The NALCN ion channel is activated by M3 muscarinic receptors in a pancreatic β-cell line

A previously uncharacterized putative ion channel, NALCN (sodium leak channel, non-selective), has been recently shown to be responsible for the tetrodotoxin (TTX)-resistant sodium leak current implicated in the regulation of neuronal excitability. Here, we show that NALCN encodes a current that is activated by M3 muscarinic receptors (M3R) in a pancreatic β-cell line. This current is primarily permeant to sodium ions, independent of intracellular calcium stores and G proteins but dependent on Src activation, and resistant to TTX. The current is recapitulated by co-expression of NALCN and M3R in human embryonic kidney-293 cells and in Xenopus oocytes. We also show that NALCN and M3R belong to the same protein complex, involving the intracellular I–II loop of NALCN and the intracellular i3 loop of M3R. Taken together, our data show the molecular basis of a muscarinic-activated inward sodium current that is independent of G-protein activation, and provide new insights into the properties of NALCN channels

Particulate matter exposure during pregnancy is associated with birth weight, but not gestational age, 1962-1992: a cohort study

<p>Abstract</p> <p>Background</p> <p>Exposure to air pollutants is suggested to adversely affect fetal growth, but the evidence remains inconsistent in relation to specific outcomes and exposure windows.</p> <p>Methods</p> <p>Using birth records from the two major maternity hospitals in Newcastle upon Tyne in northern England between 1961 and 1992, we constructed a database of all births to mothers resident within the city. Weekly black smoke exposure levels from routine data recorded at 20 air pollution monitoring stations were obtained and individual exposures were estimated via a two-stage modeling strategy, incorporating temporally and spatially varying covariates. Regression analyses, including 88,679 births, assessed potential associations between exposure to black smoke and birth weight, gestational age and birth weight standardized for gestational age and sex.</p> <p>Results</p> <p>Significant associations were seen between black smoke and both standardized and unstandardized birth weight, but not for gestational age when adjusted for potential confounders. Not all associations were linear. For an increase in whole pregnancy black smoke exposure, from the 1^st(7.4 μg/m³) to the 25^th(17.2 μg/m³), 50^th(33.8 μg/m³), 75^th(108.3 μg/m³), and 90^th(180.8 μg/m³) percentiles, the adjusted estimated decreases in birth weight were 33 g (SE 1.05), 62 g (1.63), 98 g (2.26) and 109 g (2.44) respectively. A significant interaction was observed between socio-economic deprivation and black smoke on both standardized and unstandardized birth weight with increasing effects of black smoke in reducing birth weight seen with increasing socio-economic disadvantage.</p> <p>Conclusions</p> <p>The findings of this study progress the hypothesis that the association between black smoke and birth weight may be mediated through intrauterine growth restriction. The associations between black smoke and birth weight were of the same order of magnitude as those reported for passive smoking. These findings add to the growing evidence of the harmful effects of air pollution on birth outcomes.</p

Treatment of -mutated advanced or metastatic melanoma: rationale, current trials and evidence to date

The disease course of BRAF (v-raf murine sarcoma viral oncogene homolog B1)-mutant melanoma has been drastically improved by the arrival of targeted therapies. NRAS (neuroblastoma RAS viral oncogene homolog)-mutated melanoma represents 15–25% of all metastatic melanoma patients. It currently does not have an approved targeted therapy. Metastatic patients receive immune-based therapies as first-line treatments, then cytotoxic chemotherapy like carboplatin/paclitaxel (C/P), dacarbazine (DTIC) or temozolomide (TMZ) as a second-line treatment. We will review current preclinical and clinical developments in NRAS -mutated melanoma, and analyze ongoing clinical trials that are evaluating the benefit of different targeted and immune-based therapies, either tested as single agents or in combination, in NRAS -mutant melanoma

Évaluation du kit de PCR en temps réel MycoGENIE ® Pneumocystis jirovecii pour le diagnostic moléculaire de la pneumocystose

International audiencePneumocystis jirovecii (Pj) est un champignon pathogène opportuniste strictement humain. Il colonise transitoirement et sans symptôme l’arbre respiratoire des patients immunocompétents, et peut être responsable d’infections pulmonaires sévères chez les patients immunodéprimés. Le diagnostic biologique de pneumocystose repose sur la mise en évidence du pathogène à l’examen direct d’un prélèvement pulmonaire profond. Si l’examen direct manque souvent de sensibilité, de nombreuses techniques de PCR se sont développées pour améliorer la sensibilité de détection du champignon. Les techniques de PCR quantitatives (qPCR) permettent de quantifier la charge fongique dans le prélèvement, et potentiellement de différencier la colonisation de l’arbre respiratoire par Pj d’une véritable infection généralement associée à une charge fongique plus élevée.L’objectif de notre travail était d’évaluer les performances du kit MycoGENIE® P. jirovecii (Ademtech) dans le diagnostic de la pneumocystose en comparaison avec la technique de qPCR utilisée en routine au laboratoire de parasitologie-mycologie du CHU de Dijon.Soixante-dix-huit échantillons respiratoires (72 LBA, 3 ATP, 3 crachats) ont été testés en parallèle : 46 échantillons rétrospectifs (prélevés entre janvier 2016 et juin 2016 et conservés à −80 °C) et 32 échantillons prospectifs prélevés entre juillet 2016 et septembre 2016. Les 78 échantillons ont été extraits par l’automate Magtration System 12GC® (Bionobis). Les deux techniques de PCR utilisées dans notre étude ciblaient la même séquence d’ADN de 79 pb du gène mitochondrial codant la grosse sous-unité de l’ARN ribosomal (mtLSU rRNA). Les PCR ont été réalisées sur l’automate LC480 II (Roche Diagnostics). Le test MycoGENIE® Pj contenait une gamme étalon permettant de quantifier la charge fongique du prélèvement en nombre de copies/mL ainsi qu’un contrôle interne d’inhibition. Le kit est marqué CE-IVD.Sur les 46 échantillons testés rétrospectivement, 36 étaient positifs et 9 étaient négatifs par les deux techniques. Un seul résultat était discordant (négatif avec la technique de notre laboratoire et positif avec le kit MycoGENIE® Pj). Les résultats des 32 échantillons testés prospectivement étaient concordants (12 positifs et 20 négatifs). La concordance entre les 2 tests est donc de 98,7 %. Aucun inhibiteur n’a été détecté avec le kit MycoGENIE® Pj. La détection de l’ADN a été plus précoce pour l’ensemble des échantillons positifs avec le kit MycoGENIE® Pj, avec un seuil de détection plus précoce en moyenne de 1,2 cycle. Le résultat discordant concernait un patient atteint d’une leucémie aiguë myéloblastique en aplasie post-inductive intensive pour lequel la radiologie et la clinique orientaient le diagnostic vers une pneumopathie interstitielle hypoxémiante et bilatérale.La quantification de l’ADN n’a pas permis de définir de seuil pour différencier pneumocystose maladie et colonisation : des taux très faibles ont été retrouvés chez des patients dont le diagnostic retenu était une pneumocystose ; à l’inverse, des taux élevés d’ADN de Pj étaient observés chez des patients considérés comme colonisés. D’autres éléments comme le terrain, l’aspect radiologique, la clinique et d’autres marqueurs biologiques (ex. β-glucanes) devraient être pris en compte pour préciser le diagnostic de pneumocystose ou de colonisation à Pj

Association of Immunotherapy With Overall Survival in Elderly Patients With Melanoma

International audienceIMPORTANCE Melanoma treatment has been revolutionized with the development of immune-based therapies that offer durable clinical responses in a subset of patients. Clinical outcomes after treatment by immunotherapy can be influenced by the host's immune system. The immune system is modified with age by age-related immune dysfunction. OBJECTIVE To evaluate if age influences clinical outcome and immune adverse events in patients treated by immunotherapy for metastatic melanoma. DESIGN. SETTING, AND PARTICIPANTS This was a single-center cohort analysis in patients treated with immunotherapy for metastatic melanoma between January 2007 and February 2016, in the Lyon Sud Hospital, France, A total of 92 patients with metastatic melanoma treated with ipilimumab, nivoiumab,or pembrolizumab were retrospectively analyzed. MAIN OUTCOMES AND MEASURES Overall survival, progression-free survival, and immune-related adverse events were evaluated for each treatment line according to the patients' age. RESULTS A total of 92 patients were eligible and included in this study for a total of 120 lines of treatment. Fifty-four patients were included in the cohort that was 65 years or younger (24 [44%] were female; mean [SD] age, 48.1 [12.5] years), and 38 patients were included in the cohort that was older than 65 years (12 [34%] were female; mean [SD] age, 74.8 [6.9] years). Mean follow-up duration starting at treatment initiation was 12.5 months. Patients older than 65 years treated with immunotherapy had a better mean progression-free survival (4.8 vs 3.4 months; P = .04) and overall survival (not reached vs 10.1 months; P = .009) than younger patients in univariate analysis, and after adjusting on prognosis covariates. This was particularly true with patients treated with anti-programmed cell death protein1. Common immune-related adverse effects were similar in both cohorts. CONCLUSIONSAND RELEVANCE Age might be associated with a better clinical outcome after treatment with immunotherapy in the real-life setting. In our cohort, older patients did not have more immune-related adverse events. Further studies are warranted to confirm our results and describe the underlying mechanisms involved

Graves’ Disease during Immune Checkpoint Inhibitor Therapy (A Case Series and Literature Review)

International audienceThyrotoxicosis is an adverse event associated with immune checkpoint inhibitors (ICPis) that occurs in 0.6 to 3.2% of treated patients, depending on ICPi class. Presentation usually consists of a biphasic thyroiditis with transient thyrotoxicosis and secondary hypothyroidism. ICPi-induced Graves’ disease (GD), due to the stimulating activity of TSH-receptor autoantibodies (TRAb), is extremely rare. The aim of this retrospective study was to describe the characteristics and evolution of GD during ICPi therapy. Five among 243 patients followed for ICPi-induced thyrotoxicosis showed TRAb positivity (2% of the cohort). GD occurred quickly after initiation of ICPis; its course was typical for two patients, with prolonged requirement for antithyroid drug treatment (ATD). The three other patients experienced biphasic thyroiditis with secondary hypothyroidism requiring long-term substitution. Three other patients had a diagnosis of GD before starting ICPis; they evolved toward hypothyroidism with early cessation of ATD and long-term substitution treatment during ICPi treatment. None developed significant Graves’ orbitopathy. ICPi treatment was not interrupted for thyroid dysfunction. In conclusion, GD is a rare, immune-related adverse event of ICPis with an unusual course and frequent evolution to biphasic thyroiditis. In the case of ICPi-induced thyrotoxicosis in the presence of TRAb, observing the spontaneous evolution and performing a scintigraphy are useful before starting ATD treatment. Pre-existing GD is not exacerbated by ICPis and tends to evolve towards hypothyroidism. ICPi treatment can be maintained with adequate biochemical surveillance

Preclinical studies on the mechanism of action and the anti-lymphoma activity of the novel anti-CD20 antibody GA101.

International audienceGA101 is a novel glycoengineered Type II CD20 monoclonal antibody. When compared with rituximab, it mediates less complement-dependent cytotoxicity (CDC). As expected for a Type II antibody, GA101 appears not to act through CDC and is more potent than the Type I antibody rituximab in inducing cell death via nonclassical induction of apoptosis cytotoxicity, with more direct cytotoxicity and more antibody-dependent cell-mediated cytotoxicity. We evaluated the antitumor activity of GA101 against the human-transformed follicular lymphoma RL model in vivo in severe combined immunodeficient mice (SCID) mice. GA101 induced stronger inhibition of tumor growth than rituximab. Combination of GA101 with cyclophosphamide in vivo confirmed the superiority of GA101 over rituximab. Neutralizing the complement system with cobra venom factor partially impaired the antitumor activity of rituximab, but had no impact on the efficacy of GA101. In vitro GA101 more potently induced cell death of RL cells than rituximab. The expression of a limited number of genes was found to be induced by both antibodies after exposure in vitro. Among these, early growth response 1 and activation transcription factor 3 were confirmed to be increased at the protein level, suggesting a possible role of these proteins in the apoptotic signalling of anti-CD20 antibodies. These data imply that GA101 is superior to rituximab not only as a single agent, but also in combination with chemotherapy. These data suggest the presence of novel signalization pathways activated after exposure to anti-CD20 antibodies

Management of adjuvant settings for Stage III melanoma patients in France prior to checkpoint inhibitors: epidemiological data from the RIC-Mel database

International audienceBackground: Targeted therapies such as BRAF and MEK inhibitors and immunotherapies have been made available to treat melanoma.Objectives: To provide an overview of the management of the French Stage III melanoma population after complete lymph node resection prior to new adjuvant therapies.Materials and methods: A subgroup data analysis.Results: Data from 1,835 patients were analysed (15.58% Stage IIIA, 39.24% Stage IIIB, 43.92% Stage IIIC and 1.25% Stage IIID). Superficial spreading melanoma was the most frequent (70.98% in Stage IIIA for whom mutation analysis was performed; BRAF mutation was identified in up to 62% Stage IIIA patients). Sentinel lymph node biopsy was performed in 88.46% of Stage IIIA patients, 42.36% of Stage IIIB, 53.97% of Stage IIIC and 34.78% of Stage IIID. Up to 80% of Stage IIIA patients had no adjuvant treatment follow-up. Ulceration (p = 0.004; RR: 2.98; 95% CI: 1.4-6.3) and age at diagnosis (p = 0.0002; RR: 1.04; 95% CI: 1.02-1.06) were significant predictive factors for survival. Adjuvant interferon-α was administered in up to 13.04% of Stage IIID patients.Conclusion: Only a small number of Stage III melanoma patients were treated with interferon-α in adjuvant settings. New adjuvant therapies are currently having an effect on clinical practice in France, increasing survival and decreasing cost

A targeted genomic alteration analysis predicts survival of melanoma patients under BRAF inhibitors

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