1,721,202 research outputs found
HSP90 inhibition as a means of radiosensitizing resistant, aggressive soft tissue sarcomas
No full textRadiotherapy is an essential part of multi-modal treatment for soft tissue sarcomas. Treatment failure is commonly attributed to radioresistance, but comprehensive analyses of radiosensitivity are not available, and suitable biomarkers or candidates for targeted radiosensitization are scarce. Here, we systematically analyzed the intrinsic radioresistance of a panel of soft tissue sarcoma cell lines, and extracted scores of radioresistance by principal component analysis (PCA). To identify molecular markers of radioresistance, transcriptomic profiling of DNA damage response regulators was performed. The expression levels of HSP90 and its clients ATR, ATM, and NBS1 revealed strong, positive correlations with the PCA-derived radioresistance scores. Their functional involvement was addressed by HSP90 inhibition, which preferentially sensitized radioresistant sarcoma cells and was accompanied by delayed γ-H2AX foci clearance and HSP90 client protein degradation. The induction of apoptosis and necrosis was not significantly enhanced, but increased levels of basal and irradiation-induced senescence upon HSP90 inhibition were detected. Finally, evaluation of our findings in the TCGA soft tissue sarcoma cohort revealed elevated expression levels of HSP90, ATR, ATM, and NBS1 in a relevant subset of cases with particularly poor prognosis, which might preferentially benefit from HSP90 inhibition in combination with radiotherapy in the future
Development of chemical protein glycosylation, multivalent carbohydrates, and folded peptide-PNA catalyst
No full textThis Ph.D. thesis is composed by three chapters based on different interests in chemical biology. The first chapter describes my work on the chemical glycosylation of proteins by cell compatible biorthogonal ligation. Current existing methods for the chemical glycosylation of proteins are powerful, however they are limited for purified proteins. Also, multiple synthetic steps are required to functionalize glycans of interest. The author has developed new synthetic methods to functionalize a series of native bioactive oligosaccharides in 1-2 steps. These neo glycans have enabled the glycosylation of proteins and other biomolecules in the ways that are applicable in live cells, which are tetrazine-strain alkene/alkyne cycloaddition and chelation assisted CuAAC. The second chapter describes my work on inhibition of multivalent interactions between glycans and pathogenic bacterial lectin. While multivalent interactions have been successfully blocked by glycopolymer and glycodendrimer approaches, a structurally well-designed ligand is required to enhance the potency and selectivity. A PNA encoded heteroglycan library was investigated to discover various sets of divalent glycan inhibitors for corresponding pathogenic lectins and DNA display showed structural information of ligands for preferential lectins. Furthermore, the interaction was also inhibited by PNA-programed dynamic glycan assembly. The dynamic glycan assembly was stabilized only when multivalent lectin is exists and showed nanomolar affinity to pathogenic lectin BambL. The assembly also successfully blocked the invasion of BambL to human lung epithelial cell H1299 in 723 fold effectively compared with monomeric glycan. 6 The third chapter describes my work on the development of constrained peptide catalysts folded through PNA hybridization. Folded short peptide catalysts have great potential to imitate enzyme active site and perform enzyme-like functions and ground breaking peptide catalysts have been reported to perform the diverse stereo and regio selective reactions with rate acceleration. The 1,000 member folded peptide-PNA library was screened to observe enhanced catalytic activity for phosphate bond hydrolysis. Identified sequence had >25 fold increased activity when the peptide structure was folded through PNA hybridization and the activity was moderated by strand displacement. Furthermore, to apply this concept for more broad chemistry, metallo peptide-PNA catalyst was designed and synthesized to hydrolyse metal chelating ester
From privileged natural product scaffolds to PNA-encoded chemical libraries
No full textEn croisant notre connaissance en synthèse de collections encodées par APN, notre expérience obtenue lors de la synthèse des quelques membres de la famille des lactones d'acide résorcylique, et notre constant intérêt dans l'inhibition de kinases et chaperonnes moléculaires, nous avons construit plusieurs collections moléculaires encodées par APN avec des pharmacophores variés incluant des hétérocycles, des acides aminés, des dérivés synthétiques de lactones d'acide résorcylique, et des médicaments autorisés par la FDA ou des fragments de ceux-ci, comme autant de points de diversité, dirigés contre différentes protéines cibles. La chaperonne moléculaire HSP70, les protéines contenant un bromodomaine et d'autres kinases ont toutes été exposées à ces collections encodées par APN et résolues spatialement sur micropuce ADN, démontrant ainsi la robustesse et la puissance de cette méthodologie de criblage dans une tentative de dépasser l'effet de goulot en découverte médicamenteuse
Extending the diversity of privileged natural product motifs : synthesis of a library of resorcylic acid lactones and studies towards the guaianes and pseudoguaianes
No full textLes macrolides du résorcinol présentant une fonction cis-enone se sont révélés être de puissants et irréversibles inhibiteurs de protéines kinases, des enzymes impliquées dans toutes les transductions de signaux dont le dysfonctionnement est à l’origine de pathologies telles que le cancer, les inflammations ou les maladies neurodégénératives. Attiré par leur activité biologique, notre laboratoire a développé une voie synthétique pour ces composés, basée sur la technologie des tags fluorés. La première partie de ma thèse a consisté à appliquer cette synthèse à l’élaboration d’une chimiothèque de 51 macrocycles. Cette dernière a permis d’identifier deux modifications permettant d’accroître l’activité biologique : l’ajout d’un carbone supplémentaire dans le macrocycle et d’un groupement hydroxyl en position β du diol. La seconde partie de ma thèse a consisté à développer une voie de synthèse générale permettant d’accéder à divers membres d’une autre classe de puissants inhibiteurs irréversibles : les lactones sesquiterpéniques. Ces composés présentent un large spectre de propriétés biologiques comme des propriétés cytotoxiques, anti-tumorales et anti-inflammatoires. Malheureusement la plupart des cibles biologiques et mécanismes associés à ces propriétés n’ont pas encore été identifiés. La voie de synthèse élaborée, basée sur un intermédiaire central bicyclique simple accessible en 13 étapes (avec pour étape clé une métathèse domino d’enyne) et inspirée de la biosynthèse, devrait permettre de développer des « composés outils » pour lever ces interrogations. Cette synthèse a notamment pu être appliquée à la préparation des molécules naturelles geigérine et 6-deoxy-geigérine.The resorcylic acid lactones bearing a cis-enone functionality proved to be potent and irreversible inhibitors of protein kinases, a class of enzymes implicated in every transduction pathways whose dysfunction is at the origin of pathologies ranging from oncology to inflammation and neurodegenerative diseases. Attracted by their important biological activity, our laboratory developed a synthetic pathway to these pharmacophores, based on the fluorous tags technology. The first part of my thesis consisted in applying this synthesis to the elaboration of a library of 51 macrocycles. This library allowed to identify two modifications as increasing the biological activity : the introduction of an extra carbon in the macrocycle and of a hydroxyl group in β position of the diol. The second part of my thesis consisted in developing a general synthetic pathway to access various members of another class of potent irreversible inhibitors : the sesquiterpene lactones. These compounds exibit a wide spectrum of biological activity including cytotoxic, anti-tumor and anti-inflammatory properties. Unfortunately most of the targets and mode of action associated with these properties have not been identified yet. The synthetic pathway developed, based on a simple central bicyclic intermediate achievable in 13 steps (with the key step being a domino enyne metathesis) and inspired by the biogenesis, should allow the development of "tool compounds" to address these questions. This synthesis has been applied in particular to the preparation of the natural products geigerin and 6-deoxy-geigerin
Diversity-oriented synthesis of pochonins: a privilege scaffold for ATPase and kinase inhibition
No full textLes pochonines A-F sont six molécules récemment décrites appartenant à la famille des lactones résorcyliques à 14 chaînons et qui présentent une activité contre le virus de l’herpès. Notre intérêt initial pour ces dérivés était fondé sur l’observation que plusieurs autres lactones résorcyliques sont connues pour leur potentiel biologique comme inhibiteurs de kinases ou d’ATPases. Par ailleurs le radicicol, de structure proche, est un puissant inhibiteur de la protéine de choc thermique HSP90, cible privilégiée pour la chimiothérapie en raison de son rôle fondamental dans la maturation fonctionnelle de nombreux oncogènes. Une synthèse de la pochonine C et sa conversion en radicicol ont été réalisées en respectivement sept et huit étapes, à partir de trois fragments facilement accessibles. Des synthèses alternatives de ces composés ont également été réalisées en utilisant des réactifs supportés et des réactions sur phase solide. A partir d’une analyse théorique de la dynamique moléculaire du radicicol et de ses analogues connus, une corrélation entre l’inhibition de HSP90 et l’énergie libre du conformère bioactif a été établie, permettant ainsi l’identification de la pochonine D comme inhibiteur potentiel de cette protéine. La synthèse totale de la pochonine D en utilisant des réactifs supportés a permis de confirmer cette hypothèse et de montrer que celle-ci est quasiment aussi active contre HSP90 que le radicicol (IC50 (radicicol) = 20 nM, IC50 (pochonine D) = 80 nM). Considérant les similarités structurales entre les pochonines D et A, nous avons également synthétisé la pochonine A pour son évaluation en tant qu’inhibiteur de HSP90, celle-ci présentant effectivement une activité de l’ordre du nanomolaire. La chimie développée à l’aide de réactifs supportés pour la synthèse des pochonines naturelles (A et D en particulier) nous a ensuite permis de préparer une banque de molécules fondée sur le même squelette et présentant cinq points de diversité, de manière à étudier les relations structure-activité sur différentes cibles. L’évaluation biologique de cette bibliothèque de 100 molécules pour l’inhibition d’un panel de 24 kinases et de HSP90 a révélé plusieurs composés actifs et sélectifs, démontrant ainsi le potentiel de la famille des lactones résorcyliques pour l’inhibition de kinases et d’ATPases.Pochonins A − F are six new members of the 14-membered resorcylic acid lactones (RALs) family which were identified in a Herpes Simplex Virus replication assay. Our interest in the pochonins stems from the observation that several other RALs are known to inhibit ATPases or kinases. Among them, radicicol was shown to be a potent inhibitor of HSP90 (Heat Shock Protein 90) whose activity is required for the functional maturation of a number of oncogenes, thus making this protein an attractive target for chemotherapy. A modular synthesis of pochonin C and its conversion to radicicol is presented. Both natural products are prepared in seven and eight steps, respectively, from three readily available fragments. Alternative syntheses of these compounds were also achieved using a combination of polymer-bound reagents and reactions on solid phase. Based on a molecular dynamics/minimization of radicicol and several known analogs, a correlation between the HSP90-inhibitory activity and the free energy of the bioactive conformer was established, leading to the identification of pochonin D as a potential inhibitor of HSP90. Its synthesis using polymer-bound reagents allowed us to confirm this finding and pochonin D was shown to be nearly as potent an HSP90 inhibitor as radicicol. Considering their closely-related structure, pochonin A was also synthesized and tested for HSP90 inhibition. The polymer-assisted chemistry developed for the pochonins syntheses enabled us to prepare a library based on the pochonin scaffold bearing five points of diversity, allowing us to extend beyond the modifications of the natural resorcylic acid lactones. Testing the library for its inhibition against a panel of 24 kinases at 10 µM and against HSP90 reveals several hits, thereby demonstrating the potential of the resorcylides towards the inhibition of therapeutically relevant kinases and ATPases. These syntheses of the pochonins were also the first reported in the literature allowing us to confirm structural assignments and to provide optical rotations as well as to define the stereochemistry of pochonin C’s carbon bearing the chlorine atom
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Programming ligand interaction and reactions with nucleic acid templated processes
No full textThis PhD thesis is focused on DNA and RNA templated reactions, using PNAs as reacting partners. The work deals with the application and improvement of a previously known reaction, in which a phenylazide is reduced by a Ru(II) photocatalyst in presence of a sacrificial reductant. We first demonstrated that this reaction can serve as a method to sense oligonucleotides in vivo. The same chemistry was applied to the development of a lateral flow assay for the enzyme-free detection of DNA from food samples. The combination of the use of a new substrate for the photoreduction and a precipitating dye enabled precise localization of the fluorophore and high speed of release. Bioluminescence was used as the source of photons to enable Ru(II) photocatalysis via BRET, for the release of small molecules at biological effective concentrations. Finally, a novel Ru(II) photocatalyst was prepared from an inactive Ru(II) precursor via oligonucleotide templated ligation
Peptide nucleic acids: synthesis, cellular uptake, encoding and templated reaction
No full textIn this work, a PNA synthetic methodology taking advantage of the Ugi four component reaction is described. Novel strategies for improving PNA cellular uptake are explored. PNA encoded libraries were screened against p97 and Akt/mSin1. Finally protein kinases were used to template an abiotic reaction
Supramolecular Networks Responsive to Proteins and Small Molecules
No full textThe thesis describes a broad range of responsive nanoscale devices and their application to biological systems. This thesis is composed of five projects aiming to develop responsive devices applicable in biological settings. The first four projects involve the design and application of supramolecular networks able to translate biological inputs, such as the presence of membrane proteins, luciferases or miRNA sequences, into functional outputs. The construction of the networks relies on peptide nucleic acid (PNA) technology, with a focus on PNA templated reactions capitalizing on the ruthenium catalysed photoreduction of picolinium immolative linkers. The last project describes the discovery of a small-molecule caged fluorophore that interacts with the motor protein kinesin-1 and responds by eliciting a fluorescent output. The systems described herein enable the visualisation of important biological processes such as motor protein movement, as well as cancer cell detection and targeted drug release
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