Modulation de l'efficacité des hormones gonadotropes par l'intermédiaire d'anticorps potentialisants ou de modifications chimiques du ligand

Thèse confidentielle Diplôme : Dr. d'Universit

Modulation of gonadotrophin's biological efficacity by interaction with an enhancing antibody or by chemical modifications of agonist

L’hormone lutéinisante équine (eLH) et l’hormone choriogonadotrope équine (eCG) ont la particularité d’activer à la fois les récepteurs LH et FSH (hormone folliculo-stimulante) d’origine non-équine. Des modifications structurales des chaînes glycanniques de la eLH ou des changements conformationnels de la eCG induits par la liaison d’anticorps à effet potentialisant entraînent une modulation de leur efficacité biologique. Dans le but de comprendre les mécanismes moléculaires mis en place par ces 2 types de ligands, nous avons étudié leur impact sur les 2 voies de signalisation induites lors de l’activation du récepteur FSH (RFSH) : la voie dépendante des protéines G et la voie dépendante des b-arrestines. Nos résultats mettent en évidence que chaque type de nouvel agoniste peut entraîner un biais dans la signalisation de l’une ou des 2 voies impliquées et induire des effets sélectifs au niveau des voies de signalisation. Ces résultats ouvrent sur la perspective d’une nouvelle pharmacologie des hormones gonadotropes qui pourraient être de potentiels candidats thérapeutiques en médecine vétérinaire et humaine.The equine luteinizing hormone (eLH) and the equine chorionic gonadotrophin (eCG) activate cell signaling from two distinct GPCRs in non-equine species: the LH receptor (LHR) and the FSH (Follicle stimulating Hormone) receptor (FSHR). Structural modifications of glycanic chains of eLH or conformational modification of eCG consecutive to its interaction with an antibody (Ab) both led to a modulation of the native hormone’s biological efficacy. In order to understand the molecular mechanisms induced by the 2 type of hormone modifications, we have studied their impact on 2 of the signaling pathways triggered by FSHR: the G protein-and the b-arrestin-dependent pathway. Our results demonstrate that modifications induce signaling bias. We conclude that our finding might lead the way to a new generation of gonadotrophins’ modulating drugs with promising applications in animal breeding and in Human medicine

Bases moléculaires de la modulation de la réponse FSH et LH de la eCG induite par des anticorps anti-eCG potentialisants

National audienc

Selective modulation of follicle-stimulating hormone signaling pathways with enhancing equine chorionic gonadotropin/antibody immune complexes

 The injection of equine chorionic gonadotropin(eCG) in dairy goats induces the production of anti-eCG antibodies (Abs) in some females. We have previously shown that Abs negatively modulate the LH and FSH-like bioactivities of eCG, in most cases, compromising fertility in treated females. Surprisingly, we found out that some anti-eCG Abs improved fertility and prolificity of the treated females, in vivo. These Abs, when complexed with eCG, enhanced LH and FSH ability to induce steroidogenesis on specific target cells, in vitro. In the present study, we analyzed the impact of three eCG/anti-eCG Ab-enhancing complexes on two transduction mechanisms triggered by the FSH receptor: guanine nucleotide-binding protein alpha S-subunit/cAMP/protein kinase A (PKA) and beta-arrestin-dependent pathways, respectively. In all cases, significant enhancing effects were observed on ERK phosphorylation compared with eCG alone. However, cAMP production and PKA activation induced by eCG could be differently modulated by Abs. By using a pharmacological inhibitor of PKA and small interfering RNA-mediated knock-down of endogenous beta-arrestin 1 and 2, we demonstrated that signaling bias was induced and was clearly dependent on the complexed Ab. Together, our data show that eCG/anti-eCG Ab-enhancing complexes can differentially modulate cAMP/PKA and beta-arrestin pathways as a function of the complexed Ab. We hypothesize that enhancing Abs may change the eCG conformation, the immune complex acquiring new "biased" pharmacological properties ultimately leading to the physiological effects observed in vivo. The modulation of ligand pharmacological properties by Abs opens promising research avenues towards the optimization of glycoprotein hormone biological activities and, more generally, the development of new therapeutics. (Endocrinology 151: 2788-2799, 2010

Structure–function relationships of the variable domains of monoclonal antibodies approved for cancer treatment

Due to their exquisite specificity for a given epitope on the target antigen, recombinant monoclonal antibodies (rmAb) can deliver “targeted therapy” in oncology. This review focuses on the structural bases of “antigen specificity” to aid clinical researchers and pharmacologists in managing these new drugs. The fine structure of the Fv (Fragment variable) module (combination of VH and VL domains) from the five unconjugated antibodies currently approved for cancer treatment, namely rituximab, cetuximab, alemtuzumab, trastuzumab and bevacizumab, is presented and analysed. Co-crystal and functional studies are reviewed to define rmAb residues contributing to antigen binding site (paratope)–epitope interfaces. The genetic origin of these recombinant monoclonal antibodies, determined through the IMGT/3Dstructure-DB database and IMGT/V-QUEST (http://imgt.cines.fr), is presented, allowing the evaluation of homologies between antibodies and their closest germline human counterparts and hence their possible immunogenicity. Overall, the IMGT standards appear as a first and crucial step in the evaluation of recombinant antibodies

Selective ß-arrestin-dependent signalling by FSHR mutants

International audienc

Dual role of mitochondria in producing melatonin and driving GPCR signaling to block cytochrome c release

G protein-coupled receptors (GPCRs) are classically characterized as cell-surface receptors transmitting extracellular signals into cells. Here we show that central components of a GPCR signaling system comprised of the melatonin type 1 receptor (MT), its associated G protein, and β-arrestins are on and within neuronal mitochondria. We discovered that the ligand melatonin is exclusively synthesized in the mitochondrial matrix and released by the organelle activating the mitochondrial MT signal-transduction pathway inhibiting stress-mediated cytochrome release and caspase activation. These findings coupled with our observation that mitochondrial MT overexpression reduces ischemic brain injury in mice delineate a mitochondrial GPCR mechanism contributing to the neuroprotective action of melatonin. We propose a new term, automitocrine, analogous to autocrine when a similar phenomenon occurs at the cellular level, to describe this unexpected intracellular organelle ligand-receptor pathway that opens a new research avenue investigating mitochondrial GPCR biology