1,721,037 research outputs found
Production of antibody derivatives in the methylotrophic yeast Pichia pastoris
No full textNew antibody derivatives are continuously being generated to interact with a range of therapeutic targets. The cost-effective and efficient production of these and other antibody derivatives is crucial for their further success. Here, we describe the construction of the expression vectors needed for heterologous expression of a Fab fragment in the yeast Pichia pastoris. The experimental conditions for lab-scale expressions are discussed, and an overview of an efficient purification strategy is presented
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
Generation and study of the mode of action immunomodulatory antibodies multispecies derived from nanobodies targeting inhibitory receptors CD96, TIGIT and PD-L1
No full textLa compréhension des interactions entre les cellules tumorales et leur environnement immunitaire a conduit au développement remarquable de nouveaux agents thérapeutiques basés sur des anticorps bloquants, ciblant des récepteurs inhibiteurs des lymphocytes T tels que CTLA-4 et PD-1. Le succès de ces thérapies est cependant limité par la proportion de patients y répondant. D'autres récepteurs modulateurs des cellules T et Natural Killer (NK), dont TIGIT et CD96, font l'objet de recherches afin d'augmenter l'efficacité et les taux de réponse des thérapies ciblant les checkpoint immunitaires.TIGIT et CD96 sont des récepteurs exprimés par les T et les NK infiltrant les tumeurs et ont le même ligand, PVR. Leur expression est associé à l'exhaustion des cellules immunitaires effectrices et à la progression tumorale.Dans ce projet, l'objectif était de développer de nouveaux formats de molécules multispécifiques compactes basés sur des nanobodies, afin de cibler simultanément plusieurs récepteurs immunomodulateurs.Nous avons généré un panel de nanobodies ciblant TIGIT, CD96 et PD-L1 et les avons utilisés en tant que briques de constructions pour produire différentes molécules bispécifiques. En particulier, nous avons produit des tandem bispécifiques TIGIT x CD96 qui conservent les propriétés d'adhésion des deux nanobodies impliqués; ainsi que des molécules anti-TIGIT ou TIGIT x PD-L1 contenant un fragment Fc afin d'induire une forte lyse anticorps dépendante (ADCC) contre les cellules PD-L1+ et les Tregs.The understanding of interactions between tumor cells and theirimmune environment had led to the outstanding development of newcancer therapies based on blocking antibody targeting T cell immunecheckpoint inhibitory receptors such as PD-1 and CTLA-4. Thesuccess of these therapies is unfortunately limited by the proportion ofresponding patients. Other T and natural killer (NK) cells modulatoryreceptors are now being investigated to increase the efficacy andresponse rate of immune checkpoint therapies and propose newcombination therapies, among which TIGIT and CD96.TIGIT and CD96 are two receptors expressed by tumor-infiltrated NKand T cells and bind to PVR. They are associated to exhaustion andtumor progression.In this study we aimed at developing new nanobody-based formats ofmultispecific molecules to simultaneously target several immunecheckpoint receptors with compact molecules. We obtained a panel ofTIGIT, CD96 and PD-L1 binding nanobodies and used them asbuilding brick to generate bispecific molecules. Particularly weproduced anti TIGIT x CD96 tandems that retained their bindingability on both targets as well as Anti TIGIT x PD-L1-Fc and antiTIGIT-Fc formats that were able to induce strong ADCC on PD-L1+cells and Tregs, respectively
Nanobody
Full text linkNanobodies have become outstanding tools for biomedical research, diagnostics and therapy. Recent advances in the identification and functionalization of target-specific nanobodies now make nanobody-based approaches broadly available to many researches in the field. This book provides a compilation of original research articles and comprehensive reviews covering important and up to date aspects of research on nanobodies and their applications for immunoassays, proteomics, protein crystallization and in vitro and in vivo imaging
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Development of tools for the study of G protein-coupled receptor-mediated signaling based on the use of lama single domain antibodies
No full textL'objectif principal de ma thèse était de développer de nouvelles technologies et des outils pour l’étude de l’activation des récepteurs couplés aux protéines G (GPCR).À la surface de la cellule se trouve une multitude de récepteurs qui jouent un rôle critique dans la communication cellule-cellule, dont les GPCR, une famille de récepteur utilisant les protéines G intracellulaires pour transmettre leurs signaux. Le ciblage de ces récepteurs à des fins thérapeutique est innovant et très prometteur. Mais à ce jour seuls quelques médicaments ciblant les GPCR ont été mis sur le marché, en partie en raison d'un manque d'outils permettant le suivi de leur action sur les cellules natives.L’objectif de cette thèse est donc de développer des tests simples pour suivre l’activation de n’importe quel GPCR. Pour développer ce type de test, nous avons décidé d'utiliser des fragments d'anticorps appelés nanobodies. Les anticorps sont des protéines du sang produites en réponse à un antigène spécifique qui sont capable de le neutraliser. Les nanobodies correspondent au domaine variable de certains anticorps de camélidés. En raison de leur faible taille (13 kDa) et de leur site de liaison à l'antigène réduit, les nanobodies se lient souvent à des cavités et présentent une grande sensibilité aux changements de conformation de l'antigène.The main objective of my thesis was to develop technologies and tools to study activation of G protein-coupled receptors (GPCRs).The cell surface is displaying a multitude of receptors, who play critical roles in cell-cell communication. Among them, GPCRs represent a large family relying on the use of intracellular G proteins for their signaling. Targeting these receptors for therapies is very promising and innovative. So far, only few new drugs have been put on the market, partly due to a lack of tools enabling the follow-up of their action on native cells.The aim of this thesis is thus to develop simple assays to study activation of any GPCRs. To develop this kind of test, we used antibody fragments called nanobodies. Antibodies are blood protein produced in response to and counteracting a specific antigen. Nanobodies correspond to antibody fragments derived from the variable domain of a special class of camelid antibodies. Because of their small size (13 kDa) and reduced antigen binding site, nanobodies often bind cavities and show a high sensitivity to antigen conformational changes
Generation of nanobodies for immunotherapy applications
No full textL’immunothérapie est une approche thérapeutique qui consiste à restaurer les fonctions anti-tumorales du système immunitaire pour qu’il soit à nouveau capable d’éliminer les cellules cancéreuses. Pour cela, différentes stratégies sont étudiées : l’une d’entre elles consiste à cibler des récepteurs inhibiteurs présents à la surface des cellules immunitaires infiltrées dans les tumeurs de façon à réactiver leurs fonctions. L’entreprise Innate Pharma développe des anticorps monoclonaux capables de restaurer les fonctions immunitaires de cellules effectrices de l’immunité innée. Le but de la collaboration était de générer des sdAbs (single-domain Antibodies), fragments d’anticorps dérivés de camélidés, capables de bloquer des interactions de type « immune checkpoint ». Ces anticorps à domaine unique sont notamment capables de lier des épitopes inaccessibles aux anticorps conventionnels tels que des cavités. Dans ce manuscrit, les résultats concernant deux de ces projets sont exposés : la modulation de l’axe inhibiteur SIRPα/CD47 et le blocage de l’interaction réalisée entre les récepteurs Siglec-7/-9 et leurs ligands sialylés. Lors de ces travaux, plusieurs sdAbs ciblant les récepteurs d’intérêt ont été sélectionnés grâce à la technologie du phage display. Ces molécules monovalentes ont été caractérisées puis clonées en différents formats multivalents de façon à améliorer leur affinité par effet d’avidité, et ainsi potentialiser leur efficacité de blocage. Enfin, plusieurs tests fonctionnels ont été réalisés pour évaluer l’effet de ces molécules sur les fonctions effectrices de différentes cellules du système immunitaire.Immunotherapy is a therapeutic approach which consists in restoring anti-tumoral functions of the immune system for eliminating cancer cells. For this, several strategies are developed: one of them is to target inhibitory receptors at the surface of effector cells in order to reactivate their functions in the tumor microenvironment.The company Innate pharma develops monoclonal antibodies able to restor immune functions of innate effector cells. The aim of the collaboration was to generate single-domain Antibodies (sdAbs), antibody fragments derived from camelids, which have the capacity of blocking interactions such as immune checkpoints. These sdAbs have several useful characteristics in terms of stability, production and especially in epitope binding. Indeed, because of their small size, they are able to bind on epitopes which are not accessible to conventional antibodies.In this manuscript, the results of two projets are reported: the modulation of the inhibitory axis SIRPα/CD47 and the blocking of the interaction between the receptors Siglec-7/-9 and their sialylated ligands. In these studies, several sdAbs targeting the receptors of interest were isolated by selection using phage display technology. These monovalent molecules were characterized to determine their specificity and estimate their binding and blocking capacities. Best candidates were cloned into several multivalent formats to optimize their affinity by avidity effect and to potent their blocking efficacy. Finally, several functional assays were performed to evaluate the efficacy of these multivalent constructions to restore immune functions of several effector cells
Nanobody
No full textNanobodies have become outstanding tools for biomedical research, diagnostics and therapy. Recent advances in the identification and functionalization of target-specific nanobodies now make nanobody-based approaches broadly available to many researches in the field. This book provides a compilation of original research articles and comprehensive reviews covering important and up to date aspects of research on nanobodies and their applications for immunoassays, proteomics, protein crystallization and in vitro and in vivo imaging
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