstrated. The extremely low infectivity under natural conditions of most non-prion amyloids is far below the capacity to spread exhibited by prions. Nonetheless, it is important to elucidate the key factors that cause non-prion amyloids to become infectious agents. In recent years, important advances in our understanding of the amyloid processes of amyloid-like proteins and unrelated prions (i.e., yeast and fungal prions) have yielded essential information that can shed light on the prion phenomenon in mammals and humans. As shown in this review, recent evidence suggests that there are key factors that could dramatically modulate the prion capacity of proteins in the amyloid conformation. The concentration of nuclei, the presence of oligomers, and the toxicity, resistance and localization of these aggregates could all be key factors affecting their spread. In short, those factors that favor the high concentration of extracellular nuclei or oligomers, characterized by small size, with a low toxicity could dramatically increase prion propensity; whereas low concentrations of highly toxic intracellular amyloids, with a large size, would effectively prevent infectivity.
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http://dx.doi.org/10.3389/fnmol.2016.00029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4840800PMC
May 2016

Histidine-Rich Oligopeptides To Lessen Copper-Mediated Amyloid-β Toxicity.

Chemistry 2016 05 13;22(21):7268-80. Epub 2016 Apr 13.

Department of Inorganic and Organic Chemistry, University of Barcelona, Martí i Franquès 1-11, 08028, Barcelona, Spain.

Brain copper imbalance plays an important role in amyloid-β aggregation, tau hyperphosphorylation, and neurotoxicity observed in Alzheimer's disease (AD). Therefore, the administration of biocompatible metal-binding agents may offer a potential therapeutic solution to target mislocalized copper ions and restore metallostasis. Histidine-containing peptides and proteins are excellent metal binders and are found in many natural systems. The design of short peptides showing optimal binding properties represents a promising approach to capture and redistribute mislocalized metal ions, mainly due to their biocompatibility, ease of synthesis, and the possibility of fine-tuning their metal-binding affinities in order to suppress unwanted competitive binding with copper-containing proteins. In the present study, three peptides, namely HWH, HK(C) H, and HAH, have been designed with the objective of reducing copper toxicity in AD. These tripeptides form highly stable albumin-like complexes, showing higher affinity for Cu(II) than that of Aβ(1-40). Furthermore, HWH, HK(C) H, and HAH act as very efficient inhibitors of copper-mediated reactive oxygen species (ROS) generation and prevent the copper-induced overproduction of toxic oligomers in the initial steps of amyloid aggregation in the presence of Cu(II) ions. These tripeptides, and more generally small peptides including the sequence His-Xaa-His at the N-terminus, may therefore be considered as promising motifs for the future development of new and efficient anti-Alzheimer drugs.
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http://dx.doi.org/10.1002/chem.201600286DOI Listing
May 2016

Natural Xanthones from Garcinia mangostana with Multifunctional Activities for the Therapy of Alzheimer's Disease.

Neurochem Res 2016 Jul 2;41(7):1806-17. Epub 2016 Apr 2.

School of Pharmaceutical Sciences, Sun Yat-Sen University, Guangzhou, 510006, China.

Natural xanthones have diversity pharmacological activities. Here, a series of xanthones isolated from the pericarps of Garcinia mangostana Linn, named α-Mangostin, 8-Deoxygartanin, Gartanin, Garciniafuran, Garcinone C, Garcinone D, and γ-Mangostin were investigated. Biological screening performed in vitro and in Escherichia coli cells indicated that most of the xanthones exhibited significant inhibition of self-induced β-amyloid (Aβ) aggregation and also β-site amyloid precursor protein-cleaving enzyme 1, acted as potential antioxidants and biometal chelators. Among these compounds, α-Mangostin, Gartanin, Garcinone C and γ-Mangostin showed better antioxidant properties to scavenge Diphenyl-1-(2,4,6-trinitrophenyl) hydrazyl (DPPH) free radical than Trolox, and potent neuroprotective effects against glutamate-induced HT22 cell death partly by up-regulating HO-1 protein level and then scavenging reactive oxygen species. Moreover, Gartanin, Garcinone C and γ-Mangostin could be able to penetrate the blood-brain barrier (BBB) in vitro. These findings suggest that the natural xanthones have multifunctional activities against Alzheimer's disease (AD) and could be promising compounds for the therapy of AD.
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http://dx.doi.org/10.1007/s11064-016-1896-yDOI Listing
July 2016

Ultra rapid in vivo screening for anti-Alzheimer anti-amyloid drugs.

Sci Rep 2016 Mar 22;6:23349. Epub 2016 Mar 22.

Department of Physical Chemistry, School of Pharmacy, and Institute of Nanoscience and Nanotechnology (IN2UB), University of Barcelona, Spain.

More than 46 million people worldwide suffer from Alzheimer's disease. A large number of potential treatments have been proposed; among these, the inhibition of the aggregation of amyloid β-peptide (Aβ), considered one of the main culprits in Alzheimer's disease. Limitations in monitoring the aggregation of Aβ in cells and tissues restrict the screening of anti-amyloid drugs to in vitro studies in most cases. We have developed a simple but powerful method to track Aβ aggregation in vivo in real-time, using bacteria as in vivo amyloid reservoir. We use the specific amyloid dye Thioflavin-S (Th-S) to stain bacterial inclusion bodies (IBs), in this case mainly formed of Aβ in amyloid conformation. Th-S binding to amyloids leads to an increment of fluorescence that can be monitored. The quantification of the Th-S fluorescence along the time allows tracking Aβ aggregation and the effect of potential anti-aggregating agents.
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http://dx.doi.org/10.1038/srep23349DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4802339PMC
March 2016

Magnetic Nanoparticles Cross the Blood-Brain Barrier: When Physics Rises to a Challenge.

Nanomaterials (Basel) 2015 Dec 11;5(4):2231-2248. Epub 2015 Dec 11.

Department of Physical Chemistry, Faculty of Pharmacy, University of Barcelona and Institute of Nanoscience and Nanotechnology (IN2UB), Avda. Joan XXIII, 08028 Barcelona, Spain.

The blood-brain barrier is a physical and physiological barrier that protects the brain from toxic substances within the bloodstream and helps maintain brain homeostasis. It also represents the main obstacle in the treatment of many diseases of the central nervous system. Among the different approaches employed to overcome this barrier, the use of nanoparticles as a tool to enhance delivery of therapeutic molecules to the brain is particularly promising. There is special interest in the use of magnetic nanoparticles, as their physical characteristics endow them with additional potentially useful properties. Following systemic administration, a magnetic field applied externally can mediate the capacity of magnetic nanoparticles to permeate the blood-brain barrier. Meanwhile, thermal energy released by magnetic nanoparticles under the influence of radiofrequency radiation can modulate blood-brain barrier integrity, increasing its permeability. In this review, we present the strategies that use magnetic nanoparticles, specifically iron oxide nanoparticles, to enhance drug delivery to the brain.
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http://dx.doi.org/10.3390/nano5042231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5304810PMC
December 2015

Amyloids in solid-state nuclear magnetic resonance: potential causes of the usually low resolution.

Int J Nanomedicine 2015 9;10:6975-83. Epub 2015 Nov 9.

Department of Physical Chemistry, School of Pharmacy, Institute of Nanoscience and Nanotechnology (IN UB), University of Barcelona, Barcelona, Spain.

Amyloids are non-crystalline and insoluble, which imply that the classical structural biology tools, ie, X-ray crystallography and solution nuclear magnetic resonance (NMR), are not suitable for their analysis. In the last years, solid-state NMR (ssNMR) has emerged as an alternative tool to decrypt the structural signatures of amyloid fibrils, providing major contributions to our understanding of molecular structures of amyloids such as β-amyloid peptide associated with Alzheimer's disease or fungal prions, among others. Despite this, the wide majority of amyloid fibrils display low resolution by ssNMR. Usually, this low resolution has been attributed to a high disorder or polymorphism of the fibrils, suggesting the existence of diverse elementary β-sheet structures. Here, we propose that a single β-sheet structure could be responsible for the broadening of the line widths in the ssNMR spectra. Although the fibrils and fibers consist of a single elementary structure, the angle of twist of each individual fibril in the mature fiber depends on the number of individual fibrils as well as the fibril arrangement in the final mature fiber. Thus, a wide range of angles of twist could be observed in the same amyloid sample. These twist variations involve changes in amino acid alignments that could be enough to limit the ssNMR resolution.
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http://dx.doi.org/10.2147/IJN.S89385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4646584PMC
September 2016

In vivo amyloid aggregation kinetics tracked by time-lapse confocal microscopy in real-time.

Biotechnol J 2016 Jan 5;11(1):172-7. Epub 2016 Jan 5.

Departament de Fisicoquímica, Facultat de Farmàcia, Universitat de Barcelona, Barcelona, Spain.

Amyloid polymerization underlies an increasing number of human diseases. Despite this process having been studied extensively in vitro, aggregation is a difficult process to track in vivo due to methodological limitations and the slow kinetics of aggregation reactions in cells and tissues. Herein we exploit the amyloid properties of the inclusions bodies (IBs) formed by amyloidogenic proteins in bacteria to address the kinetics of in vivo amyloid aggregation. To this aim we used time-lapse confocal microscopy and a fusion of the amyloid-beta peptide (A β42) with a fluorescent reporter. This strategy allowed us to follow the intracellular kinetics of amyloid-like aggregation in real-time and to discriminate between variants exhibiting different in vivo aggregation propensity. Overall, the approach opens the possibility to assess the impact of point mutations as well as potential anti-aggregation drugs in the process of amyloid formation in living cells.
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http://dx.doi.org/10.1002/biot.201500252DOI Listing
January 2016

Novel Levetiracetam Derivatives That Are Effective against the Alzheimer-like Phenotype in Mice: Synthesis, in Vitro, ex Vivo, and in Vivo Efficacy Studies.

J Med Chem 2015 Aug 27;58(15):6018-32. Epub 2015 Jul 27.

†Laboratori de Química Farmacèutica (Unitat Associada al CSIC), Facultat de Farmàcia, and Institut de Biomedicina (IBUB), Universitat de Barcelona, Av. Joan XXIII 27-31, E-08028, Barcelona, Spain.

We have synthesized a series of heptamethylene-linked levetiracetam-huprine and levetiracetam-(6-chloro)tacrine hybrids to hit amyloid, tau, and cholinergic pathologies as well as β-amyloid (Aβ)-induced epileptiform activity, some of the mechanisms that eventually lead to cognitive deficits in Alzheimer's disease patients. These hybrids are potent inhibitors of human acetylcholinesterase and butyrylcholinesterase in vitro and moderately potent Aβ42 and tau antiaggregating agents in a simple E. coli model of amyloid aggregation. Ex vivo determination of the brain acetylcholinesterase inhibitory activity of these compounds after intraperitoneal injection to C57BL6J mice has demonstrated their ability to enter the brain. The levetiracetam-huprine hybrid 10 significantly reduced the incidence of epileptic seizures, cortical amyloid burden, and neuroinflammation in APP/PS1 mice after a 4-week treatment with a 5 mg/kg dose. Moreover, the hybrid 10 rescued transgenic mice from cognitive deficits, thereby emerging as an interesting disease-modifying anti-Alzheimer drug candidate.
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http://dx.doi.org/10.1021/acs.jmedchem.5b00624DOI Listing
August 2015

Could α-synuclein amyloid-like aggregates trigger a prionic neuronal invasion?

Biomed Res Int 2015 19;2015:172018. Epub 2015 Mar 19.

Department of Physical Chemistry, Faculty of Pharmacy, University of Barcelona, and Institute of Nanoscience and Nanotechnology of the University of Barcelona (IN2UB), Avenue Joan XXIII 27-31, Barcelona, 08028 Catalonia, Spain.

Parkinson's disease (PD), a progressive neurodegenerative disease primarily affecting voluntary and controlled movement, is characterized by abnormal accumulations of α-synuclein (α-syn) in intraneuronal Lewy bodies. In the last years, the increased number of evidences from both the in vitro and in vivo studies has shown the ability of α-syn to misfold in amyloid conformations and to spread via neuron-to-neuron transmission, suggesting a prion-like behaviour. However, in contrast to prion protein (PrP), α-syn transmission is far from neuronal invasion. The high neuronal toxicity of both mature fibres and oligomeric species, as well as the intracellular localization of the protein and the difficulty to be secreted, could be key factors impeding the prion ability of α-syn aggregates.
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http://dx.doi.org/10.1155/2015/172018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4383319PMC
December 2015

Predicting the aggregation propensity of prion sequences.

Virus Res 2015 Sep 6;207:127-35. Epub 2015 Mar 6.

Department of Physical Chemistry, Faculty of Pharmacy, University of Barcelona, Avda. Joan XXIII 27-31, E-08028 Barcelona, Spain; Institute of Nanoscience and Nanotechnology of the University of Barcelona (IN(2)UB), Spain. Electronic address:

The presence of prions can result in debilitating and neurodegenerative diseases in mammals and protein-based genetic elements in fungi. Prions are defined as a subclass of amyloids in which the self-aggregation process becomes self-perpetuating and infectious. Like all amyloids, prions polymerize into fibres with a common core formed of β-sheet structures oriented perpendicular to the fibril axes which form a structure known as a cross-β structure. The intermolecular β-sheet propensity, a characteristic of the amyloid pattern, as well as other key parameters of amyloid fibril formation can be predicted. Mathematical algorithms have been proposed to predict both amyloid and prion propensities. However, it has been shown that the presence of amyloid-prone regions in a polypeptide sequence could be insufficient for amyloid formation. It has also often been stated that the formation of amyloid fibrils does not imply that these are prions. Despite these limitations, in silico prediction of amyloid and prion propensities should help detect potential new prion sequences in mammals. In addition, the determination of amyloid-prone regions in prion sequences could be very useful in understanding the effect of sporadic mutations and polymorphisms as well as in the search for therapeutic targets.
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http://dx.doi.org/10.1016/j.virusres.2015.03.001DOI Listing
September 2015

Shogaol-huprine hybrids: dual antioxidant and anticholinesterase agents with β-amyloid and tau anti-aggregating properties.

Bioorg Med Chem 2014 Oct 7;22(19):5298-307. Epub 2014 Aug 7.

Laboratori de Química Farmacèutica (Unitat Associada al CSIC), Facultat de Farmàcia, and Institut de Biomedicina (IBUB), Universitat de Barcelona, Av. Joan XXIII, 27-31, E-08028 Barcelona, Spain. Electronic address:

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 [4]-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.
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http://dx.doi.org/10.1016/j.bmc.2014.07.053DOI Listing
October 2014

Tetrahydrobenzo[h][1,6]naphthyridine-6-chlorotacrine hybrids as a new family of anti-Alzheimer agents targeting β-amyloid, tau, and cholinesterase pathologies.

Eur J Med Chem 2014 Sep 7;84:107-17. Epub 2014 Jul 7.

Laboratori de Química Farmacèutica (Unitat Associada al CSIC), Facultat de Farmàcia, and Institut de Biomedicina (IBUB), Universitat de Barcelona, Av. Joan XXIII, 27-31, E-08028 Barcelona, Spain. Electronic address:

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.
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http://dx.doi.org/10.1016/j.ejmech.2014.07.021DOI Listing
September 2014

Screening for amyloid aggregation: in-silico, in-vitro and in-vivo detection.

Curr Protein Pept Sci 2014 ;15(5):477-89

Departament de Bioquímica i Biologia Molecular, Facultat de Biociències, Universitat Autònoma de Barcelona, E-08193 Bellaterra, Spain.

Protein misfolding and aggregation into amyloid structures is linked with an increasing number of nonneuropathic (either localized or systemic) and neurodegenerative human disorders. In the present review, we compile and describe methods, which have been developed to predict, detect and characterize amyloid and amyloid-like protein deposits. We focus in the state-of-the-art methodologies to study and image amyloid aggregation in-vitro, from qualitative and low-resolution techniques to methods addressed to resolve protein structures at atomic level. We also recapitulate the most relevant literature describing approaches that have been demonstrated to be able to detect and characterize protein aggregation in cells and living organisms, as well as methodologies to report cytotoxicity associated to amyloid formation. Overall, the aim of this review is to illustrate computational and experimental methods to characterize and predict in-vitro and in-vivo amyloid aggregation, providing the readers valuable information to elect the most appropriate techniques at their convenience.
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http://dx.doi.org/10.2174/1389203715666140221101038DOI Listing
June 2015

Discovery of novel inhibitors of amyloid β-peptide 1-42 aggregation.

J Med Chem 2012 Nov 22;55(22):9521-30. Epub 2012 Oct 22.

Departamento de Bioquímica y Biología Molecular y Celular, Facultad de Ciencias, Universidad de Zaragoza, Pedro Cerbuna 12, 50009 Zaragoza, Spain.

Alzheimer's disease, characterized by deposits of amyloid β-peptide (Aβ), is the most common neurodegenerative disease, but it still lacks a specific treatment. We have discovered five chemically unrelated inhibitors of the in vitro aggregation of the Aβ17-40 peptide by screening two commercial chemical libraries. Four of them (1-4) exhibit relatively low MCCs toward HeLa cells (17-184 μM). The usefulness of compounds 1-4 to inhibit the in vivo aggregation of Aβ1-42 has been demonstrated using two fungi models, Saccharomyces cerevisiae and Podospora anserina, previously transformed to express Aβ1-42. Estimated IC(50)s are around 1-2 μM. Interestingly, addition of any of the four compounds to sonicated preformed P. anserina aggregates completely inhibited the appearance of SDS-resistant oligomers. This combination of HTP in vitro screening with validation in fungi models provides an efficient way to identify novel inhibitory compounds of Aβ1-42 aggregation for subsequent testing in animal models.
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http://dx.doi.org/10.1021/jm301186pDOI Listing
November 2012

Thioflavin-S staining coupled to flow cytometry. A screening tool to detect in vivo protein aggregation.

Mol Biosyst 2012 Nov 7;8(11):2839-44. Epub 2012 Aug 7.

Institut de Biotecnologia i Biomedicina, Universitat Autònoma de Barcelona, 08193-Bellaterra, Barcelona, Spain.

Amyloid deposits are associated with an increasing number of human disorders, including Alzheimer's and Parkinson's diseases. Recent studies provide compelling evidence for the existence of amyloid-like conformations in the insoluble bacterial inclusion bodies (IBs) produced during the recombinant expression of amyloidogenic proteins. This makes prokaryotic cells a physiologically relevant system to study the mechanisms of in vivo amyloid deposition. We show here that the application of flow cytometry to detect Thioflavin-S (Th-S) fluorescence provides a fast, robust, quantitative, non-invasive method to screen for the presence of in vivo intracellular amyloid-like aggregates in bacteria, with potential application in the analysis of the impact of genetic mutations or chemical compounds on the aggregation of disease-associated polypeptides.
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http://dx.doi.org/10.1039/c2mb25214gDOI Listing
November 2012

Yeast prions form infectious amyloid inclusion bodies in bacteria.

Microb Cell Fact 2012 Jun 25;11:89. Epub 2012 Jun 25.

Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, Bellaterra, Spain.

Background: Prions were first identified as infectious proteins associated with fatal brain diseases in mammals. However, fungal prions behave as epigenetic regulators that can alter a range of cellular processes. These proteins propagate as self-perpetuating amyloid aggregates being an example of structural inheritance. The best-characterized examples are the Sup35 and Ure2 yeast proteins, corresponding to [PSI+] and [URE3] phenotypes, respectively.

Results: Here we show that both the prion domain of Sup35 (Sup35-NM) and the Ure2 protein (Ure2p) form inclusion bodies (IBs) displaying amyloid-like properties when expressed in bacteria. These intracellular aggregates template the conformational change and promote the aggregation of homologous, but not heterologous, soluble prionogenic molecules. Moreover, in the case of Sup35-NM, purified IBs are able to induce different [PSI+] phenotypes in yeast, indicating that at least a fraction of the protein embedded in these deposits adopts an infectious prion fold.

Conclusions: An important feature of prion inheritance is the existence of strains, which are phenotypic variants encoded by different conformations of the same polypeptide. We show here that the proportion of infected yeast cells displaying strong and weak [PSI+] phenotypes depends on the conditions under which the prionogenic aggregates are formed in E. coli, suggesting that bacterial systems might become useful tools to generate prion strain diversity.
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http://dx.doi.org/10.1186/1475-2859-11-89DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3520751PMC
June 2012

Native structure protects SUMO proteins from aggregation into amyloid fibrils.

Biomacromolecules 2012 Jun 25;13(6):1916-26. Epub 2012 May 25.

Institut de Biotecnologia i de Biomedicina, Universitat Autònoma de Barcelona, 08193 Bellaterra (Barcelona), Spain.

SUMO proteins belong to the Ubiquitin-like protein family, all sharing a common fold and a similar mechanism of conjugation to target polypeptides. SUMO is ubiquitous in all eukaryotes and participates in many crucial pathways. Native SUMO proteins are highly soluble, a property that is exploited in biotechnology. Moreover, SUMO regulates the solubility of aggregation-prone proteins in neurodegenerative disorders. Despite these properties, we show here that human SUMO1, SUMO2, and SUMO3 proteins are at risk of aggregation into amyloid structures if their native conformation is perturbed. Aggregation is mediated by specific regions, which overlap with SUMO functional interfaces, illustrating a competition between function and aggregation. Aggregation of SUMOs might have important physiological implications because disruption of the SUMO pathway is lethal in different organisms. It appears that functional constraints make it difficult to avoid the competition between productive folding and deleterious aggregation in globular proteins, even for essential polypeptides.
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http://dx.doi.org/10.1021/bm3004385DOI Listing
June 2012
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