HAL Portal ESPCI (Ecole Supérieure de Physique et de Chimie Industrielles)
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Identification of Cellular Protein Targets of a Half-Sandwich Iridium(III) Complex Reveals Its Dual Mechanism of Action via Both Electrophilic and Oxidative Stresses
No full textInternational audienceIdentification of intracellular targets of anticancer drug candidates provides key information on their mechanism of action. Exploiting the ability of the anticancer (C∧N)-chelated half-sandwich iridium(III) complexes to covalently bind proteins, click chemistry with a bioorthogonal azido probe was used to localize a phenyloxazoline-chelated iridium complex within cells and profile its interactome at the proteome-wide scale. Proteins involved in protein folding and actin cytoskeleton regulation were identified as high-affinity targets. Upon iridium complex treatment, the folding activity of Heat Shock Protein HSP90 was inhibited in vitro and major cytoskeleton disorganization was observed. A wide array of imaging and biochemical methods validated selected targets and provided a multiscale overview of the effects of this complex on live human cells. We demonstrate that it behaves as a dual agent, inducing both electrophilic and oxidative stresses in cells that account for its cytotoxicity. The proposed methodological workflow can open innovative avenues in metallodrug discovery
High-resolution cryo-EM of the human CDK-activating kinase for structure-based drug design
No full textInternational audienceRational design of next-generation therapeutics can be facilitated by high-resolution structures of drug targets bound to small-molecule inhibitors. However, application of structure-based methods to macromolecules refractory to crystallization has been hampered by the often-limiting resolution and throughput of cryogenic electron microscopy (cryo-EM). Here, we use high-resolution cryo-EM to determine structures of the CDK-activating kinase, a master regulator of cell growth and division, in its free and nucleotide-bound states and in complex with 15 inhibitors at up to 1.8 Å resolution. Our structures provide detailed insight into inhibitor interactions and networks of water molecules in the active site of cyclin-dependent kinase 7 and provide insights into the mechanisms contributing to inhibitor selectivity, thereby providing the basis for rational design of next-generation therapeutics. These results establish a methodological framework for the use of high-resolution cryo-EM in structure-based drug design
Switchable molecular tweezers: design and applications
No full textInternational audienceSwitchable molecular tweezers are a unique class of molecular switches that, like their macroscopic analogs, exhibit mechanical motion between an open and closed conformation in response to stimuli. Such systems constitute an essential component of artificial molecular machines. This review will present selected examples of switchable molecular tweezers and their potential applications. The first part will be devoted to chemically responsive tweezers, including stimuli such as pH, metal coordination, and anion binding. Then, redox-active and photochemical tweezers will be presented
Influence of microgravity on spontaneous calcium activity of primary hippocampal neurons grown in microfluidic chips
No full textInternational audienceAbstract The influence of variations of gravity, either hypergravity or microgravity, on the brain of astronauts is a major concern for long journeys in space, to the Moon or to Mars, or simply long-duration missions on the ISS (International Space Station). Monitoring brain activity, before and after ISS missions already demonstrated important and long term effects on the brains of astronauts. In this study, we focus on the influence of gravity variations at the cellular level on primary hippocampal neurons. A dedicated setup has been designed and built to perform live calcium imaging during parabolic flights. During a CNES (Centre National d’Etudes Spatiales) parabolic flight campaign, we were able to observe and monitor the calcium activity of 2D networks of neurons inside microfluidic devices during gravity changes over different parabolas. Our preliminary results clearly indicate a modification of the calcium activity associated to variations of gravity
Ring‐opening (co)polymerization of cardanol glycidyl ether
No full textInternational audienceThe anionic ring‐opening copolymerization of commercially available bio‐based cardanol glycidyl ether (CGE) was investigated without any prior purification. As a first step, anionic‐ring‐opening homopolymerization was attempted through active chain‐end and monomer‐activated mechanisms. Both strategies were unsuccessful. Conversely, in a second step, the anionic alternating ring‐opening copolymerization (AAROP) of CGE with the renewable N ‐acetyl homocysteine thiolactone (NHTL) was successfully carried out in the presence of a strong base. Anisole, a solvent classified as sustainable and rarely used in anionic ring‐opening polymerization, proved to be a suitable for the AAROP. This polymerization is an unusual example of synthesis of linear polyesters with cardanol‐based monomers. The copolymers were carefully characterized by 1 H, 13 C, COSY, HSQC, 1 H‐ 15 N NMR and MALDI‐ToF, demonstrating an alternating structure. Then, CGE was copolymerized with NHTL and another additional epoxide. The cardanol‐derived monomers enable the preparation of functionalizable poly(ester‐ alt ‐thioether) bearing multiple allyl and alkene groups. The AAROP method in anisole offers new opportunity for green anionic polymerization through the use of sustainable chemicals, witnessed by the valorization of cardanol‐derived compounds and expands the scope of synthesized renewable polyesters
Flexible implementation of modulated localization microscopy based on DMD
No full textInternational audienceLocalization microscopy of individual molecules allows one to bypass the diffraction limit, revealing cellular organization on a nanometric scale. This method, which relies on spatial analysis of the signal emitted by molecules, is often limited to the observation of biological objects at shallow depths, or with very few aberrations. The introduction of a temporal parameter into the localization process through a time modulated excitation was recently proposed to address these limitations. This method called ModLoc, is demonstrated here with an alternative flexible strategy. In this implementation, to encode the time modulated excitation a digital micromirror device is used in combination with a fast demodulation approach, and provides a twofold enhancement in localization precision
GWTC-2.1: Deep extended catalog of compact binary coalescences observed by LIGO and Virgo during the first half of the third observing run
No full textInternational audienceThe second Gravitational-Wave Transient Catalog, GWTC-2, reported on 39 compact binary coalescences observed by the Advanced LIGO and Advanced Virgo detectors between 1 April 2019 15∶00 UTC and 1 October 2019 15∶00 UTC. Here, we present GWTC-2.1, which reports on a deeper list of candidate events observed over the same period. We analyze the final version of the strain data over this period with improved calibration and better subtraction of excess noise, which has been publicly released. We employ three matched-filter search pipelines for candidate identification, and estimate the probability of astrophysical origin for each candidate event. While GWTC-2 used a false alarm rate threshold of 2 per year, we include in GWTC-2.1, 1201 candidates that pass a false alarm rate threshold of 2 per day. We calculate the source properties of a subset of 44 high-significance candidates that have a probability of astrophysical origin greater than 0.5. Of these candidates, 36 have been reported in GWTC-2. We also calculate updated source properties for all binary black hole events previously reported in GWTC-1. If the eight additional high-significance candidates presented here are astrophysical, the mass range of events that are unambiguously identified as binary black holes (both objects ≥3M⊙) is increased compared to GWTC-2, with total masses from ∼14M⊙ for GW190924_021846 to ∼182M⊙ for GW190426_190642. Source properties calculated using our default prior suggest that the primary components of two new candidate events (GW190403_051519 and GW190426_190642) fall in the mass gap predicted by pair-instability supernova theory. We also expand the population of binaries with significantly asymmetric mass ratios reported in GWTC-2 by an additional two events (the mass ratio is less than 0.65 and 0.44 at 90% probability for GW190403_051519 and GW190917_114630 respectively), and find that two of the eight new events have effective inspiral spins χeff>0 (at 90% credibility), while no binary is consistent with χeff<0 at the same significance. We provide updated estimates for rates of binary black hole and binary neutron star coalescence in the local Universe
Adsorption of sodium alginate on calcium carbonate microparticles: Effect of molar mass and composition
No full textInternational audienceSodium alginate and calcium carbonate are natural materials with a high compositional variability. The molar mass and structural composition of sodium alginate and the crystalline structure of calcium carbonate can affect their interactions in pure water. Herein, we studied the adsorption of sodium alginate onto calcium carbonate microparticles in pure water. The adsorption appears to be driven by electrostatic interactions stabilized by calcium ions at the surface of the particles and in solution due to the dissolution of calcium carbonate in pure water. We found that the adsorption of sodium alginate is favored onto microparticles with more calcite. We also found that the adsorption of sodium alginate polymers with low guluronate content (∼ 30%) onto calcium carbonate particles tends to increase for polymers with lower average molar mass and more flexible polymer chains. Polymers with high guluronate content (∼ 65%), exhibiting a more rigid conformation, tend to adsorb more onto calcite when they present a lower average molar mass polymers as well as a higher proportion of guluronate monomers
Electromediated Alcohol-Based Passerini-Type Reaction
No full textInternational audienceAn electrochemical variant of the alcohol-based oxidative Passerini reaction is reported here. It relies on an indirect anodic oxidation process followed by a three-component coupling, in which TEMPO serves as a key redox mediator. This electrochemical approach permits to operate without the need for a metal catalyst nor oxygen atmosphere and allows the use of nonactivated alcohols as reaction partners. It could be applied to the preparation of good variety of α-acyloxy-carboxamides in yields ranging from 24% to 80%
Développement de méthodes de traitement de l’échantillon et d’analyse des hydrocarbures aromatiques polycycliques dans le sérum et le placenta humain
No full textPolycyclic aromatic hydrocarbons (PAHs) are persistent organic pollutants resulting from incomplete combustion of organic substances emitted into the environment through natural processes (forest fires, volcanic eruptions) and human activities (industries, road traffic, heating…) processes. These ubiquitous environmental pollutants are found in air, soil, water, sediment and food, leading to constant human exposure through skin contact, inhalation, or ingestion. PAHs are recognized as carcinogenic, mutagenic, and reprotoxic substances. Due to their high toxicity, 24 PAHs are regulated and monitored in the environment and/or food. Exposure to these compounds has deleterious effects on human health, particularly in at-risk populations such as pregnant women and their fetuses. Exposure to PAHs during pregnancy is linked to various pathologies (abortions, fetal malformations, prematurity), as well as disorders occurring in children and adults (delayed psychomotor development, asthma). To date, studies assessing maternal-fetal exposure are mainly carried out in blood and placental samples. They require both large sample quantities (0.5 to 10 mL of blood and 0.5 to 10g of placenta) and large volumes of solvents (> 10 mL), often toxic. Additionally, they are limited to the PAHs regulated in the environment (16 PAHs US-EPA). Monitoring these compounds at trace amounts in serum or placenta requires developing efficient extraction methods coupled with a resolution separation and a detection mode that is both sensitive and specific. The main challenge of this thesis was to design an extraction method for the 24 regulated PAHs, characterized by a wide range of polarities (3.3-7.7), from small sample quantities (100 µL of serum and 100 mg of placenta). The goal was to access epidemiological cohorts while minimizing extract dilution to maintain sensitivity. To achieve this, an approach combining a liquid-liquid extraction method using the salting-out effect (SALLE) followed by a dispersive liquid-liquid microextraction method based on the solidification of a floating organic drop (DLLME-SFO) was chosen. Samples were analyzed by liquid chromatography coupled with UV and fluorescence detection (LC-UV/FD). Parameters influencing the extraction yields of DLLME-SFO were studied on a miniaturized scale (1 mL of ultra-pure water). To remove proteins and other biological interferences from serum, SALLE was performed prior to the application of DLLME-SFO, thus purifying and preconcentrating the sample. The developed SALLE-DLLME-SFO method in serum was then adapted to the placenta. This required the development of a sample pretreatment step to extract PAHs from placenta which is semi-solid tissue. To assess the effectiveness of the developed methods, sensitivity, extraction yields, and method repeatability were determined in human serum and placenta enriched with PAHs. The quantification limits measured in serum and placenta allow for the quantification of concentrations observed in the literature, demonstrating the potential of these methods in studying PAH exposure.Les hydrocarbures aromatiques polycycliques (HAP) sont des polluants organiques persistants issus de la combustion incomplète de substance organique émis dans l'environnement par des processus naturels (feux de forêts, éruptions volcaniques) et anthropiques (industries, trafic routier, chauffage, …). Ces polluants ubiquitaires de l'environnement sont retrouvés dans l'air, les sols, les eaux et les aliments entrainant une exposition constante de l'Homme par contact cutané, inhalation ou ingestion. Les HAP sont reconnus comme des substances cancérigènes, mutagènes et reprotoxiques. En raison de leur forte toxicité, 24 HAP sont réglementés et surveillés dans l'environnement et/ou l'alimentation. L'exposition à ces composés à des effets néfastes sur la santé humaine en particulier au sein de populations à risque telles que celle des femmes enceintes et leur fœtus. L'exposition aux HAP au cours de la grossesse est à l'origine de nombreuses pathologies (avortements, malformations fœtales, prématurité, …) mais aussi des troubles survenant plus chez l'enfant et à l'âge adulte (retard du développement psychomoteur, asthme, …). A ce jour, les études menées pour évaluer l'exposition materno-fœtale sont principalement menées dans le sang et le placenta. Elles nécessitent à la fois de grande quantité d'échantillon (0,5 à 10 mL de sang et 0,5 à 10g de placenta) et consomment de grand volume de solvants (> 10 mL), le plus souvent toxiques. Aussi, elles se limitent seulement aux HAP réglementés dans l'environnement (16 HAP US-EPA). Leur suivi à l'état de traces dans le sérum ou le placenta impose de développer des méthodes d'extraction performantes et de les coupler à une séparation résolutive et à un mode de détection à la fois sensible et spécifique. Le principal défi de cette thèse a été de concevoir une méthode d'extraction pour les 24 HAP réglementés, caractérisés par une large gamme de polarités (3,3-7,7), à partir de petites quantités d'échantillons (100 µL de sérum et 100 mg de placenta). L'objectif était d'accéder à des cohortes épidémiologiques tout en limitant au maximum la dilution de l'extrait pour maintenir la sensibilité. Pour ce faire, une approche combinant une méthode d'extraction liquide-liquide par effet de sel (SALLE) suivie d'une méthode de microextraction liquide-liquide en mode dispersif basée sur la solidification d'une goutte flottante organique (DLLME-SFO) a été choisie. L'analyse des échantillons a été réalisée par chromatographie en phase liquide couplée à une détection UV et par fluorescence (LC-UV/FD). Les paramètres influençant les rendements d'extraction de la DLLME-SFO ont été étudiés à une échelle miniaturisée (1 mL d'eau ultra-pure). Pour éliminer les protéines et autres interférents biologiques du sérum, une SALLE a été effectuée avant l'application de la DLLME-SFO permettant ainsi de purifier et préconcentrer l'échantillon. La méthode développée de SALLE-DLLME-SFO dans le sérum a ensuite été adaptée au placenta. Cela a nécessité le développement d'une étape de prétraitement de l'échantillon permettant d'extraire les HAP du placenta qui est un tissu semi-solide. Pour évaluer l'efficacité des méthodes développées, la sensibilité, les rendements d'extraction et la répétabilité de la méthode ont été déterminés dans le sérum et le placenta humain enrichi en HAP. Les limites de quantification mesurées dans le sérum et le placenta permettent de quantifier les concentrations observées dans la littérature, démontrant ainsi le potentiel de ces méthodes dans l'étude de l'exposition aux HAP