Kororamides, convolutamines, and indole derivatives as possible tau and dual specificity kinases inhibitors for Alzheimer's Disease

Alzheimer's disease (AD) is becoming one of the most disturbing health and socioeconomic problems nowadays, as it is a neurodegenerative pathology with no treatment, which is expected to grow further due to population ageing. Actual treatments for AD produce only a modest amelioration of symptoms, although there is a constant ongoing research of new therapeutic strategies oriented to improve the amelioration of the symptoms, and even to completely cure the disease. A principal feature of AD is the presence of neurofibrillary tangles (NFT) induced by the aberrant phosphorylation of the microtubule-associated protein tau in the brains of affected individuals. Glycogen synthetase kinase-3 beta (GSK3β), casein kinase 1 delta (CK1δ), dual-specificity tyrosine phosphorylation regulated kinase 1A (DYRK1A) and dual-specificity kinase cdc2-like kinase 1 (CLK1) have been identified as the principal proteins involved in this process. Due to this, the inhibition of these kinases has been proposed as a plausible therapeutic strategy to fight AD. In this study, we tested in silico the inhibitory activity of different marine natural compounds, as well as newly-designed molecules from some of them, over the mentioned protein kinases, finding some new possible inhibitors with potential therapeutic application

Meridianins and Lignarenone B as Potential GSK3β Inhibitors and Inductors of Structural Neuronal Plasticity

Glycogen Synthase Kinase 3 (GSK3) is an essential protein, with a relevant role in many diseases such as diabetes, cancer and neurodegenerative disorders. Particularly, the isoform GSK3β is related to pathologies such as Alzheimer's disease (AD). This enzyme constitutes a very interesting target for the discovery and/or design of new therapeutic agents against AD due to its relation to the hyperphosphorylation of the microtubule-associated protein tau (MAPT), and therefore, its contribution to neurofibrillary tangles (NFT) formation. An in silico target profiling study identified two marine molecular families, the indole alkaloids meridianins from the tunicate genus Aplidium, and lignarenones, the secondary metabolites of the shelled cephalaspidean mollusc Scaphander lignarius, as possible GSK3β inhibitors. The analysis of the surface of GSK3β, aimed to find possible binding regions, and the subsequent in silico binding studies revealed that both marine molecular families can act over the ATP and/or substrate binding regions. The predicted inhibitory potential of the molecules from these two chemical families was experimentally validated in vitro by showing a ~50% of increased Ser9 phosphorylation levels of the GSK3β protein. Furthermore, we determined that molecules from both molecular families potentiate structural neuronal plasticity in vitro. These results allow us to suggest that meridianins and lignarenone B could be used as possible therapeutic candidates for the treatment of GSK3β involved pathologies, such as AD

Silibinin is a direct inhibitor of STAT3.

We herein combined experimental and computational efforts to delineate the mechanism of action through which the flavonolignan silibinin targets STAT3. Silibinin reduced IL-6 inducible, constitutive, and acquired feedback activation of STAT3 at tyrosine 705 (Y705). Silibinin attenuated the inducible phospho-activation of Y705 in GFP-STAT3 genetic fusions without drastically altering the kinase activity of the STAT3 upstream kinases JAK1 and JAK2. A comparative computational study based on docking and molecular dynamics simulation over 14 different STAT3 inhibitors (STAT3i) predicted that silibinin could directly bind with high affinity to both the Src homology-2 (SH2) domain and the DNA-binding domain (DBD) of STAT3. Silibinin partially overlapped with the cavity occupied by other STAT3i in the SH2 domain to indirectly prevent Y705 phosphorylation, yet showing a unique binding mode. Moreover, silibinin was the only STAT3i predicted to establish direct interactions with DNA in its targeting to the STAT3 DBD. The prevention of STAT3 nuclear translocation, the blockade of the binding of activated STAT3 to its consensus DNA sequence, and the suppression of STAT3-directed transcriptional activity confirmed silibinin as a direct STAT3i. The unique characteristics of silibinin as a bimodal SH2- and DBD-targeting STAT3i make silibinin a promising lead for designing new, more effective STAT3i.This work was supported by grants from the Ministerio de Ciencia e Innovacion (Grant SAF2016-80639-P to J. A. Menendez), Plan Nacional de I + D + I, Spain, and the Agencia de Gestio d'Ajuts Universitaris i de Recerca (AGAUR) (Grant 2014 SGR229 to J. A. Menendez). This study was supported also by unrestricted research grants from Roche Pharma (Spain) and Astellas Pharma (Spain) to the Program Against Cancer Therapeutic Resistance (ProCURE, Catalan Institute of Oncology). Joaquim Bosch-Barrera is supported by SEOM, Pfizer (Grant WI190764), Boehringer Ingelheim, Meda Pharma, and Pla strategic de recerca i innovacio en salut 2016-2020 de la Generalitat de Catalunya (SLT006/17/114). Elisabet Cuyas is supported by a Sara Borrell postdoctoral contract (CD15/00033) from the Ministerio de Sanidad y Consumo, Fondo de Investigacion Sanitaria (FIS), Spain. The authors would like to thank Dr. Kenneth McCreath for editorial support.S

Metformin Is a Direct SIRT1-Activating Compound: Computational Modeling and Experimental Validation

Metformin has been proposed to operate as an agonist of SIRT1, a nicotinamide adenine dinucleotide (NAD(+))-dependent deacetylase that mimics most of the metabolic responses to calorie restriction. Herein, we present an in silico analysis focusing on the molecular docking and dynamic simulation of the putative interactions between metformin and SIRT1. Using eight different crystal structures of human SIRT1 protein, our computational approach was able to delineate the putative binding modes of metformin to several pockets inside and outside the central deacetylase catalytic domain. First, metformin was predicted to interact with the very same allosteric site occupied by resveratrol and other sirtuin-activating compounds (STATCs) at the amino-terminal activation domain of SIRT1. Second, metformin was predicted to interact with the NAD(+) binding site in a manner slightly different to that of SIRT1 inhibitors containing an indole ring. Third, metformin was predicted to interact with the C-terminal regulatory segment of SIRT1 bound to the NAD(+) hydrolysis product ADP-ribose, a "C-pocket"-related mechanism that appears to be essential for mechanism-based activation of SIRT1. Enzymatic assays confirmed that the net biochemical effect of metformin and other biguanides such as a phenformin was to improve the catalytic efficiency of SIRT1 operating in conditions of low NAD(+) in vitro. Forthcoming studies should confirm the mechanistic relevance of our computational insights into how the putative binding modes of metformin to SIRT1 could explain its ability to operate as a direct SIRT1 -activating compound. These findings might have important implications for understanding how metformin might confer health benefits via maintenance of SIRT1 activity during the aging process when NAD(+) levels decline

Extra Virgin Olive Oil Contains a Phenolic Inhibitor of the Histone Demethylase LSD1/KDM1A

The lysine-specific histone demethylase 1A (LSD1) also known as lysine (K)-specific demethylase 1A (KDM1A) is a central epigenetic regulator of metabolic reprogramming in obesity-associated diseases, neurological disorders, and cancer. Here, we evaluated the ability of oleacein, a biophenol secoiridoid naturally present in extra virgin olive oil (EVOO), to target LSD1. Molecular docking and dynamic simulation approaches revealed that oleacein could target the binding site of the LSD1 cofactor flavin adenosine dinucleotide with high affinity and at low concentrations. At higher concentrations, oleacein was predicted to target the interaction of LSD1 with histone H3 and the LSD1 co-repressor (RCOR1/CoREST), likely disturbing the anchorage of LSD1 to chromatin. AlphaScreen-based in vitro assays confirmed the ability of oleacein to act as a direct inhibitor of recombinant LSD1, with an IC50 as low as 2.5 umol/L. Further, oleacein fully suppressed the expression of the transcription factor SOX2 (SEX determining Region Y-box 2) in cancer stem-like and induced pluripotent stem (iPS) cells, which specifically occurs under the control of an LSD1-targeted distal enhancer. Conversely, oleacein failed to modify ectopic SOX2 overexpression driven by a constitutive promoter. Overall, our findings provide the first evidence that EVOO contains a naturally occurring phenolic inhibitor of LSD1, and support the use of oleacein as a template to design new secoiridoid-based LSD1 inhibitors.Work in the Menendez laboratory is supported by the Spanish Ministry of Science and Innovation (Grant SAF2016-80639-P, Plan Nacional de l+D+I, founded by the European Regional Development Fund, Spain) and by an unrestricted research grant from the Fundació Oncolliga Girona (Lliga catalana d’ajuda al malalt de càncer, Girona). The Spanish Ministry of Economy and Competitiveness (MINECO, Project RTI2018-096724-B-C21) and the Generalitat Valenciana (PROMETEO/2016/006) supports work in the Encinar laborator

Metformin Is a Direct SIRT1-Activating Compound: Computational Modeling and Experimental Validation

Metformin has been proposed to operate as an agonist of SIRT1, a nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase that mimics most of the metabolic responses to calorie restriction. Herein, we present an in silico analysis focusing on the molecular docking and dynamic simulation of the putative interactions between metformin and SIRT1. Using eight different crystal structures of human SIRT1 protein, our computational approach was able to delineate the putative binding modes of metformin to several pockets inside and outside the central deacetylase catalytic domain. First, metformin was predicted to interact with the very same allosteric site occupied by resveratrol and other sirtuin-activating compounds (STATCs) at the amino-terminal activation domain of SIRT1. Second, metformin was predicted to interact with the NAD+ binding site in a manner slightly different to that of SIRT1 inhibitors containing an indole ring. Third, metformin was predicted to interact with the C-terminal regulatory segment of SIRT1 bound to the NAD+ hydrolysis product ADP-ribose, a “C-pocket”-related mechanism that appears to be essential for mechanism-based activation of SIRT1. Enzymatic assays confirmed that the net biochemical effect of metformin and other biguanides such as a phenformin was to improve the catalytic efficiency of SIRT1 operating in conditions of low NAD+ in vitro. Forthcoming studies should confirm the mechanistic relevance of our computational insights into how the putative binding modes of metformin to SIRT1 could explain its ability to operate as a direct SIRT1-activating compound. These findings might have important implications for understanding how metformin might confer health benefits via maintenance of SIRT1 activity during the aging process when NAD+ levels decline

Computer-Aided Drug Design applied to marine drug discovery = Disseny de fàrmacs assistit per ordinador aplicat a la cerca de possibles fàrmacs marins

[eng] The potential of natural products in general, and marine natural products in particular, as pharmacological entities has been widely demonstrated in recent years. Marine benthic ecosystems contain an extraordinary range of diverse organisms that possess bioactive natural compounds, which are commonly used as defensive or protective chemical mechanisms. These effective defensive strategies are based on secondary metabolites that are crucial for the species survival. The pharmacological properties of these unique chemical compounds constitute an interesting and emerging hot research line, based upon exploiting them for the development of new drugs. The evolution, biodiversity, and specific environmental conditions found in marine ecosystems, such as Antarctica and the Mediterranean Sea, make them an amazing source of potential therapeutic agents. Interestingly, some of these natural products are capable to modulate protein functions in pathogenesis-related pathways. The process of discovery and development of new drugs, for instance small molecules, with the aforementioned capacity to modulate protein functions, is a tedious procedure that requires economic resources and time. To reduce these drawbacks, computer-aided drug design (CADD) has emerged as one of the most effective methods. A rapid exploration of the chemical space can be done with computational methods, and they are very interesting and useful complementary approaches to experimental methods. CADD techniques can be applied in different steps of the drug discovery pipeline, and also, can cover several phases of this pipeline. To that end, several objectives have been set and reached in this thesis: 1. To find possible therapeutic activities and to establish the capability to modulate protein functions in pathogenesis-related pathways from marine molecules by using different CADD tools and techniques: I. Improve the drug discovery pipeline by the elucidation of the possible therapeutic potential of a set of marine molecules against a list of targets related to different pathologies. II. Elucidation of different pharmacophoric features of marine compounds and a precise in silico binding study, highlighting the power of CADD techniques, and reporting the inhibitory activity of different natural products and indole scaffold derivatives as GSK3β, CK1δ, DYRK1A, and CLK1 inhibitors. III. Computational study and an experimental validation of meridianins and lignarenones as possible ATP and/or substrate inhibitors of GSK3β. The main conclusions of this thesis are that marine molecules can be used as therapeutic agents against protein kinases related to the AD, and the exemplification of CADD potential applied to marine drug discovery.[cat] El potencial dels productes naturals en general, i els productes naturals marins en particular, com a entitats farmacològiques ha quedat demostrat al llarg dels últims anys. Els ecosistemes bentònics marins contenen una extraordinària diversitat d'organismes que posseeixen compostos naturals bioactius, que utilitzen com mecanismes químics defensius i de protecció. Aquestes efectives estratègies defensives es basen en metabòlits secundaris, crucials per a la supervivència de les espècies. Tenint en compte les propietats farmacològiques d'aquests compostos químics únics, utilitzar-los per al desenvolupament de nous fàrmacs constitueix una línia interessant de recerca emergent. L'evolució, la biodiversitat i les condicions específiques que es troben en els ecosistemes marins, com ara l'Antàrtida i el mar Mediterrani, els converteixen en una font increïble de possibles agents terapèutics, capaços de modular funcions de proteïnes involucrades en determinades patologies. El procés de descobriment i desenvolupament de nous fàrmacs, per exemple, molècules petites, és un procediment tediós que requereix de recursos econòmics i de temps. Per reduir aquests inconvenients, el disseny de fàrmacs assistit per ordinador (DFAO) ha sorgit com un dels mètodes principals i més eficaços. Es pot fer una exploració ràpida de l'espai químic amb mètodes computacionals i a més, són aproximacions complementàries als mètodes experimentals molt interessants i útils. Les tècniques de DFAO es poden aplicar en diferents passos del procés de descobriment de fàrmacs, i també, poden cobrir diverses fases d'aquest pipeline. Amb aquesta finalitat, es varen establir diversos objectius en aquesta tesi: 1. Dilucidar la possible activitat terapèutica i la capacitat per modular les funcions de proteïnes que estan relacionades amb una determinada patologia de les molècules marines mitjançant l'ús de diferents eines i tècniques de DFAO: I. millorar el pipeline de descobriment de fàrmacs mitjançant l'elucidació del possible potencial terapèutic d'un conjunt de molècules marines enfront d'una llista de dianes relacionades amb diferents patologies. II. Dilucidació de les diferents característiques farmacofóriques dels compostos marins i en un precís estudi d’unió in silico, destacant el poder de les tècniques de DFAO, i avaluar l'activitat inhibidora de diferents productes naturals i derivats d’esquelets indòlics com inhibidors de GSK3β, CK1δ, DYRK1A i CLK1. III. Estudi computacional i validació experimental de meridianines i lignarenones com a possibles inhibidors de GSK3β mitjançant la unió a la cavitat de l'ATP i/o del substrat. En relació amb aquests objectius, les conclusions principals d'aquesta tesi són, que les molècules marines poden ser utilitzades com a agents terapèutics contra proteïnes quinases relacionades amb la malaltia d’Alzheimer, i l'exemplificació del potencial de les tècniques de DFAO aplicat al descobriment de fàrmacs marins

Computer-Aided Drug Design Applied to Marine Drug Discovery: Meridianins as Alzheimer's Disease Therapeutic Agents

Computer-aided drug discovery/design (CADD) techniques allow the identification of natural products that are capable of modulating protein functions in pathogenesis-related pathways, constituting one of the most promising lines followed in drug discovery. In this paper, we computationally evaluated and reported the inhibitory activity found in meridianins A-G, a group of marine indole alkaloids isolated from the marine tunicate Aplidium, against various protein kinases involved in Alzheimer's disease (AD), a neurodegenerative pathology characterized by the presence of neurofibrillary tangles (NFT). Balance splitting between tau kinase and phosphate activities caused tau hyperphosphorylation and, thereby, its aggregation and NTF formation. Inhibition of specific kinases involved in its phosphorylation pathway could be one of the key strategies to reverse tau hyperphosphorylation and would represent an approach to develop drugs to palliate AD symptoms. Meridianins bind to the adenosine triphosphate (ATP) binding site of certain protein kinases, acting as ATP competitive inhibitors. These compounds show very promising scaffolds to design new drugs against AD, which could act over tau protein kinases Glycogen synthetase kinase-3 Beta (GSK3β) and Casein kinase 1 delta (CK1δ, CK1D or KC1D), and dual specificity kinases as dual specificity tyrosine phosphorylation regulated kinase 1 (DYRK1A) and cdc2-like kinases (CLK1). This work is aimed to highlight the role of CADD techniques in marine drug discovery and to provide precise information regarding the binding mode and strength of meridianins against several protein kinases that could help in the future development of anti-AD drugs

Machine-Learning QSAR Model for Predicting Activity against Malaria Parasite's Ion Pump PfATP4 and In Silico Binding Assay Validation

Malaria is a mosquito-borne infectious disease caused by parasitic protozoans of the genus Plasmodium. Although different effective antimalarial medicines have been developed, there is serious concern that parasites are developing widespread resistance to these drugs. To avoid this, now the efforts are concentrated on treating the malaria inside the Anopheles mosquito

Machine-Learning QSAR Model for Predicting Activity against Malaria Parasite’s Ion Pump PfATP4 and In Silico Binding Assay Validation

Malaria is a mosquito-borne infectious disease caused by parasitic protozoans of the genus Plasmodium. [...