RESUMEN
Several anthropogenic contaminants have been identified as competing with the thyroid hormone thyroxine (T4) for binding to transport proteins as transthyretin (TTR). This binding can potentially create toxicity mechanisms posing a threat to human health. Many organic UV filters (UVFs) and paraben preservatives (PBs), widely used in personal care products, are chemicals of emerging concern due to their adverse effects as potential thyroid-disrupting compounds. Recently, organic UVFs have been found in paired maternal and fetal samples and PBs have been detected in placenta, which opens the possibility of the involvement of TTR in the transfer of these chemicals across physiological barriers. We aimed to investigate a discrete set of organic UVFs and PBs to identify novel TTR binders. The binding affinities of target UVFs towards TTR were evaluated using in vitro T4 competitive binding assays. The ligand-TTR affinities were determined by isothermal titration calorimetry (ITC) and compared with known TTR ligands. In parallel, computational studies were used to predict the 3-D structures of the binding modes of these chemicals to TTR. Some organic UVFs, compounds 2,2',4,4'-tetrahydroxybenzophenone (BP2, Kd = 0.43 µM); 2,4-dihydroxybenzophenone (BP1, Kd = 0.60 µM); 4,4'-dihydroxybenzophenone (4DHB, Kd = 0.83 µM), and 4-hydroxybenzophenone (4HB, Kd = 0.93 µM), were found to display a high affinity to TTR, being BP2 the strongest TTR binder (ΔH = -14.93 Kcal/mol). Finally, a correlation was found between the experimental ITC data and the TTR-ligand docking scores obtained by computational studies. The approach integrating in vitro assays and in silico methods constituted a useful tool to find TTR binders among common organic UVFs. Further studies on the involvement of the transporter protein TTR in assisting the transplacental transfer of these chemicals across physiological barriers and the long-term consequences of prenatal exposure to them should be pursued.
Asunto(s)
Prealbúmina , Hormonas Tiroideas , Embarazo , Femenino , Humanos , Prealbúmina/química , Prealbúmina/metabolismo , Ligandos , Hormonas Tiroideas/metabolismo , Tiroxina , Proteínas PortadorasRESUMEN
Cyclin-dependent kinase 5 (CDK5) is a serine/threonine kinase that has emerged as a key regulator of neurotransmission in complex cognitive processes. Its expression is altered in treated schizophrenia patients, and cannabinoids modulate CDK5 levels in the brain of rodents. However, the role of this kinase, and its interaction with cannabis use in first-episode psychosis (FEP) patients is still not known. Hence, we studied the expression changes of CDK5 and its signaling partner, postsynaptic density protein 95 (PSD95) in olfactory neuroepithelial (ON) cells of FEP patients with (FEP/c) and without (FEP/nc) prior cannabis use, and in a dual-hit mouse model of psychosis. In this model, adolescent mice were exposed to the cannabinoid receptor 1 agonist (CB1R) WIN-55,212-2 (WIN: 1 mg/kg) during 21 days, and to the N-methyl-d-aspartate receptor (NMDAR) blocker phencyclidine (PCP: 10 mg/kg) during 10 days. FEP/c showed less social functioning deficits, lower CDK5 and higher PSD95 levels than FEP/nc. These changes correlated with social skills, but not cognitive deficits. Consistently, exposure of ON cells from FEP/nc patients to WIN in vitro reduced CDK5 levels. Convergent results were obtained in mice, where PCP by itself induced more sociability deficits, and PSD95/CDK5 alterations in the prefrontal cortex and hippocampus than exposure to PCP-WIN. In addition, central blockade of CDK5 activity with roscovitine in PCP-treated mice restored both sociability impairments and PSD95 levels. We provide translational evidence that increased CDK5 could be an early indicator of psychosis associated with social deficits, and that this biomarker is modulated by prior cannabis use.
Asunto(s)
Cannabinoides , Trastornos Psicóticos , Esquizofrenia , Ratones , Animales , Quinasa 5 Dependiente de la Ciclina/metabolismo , Trastornos Psicóticos/tratamiento farmacológico , Fenciclidina/farmacología , Agonistas de Receptores de Cannabinoides , Homólogo 4 de la Proteína Discs LargeRESUMEN
Transthyretin (TTR) has a well-established role in neuroprotection in Alzheimer's Disease (AD). We have setup a drug discovery program of small-molecule compounds that act as chaperones enhancing TTR/Amyloid-beta peptide (Aß) interactions. A combination of computational drug repurposing approaches and in vitro biological assays have resulted in a set of molecules which were then screened with our in-house validated high-throughput screening ternary test. A prioritized list of chaperones was obtained and corroborated with ITC studies. Small-molecule chaperones have been discovered, among them our lead compound Iododiflunisal (IDIF), a molecule in the discovery phase; one investigational drug (luteolin); and 3 marketed drugs (sulindac, olsalazine and flufenamic), which could be directly repurposed or repositioned for clinical use. Not all TTR tetramer stabilizers behave as chaperones in vitro. These chemically diverse chaperones will be used for validating TTR as a target in vivo, and to select one repurposed drug as a candidate to enter clinical trials as AD disease-modifying drug.
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Enfermedad de Alzheimer/tratamiento farmacológico , Descubrimiento de Drogas , Chaperonas Moleculares/farmacología , Prealbúmina/antagonistas & inhibidores , Bibliotecas de Moléculas Pequeñas/farmacología , Enfermedad de Alzheimer/metabolismo , Calorimetría , Relación Dosis-Respuesta a Droga , Humanos , Modelos Moleculares , Chaperonas Moleculares/química , Estructura Molecular , Prealbúmina/metabolismo , Bibliotecas de Moléculas Pequeñas/química , Programas Informáticos , Relación Estructura-ActividadRESUMEN
Pre-competitive data sharing can offer the pharmaceutical industry significant benefits in terms of reducing the time and costs involved in getting a new drug to market through more informed testing strategies and knowledge gained by pooling data. If sufficient data is shared and can be co-analyzed, then it can also offer the potential for reduced animal usage and improvements in the in silico prediction of toxicological effects. Data sharing benefits can be further enhanced by applying the FAIR Guiding Principles, reducing time spent curating, transforming and aggregating datasets and allowing more time for data mining and analysis. We hope to facilitate data sharing by other organizations and initiatives by describing lessons learned as part of the Enhancing TRANslational SAFEty Assessment through Integrative Knowledge Management (eTRANSAFE) project, an Innovative Medicines Initiative (IMI) partnership which aims to integrate publicly available data sources with proprietary preclinical and clinical data donated by pharmaceutical organizations. Methods to foster trust and overcome non-technical barriers to data sharing such as legal and IPR (intellectual property rights) are described, including the security requirements that pharmaceutical organizations generally expect to be met. We share the consensus achieved among pharmaceutical partners on decision criteria to be included in internal clearance procedures used to decide if data can be shared. We also report on the consensus achieved on specific data fields to be excluded from sharing for sensitive preclinical safety and pharmacology data that could otherwise not be shared.
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Minería de Datos , Difusión de la Información , Animales , Simulación por Computador , Industria FarmacéuticaRESUMEN
In the past few years, attempts have been made to use decision criteria beyond Lipinski's guidelines (Rule of five) to guide drug discovery projects more effectively. Several variables and formulations have been proposed and investigated within the framework of multiparameter optimization methods to guide drug discovery. In this context, the combination of Ligand Efficiency Indices (LEI) has been predominantly used to map and monitor the drug discovery process in a retrospective fashion. Here we provide an example of the use of a novel application of the LEI methodology for prospective lead optimization by using the transthyretin (TTR) fibrillogenesis inhibitor iododiflunisal (IDIF) as example. Using this approach, a number of compounds with theoretical efficiencies higher than the reference compound IDIF were identified. From this group, ten compounds were selected, synthesized and biologically tested. Half of the compounds (5, 6, 7, 8 and 10) showed potencies in terms of IC50 inhibition of TTR aggregation equal or higher than the lead compound. These optimized compounds mapped within the region of more efficient candidates in the corresponding experimental nBEI-NSEI plot, matching their position in the theoretical optimization plane that was used for the prediction. Due to their upstream (North-Eastern) position in the progression lines of NPOLâ¯=â¯3 or 4 of the nBEI-NSEI plot, three of them (5, 6 and 8) are more interesting candidates than iododiflunisal because they have been optimized in the three crucial LEI variables of potency, size and polarity at the same time. This is the first example of the effectiveness of using the combined LEIs within the decision process to validate the application of the LEI formulation for the prospective optimization of lead compounds.
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Ligandos , Prealbúmina/metabolismo , Diflunisal/análogos & derivados , Diflunisal/farmacología , Humanos , Cinética , Mutagénesis Sitio-Dirigida , Prealbúmina/antagonistas & inhibidores , Prealbúmina/genética , Unión Proteica , Multimerización de Proteína/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
Transthyretin (TTR) is a homotetrameric protein involved in human amyloidosis, including familial amyloid polyneuropathy (FAP). Discovering small-molecule stabilizers of the TTR tetramer is a therapeutic strategy for these diseases. Tafamidis, the only approved drug for FAP treatment, is not effective for all patients. Herein, we discovered that benzbromarone (BBM), a uricosuric drug, is an effective TTR stabilizer and inhibitor against TTR amyloid fibril formation. BBM rendered TTR more resistant to urea denaturation, similarly to iododiflunisal (IDIF), a very potent TTR stabilizer. BBM competes with thyroxine for binding in the TTR central channel, with an IC50 similar to IDIF and tafamidis. Results obtained by isothermal titration calorimetry (ITC) demonstrated that BBM binds TTR with an affinity similar to IDIF, tolcapone and tafamidis, confirming BBM as a potent binder of TTR. The crystal structure of the BBM-TTR complex shows two molecules binding deeply in the thyroxine binding channel, forming strong intermonomer hydrogen bonds and increasing the stability of the TTR tetramer. Finally, kinetic analysis of the ability of BBM to inhibit TTR fibrillogenesis at acidic pH and comparison with other stabilizers revealed that benzbromarone is a potent inhibitor of TTR amyloidogenesis, adding a new interesting scaffold for drug design of TTR stabilizers.
Asunto(s)
Benzbromarona/química , Reposicionamiento de Medicamentos , Fármacos Neuroprotectores/química , Prealbúmina/química , Tiroxina/química , Amiloide/antagonistas & inhibidores , Benzbromarona/metabolismo , Benzoxazoles/química , Benzoxazoles/metabolismo , Sitios de Unión , Unión Competitiva , Cristalografía por Rayos X , Diflunisal/análogos & derivados , Diflunisal/química , Diflunisal/metabolismo , Expresión Génica , Humanos , Enlace de Hidrógeno , Cinética , Simulación del Acoplamiento Molecular , Fármacos Neuroprotectores/metabolismo , Prealbúmina/agonistas , Prealbúmina/genética , Prealbúmina/metabolismo , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Multimerización de Proteína , Estabilidad Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Termodinámica , Tiroxina/metabolismo , Tolcapona/química , Tolcapona/metabolismoRESUMEN
BACKGROUND: Transthyretin (TTR) is a tetrameric, amyloid-ß (Aß)-binding protein, which reduces Aß toxicity. The TTR/Aß interaction can be enhanced by a series of small molecules that stabilize its tetrameric form. Hence, TTR stabilizers might act as disease-modifying drugs in Alzheimer's disease. OBJECTIVE: We monitored the therapeutic efficacy of two TTR stabilizers, iododiflunisal (IDIF), which acts as small-molecule chaperone of the TTR/Aß interaction, and tolcapone, which does not behave as a small-molecule chaperone, in an animal model of Alzheimer's disease using positron emission tomography (PET). METHODS: Female mice (AßPPswe/PS1A246E/TTR+/-) were divided into 3 groups (nâ=â7 per group): IDIF-treated, tolcapone-treated, and non-treated. The oral treatment (100âmg/Kg/day) was started at 5 months of age. Treatment efficacy assessment was based on changes in longitudinal deposition of Aß in the hippocampus (HIP) and the cortex (CTX) and determined using PET-[18F]florbetaben. Immunohistochemical analysis was performed at ageâ=â14 months. RESULTS: Standard uptake values relative to the cerebellum (SUVr) of [18F]florbetaben in CTX and HIP of non-treated animals progressively increased from ageâ=â5 to 11 months and stabilized afterwards. In contrast, [18F]florbetaben uptake in HIP of IDIF-treated animals remained constant between agesâ=â5 and 11 months and significantly increased at 14 months. In the tolcapone-treated group, SUVr progressively increased with time, but at lower rate than in the non-treated group. No significant treatment effect was observed in CTX. Results from immunohistochemistry matched the in vivo data at ageâ=â14 months. CONCLUSION: Our work provides encouraging preliminary results on the ability of small-molecule chaperones to ameliorate Aß deposition in certain brain regions.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Precursor de Proteína beta-Amiloide/antagonistas & inhibidores , Diflunisal/análogos & derivados , Hipocampo/efectos de los fármacos , Imagen Molecular/métodos , Administración Oral , Enfermedad de Alzheimer/diagnóstico por imagen , Enfermedad de Alzheimer/genética , Enfermedad de Alzheimer/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animales , Diflunisal/administración & dosificación , Femenino , Hipocampo/diagnóstico por imagen , Hipocampo/metabolismo , Estudios Longitudinales , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C3H , Ratones Transgénicos , Tomografía Computarizada por Tomografía de Emisión de Positrones/métodosRESUMEN
The protein transthyretin (TTR) modulates amyloid-ß (Aß) peptides deposition and processing and this physiological effect is further enhanced by treatment with iododiflunisal (IDIF), a small-molecule compound (SMC) with TTR tetramer stabilization properties, which behaves as chaperone of the complex. This knowledge has prompted us to design and optimize a rapid and simple high-throughput assay that relies on the ability of test compounds to form ternary soluble complexes TTR/Aß/SMC that prevent Aß aggregation. The method uses the shorter Aß(12-28) sequence which is cheaper and simpler to use while retaining the aggregation properties of their parents Aß(1-40) and Aß(1-42). The test is carried out in 96-plate wells that are UV monitored for turbidity during 6â h. Given its reproducibility, we propose that this test can be a powerful tool for efficient screening of SMCs that act as chaperones of the TTR/Aß interaction that may led to potential AD therapies.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides/química , Preparaciones Farmacéuticas , Enfermedad de Alzheimer/tratamiento farmacológico , Humanos , Prealbúmina/química , Reproducibilidad de los ResultadosRESUMEN
Transthyretin (TTR), an homotetrameric protein mainly synthesized by the liver and the choroid plexus, and secreted into the blood and the cerebrospinal fluid, respectively, has been specially acknowledged for its functions as a transporter protein of thyroxine and retinol (the latter through binding to the retinol-binding protein), in these fluids. Still, this protein has managed to stay in the spotlight as it has been assigned new and varied functions. In this review, we cover knowledge on novel TTR functions and the cellular pathways involved, spanning from neuroprotection to vascular events, while emphasizing its involvement in Alzheimer's disease (AD). We describe details of TTR as an amyloid binding protein and discuss its interaction with the amyloid Aß peptides, and the proposed mechanisms underlying TTR neuroprotection in AD. We also present the importance of translating advances in the knowledge of the TTR neuroprotective role into drug discovery strategies focused on TTR as a new target in AD therapeutics.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides/metabolismo , Sistemas de Liberación de Medicamentos , Descubrimiento de Drogas , Prealbúmina , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Humanos , Prealbúmina/antagonistas & inhibidores , Prealbúmina/metabolismoRESUMEN
Transthyretin (TTR) modulates the deposition, processing, and toxicity of Abeta (Aß) peptides. We have shown that this effect is enhanced in mice by treatment with small molecules such as iododiflunisal (IDIF, 4), a good TTR stabilizer. Here, we describe the thermodynamics of the formation of binary and ternary complexes among TTR, Aß(1-42) peptide, and TTR stabilizers using isothermal titration calorimetry (ITC). A TTR/Aß(1-42) (1:1) complex with a dissociation constant of Kd = 0.94 µM is formed; with IDIF (4), this constant improves up to Kd = 0.32 µM, indicating the presence of a ternary complex TTR/IDIF/Aß(1-42). However, with the drugs diflunisal (1) or Tafamidis (2), an analogous chaperoning effect could not be observed. Similar phenomena could be recorded with the shorter peptide Aß(12-28) (7). We propose the design of a simple assay system for the search of other chaperones that behave like IDIF and may become potential candidate drugs for Alzheimer's disease (AD).
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Péptidos beta-Amiloides/metabolismo , Benzoxazoles/metabolismo , Diflunisal/análogos & derivados , Diflunisal/metabolismo , Fragmentos de Péptidos/metabolismo , Prealbúmina/metabolismo , Multimerización de Proteína/efectos de los fármacos , Bioensayo/métodos , Calorimetría/métodos , Humanos , TermodinámicaRESUMEN
It is well settled that the amyloidogenic properties of the plasma protein transporter transthyretin (TTR) can be modulated by compounds that stabilize its native tetrameric conformation. TTR is also present in cerebrospinal fluid where it can bind to Aß-peptides and prevent Aß aggregation. We have previously shown that treatment of Alzheimer's Disease (AD) model mice with iododiflunisal (IDIF), a TTR tetramer stabilizing compound, prevents AD pathologies. This evidence positioned IDIF as a new lead drug for AD. In dissecting the mechanism of action of IDIF, we disclose here different labeling strategies for the preparation of 131I-labeled IDIF and 131I- and 124I-labeled TTR, which have been further used for the preparation of IDIF-TTR complexes labeled either on the compound or the protein. The biodistribution of all labeled species after intravenous administration has been investigated in mice using ex vivo and in vivo techniques. Our results confirm the capacity of TTR to cross the blood brain barrier (BBB) and suggest that the formation of TTR-IDIF complexes enhances BBB permeability of both IDIF and TTR. The increased TTR and IDIF brain concentrations may result in higher Aß-peptide sequestration capacity with the subsequent inhibition of AD symptoms as we have previously observed in mice.
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Encéfalo/diagnóstico por imagen , Diflunisal/análogos & derivados , Radioisótopos de Yodo/química , Prealbúmina/química , Prealbúmina/farmacocinética , Administración Intravenosa , Péptidos beta-Amiloides/metabolismo , Animales , Autorradiografía , Barrera Hematoencefálica/química , Encéfalo/metabolismo , Diflunisal/administración & dosificación , Diflunisal/química , Diflunisal/farmacocinética , Ratones , Tomografía de Emisión de Positrones , Prealbúmina/administración & dosificación , Distribución TisularRESUMEN
The European Directive 98/83/CE legislates the presence of pesticides in drinking water, but apart from a few compounds, nothing is said about which pesticides should be monitored. Nevertheless, water companies need to go beyond the accomplishment of the legislation and find out pesticide contamination in all sources of water in order to manage the hazard assessment, and to guarantee safe drinking water to all the population. The aim of this work was to develop an analytical multi-residue method for circa 100 compounds. The method analyses previously monitored compounds in Barcelona city and its metropolitan area, as well as many emerging pesticides and some transformation products. An on-line sample extraction (0.75â¯mL) coupled to fast UHPLC-MS/MS method was developed. Good linearity (r2â¯>â¯0.995, with less residuals than 15%), accuracies and precisions under 25%, and acceptable expanded uncertainties were obtained for most of the monitored compounds, according to ISO/IEC 17025, obtaining limits of quantification between 5 and 25â¯ng/L for all compounds. A monitoring campaign on natural and treated waters in the Barcelona metropolitan area was carried out during 2016-2017. Results showed that pesticide contamination at the low stretch of Llobregat River and in its aquifer is severe. The maximum concentrations were in the range of few µg/L for carbendazim, DEET, diuron and propiconazole, and in the range 0.1-0.5⯵g/L for bentazone, imidacloprid, isoproturon, simazine, metazachlor, methomyl, terbutryn and tebuconazole. However, the efficiency of advanced treatments in the DWTPs involved in drinking water production in the Barcelona metropolitan area allows the complete removal of pesticides and a safe water production for consumers. The method shows a good analytical performance for most compounds with a fast sample preparation and analysis. In addition, it has updated the knowledge about the occurrence of pesticides in the Barcelona city area.
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Agua Potable/química , Monitoreo del Ambiente , Agua Dulce/química , Plaguicidas/análisis , Contaminantes Químicos del Agua/análisis , EspañaRESUMEN
A study of organic compounds that caused a serious taste and odor episode of water supply in two residential areas in Catalonia (N.E. Spain) was carried out. Sweet and paint/solvent odor were the main descriptors used by consumers. Some cases of sickness and nausea were also associated with drinking water consumption by the consumers. Closed-loop stripping analysis (CLSA) combined with sensory gas chromatography and gas chromatography mass spectrometry detection were used to study the problem. As a result, 3-(trifluoromethyl)phenol (CAS number 98-17-9) was for the first time identified as a responsible of an odor incident in drinking water. Concentration levels of this compound were up to 17,000 ng/L in groundwater and up to 600 ng/L in distributed water. Odor threshold in water for 3-(trifluoromethyl)phenol was determined as 13 ng/L (45 °C).
Asunto(s)
Cresoles/análisis , Agua Potable/análisis , Odorantes/análisis , Contaminantes Químicos del Agua/análisis , Adulto , Agua Potable/química , Femenino , Cromatografía de Gases y Espectrometría de Masas/métodos , Humanos , Extracción Líquido-Líquido , Masculino , Umbral Sensorial , España , Gusto , Abastecimiento de AguaRESUMEN
Compound screening in biological assays and subsequent optimization of hits is indispensable for the development of new molecular research tools and drug candidates. To facilitate such discoveries, the European Research Infrastructure EU-OPENSCREEN was founded recently with the support of its member countries and the European Commission. Its distributed character harnesses complementary knowledge, expertise, and instrumentation in the discipline of chemical biology from 20 European partners, and its open working model ensures that academia and industry can readily access EU-OPENSCREEN's compound collection, equipment, and generated data. To demonstrate the power of this collaborative approach, this perspective article highlights recent projects from EU-OPENSCREEN partner institutions. These studies yielded (1) 2-aminoquinazolin-4(3 H)-ones as potential lead structures for new antimalarial drugs, (2) a novel lipodepsipeptide specifically inducing apoptosis in cells deficient for the pVHL tumor suppressor, (3) small-molecule-based ROCK inhibitors that induce definitive endoderm formation and can potentially be used for regenerative medicine, (4) potential pharmacological chaperones for inborn errors of metabolism and a familiar form of acute myeloid leukemia (AML), and (5) novel tankyrase inhibitors that entered a lead-to-candidate program. Collectively, these findings highlight the benefits of small-molecule screening, the plethora of assay designs, and the close connection between screening and medicinal chemistry within EU-OPENSCREEN.
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Conducta Cooperativa , Descubrimiento de Drogas/métodos , Evaluación Preclínica de Medicamentos , Europa (Continente) , Ensayos Analíticos de Alto Rendimiento , Humanos , Relación Estructura-ActividadRESUMEN
In silico methods are increasingly being used for assessing the chemical safety of substances, as a part of integrated approaches involving in vitro and in vivo experiments. A paradigmatic example of these strategies is the eTOX project http://www.etoxproject.eu, funded by the European Innovative Medicines Initiative (IMI), which aimed at producing high quality predictions of in vivo toxicity of drug candidates and resulted in generating about 200 models for diverse endpoints of toxicological interest. In an industry-oriented project like eTOX, apart from the predictive quality, the models need to meet other quality parameters related to the procedures for their generation and their intended use. For example, when the models are used for predicting the properties of drug candidates, the prediction system must guarantee the complete confidentiality of the compound structures. The interface of the system must be designed to provide non-expert users all the information required to choose the models and appropriately interpret the results. Moreover, procedures like installation, maintenance, documentation, validation and versioning, which are common in software development, must be also implemented for the models and for the prediction platform in which they are implemented. In this article we describe our experience in the eTOX project and the lessons learned after 7 years of close collaboration between industrial and academic partners. We believe that some of the solutions found and the tools developed could be useful for supporting similar initiatives in the future.
RESUMEN
Several strategies against Alzheimer disease (AD) are directed to target Aß-peptides. The ability of transthyretin (TTR) to bind Aß-peptides and the positive effect exerted by some TTR stabilizers for modulating the TTR-Aß interaction have been previously studied. Herein, key structural features of the interaction between TTR and the Aß(12-28) peptide (3), the essential recognition element of Aß, have been unravelled by STD-NMR spectroscopy methods in solution. Molecular aspects related to the role of the TTR stabilizer iododiflunisal (IDIF, 5) on the TTR-Aß complex have been also examined. The NMR results, assisted by molecular modeling protocols, have provided a structural model for the TTR-Aß interaction, as well as for the ternary complex formed in the presence of IDIF. This basic structural information could be relevant for providing light on the mechanisms involved in the ameliorating effects of AD symptoms observed in AD/TTR± animal models after IDIF treatment and eventually for designing new molecules toward AD therapeutic drugs.
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Péptidos beta-Amiloides/metabolismo , Diflunisal/análogos & derivados , Prealbúmina/metabolismo , Mapas de Interacción de Proteínas/efectos de los fármacos , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/química , Cristalografía por Rayos X , Diflunisal/química , Diflunisal/farmacología , Humanos , Espectroscopía de Resonancia Magnética , Simulación del Acoplamiento Molecular , Prealbúmina/químicaRESUMEN
1,4-Dihydropyridines are well-known calcium channel blockers, but variations in the substituents attached to the ring have resulted in their role reversal making them calcium channel activators in some cases. We describe the microwave-assisted eco-friendly approach for the synthesis of pyranopyrazole-1,4-dihydropyridines, a new class of 1,4-DHPs, under solvent-free conditions in good yield, and screen them for various in silico, in vitro and in vivo activities. The in vivo experimentation results show that the compounds possess positive inotropic effect, and the docking results validate their good binding with calcium channels. Compounds 7c, 7g and 7i appear to be the most effective positive inotropes, even at low doses, and bind with the calcium channels even more strongly than Bay K 8644, a well-known calcium channel activator. The chronotropic effect for the new compounds was also studied. The target and off-target affinity profiling supported the in vivo results and revealed that the hybridized pyranopyrazole dihydropyridine scaffold has delivered new moderate hits, to be optimized, for the cytochrome P450 3A4 enzymes, opening avenues for combined pharmacological activity through standard structural modification.
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Agonistas de los Canales de Calcio/administración & dosificación , Agonistas de los Canales de Calcio/síntesis química , Dihidropiridinas/administración & dosificación , Dihidropiridinas/síntesis química , Animales , Presión Sanguínea/efectos de los fármacos , Agonistas de los Canales de Calcio/química , Agonistas de los Canales de Calcio/farmacología , Dihidropiridinas/química , Dihidropiridinas/farmacología , Relación Dosis-Respuesta a Droga , Frecuencia Cardíaca/efectos de los fármacos , Ratones , Microondas , Modelos Moleculares , Simulación del Acoplamiento Molecular , Estructura MolecularRESUMEN
A study has been carried out to identify the origin of the odorous compounds at trace levels detected in surface waters and in Barcelona's tap water (NE Spain) which caused consumer complaints. The malodorous compounds were 2,5,5-trimethyl-1,3-dioxane (TMD) and 2-ethyl-5,5-dimethyl-1,3-dioxane (2EDD) which impart a distinctive sickening or olive-oil odor to drinking water at low ng/L levels. Flavor proï¬le analysis (FPA) or threshold odor number (TON) were used for organoleptic purposes. Levels up to 749 ng/L for TMD and 658 ng/L for 2EDD were measured at the entrance of the drinking water treatment plant. Three wastewater treatment plants (WWTPs) using industrial byproducts coming from resin manufacturing plants to promote codigestion were found to be the origin of the event. Corrective measures were applied, including the prohibition to use these byproducts for codigestion in the WWTPs involved. A similar event was already recorded in the same area 20 years ago.
Asunto(s)
Dioxanos/análisis , Odorantes/análisis , Purificación del Agua , Agua Potable/análisis , Ríos/química , España , Espectrometría de Fluorescencia , Aguas Residuales/análisis , Contaminantes Químicos del Agua/análisisRESUMEN
This work describes a collaborative effort to define and apply a protocol for the rational selection of a general-purpose screening library, to be used by the screening platforms affiliated with the EU-OPENSCREEN initiative. It is designed as a standard source of compounds for primary screening against novel biological targets, at the request of research partners. Given the general nature of the potential applications of this compound collection, the focus of the selection strategy lies on ensuring chemical stability, absence of reactive compounds, screening-compliant physicochemical properties, loose compliance to drug-likeness criteria (as drug design is a major, but not exclusive application), and maximal diversity/coverage of chemical space, aimed at providing hits for a wide spectrum of drugable targets. Finally, practical availability/cost issues cannot be avoided. The main goal of this publication is to inform potential future users of this library about its conception, sources, and characteristics. The outline of the selection procedure, notably of the filtering rules designed by a large committee of European medicinal chemists and chemoinformaticians, may be of general methodological interest for the screening/medicinal chemistry community. The selection task of 200K molecules out of a pre-filtered set of 1.4M candidates was shared by five independent European research groups, each picking a subset of 40K compounds according to their own in-house methodology and expertise. An in-depth analysis of chemical space coverage of the library serves not only to characterize the collection, but also to compare the various chemoinformatics-driven selection procedures of maximal diversity sets. Compound selections contributed by various participating groups were mapped onto general-purpose self-organizing maps (SOMs) built on the basis of marketed drugs and bioactive reference molecules. In this way, the occupancy of chemical space by the EU-OPENSCREEN library could be directly compared with distributions of known bioactives of various classes. This mapping highlights the relevance of the selection and shows how the consensus reached by merging the five different 40K selections contributes to achieve this relevance. The approach also allows one to readily identify subsets of target- or target-class-oriented compounds from the EU-OPENSCREEN library to suit the needs of the diverse range of potential users. The final EU-OPENSCREEN library, assembled by merging five independent selections of 40K compounds from various expert groups, represents an excellent example of a Europe-wide collaborative effort toward the common objective of building best-in-class European open screening platforms.
Asunto(s)
Evaluación Preclínica de Medicamentos , Unión EuropeaRESUMEN
The amyloidogenic protein transthyretin (TTR) is thought to aggregate into amyloid fibrils by tetramer dissociation which can be inhibited by a number of small molecule compounds. Our analysis of a series of crystallographic protein-inhibitor complexes has shown no clear correlation between the observed molecular interactions and the in vitro activity of the inhibitors. From this analysis, it emerged that halogen bonding (XB) could be mediating some key interactions. Analysis of the halogenated derivatives of two well-known TTR inhibitors has shown that while flufenamic acid affinity for TTR was unchanged by halogenation, diflunisal gradually improves binding up to 1 order of magnitude after iodination through interactions that can be interpreted as a suboptimal XB (carbonyl Thr106: I...O distance 3.96-4.05 Å; C-I...O angle 152-156°) or as rather optimized van der Waals contacts or as a mixture of both. These results illustrate the potential of halogenation strategies in designing and optimizing TTR fibrillogenesis inhibitors.