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International audience; Phytochemicals contribute to the health benefits of plant-rich diets, notably through their antioxidant and anti-inflammatory effects. However, recommended daily amounts of the main dietary phytochemicals remain undetermined. We aimed to estimate the amounts of phytochemicals in a well-balanced diet. A modelled diet was created, containing dietary reference intakes for adults in France. Two one-week menus (summer and winter) were devised to reflect typical intakes of plant-based foods. Existing databases were used to estimate daily phytochemical content for seven phytochemical families: phenolic acids, flavonoids (except anthocyanins), anthocyanins, tannins, organosulfur compounds, carotenoids, and caffeine. The summer and winter menus provided 1607 and 1441 mg/day, respectively, of total polyphenols (phenolic acids, flavonoids, anthocyanins, and tannins), the difference being driven by reduced anthocyanin intake in winter. Phenolic acids, flavonoids (including anthocyanins), and tannins accounted for approximately 50%, 25%, and 25% of total polyphenols, respectively. Dietary carotenoid and organosulfur compound content was estimated to be approximately 17 and 70 mg/day, respectively, in both seasons. Finally, both menus provided approximately 110 mg/day of caffeine, exclusively from tea and coffee. Our work supports ongoing efforts to define phytochemical insufficiency states that may occur in individuals with unbalanced diets and related disease risk factors.

Introduction: In the JAVELIN Lung 200 trial, avelumab (anti-programmed death-ligand 1 [PD-L1] antibody) did not significantly prolong overall survival (OS) versus docetaxel in patients with platinum-treated PD-L1+ NSCLC. We report greater than 2-year follow-up data. Methods: Patients with stage IIIB or IV or recurrent NSCLC with disease progression after platinum-doublet chemotherapy were randomized 1:1 to avelumab 10 mg/kg every 2 weeks or docetaxel 75 mg/m(2) every 3 weeks. The primary end point was OS in patients with PD-L1+ tumors (greater than or equal to 1% tumor cell expression; IHC 73-10 pharmDx assay). Results: Of 792 patients, 529 had PD-L1+ tumors (264 versus 265 in the avelumab versus docetaxel arms, respectively). As of March 4, 2019, median duration of follow-up for OS in the PD L1+ population was 35.4 months in the avelumab arm and 34.7 months in the docetaxel arm; study treatment was ongoing in 25 (9.5%) versus 0 patients, respectively. In the PD-L1+ population, 2-year OS rates (95% confidence interval [CI]) with avelumab versus docetaxel were 29.9% (24.5%-35.5%) versus 20.5% (15.6%-25.8%); in greater than or equal to 50% PD L1+ subgroups, 2-year OS rates were 36.4% (29.1%-43.7%) versus 17.7% (11.8%-24.7%) and in the greater than or equal to 80% subgroup were 40.2% (31.3%-49.0%) versus 20.3% (12.9%-28.8%), respectively. Median duration of response (investigator assessed) was 19.1 months (95% CI: 10.8-34.8) versus 5.7 months (95% CI: 4.1-8.3). Safety profiles for both arms were consistent with the primary analysis. Conclusions: Although the JAVELIN Lung 200 primary analysis (reported previously) revealed that avelumab did not significantly prolong OS versus docetaxel in patients with platinum-treated PD-L1+ NSCLC, posthoc analyses at 2 years of follow-up revealed that 2-year OS rates were doubled with avelumab in subgroups with higher PD-L1 expression (greater than or equal to 50% and greater than or equal to 80%). (C) 2021 International Association for the Study of Lung Cancer. Published by Elsevier Inc., EMD Serono Research & Development Institute, Inc., an affiliate of Merck KGaA, Darmstadt, Germany; Pfizer; Merck KGaA, LL This study was sponsored by EMD Serono Research & Development Institute, Inc., an affiliate of Merck KGaA, Darmstadt, Germany, as part of an alliance between Merck KGaA and Pfizer. Employees of the sponsor are coauthors of this manuscript and contributed to the design, execution, and interpretation of the analyses being reported, writing the report, and the decision to submit the article for publication, along with the other coauthors. The authors thank the patients and their families; the investigators, coinvestigators, and study teams at each participating center and at Merck KGaA, Darmstadt, Germany, and EMD Serono Research & Development Institute, Inc., Billerica, MA, an affiliate of Merck KGaA, Darmstadt, Germany, and Quintiles (Durham, NC) . Medical writing support was provided by Abhijith Thippeswamy of ClinicalThinking and funded by Merck KGaA and Pfizer.

International audience; Background : Chronic hand eczema (CHE) is the most common skin disorder affecting the hands. It causes major physical and psychological burden for patients. Classification of CHE remains challenging because of its aetiological and clinical heterogeneity. Objectives : Using latent class analysis (LCA) on a large categorical data set, our aim was to identify distinct phenotypes in a cohort of unselected CHE patients based upon clinical, genetic, molecular and physical parameters of the affected skin. Methods : We performed two independent LCA on a cohort of 71 well‐characterized patients that initially integrated clinical severity, total immunoglobulin E plasma level, transepidermal water loss, hydration index, interleukin(IL)‐8 lesional skin level, Staphylococcus (aureus and epidermidis) colonization, FLG genotype and the expression (mRNA) of genes involved either in the filaggrin degradation and the natural moisturizing factor synthesis, the cornified envelope formation, the tight junctions’ structure and the desquamation process, or encoding antimicrobial peptides and chemokines. Results : The first LCA categorized patients into a group displaying high severity of CHE, high skin barrier impairment, high Staphylococcus colonization, high IL‐8 skin level and high frequency of mutation in the FLG gene and a second group with opposite characteristics. The second LCA identified two independent groups of patients categorized by their low or high level of skin barrier impairment and corresponding changes in the expression of the related genes.Conclusions : Our study suggests that the degree of skin barrier dysfunction is the most important parameter to discriminate CHE patients and probably plays a pivotal role in the pathogenesis of the disease whatever the aetiological factors. As far as we know, this is the first study to address this topic using a statistical categorization method without preconception.

International audience; Advances in sequencing, bioinformatics and analytics now allow the structure, function and interrelations of whole microbial communities to be studied in greater detail. Collaborative efforts and multidisciplinary studies, crossing the boundary between environmental and medical microbiology, have allowed specific environmental, animal and human microbiomes to be characterized. One of the main challenges for microbial ecology is to link the phylogenetic diversity of host-associated microbes to their functional roles within the community. Much remains to be learned on the way microbes colonize the skin of different living organisms and the way the skin microbiome reacts to the surrounding environment (air, water, etc.). In this review, we discuss examples of recent studies that have used modern technology to provide insights into microbial communities in water and on skin, such as those in natural resources (thermal spring water), large mammals (humpback whales) and humans (the skin microbiome). The results of these studies demonstrate how a greater understanding of the structure and functioning of microbiota, together with their interactions with the environment, may facilitate the discovery of new probiotics or postbiotics, provide indicators for the quality of the environment, and show how changes in lifestyle and living environment, such as urbanization, can impact on the skin microbiome and skin health and disease in humans. Key Points Multidisciplinary approaches and advances in sequenc-ing, bioinformatics and analytics have allowed the diversity of whole microbial communities to be investigated. Studies of the microbiomes of natural resources, such as spring water, have the potential to uncover targets for the development of new probiotics or postbiotics. Analyses of the microbiomes of animals can provide vital information about the health status of the planet, whereas studies of the human microbiome can help determine the impact of living environments on human health and disease.

International audience; Background: Chronic hand eczema (CHE) is a major burden for patients. Mainte-nance treatment involves prevention measures limiting detrimental behaviour and aggravating factors. Objective: To evaluate the effect of a standardised care program including therapeu-tic patient education (TPE) on hand care behaviours, clinical severity, quality of life, and work productivity.Methods: A single-centre study was conducted prospectively. Together with the pre-scription of a topical steroid, patients participated in individual TPE sessions. Evalua-tions were performed initially and repeated three months after the therapeutic intervention. They included a structured analysis of hand care behaviours, the assess-ment of the mTLSS (modified Total Lesion Symptom Score), DLQI (Dermatology Life Quality Index), and WPAI (Work Productivity and Activity Impairment). Results: Seventy-one patients were included (30 men, 42.3%). Three months after completion of the standardised care program, hand care behaviours such as hand washing and rinsing, hand drying, wearing protective gloves, using moisturizing cre-ams, and following specific treatments and recommendations for CHE improved sig-nificantly in the 58 patients who completed the study and were associated with a significant improvement in the mTLSS, DLQI, and WPAI scores. Conclusions: TPE helps patients change their hand care behaviours and adopt skin protection measures, and may improve CHE severity, quality of life, and work productivity.

International audience; Background: A variety of human skin disorders is characterized by defects in the epidermal barrier, leading to dehydration, itchiness, and rashes. Previously published literature suggests that microtubule stabilization at the cortex of differentiating keratinocytes is necessary for the formation of the epidermal barrier. Objectives: We tested whether stabilization of microtubules with paclitaxel or epothilone B can repair barrier defects that were experimentally induced in three-dimensional culture models of epidermis. Methods: We established two models of defective epidermis in vitro, using three-dimensional cultures of primary human keratinocytes on filter supports: immature reconstructed human epidermis (RHE), and RHE that was compromised by treatment with inflammatory cytokines, the latter mimicking defects seen in atopic dermatitis. Results: Both paclitaxel and epothilone B promoted keratinocyte differentiation, accumulation of junctional proteins at the cell cortex, and the early appearance of lamellar bodies in immature RHE, whereas destabilization of microtubules by nocodazole had the reverse effect. Moreover, stabilization of microtubules rescued the barrier after cytokine treatment. The rescued barrier function correlated with the restoration of filaggrin and loricrin protein levels, the cortical accumulation of junctional proteins (E-cadherin, β-catenin, and claudin-1), and with the secretion of lamellar bodies. Conclusions: Our data suggest that the microtubule network is important for the formation of the epidermis, and that stabilization of microtubules promotes barrier formation. Microtubule stabilization may support regeneration of damaged skin, by restoring or improving the barrier.

The formation of DNA photoproducts caused by solar UVR exposure needs to be investigated in-vivo and in particular in order to assess sunscreens' level of protection against solar genotoxicity. The study's purposes were: i) to evaluate if the roof of suction blisters is an appropriate sampling method for measuring photoproducts, and ii) to measure in-vivo sunscreen protection against cyclobutane pyrimidine dimers. Skin areas on the interior forearms of eight healthy volunteers were exposed in-vivo to 2 MED of simulated solar radiation (SSR) and to 15 MED on a sunscreen protected area. After irradiation, six suction blisters were induced and the blister roofs were collected. Analysis of SSR-induced CPDs was performed by two independent methods: a chromatography coupled to mass spectroscopy (HPLC-MS/MS) approach and a 3D-imaging of CPD immunostaining by multiphoton microscopy on floating epidermal sheets. HPLC-MS/MS analyses showed that SSR-unexposed skin presented no CPD dimers, whereas 2 MED SSR-exposed skin showed a significant number of TT-CPD. The sunscreen covered skin exposed to 15 MED appeared highly protected from DNA damage, as the amount of CPD-dimers remained below the detection limit. The multiphoton-immunostaining analysis consistently showed that no CPD staining was observed on the non-SSR-exposed skin. A significant increase of CPD staining intensity and number of CPD-positive cells were observed on the 2 MED SSR-exposed skin. Sunscreen protected skin presented a very low staining intensity and the number of CPD-positive cells remained very close to non-SSR-exposed skin. This study showed that suction blister samples are very appropriate for measuring CPD dimers in-vivo, and that sunscreens provide high protection against UVR-induced DNA damage.

AIMS: Vinflunine is a novel tubulin‐targeted inhibitor indicated as a single agent for the treatment of bladder cancers after failure of prior platinum‐based therapy. Its pharmacokinetics (PK) and pharmacodynamics (PD) have been independently characterized through several phase I and phase II studies. However, no global pharmacometric analysis had been conducted as yet. METHODS: Vinflunine concentrations and safety data from 18 phase I and phase II studies were used to conduct population PK and PK/PD analysis, using Nonmem. A four‐compartment model was used to describe vinflunine PK and several covariates were tested to explain interindividual variability. In terms of PK/PD relationship, a semiphysiological population PK/PD model was applied to describe time course of absolute neutrophil counts (ANC) after vinflunine administration and logistic regression models were used to test the relationship between vinflunine exposure and toxicities. RESULTS: Vinflunine clearance is explained by creatinine clearance, body surface area and combination with PEGylated doxorubicin, leading to a decrease from 28.2 to 25.3% of the interindividual variability. When vinflunine dose is decreased, simulations of ANC time course (via a semiphysiological model) after vinflunine administration show a risk of neutropenia grade 3–4 at cycle 2 always lower than when dose is delayed. As an example, for moderate renal impaired patients, the risk is 42.1% when vinflunine is dosed at 320 mg m(–2) once every 4 weeks vs. 23.3% for 280 mg m(–2) once every 3 weeks. CONCLUSIONS: We propose for the first time a global comprehensive clinical pharmacological analysis for intravenous vinflunine that may help drive dose adjustment.

International audience; Allergic contact dermatitis is regarded as the most frequent expression of immunotoxicity in humans. Many odorant terpenes commonly used in fragrance compositions are considered as weak skin sensitizers, whereas some of their autoxidation products, allylic hydroperoxides, are classified as strong sensitizers according to the local lymph node assay. However, the mechanism of their effects on the immune system remains unclear. Since dendritic cells play a key role in allergic contact dermatitis, we studied their activation by the frequently used linalool (LINA) and limonene (LIMO), and their respective sensitizing allylic hydroperoxides (LINA-OOH, LIMO-OOH). The THP-1 cell-line was used as a surrogate for dendritic cells, the model currently employed in the validated h-CLAT in vitro test. Our data showed that allylic hydroperoxides behave differently. Both LINA-OOH and LIMO-OOH oxidized cell surface thiols 30 min after stimulation. However, the oxidative stress induced by LINA-OOH was stronger, with a higher decreased GSH/GSSG ratio and a stronger reactive species production. Moreover, LINA-OOH induced a stronger Nrf2 accumulation in correlation with nqo1 and ho-1 gene expression, 2 Nrf2 target genes. Regarding signaling pathways involved in these effects, P38 mitogen-activated protein kinase and P-ERK were activated in response to LINA-OOH but not with LIMO-OOH. CD54 and CD86 were induced 24-h postexposure. In contrast, LINA and LIMO did not modify THP-1 phenotype. This work underlies that autoxidation forming allylic hydroperoxide (ROOH) does not lead to equal chemical reactivity since LINA-OOH appears to be a stronger activator than LIMO-OOH, in regard to oxidative stress and Nrf2 pathway activation.

International audience; Highlights• The signal statistics in 50 MHz high frequency ultrasound images of the dermis are studied accurately layer by layer.• An automatic, accurate segmentation method is developed, which is highly resistant to artifacts thanks to the use of elaborate non-linear filters and a multiple loss level-set segmentation algorithm.• Based on fits to a Nakagami law, a score is computed for characterizing skin photo aging.• The joint segmentation and characterization algorithm is designed to be applied to 2D and 3D images.AbstractWe propose a novel joint segmentation and characterization algorithm for the assessment of skin aging using 50 MHz high-frequency ultrasound images. The proposed segmentation method allows a fine determination of the envelope signal's statistics in the dermis as a function of depth. The sequence of statistical estimates obtained is then combined into a single aging score. The segmentation is based on tailored recursive non-linear filters. The epidermis and the dermis are jointly segmented with a non-parametric active contour combining a texture criterion, an epidermis indicator map and the geometric constraint of horizontal continuity. The algorithm is designed to apply to 2D and 3D images as well. We evaluated skin photo-aging on ultrasound images with an experimental study on a cohort of 76 women separated into 2 groups of different ages. Two aging scores are computed from the images: local dermal contrast and skin roughness. We show that these scores are much better at identifying the two groups (p-value) than the previously used indicator (p-value 0.046). Moreover, we find that a combined score more reliably evaluates skin photo-aging, with 84% success, than a scoring of the ultrasound images by 4 experts.

International audience; The objective of the work was to check the presence of Non-Intended Added Substances (NIAS) with hormonal activities in aluminium coatings extracts coded: AA, BBF, MC and RR, furnished by four different suppliers. Water samples were prepared at room temperature or at 40°C for three months to verify the storage effect on the coatings. Solid phase extraction was used to concentrate and to extract coating substances. Hormonal activities were checked in vitro using reporter gene bioassays. Except BBF, all extracts induced a weak but significant estrogenic agonist activity in the human cell line. Using an estrogenic antagonist (ICI-182, 780), the answer was demonstrated specific in the bioassay. RR was the only extract to induce a concentration dependent anti-androgenic response in the MDA-KB2 cell line. Analysis performed using GC-MS and HPLC-MS detected 12 substances in most of the extracts. 8 NIAS were present. Among them, 4 were identified with certainty: HMBT, BGA, DCU and BPA. Estrogenic potency was BPA>DCU>BGA>HMBT. HMBT was also anti-androgenic at high concentration. Combining chemical analysis and bioassays data, we demonstrated that in the RR and the RR40 extracts, the observed estrogenic response was mainly due to BPA, the anti-androgenic activity of RR could be due to a synergism between HMBT and BPA. For MC and AA, estrogenic responses appear to be due to the presence of DCU. Except BBF, storage conditions tended to increase estrogenic activities in all extracts. However, in term of risk assessment, activities observed were negligible. This work demonstrated that sensitive bioassays are pertinent tools in complement to chemical analysis to monitor and check the presence of NIAS with hormonal activity in coating extracts.

International audience; Atopic dermatitis (AD) is a common skin inflammatory disease characterized by the production of thymic stromal lymphopoietin (TSLP) and marked TH 2 polarization. Recent studies suggest that IL-1β contributes to the development of AD skin inflammation. Here, we have investigated the impact of IL-1β signalling on the epidermal homeostasis of both healthy subjects and AD patients [with functional filaggrin (FLG) alleles], with particular attention to TSLP production and keratinocyte differentiation. In healthy reconstructed human epidermis (RHE), IL-1β promoted (i) robust secretion of TSLP in an NF-\kappaB-dependent manner and (ii) a significant decrease in the expression of filaggrin and other proteins of the epidermal differentiation complex. These effects were prevented by treatment of RHE with the anti-IL-1β mAb canakinumab and by the IL-1 receptor antagonist anakinra. Interestingly, RHE generated from AD donors behaved like that of healthy individuals and showed comparable responses to IL-1β signals. Collectively, our results suggest that IL-1β may be an early key mediator for the acquisition of an AD phenotype through induction of TSLP and alteration of the epidermal homeostasis. Copyright \textcopyright 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

International audience; Background/purpose : The purpose of this clinical study was to identify suitable biomarkers for a better understanding of the molecular and organizational changes in human dermis during intrinsic and extrinsic ageing. Methods : Sun-exposed and non-exposed skin biopsies were collected from twenty-eight women devised in two groups (20-30 and ≥60 years old). The hydric organization and thermal transitions were determined by Differential Scanning Calorimetry (DSC). Fourier Transform Infrared spectroscopy (FTIR) was used to identify the absorption bands of the dermis and to quantify the different absorbance ratio. Results : The amounts of total, freezable and unfreezable water were determined. A significant increasing amount of freezable water is evidenced in sun-exposed area skin of aged group compared with young group (P=.0126). Another significant effect of extrinsic ageing (P=.0489) is the drastic decrease of fibrillary collagen, the main protein component of dermis. The only significant effect of intrinsic ageing (P=.0184) is an increase of the heat-stable fraction of collagens in dermis. Conclusion : DSC and FTIR are well-suited techniques to characterize human skin, giving accurate results with a high reproducibility. The combination of these techniques is useful for a better understanding of human skin modifications with intrinsic and extrinsic ageing.

In this study, a comprehensive characterization of xenobiotic metabolizing enzymes (XMEs) based on gene expression and enzyme functionality was made in a reconstructed skin epidermal model derived from the outer root sheath (ORS) of hair follicles (ORS-RHE). The ORS-RHE model XME gene profile was consistent with native human skin. Cytochromes P450 (CYPs) consistently reported to be detected in native human skin were also present at the gene level in the ORS-RHE model. The highest Phase I XME gene expression levels were observed for alcohol/aldehyde dehydrogenases and (carboxyl) esterases. The model was responsive to the CYP inducers, 3-methylcholanthrene (3-MC) and β-naphthoflavone (βNF) after topical and systemic applications, evident at the gene and enzyme activity level. Phase II XME levels were generally higher than those of Phase I XMEs, the highest levels were GSTs and transferases, including NAT1. The presence of functional CYPs, UGTs and SULTs was confirmed by incubating the models with 7-ethoxycoumarin, testosterone, benzo(a)pyrene and 3-MC, all of which were rapidly metabolized within 24h after topical application. The extent of metabolism was dependent on saturable and non-saturable metabolism by the XMEs and on the residence time within the model. In conclusion, the ORS-RHE model expresses a number of Phase I and II XMEs, some of which may be induced by AhR ligands. Functional XME activities were also demonstrated using systemic or topical application routes, supporting their use in cutaneous metabolism studies. Such a reproducible model will be of interest when evaluating the cutaneous metabolism and potential toxicity of innovative dermo-cosmetic ingredients.

DNA, RNA and histone methylation is implicated in various human diseases such as cancer or viral infections, playing a major role in cell process regulation, especially in modulation of gene expression. Here we developed a convergent synthetic pathway starting from a protected bromomethylcytosine derivative to synthesize transition state analogues of the DNA methyltransferases. This approach led to seven 5-methylcytosine-adenosine compounds that were, surprisingly, inactive against hDNMT1, hDNMT3Acat, TRDMT1 and other RNA human and viral methyltransferases. Interestingly, compound 4 and its derivative 2 showed an inhibitory activity against PRMT4 in the micromolar range. Crystal structures showed that compound 4 binds to the PRMT4 active site, displacing strongly the S -adenosyl- l -methionine cofactor, occupying its binding site, and interacting with the arginine substrate site through the cytosine moiety that probes the space filled by a substrate peptide methylation intermediate. Furthermore, the binding of the compounds induces important structural switches. These findings open new routes for the conception of new potent PRMT4 inhibitors based on the 5-methylcytosine-adenosine scaffold. This article is part of a discussion meeting issue ‘Frontiers in epigenetic chemical biology’.

International audience; PURPOSE: The Gastro-Intestinal Working Party of the EORTC Radiation Oncology Group (GIWP-ROG) developed guidelines for target volume definition in neoadjuvant radiation of adenocarcinomas of the gastroesophageal junction (GEJ) and the stomach. METHODS AND MATERIALS: Guidelines about the definition of the clinical target volume (CTV) are based on a systematic literature review of the location and frequency of local recurrences and lymph node involvement in adenocarcinomas of the GEJ and the stomach. Therefore, MEDLINE was searched up to August 2008. Guidelines concerning prescription, planning and treatment delivery are based on a consensus between the members of the GIWP-ROG. RESULTS: In order to support a curative resection of GEJ and gastric cancer, an individualized preoperative treatment volume based on tumour location has to include the primary tumour and the draining regional lymph nodes area. Therefore we recommend to use the 2nd English Edition of the Japanese Classification of Gastric Carcinoma of the Japanese Gastric Cancer Association which developed the concept of assigning tumours of the GEJ and the stomach to anatomically defined sub-sites corresponding respectively to a distinct lymphatic spread pattern. CONCLUSION: The GIWP-ROG defined guidelines for preoperative irradiation of adenocarcinomas of the GEJ and the stomach to reduce variability in the framework of future clinical trials.

International audience; BACKGROUND: Use of cetuximab, a monoclonal antibody targeting the epidermal growth factor receptor (EGFR), has the potential to increase survival in patients with advanced non-small-cell lung cancer. We therefore compared chemotherapy plus cetuximab with chemotherapy alone in patients with advanced EGFR-positive non-small-cell lung cancer. METHODS: In a multinational, multicentre, open-label, phase III trial, chemotherapy-naive patients (>or=18 years) with advanced EGFR-expressing histologically or cytologically proven stage wet IIIB or stage IV non-small-cell lung cancer were randomly assigned in a 1:1 ratio to chemotherapy plus cetuximab or just chemotherapy. Chemotherapy was cisplatin 80 mg/m(2) intravenous infusion on day 1, and vinorelbine 25 mg/m(2) intravenous infusion on days 1 and 8 of every 3-week cycle) for up to six cycles. Cetuximab-at a starting dose of 400 mg/m(2) intravenous infusion over 2 h on day 1, and from day 8 onwards at 250 mg/m(2) over 1 h per week-was continued after the end of chemotherapy until disease progression or unacceptable toxicity had occurred. The primary endpoint was overall survival. Analysis was by intention to treat. This study is registered with ClinicalTrials.gov, number NCT00148798. FINDINGS: Between October, 2004, and January, 2006, 1125 patients were randomly assigned to chemotherapy plus cetuximab (n=557) or chemotherapy alone (n=568). Patients given chemotherapy plus cetuximab survived longer than those in the chemotherapy-alone group (median 11.3 months vs 10.1 months; hazard ratio for death 0.871 [95% CI 0.762-0.996]; p=0.044). The main cetuximab-related adverse event was acne-like rash (57 [10%] of 548, grade 3). INTERPRETATION: Addition of cetuximab to platinum-based chemotherapy represents a new treatment option for patients with advanced non-small-cell lung cancer. FUNDING: Merck KGaA.

International audience; Ca(2+) is an essential factor inducing keratinocyte differentiation due to the natural Ca(2+) gradient in the skin. However, the membrane mechanisms that mediate calcium entry and trigger keratinocyte differentiation had not previously been elucidated. In this study we demonstrate that Ca(2+)-induced differentiation up-regulates both mRNA and protein expression of a transient receptor potential highly Ca(2+)-selective channel, TRPV6. The latter mediates Ca(2+) uptake and accounts for the basal [Ca(2+)](i) in human keratinocytes. Our results show that TRPV6 is a prerequisite for keratinocyte entry into differentiation, because the silencing of TRPV6 in human primary keratinocytes led to the development of impaired differentiated phenotype triggered by Ca(2+). The expression of such differentiation markers as involucrin, transglutaminase-1, and cytokeratin-10 was significantly inhibited by small interfering RNA-TRPV6 as compared with differentiated control cells. TRPV6 silencing affected cell morphology and the development of intercellular contacts, as well as the ability of cells to stratify. 1,25-Dihydroxyvitamin D3, a cofactor of differentiation, dose-dependently increased TRPV6 mRNA and protein expression in human keratinocytes. This TRPV6 up-regulation led to a significant increase in Ca(2+) uptake in both undifferentiated and differentiated keratinocytes. We conclude that TRPV6 mediates, at least in part, the pro-differentiating effects of 1,25-dihydroxyvitamin D3 by increasing Ca(2+) entry, thereby promoting differentiation. Taken together, these data suggest that the TRPV6 channel is a key element in Ca(2+)/1,25-dihydroxyvitamin D3-induced differentiation of human keratinocytes.

International audience; Background: Peptidylarginine deiminases (PADs) catalyze deimination (or citrullination), a calcium-dependent post-translational modification involved in several physiological processes and human diseases, such as rheumatoid arthritis and cancer. Deimination of filaggrin (FLG) by PAD1 and PAD3 during the last steps of keratinocyte differentiation is a crucial event for the epidermis function and homeostasis. This allows the complete degradation of FLG, leading to the production of free amino acids and their derivatives that are essential for epidermal photoprotection and moisturizing of the stratum corneum. Objective: To increase the flux of this catabolic pathway, we searched for activators of PADs. Methods: A large chemical library was screened first in silico and then by using an automated assay based on an indirect colorimetric measurement of recombinant human PAD activity. Potential activators were then confirmed using a recombinant human FLG as a substrate, and secondly after topical application at the surface of three-dimensional reconstructed human epidermis. Results: The data obtained after the library screening pointed to xanthine derivatives as potential PAD activators. Among seven xanthine derivatives tested at 50-300μM, caffeine, theobromine and acefylline proved to be the most potent enhancers of in vitro deimination of FLG by PAD1 and PAD3. After topical application of a gel formulation containing 3% acefylline at the surface of reconstructed epidermis, immunoblotting analysis showed an increase in the total amount of deiminated proteins, and confocal microscopy showed an enhanced deimination in the stratum corneum. This demonstrated the activation of PADs in living cells. Conclusion: As a PAD activator, acefylline will be useful to study the role of deimination and could be proposed to increase or correct the hydration of the cornified layers of the epidermis.

Background Long acting bronchodilators are the standard of care in the management of chronic obstructive pulmonary disease (COPD). The aim of this study was to investigate the efficacy and safety of V0162, a novel anticholinergic agent with bronchodilator properties, in preclinical models and in patients with COPD. Methods Guinea pigs were used to evaluate the impact of V0162 on the acetylcholine or histamine-induced bronchoconstriction. V0162 was also investigated in an allergic asthma model on ovalbumin-sensitized guinea pig. For clinical investigations, healthy volunteers were included in a dose-escalation, randomized, placebo-controlled phase I study to determine the maximal tolerated dose, followed by a randomized, placebo-controlled, cross-over phase II study in patients with COPD. V0162 was given via inhalation route. The objectives of the phase I/II study were to assess the safety and efficacy of V0162, in terms of bronchodilation and reduction in hyperinflation. Results Preclinical results showed that V0162 was able to prevent bronchoconstriction induced either by acetylcholine or histamine. V0162 reversed the bronchoconstriction and airway inflammation caused by ovalbumin challenge in sensitized guinea pigs. In the healthy volunteers study, 88 subjects were enrolled: 66 received V0162 and 22 received placebo. No particular safety concerns were raised. The maximal tolerated dose was not reached and the dose escalation was stopped at 2400 μg. A total of 20 patients with COPD were then enrolled. All patients received a single-dose of V0162 1600 μg and of placebo in two alternating periods. In COPD patients, V0162 demonstrated a significant increase in FEV1 compared with placebo (148 ± 137 ml vs. 36 ± 151 ml, p = 0.003). This bronchodilatory effect was corroborated by a reduction in hyperinflation. There was a trend toward dyspnea relief (change in visual analog scale at 22 h, −15.1 ± 26.0 mm vs.- 5.3 ± 28.8 mm with placebo, p = 0.054). No serious adverse events (AEs) were reported. Most common AEs were productive and non-productive cough, dyspnea and pruritus. Conclusions V0162 improved pulmonary function and tended to improve dyspnea in patients with COPD over more than 24 h. The slight plasmatic exposure observed might support the good safety profile. Trial registration ClinicalTrials.gov identifier: NCT01348555

International audience; Induction of skin cancer is the most deleterious effect of excessive exposure to sunlight. Accurate evaluation of sunscreens to protect the genome is thus of major importance. In particular, the ability. of suncare products to prevent the formation of DNA damage should be evaluated more directly since the Sun Protection Factor is only related to erythema induction. For this purpose, we developed an in vitro approach using a recently characterized reconstituted human epidermis (RHE) model engineered from hair follicle. The relevance of this skin substitute in terms of UV-induced genotoxicity was compared to ex vivo explants exposed to solar-simulated radiation (SSR). The yield of bipyrimidine photoproducts, their rate of repair, and the induction of apoptosis were very similar in both types of skin samples. In order to evaluate the protection afforded by sunscreen against DNA damage, bipyrimidine photoproducts were quantified in tissue models following SSR exposure in the presence or absence of a SPF50+ formula. A rather high DNA protection factor of approximately 20 was found in RHE, very similar to that determined for explants. Thus, RHE is a good surrogate to human skin, and also a convenient and useful tool for investigation of the genoprotection of sunscreens. (C) 2015 Elsevier B.V. All rights reserved.

International audience; Interactions between the immune system and skin bacteria are of major importance in the pathophysiology of atopic dermatitis (AD), yet our understanding of them is limited. From a cohort of very young AD children (1 to 3 years old), sensitized to Dermatophagoides pteronyssinus allergens (Der p), we conducted culturomic analysis of skin microbiota, cutaneous transcript profiling and quantification of anti-Der p CD4+ T cells. This showed that the presence of S. aureus in inflamed skin of AD patients was associated with a high IgE response, increased expression of inflammatory and Th2/Th22 transcripts and the prevalence of a peripheral Th2 anti-Der p response. Monocyte-derived dendritic cells (moDC) exposed to the S. aureus and S. epidermidis secretomes were found to release pro-inflammatory IFN-γ and anti-inflammatory IL-10, respectively. Allogeneic moDC exposed to the S. aureus secretome also induced the proliferation of CD4+ T cells and this effect was counteracted by concurrent exposure to the S. epidermidis secretome. In addition, whereas the S. epidermidis secretome promoted the activity of regulatory T cells (Treg) in suppressing the proliferation of conventional CD4+ T cells, the Treg lost this ability in the presence of the S. aureus secretome. We therefore conclude that S. aureus may cause and promote inflammation in the skin of AD children through concomitant Th2 activation and the silencing of resident Treg cells. Commensals such as S. epidermidis may counteract these effects by inducing the release of IL-10 by skin dendritic cells.

The role of Beta-adrenergic receptors (B-ARs) remains controversial in normal and tumor breast. Herein we explore the cAMP signaling involved in B-AR-dependent control of proliferation and adhesion of nontumor human breast cell line MCF-10A. Low concentrations of a B-agonist, isoproterenol (ISO), promote cell adhesion (87.5% cells remaining adherent to the plastic dishes following specific detachment vs. 35.0% in control, P 0.001), while increasing concentrations further engages an additional 36% inhibition of Erk1/2 phosphorylation (p-Erk1/2) -dependent cell proliferation (P 0.01). Isoproterenol dose response on cell adhesion was fitted to a 2-site curve (EC50(1): 16.5 +/-11.5 fM, EC50(2): 4.08 +/- 3.09 nM), while ISO significantly inhibited p-Erk1/2 according to a 1-site model (EC50: 0.25 +/- 0.13 nM). Using B-AR-selective agonist/antagonists and cAMP analogs/inhibitors, we identified a dosage-dependent signaling in which low ISO concentrations target a B2-AR population localized in raft microdomains and stimulate a Gs/cAMP/Epac/adhesion-signaling module, while higher concentrations engage a concomitant activation of another B2-AR population outside rafts and inhibit the proliferation by a Gs/cAMP/PKA-dependent signaling module. Our data provide a new molecular basis for the dose-dependent switch of B-AR signaling. This study also sheds light on a new cAMP pathway core mechanism with a single receptor triggering dual cAMP signaling controlled by PKA or Epac but with different cellular outputs. Fil: Bruzzone, Ariana. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentina Fil: Saulière, Aude. Inserm; Francia Fil: Finana, Frédéric . Institut de Recherche Pierre Fabre; Francia Fil: Sénard, Jean Michel. Inserm; Francia. Toulouse University Hospital; Francia Fil: Luthy, Isabel Alicia. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Biología y Medicina Experimental (i); Argentina Fil: Galés, Céline. Inserm; Francia

International audience; Elastin is a long-lived protein and a key component of connective tissues. The tissular elastin content decreases during chronological aging, and the mechanisms underlying its slow repair are not known. Lipid oxidation products that accumulate in aged tissues may generate protein dysfunction. We hypothesized that 4-hydroxynonenal (4-HNE), a highly reactive α,β-aldehydic product generated from polyunsaturated fatty acid peroxidation, could contribute to inhibiting elastin repair by antagonizing the elastogenic signaling of transforming growth factor-β1 (TGF-β1) in skin fibroblasts. We report that a low 4-HNE concentration (2µmol/L) inhibits the upregulation of tropoelastin expression stimulated by TGF-β1 in human and murine fibroblasts. The study of signaling pathways potentially involved in the regulation of elastin expression showed that 4-HNE did not block the phosphorylation of Smad3, an early step of TGF-β1 signaling, but inhibited the nuclear translocation of Smad2. Concomitantly, 4-HNE modified and stimulated the phosphorylation of the epidermal growth factor receptor (EGFR) and subsequently ERK1/2 activation, leading to the phosphorylation/stabilization of the Smad transcriptional corepressor TGIF, which antagonizes TGF-β1 signaling. Inhibitors of EGFR (AG1478) and MEK/ERK (PD98059), and EGFR-specific siRNAs, reversed the inhibitory effect of 4-HNE on TGF-β1-induced nuclear translocation of Smad2 and tropoelastin synthesis. In vivo studies on aortas from aged C57BL/6 mice showed that EGFR is modified by 4-HNE, in correlation with an increased 4-HNE-adduct accumulation and decreased elastin content. Altogether, these data suggest that 4-HNE inhibits the elastogenic activity of TGF-β1, by modifying and activating the EGFR/ERK/TGIF pathway, which may contribute to altering elastin repair in chronological aging and oxidative stress-associated aging processes.

International audience; Functional selectivity of G protein-coupled receptor (GPCR) ligands toward different downstream signals has recently emerged as a general hallmark of this receptor class. However, pleiotropic and crosstalk signaling of GPCRs makes functional selectivity difficult to decode. To look from the initial active receptor point of view, we developed new, highly sensitive and direct bioluminescence resonance energy transfer-based G protein activation probes specific for all G protein isoforms, and we used them to evaluate the G protein-coupling activity of [(1)Sar(4)Ile(8)Ile]-angiotensin II (SII), previously described as an angiotensin II type 1 (AT(1)) receptor-biased agonist that is G protein independent but β-arrestin selective. By multiplexing assays sensing sequential signaling events, from receptor conformations to downstream signaling, we decoded SII as an agonist stabilizing a G protein-dependent AT(1A) receptor signaling module different from that of the physiological agonist angiotensin II, both in recombinant and primary cells. Thus, a biased agonist does not necessarily select effects from the physiological agonist but may instead stabilize and create a new distinct active pharmacological receptor entity.

International audience; Skin contact has been hypothesized to contribute to human exposure to bisphenol A (BPA). We examinedthe diffusion and metabolism of BPA using viable skin models: human skin explants and short-term culturesof pig ear skin, an alternative model for the study of the fate of xenobiotics following contact exposure.14C-BPA [50–800 nmol] was applied on the surface of skin models. Radioactivity distribution wasmeasured in all skin compartments and in the diffusion cells of static cells diffusion systems. BPA andmetabolites were further quantified by radio-HPLC. BPA was efficiently absorbed in short-term cultures,with no major difference between the models used in the study [viable pig ear skin: 65%; viable humanexplants: 46%; non-viable (previously frozen) pig skin: 58%]. BPA was extensively metabolized in viablesystems only. Major BPA metabolites produced by the skin were BPA mono-glucuronide and BPA monosulfate,accounting together for 73% and 27% of the dose, in pig and human, respectively. In conclusion,experiments with viable skin models unequivocally demonstrate that BPA is readily absorbed and metabolizedby the skin. The trans-dermal route is expected to contribute substantially to BPA exposure inhuman, when direct contact with BPA (free monomer) occurs.

International audience; Background: Numerous studies have demonstrated the beneficial effect of Avène Thermal Spring Water (TSW) in dermatological diseases but the molecular mechanisms remain unknown. The objective of the present study was to evaluate the effect of Avène TSW on the morphological and molecular features related to the more advanced status of differentiation of human keratinocytes.Material and methods: Normal human keratinocytes (NHK) were differentiated in medium powder reconstituted with Avène TSW and assessed by RT-PCR and immunohistochemistry. Calcium entry was measured by a Fura-2 AM probe. TRPV6 channel were detected by immunohistochemistry, RT-PCR and western blot.Results: Treatment of NHK with Avène TSW led to an enhanced constitutive calcium entry that resulted in the increased expression of involucrin and cytokeratins 1 and 10. This enhanced constitutive calcium entry in Avène TSW-treated keratinocytes was mediated by the TRPV6 calcium channel. Moreover, Avène TSW-mediated calcium entry was due to the increase in TRPV6 expression as well as the channel abundance at the cell membrane.Conclusions: An other mechanism of action of Avène TSW is described. Avène TSW treatment induced an enhanced constitutive calcium entry mediated by TRPV6 channel leading to the acceleration of human keratinocytes differentiation.

International audience; BACKGROUND: A previously published study of temozolomide concurrent with whole-brain radiation therapy (WBRT) reported significant improvement in response rates and a nonsignificant trend toward improved overall survival compared with WBRT alone in patients with brain metastases primarily from lung cancer. This study sought to confirm the benefit of adding temozolomide to WBRT in patients with non-small-cell lung cancer (NSCLC) with brain metastases. PATIENTS AND METHODS: This planned phase III study (target = 380 events) was converted to a phase II study (target = 70 events) because of poor enrollment. Patients with NSCLC and > or = 1 newly diagnosed brain lesion were randomized to WBRT (30 Gy in 10 fractions) alone or combined with temozolomide (75 mg/m2/day) for 21 or 28 days. Endpoints included overall survival and time to central nervous system (CNS) progression. RESULTS: Median overall survival and median time to CNS progression was 4.4 and 3.1 months in the WBRT + temozolomide arm (n = 47) versus 5.7 and 3.8 months in the WBRT arm (n = 48). However, there were imbalances in the percentages of patients receiving previous chemotherapy and with synchronous brain metastases. Adding temozolomide to WBRT increased the frequency of nausea, vomiting, alopecia, fatigue, anorexia, and constipation. Most adverse events were mild to moderate. CONCLUSION: The benefit of adding temozolomide to WBRT was not confirmed; however, the accrual goal for the planned phase III trial was not reached, and the study regimen differed from regimens used previously. Therefore, the role of temozolomide in treating brain metastases remains unresolved.

This study was conducted to retrospectively compare the area under the curve (AUC) and the total clearance of busulfan (Bu) following oral and intravenous (IV) administrations and to determine which intravenous dose generated equivalent exposure to that of the oral form that has been marketed for decades. Patient pharmacokinetics were assessed at dose 9 during a conditioning regimen for stem-cell transplantation and included data from 277 patients for oral Bu (71 from fixed-dose studies and 206 from studies with dose adjustment allowed) and 120 patients for IV Bu (fixed dose). AUCs were compared between patients with fixed dose of oral Bu (n = 71, 1 mg/kg) and those of IV Bu (n = 120, 0.8 mg/kg). Total clearances were calculated for all 277 patients with oral Bu and compared to those with IV Bu, with the ratio of IV-to-oral clearance representing the absolute bioavailability of the oral form. Oral and IV populations differed on disease-type distribution but presented comparable demography parameters. IV Bu dosing was mostly based on the ideal body weight index while actual body weight or adjusted ideal body weight indexes were mostly used for oral. When normalised to comparable indexes, bioequivalent AUCs were achieved between oral and IV populations. Oral Bu bioavailability was about 80% when calculated from the ratio of IV-to-oral total clearances. This retrospective study carried out on a large set of data showed that similar plasma exposures were achieved with 1.0 mg/kg oral Bu or 0.8 mg/kg IV Bu.

Purpose Oral mucositis is a common side effect induced by radio/chemotherapy in patients with head and neck cancer. Although it dramatically impairs patient quality of life, no efficient and safe therapeutic solution is available today. Therefore, we investigated the protective efficacy of a new heparan mimetic biopolymer, RGTA-OTR4131, used alone or in combination with amifostine, for oral mucositis and simultaneously evaluated its effect on tumor growth in vitro and in vivo . Methods and Materials A single dose of 16.5 Gy was selectively delivered to the snout of mice, and the effects of OTR4131 or amifostine-OTR4131 were analyzed by macroscopic scoring and histology. The effect of OTR4131 administration on tumor growth was then investigated in vitro and in xenograft models using two cell lines (HEP-2 and HT-29). Results Amifostine and OTR4131 significantly decreased the severity and duration of lip mucosal reactions. However, amifostine has to be administered before irradiation, whereas the most impressive protection was obtained when OTR4131 was injected 24 h after irradiation. In addition, OTR4131 was well tolerated, and the combination of amifostine and OTR4131 further enhanced mucosal protection. At the tumor level, OTR4131 did not modify HEP-2 cell line clonogenic survival in vitro or protect xenografted tumor cells from radiotherapy. Of interest, high doses of OTR4131 significantly decreased clonogenic survival of HT-29 cells. Conclusions RGTAs-OTR4131 is a well-tolerated, natural agent that effectively reduces radio-induced mucositis without affecting tumor sensitivity to irradiation. This suggests a possible transfer into the clinic for patients' benefit.

Purpose: The precise determination of skin's mechanical properties is still an open question. When performing an in vivo test, the piece of skin tested is not as well defined as it is in material testing. Moreover, the body zone and the body posture imply an initial stress on the skin. Consequently, a precise mechanical analysis needs a precise measurement of the natural skin tension. Methods: A new method and the relative device are presented. It is based on an extensiometry test. Skin is tested not only in traction but also in compression. The tested skin sample is well defined and protected from surrounding effects by follower tabs. The size and shape of the device have been optimised by a finite element modelisation. Results: The method was tested with elastomers pre-tensioned at different loads. It is shown that the initial tension can be retrieved with good precision. Tests were then performed in vivo on the forearm for different arm positions. It is shown that initial tension could be only clearly determined for the highest skin tension, although the skin presented very different traction behaviour with different arm positions. Conclusion: It is shown how body posture influences measurements. An innovative method for easily determining initial tension is presented. Nevertheless, further tests and device improvements are needed to apply this skin tension measurement for different body zones and body postures.

Sunscreens, whose main function is to protect the skin against the harmful effects of solar irradiation, should remain at the skin surface or impregnate the first layers of the stratum corneum only and not penetrate into the underlying living tissue. The goal of this work was to assess the penetration of titanium dioxide (TiO2) and methylene bis-benzotriazoyl tetramethylbutylphenol (MBBT), included in a broad-spectrum sunscreen formulation, into human skin in vivo, using the tape stripping method, and in vitro, using a compartmental approach. An additional objective was to propose an easy and minimally invasive approach to determining the percutaneous uptake of sunscreens following topical application. TiO2 and MBBT were quantified using colorimetric assay and HPLC analysis, respectively. The transmission electron microscopy and particle-induced X-ray emission techniques were used to localize the TiO2 in skin sections.More than 90% of both sunscreens was recovered in the first 15 tape strippings. In addition we have shown that the remaining 10% did not penetrate the viable tissue, but was localized in the furrows and in the opened infundibulum. Less than 0.1% of MBBT was detected in the receptor medium, and no TiO2 was detected in the follicle, viable epidermis or dermis. Thus, this in vivo and in vitro penetration study showed an absence of TiO2 penetration into the viable skin layers through either transcorneal or transfollicular pathways and negligible transcutaneous absorption of MBBT. However, differences in distribution within the stratum corneum reinforced the need for a complementary approach, using minimally invasive in vivo methodology and in vitro compartmental analysis.This combination represents a well-adapted method for testing the safety of topically applied sunscreen formulations in real-life conditions.

International audience; Mutagenic and carcinogenic UV-B radiation is known to damage DNA mostly through the formation of bipyrimidine photoproducts, including cyclobutane dimers (CPD) and (6-4) photoproducts ((6-4) PP). Using high-performance liquid chromatography coupled to tandem mass spectrometry, we investigated the formation and repair of thymine-thymine (TT) and thymine-cytosine (TC) CPD and (6-4) PP in the DNA of cultured human dermal fibroblasts. A major observation was that the rate of repair of the photoproducts did not depend on the identity of the modified pyrimidines. In addition, removal of CPD was found to significantly decrease with increasing applied UV-B dose, whereas (6-4) PP were efficiently repaired within less than 24 h, irrespective of the dose. As a result, a relatively large amount of CPD remained in the genome 48 h after the irradiation. Because the overall applied doses (

International audience; This study investigated the effect of seizure prophylaxis on busulfan (Bu) plasma exposure. Twenty-four adult patients received an intravenous Bu-cyclophoshamide conditioning regimen prior to bone marrow transplantation. Busilvex (0.8 mg/kg) was administered every six hours during four consecutive days. Clonazepam (0.025 to 0.03 mg/kg/day as a continuous 12-h i.v. infusion) was administered at least 12 hours prior to i.v. Bu dosing and continued until 24 hours after the last dose. Pharmacokinetic (PK) data were compared with those previously collected in patients (n=127) treated with phenytoin for seizure prophylaxis. Through population PK analysis, a 10% average increase (coefficient of variation, RSE=5.35%) in total clearance of Bu was quantified when Bu was associated with clonazepam as compared to phenytoin, which was considered as not being clinically relevant. The suspected induction on Bu metabolism by phenytoin should have resulted in the opposite effect. The patient efficacy and safety profiles were comparable between the two cohorts.