Functional characterization of the novel sequence variant p.S304R in the hinge region of TSHR in a congenital hypothyroidism patients and analogy with other formerly known mutations of this gene portion

Context: Thyroid dysgenesis may be associated with loss-of-function mutations in the thyrotropin receptor (TSHR) gene. Objectives: The aim of this study was to characterize a novel TSHR gene variant found in one patient harboring congenital hypothyroidism (CH) from a cohort of patients with various types of thyroid defects. Materials and methods: This cross-sectional cohort study involved 118 patients with CH and their family members, including 45 with familial and 73 with sporadic diseases. The thyroid gland was normal in 23 patients, 25 patients had hypoplasia, 25 hemithyroid agenesis, 21 had athyreosis, and 21 had ectopy. Genomic DNA was extracted, and 10 exons of the TSHR gene were amplified and sequenced. Mutations in other candidate genes were investigated. Ortholog alignment was performed, and TSHR functional assays were evaluated. Results: We identified one previously unknown missense variation in the hinge region (HinR) of the TSHR gene (p.S304R) in one patient with thyroid hypoplasia. This variant is conserved in our ortholog alignment. However, the p.S304R TSHR variant presented a normal glycosylation pattern and signal transduction activity in functional analysis. Conclusion: We report the ocurrence of a novel nonsynonymous substitution in the HinR of the large N-terminal extracellular domain of the TSHR gene in a patient with thyroid hypoplasia. In contrast with four others in whom TSHR mutations of the hinge portion were previously identified, the p.S304R TSHR variation neither affected TSH binding nor cAMP pathway activation. This TSHR gene variant was documented in a CH patient, but the current data do not support its role in the clinical phenotype

Les médicaments anti-angiogéniques dans le traitement du cancer

L'angiogénèse joue un rôle primordial dans les phénomènes de croissance tumorale et de formation des métastases. La première partie de cette thèse s'attache à décrire les mécanismes de l'angiogénèse physiologique chez l'Homme. La seconde partie est consacrée à l'angiogénèse tumorale, et plus particulièrement au rôle de l'hypoxie et du facteur de croissance de l'endothélium vasculaire (VEGF) dans ce processus. Enfin, la troisième partie s'intéresse aux médicaments ayant des propriétés anti-angiogéniques utilisées dans le traitement du cancer.CLERMONT FD-BCIU-Santé (631132104) / SudocLYON1-BU Santé (693882101) / SudocSudocFranceF

Modulation de l'expression de MYCN et de SNAP-25 au cours de la différenciation induite par le neuropeptide VIP dans des lignées de neuroblastome humain

Les neuroblastomes sont des tumeurs pédiatriques pouvant subir une régression spontanée ou une différenciation en leur homologue bénin. Le neuropeptide vasoactive intestinal peptide (VIP) est connu pour réguler la prolifération ou la différenciation de nombreuses cellules tumorales. In vitro, le VIP induit la différenciation de cellules de neuroblastome. Dans le travail décrit ici, les effets du VIP sur la modulation de l'expression de MYCN et de SNAP 25 (synaptosomal-associated protein of 25 kDa) ont été étudiés. Le gène MYCN code un facteur de transcription et est amplifié dans 22% des neuroblastomes. Cette amplification est associée à un mauvais pronostic. Les analyses par RT-PCR quantitative et par western-immunoblotting réalisées dans cette étude indiquent que le VIP diminue l'expression de MYCN, en particulier dans les cellules de neuroblastome humain IMR-32 présentant une amplification du gène MYCN. Cet effet du VIP a ensuite été comparé à celui de l'acide rétinoïque, molécule capable d'induire une différenciation associée à une diminution de l expression de MYCN. L'acide rétinoïque est actuellement utilisé dans la thérapie des formes les plus graves de ces cancers. Le VIP et l'acide rétinoïque diminuent l'expression de MYCN de façon complémentaire dans le temps. Des co-traitements montrent que ces deux molécules agissent en synergie sur la régulation de l expression de MYCN. SNAP-25 est une protéine SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor) impliquée dans la fusion membranaire associée à divers processus dont l'élongation neuritique. Les résultats obtenus dans la lignée de neuroblastome humain SH-SY5Y indiquent que le VIP augmente l'expression de la protéine SNAP-25 ainsi que le taux d ARNm des deux isoformes SNAP-25a et SNAP-25b. Les expériences d'interférence de l ARN réalisées mettent en évidence l'implication de SNAP-25 dans la neuritogénèse induite par le VIP. Ces travaux sur la modulation par le VIP de l'expression de MYCN et de SNAP-25 permettent une meilleure compréhension des mécanismes moléculaires de la différenciation induite par ce neuropeptide dans des cellules de neuroblastome humain. Ils suggèrent que le VIP, en favorisant une différenciation associée à une diminution de l'expression de MYCN, pourrait présenter un intérêt dans la thérapie des formes les plus graves de neuroblastomes.Neuroblastomas are pediatric tumors which can undergo spontaneous regression or differentiation into a benign form. The neuropeptide vasoactive intestinal peptide (VIP) is known to control proliferation or differentiation of numerous tumoral cells. In vitro, VIP induces differentiation of neuroblastoma cells. In the present work, the effects of VIP on the modulation of MYCN and of SNAP-25 (synaptosomal-associated protein of 25 kDa) expression were studied. The MYCN gene encodes a transcription factor and is amplified in 22% of neuroblastomas. This amplification is associated with poor prognosis. The quantitative RT-PCR and western immunoblotting analyses performed in this study indicate that VIP decreases MYCN expression, especially in the human MYCN-amplified IMR-32 cell line. This effect of VIP was next compared to that of retinoic acid, a molecule able to induce differentiation in association with a decrease of MYCN expression. Retinoic acid is at present used in therapy for aggressive forms of neuroblastoma. VIP and retinoic acid decrease MYCN expression with complementary kinetics. Cotreatments indicate that these two molecules act in synergy to regulate MYCN expression. SNAP-25 is a SNARE (soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor) protein involved in membrane fusion that is associated with various processes such as neuritic elongation. The results obtained in the human neuroblastoma cell line SH-SY5Y indicate that VIP increases the expression of SNAP-25 protein as well as the level of both SNAP-25a and SNAP-25b mRNA isoforms. RNA interference experiments show that SNAP-25 is involved in VIP-induced neuritogenesis. These studies on the modulation of MYCN and SNAP-25 expression by VIP allow a better understanding of the molecular mechanisms of VIP-induced differentiation in human neuroblastoma cells. They suggest that VIP, which induces differentiation in association with a decrease of MYCN expression, might have a therapeutic potential for the treatment of aggressive forms of neuroblastoma.POITIERS-BU Sciences (861942102) / SudocSudocFranceF

La régulation négative de la concentration sérique de la LH et de la FSH du foetus est associée à une perte de l'expression des kisspeptines et de KISS1R dans l'hypothalamus tubéral en fin de développement

National audienc

Negative fetal FSH/LH regulation in late pregnancy is associated with declined kisspeptin/KISS1R expression in the tuberal hypothalamus

Objective: Kisspeptins were recently identified as hypothalamic neuropeptides that control GnRH release at pubertal onset and in adults via the activation of KISS-1 receptor (KISS1R). Here, we have tested whether the fetal activation of the gonadotropic axis is related to the hypothalamic expression of kisspeptins and KISS1R.[br/] Design and Methods: LH and FSH levels were measured in fetal blood from the 15th week of gestation (WG) to birth. Immunohistochemistry was performed on the hypothalamus and pituitary at different developmental stages. Results: Immunostaining for kisspeptins and KISS1R appeared for both proteins in the hypothalamus as early as 15 WG and subsequently increased until 30-31 WG. In the meantime, serum LH and FSH levels decreased from postmenopausal levels in females or adult levels in males to very low levels. At full term, kisspeptin and KISS1R staining was still observed in the paraventricular, supraoptic, and ventromedial hypothalamic nuclei but not in the arcuate nucleus or median eminence. Hypothalamic GnRH staining was observed at 15 WG and did not vary after the first trimester. In an arhinencephalic fetus of 23 WG, very few GnRH neurons were observed in the hypothalamus, but serum FSH and LH levels were postmenopausal. Conclusion: Serum LH and FSH levels are independent from GnRH and kisspeptins at midgestation, and then GnRH progressively controls LH and FSH release. A shift from kisspeptin-independent to kisspeptin-dependent GnRH-induced LH and FSH release seems to occur after 30-31 WG. In addition to their function in adults, kisspeptins are also the master regulators of the gonadotropic axis activation in the fetus

PRR repeats in the intracellular domain of KISS1R are important for its export to cell membrane.: PRR repeats in intracellular domain of KISSIR

The National Institute of Health and Medical Research (INSERM, France) The Ministry of Research (France) The University Paris Diderot, Sorbonne Paris Cite (France) The National Institute of Blood Transfusion (INTS, France) The Laboratories of Excellence, GR-Ex (France)International audienceInactivating mutations of KISS-1 receptor (KISS1R) have been recently described as a rare cause of isolated hypogonadotropic hypogonadism transmitted as a recessive trait. Few mutations have been described, and the structure-function relationship of KISS1R remains poorly understood. Here, we have taken advantage of the discovery of a novel mutation of KISS1R to characterize the structure and function of an uncommon protein motif composed of 3 proline-arginine-arginine (PRR) repeats located within the intracellular domain. A heterozygous insertion of 1 PRR repeat in-frame with 3 PRR repeats leading to synthesis of a receptor bearing 4 PRR repeats (PRR-KISS1R) was found in the index case. Functional analysis of PRR-KISS1R showed a decrease of the maximal response to kisspeptin stimulation, associated to a lower cell surface expression without modification of total expression. PRR-KISS1R exerts a dominant negative effect on the synthesis of the wild-type (WT)-KISS1R. This effect was due to the nature of inserted residues but also to the difference of the length of the intracellular domain between PRR-KISS1R and WT-KISS1R. A molecular dynamic analysis showed that the additional PRR constrained this arginine-rich region into a polyproline type II helix. Altogether, this study shows that a heterozygous insertion in KISS1R may lead to hypogonadotropic hypogonadism by a dominant negative effect on the WT receptor. An additional PRR repeat into a proline-arginine-rich motif can dramatically changed the conformation of the intracellular domain of KISS1R and its probable interaction with partner proteins

Maternal residence near municipal waste incinerators and the risk of urinary tract birth defects.

International audienceOBJECTIVES: Waste incineration releases a mixture of chemicals with high embryotoxic potential, including heavy metals and dioxins/furans, into the atmosphere. In a previous ecological study we found an association between the risk of urinary tract birth defects and residence in the vicinity of municipal solid waste incinerators (MSWIs). The objective of the present study was to specifically test this association. METHODS: A population-based case-control study compared 304 infants with urinary tract birth defects diagnosed in the Rhône-Alpes region (2001-2003) with a random sample of 226 population controls frequency-matched for infant sex and year and district of birth. Exposure to dioxins in early pregnancy at the place of residence, used as a tracer of the mixture released by 21 active waste incinerators, was predicted with second-generation Gaussian modelling (ADMS3 software). Other industrial emissions of dioxins, population density and neighbourhood deprivation were also assessed. Individual risk factors including consumption of local food were obtained by interviews with 62% of the case and all control families. RESULTS: Risk was increased for mothers exposed to dioxins above the median at the beginning of pregnancy (OR 2.95, 95% CI 1.47 to 5.92 for dioxin deposits). When only interviewed cases were considered, risk estimates decreased mainly because the non-interviewed cases were more likely to live in exposed residential environments (OR 2.05, 95% CI 0.92 to 4.57). The results suggest that consumption of local food modifies this risk. CONCLUSIONS: This study confirms our previous observation of a link between the risk of urinary tract birth defects and exposure to MSWI emissions in early pregnancy and illustrates the effect of participation bias on risk estimates of environmental health impacts

Is older adults’ physical activity during transport compensated during other activities : comparing 4 study cohorts using GPS and accelerometer data

Introduction: Promoting active transport offers the potential to increase population physical activity levels. Compensation theories state that above-average physical activity in one activity is compensated in later activities; a mechanism that results in stable levels of total physical activity. Little is known about possible compensation of transport physical activity among older adults. Methods: GPS (Global Positioning System) and accelerometer data collected among older adults (65+) were pooled from four cohorts in Canada, Luxembourg, and France (n=636, collected between 2012 and 2016). Physical activity was measured as total volume of physical activity for trips and non-trip activities. Robust linear regressions on person-centered data were used to test within-person associations between transport and total physical activity. Results: 636 older adults-median age of 76 years, 49% women-provided accelerometer and GPS data for at least 4 days. 18% of the total volume of physical activity was related to transport. A positive association was found between physical activity during a trip and the physical activity during the next hour, among those with lower levels of regular physical activity. Negative associations -indicating partial compensation-were found between transport physical activity during a day, and both total physical activity during the next day and non-transport physical activity during the same day. No differences were found between the four study cohorts. Conclusions: Transport physical activity is compensated partially by older adults during nontransport physical activity. Given the presence of compensation, we strongly recommend evaluations of transport interventions to measure and analyze both non-transport and transport physical activity

Phase IB part of LOC-R01, a LOC network non-comparative randomized phase IB/II study testing R-MPV in combination with escalating doses of lenalidomide or ibrutinib for newly diagnosed primary central nervous system lymphoma (PCNSL) patients

International audienceBackground: Results of conventional induction chemotherapies in primary central nervous system lymphoma (PCNSL) need to be improved. Ibrutinib, a BTK inhibitor, and lenalidomide, an immunomodulatory drug, have shown promising results at relapse, supporting to further assess their individual use in combination with high-dose methotrexate-based chemotherapy.Methods: Patients with newly diagnosed PCNSL were randomized to receive four 28-day cycles of ibrutinib or lenalidomide in combination with R-MPV (rituximab, methotrexate, procarbazine, vincristine and prednisone) in a 3 + 3 design. Responders then received a consolidation with R-Cytarabine and an intensive chemotherapy with autologous stem cell transplantation. The objective of the phase IB study was to define the recommended phase II dose (RP2D) based on the dose-limiting toxicity (DLT) occurring during the first induction cycle.Results: Twenty-six patients (median age 52) were randomized. Four DLTs were observed: one grade 5 aspergillosis and pneumocystosis, one grade 4 catheter-related infection and two grade 3 increased alanine aminotransferase levels. RP2D of ibrutinib and lenalidomide were 560 mg daily (D3-14 and D17-28) and 15 mg daily (D1-21) respectively, in combination with R-MPV. In both arms, the most frequent grade ≥3 treatment-related adverse events were hepatic cytolysis, neutropenia and infections. One grade 4 Lyell's syndrome was reported at cycle 2 in the lenalidomide arm. After 4 induction cycles, the overall response rates were 76.9% and 83.3% in the lenalidomide and ibrutinib arm, respectively.Conclusion: Targeted induction therapies combining lenalidomide or ibrutinib with R-MPV are feasible for first-line PCNSL. The safety profile is consistent with the known safety profiles of R-MPV and both targeted therapies. The phase II part of the study is ongoing.Trial registration: NCT04446962. http://clinicaltrials.gov/show/NCT0444696