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Over the last decade, research into methodologies to identify skin sensitization hazards has led to the adoption of several non-animal methods as OECD guidelines. However, predictive accuracy beyond the chemical domains of the individual validation studies remains largely untested. In the present study, skin sensitization test results from in vitro and in chemico methods for 12 plant extracts and 15 polymeric materials are reported and compared to available in vivo skin sensitization data. Eight plant extracts were tested in the DPRA and h-CLAT, with the 2 out of 3 approach resulting in a balanced accuracy of 50%. The balanced accuracy for the 11 plant extracts assessed in the SENS-IS was 88%. Excluding 5 polymers inconclusive in vitro, the remainder, assessed using the 2 out of 3 approach, resulted in 63% balanced accuracy. The SENS-IS method, excluding one polymeric material due to technical inapplicability, showed 68% balanced accuracy. Although based on limited numbers, the results presented here indicate that some substance subgroups may not be in the applicability domains of the method used and careful analysis is required before positive or negative results can be accepted.

Serum lipids and metabolic diseases, in particular type 2 diabetes (T2D) and nonalcoholic fatty liver disease (NAFLD) , predict the atherosclerotic diseases coronary artery disease (CAD) and peripheral arterial disease (PAD) . However, it is not known in how far a more detailed characterization including serum lipids improves discrimination of PAD from CAD. A cohort of 274 statin-naïve patients with either PAD (n = 89) or stable CAD (n = 185) were referred to metabolic screening and were characterized using nuclear magnetic resonance- and liquid chromatography-tandem mass spectrometry based advanced lipid and lipoprotein analysis. Results were validated in an independent cohort of 1239 patients with PAD or CAD. We found a significant difference in T2D prevalence and in the ceramide-based lipid profile between PAD and CAD patients. However, neither cholesterol-based markers (including LDL-C, HDL-C) and detailed lipoprotein profiles nor the NAFLD status differed significantly between PAD and CAD patients (figure) . The difference between ceramide-based lipid profiles of CAD and PAD remained significant also after adjusting for body composition, smoking, inflammatory parameters, and T2D. We conclude that PAD and CAD differ in ceramide-based lipid profiles and T2D status, but not in other lipid characteristics or metabolic diseases. Disclosure A.Leiherer: None. A.Jylha: None. M.Laaperi: None. R.Laaksonen: None. W.Maerz: None. P.Fraunberger: None. M.Kleber: None. H.Drexel: None. A.Muendlein: None. E.Brandtner: None. C.H.Saely: None. A.Vonbank: None. A.Mader: None. L.Sprenger: None. M.Maechler: None. B.Larcher: None.

ZUSAMMENFASSUNGDas kolorektale Karzinom (KRK) ist einer der häufigsten malignen Tumoren in Deutschland. Einer frühzeitigen Diagnostik kommt große Bedeutung zu. Goldstandard ist hier die Koloskopie. Die aktuelle S3-Leitlinie Kolorektales Karzinom empfiehlt zum KRK-Screening den fäkalen okkulten Bluttest. Für das Monitoring von Patienten vor und nach Tumorresektion werden die Messung des Carcinoembryonalen Antigens (CEA) und der Mikrosatellitenstabilität empfohlen. Für die Auswahl der korrekten Chemotherapie scheint derzeit eine Überprüfung des Mutationsstatus, mindestens des KRAS-Gens und des BRAF-Gens, sinnvoll zu sein. Eine Reihe an neuartigen Tumormarkern befindet sich momentan in der Entwicklung, hat jedoch noch nicht die Reife für eine mögliche Anwendung in der Routinediagnostik erreicht. Den schnellsten Weg in die breite Anwendung können Next-Generation-Sequencing-basierte genetische Tests finden.

O trabalho tem por objetivo avaliar se, por meio da transparência ativa e em comparação com outras agências, a Anvisa disponibiliza informação para o cidadão sobre o processo regulatório de medicamento, de maneira visível e que possibilite a sua dedução. Para tanto, foi aplicado um instrumento de avaliação da transparência ativa no website da Anvisa, comparando-a com outras três agências reguladoras de medicamentos americanas, considerando três condicionantes: o atendimento a lei de acesso à informação, a usabilidade e navegabilidade do website e a disponibilidade da informação de maneira visível e inferível. Como resultados, o índice geral de transparência ativa da Anvisa foi o terceiro em comparação com as agências. Utilizando uma ferramenta de testagem, em uma escala que varia de 0 a 100 pontos, o website da Anvisa alcançou 61 pontos. O Parecer Público de Avaliação do Medicamento (PPAM) possui informações similares às outras agências e apresenta algumas oportunidades para o seu aprimoramento. Quanto as limitações, futuros estudos podem ser conduzidos no sentido de aperfeiçoar a metodologia proposta de avaliação e a extrapolação do seu uso em outros tipos de processos regulatórios. Como contribuições, oferece mais uma proposta metodológica para a avaliação da transparência ativa, considerando as dimensões de visibilidade e inferibilidade para transparência e em conjunto com os demais instrumentos de aprimoramento da qualidade regulatória da agência, é disponibilizada uma ferramenta para avaliação da transparência ativa, permitindo o acesso e a compreensão do processo regulatório da Agência. Purpose – Evaluate whether through active transparency and confrontation with other agencies, Anvisa provides the citizen with information on the regulatory process of medicines, visibly and allowing your deductible. Design/Methodology – It consists of the application of an instrument for assessing active transparency on the Anvisa website, and comparing Anvisa’s active transparency with three other equivalent regulatory agencies in the Americas on the basis of three conditions: compliance with the law on access to information, usability and navigability of the website, and the availability of information in accordance with the principles of transparency and open data. Findings – Anvisa's general active transparency index ranked third in comparison with the other agencies, 61 points on a scale ranging from 0 to 100 points. Public Opinion on Drug Evaluation (PPAM Anvisa) has similar information to other agencies, but features opportunities for its improvement. Research limitations – Future studies can be conducted in order to improve the proposed assessment instrument and extrapolate its use in other types of regulatory processes. Practical implications – This study offers a methodological proposal for the evaluation of active transparency, considering the dimensions of visibility and inferability for transparency. Social implications – In conjunction with the other instruments for improving the regulatory quality of the agency, the study offers a tool for assessing active transparency, permitting users access and understanding of the Agency's regulatory process. Originality – This is the first study that proposes a methodology to assess the active transparency of the regulatory process for medicines within the scope of Anvisa.

Die Anzahl an Biomarkern fur die Diagnose und Prognose von Tumorerkrankungen wachst kontinuierlich. Im klinischen Alltag wird eine Reihe von Parametern verwendet mit bekannt hoher Sensitivitat und Spezifitat. Wahrend im Stadium der initialen Diagnose die Messung der Konzentration spezifischer Tumormarker eine fruhe Tumordiagnose in Screeningverfahren ermoglichen soll zeigen andere Tumormarker ihren Wert hauptsachlich wahrend der Therapie und in der Nachsorge. Langzeitstudien haben ergeben, dass neben der Messung der Antigenkonzentration auch die Uberwachung deren Entwicklung und Dynamik sehr wichtig sein konnen. Im Folgenden sollen molekulare Biomarker auf den Ebenen der Genetik, der Epigenetik und der Proteine untersucht und deren Rolle im Bereich des Krebsscreenings und der fruhen Detektion von Rezidiven sowie als prognostische und pradiktive Faktoren diskutiert werden. Dabei wird auch immer auf die aktuellen, in Deutschland gultigen Leitlinien hingewiesen werden. Aus dem Spektrum der endokrinologischen Tumorerkrankungen soll im Besonderen auf das Kolorektale Karzinom (KRK) eingegangen sowie vielversprechende Forschungsansatze vorgestellt werden.

Background— Studies of recurrent or subsequent disease events may be susceptible to bias caused by selection of subjects who both experience and survive the primary indexing event. Currently, the magnitude of any selection bias, particularly for subsequent time-to-event analysis in genetic association studies, is unknown. Methods and Results— We used empirically inspired simulation studies to explore the impact of selection bias on the marginal hazard ratio for risk of subsequent events among those with established coronary heart disease. The extent of selection bias was determined by the magnitudes of genetic and nongenetic effects on the indexing (first) coronary heart disease event. Unless the genetic hazard ratio was unrealistically large (>1.6 per allele) and assuming the sum of all nongenetic hazard ratios was Conclusions— In most empirical settings, selection bias is expected to have a limited impact on genetic effect estimates of subsequent event risk. Nevertheless, because of undercoverage increasing with sample size, most confidence intervals will be over precise (not wide enough). When there is no effect modification by history of coronary heart disease, the false-positive rates of association tests will be close to nominal.

The applicability of the Direct Peptide Reactivity Assay (DPRA), the KeratinoSens™ assay and the human cell line activation test (OECD Test Guidelines 442C, 442D, 442E) in predicting the skin sensitising potential of nine lipid (bio)chemicals was investigated. The results from the three assays were integrated using a published prediction model (PM), by which skin sensitisation is predicted if at least two of the three assays yield positive results. Of the eight test substances that were classified as non-sensitisers using available Guinea Pig Maximisation Test (GPMT) data, only five were correctly predicted as 'negative' in the PM. (However, only two were correctly predicted as 'negative' in the murine Local Lymph Node Assay.) The one lipid (bio)chemical that tested positive in the GPMT was also positive applying the PM. Based upon the outcome of the present study, lipid (bio)chemicals with a log Kow up to 7-8 appear amenable to the three assays. However, solubility problems, that were not evident initially, affected the performance of the DPRA. Further investigations are merited to address the conclusiveness of negative test results with concurrent lack of cytotoxicity in the in vitro assays, to evaluate if poorly soluble substances come into contact with the cells.

Objective A large body of epidemiologic data strongly suggests an association between excess adiposity and coronary artery disease (CAD). Low adiponectin levels, a hormone secreted only from adipocytes, have been associated with an increased risk of CAD in observational studies. However, these associations cannot clarify whether this relationship is causal or due to a shared set of causal factors or even confounding. Genome-wide association studies have identified common variants that influence adiponectin levels, providing valuable tools to examine the genetic relationship between adiponectin and CAD. Methods Using 145 genome wide significant SNPs for adiponectin from the ADIPOGen consortium ( n = 49,891), we tested whether adiponectin-decreasing alleles influenced risk of CAD in the CARDIoGRAM consortium ( n = 85,274). Results In single-SNP analysis, 5 variants among 145 SNPs were associated with increased risk of CAD after correcting for multiple testing ( P −4 ). Using a multi-SNP genotypic risk score to test whether adiponectin levels and CAD have a shared genetic etiology, we found that adiponectin-decreasing alleles increased risk of CAD ( P = 5.4 × 10 −7 ). Conclusion These findings demonstrate that adiponectin levels and CAD have a shared allelic architecture and provide rationale to undertake a Mendelian randomization studies to understand if this relationship is causal.

Adjustment for an uncorrelated covariate in a logistic regression changes the true value of an odds ratio for a unit increase in a risk factor. Even when there is no variation due to covariates, the odds ratio for a unit increase in a risk factor also depends on the distribution of the risk factor. An instrumental variable can be used to consistently estimate a causal effect in the presence of arbitrary confounding. With a logistic outcome model, we show that the simple ratio or two-stage instrumental variable estimate is consistent for the odds ratio of an increase in the population distribution of the risk factor equal to the change due to a unit increase in the instrument divided by the average change in the risk factor due to the increase in the instrument. This odds ratio is conditional within strata of the instrumental variable, but marginal across all other covariates, and is averaged across the population distribution of the risk factor. Where the proportion of variance in the risk factor explained by the instrument is small, this is similar to the odds ratio from a randomized controlled trial without adjustment for any covariates, where the intervention corresponds to the effect of a change in the population distribution of the risk factor. This implies that the ratio or two-stage instrumental variable method is not biased, as has been suggested, but estimates a different quantity to the conditional odds ratio from an adjusted multiple regression; a quantity which has arguably more relevance to an epidemiologist or a policy-maker, especially in the context of Mendelian randomization.

Vitamin D deficiency is common and associated with higher risk for and unfavourable outcome of many diseases. Limited data exist on genetic determinants of serum 25(OH)D concentration. In a cohort of the LURIC study (n=2974, median 25(OH)D concentration 15.5 ng/ml), we tested the hypothesis that variants (SNPs, n=244) of several genes (n=15) involved in different aspects of skin pigmentation, including melanosomal biogenesis (ATP7A, DTNBP1, BLOC1S5, PLDN, PMEL), melanosomal transport within melanocytes (RAB27A, MYO5A, MLPH); or various melanocyte signaling pathways (MC1R, MITF, PAX3, SOX10, DKK1, RACK1, CNR1) are predictive of serum 25(OH)D levels. Eleven SNPs located in 6 genes were associated (p0.05) with low or high serum 25(OH)D levels, 3 out of these 11 SNPs reached the aimed significance level after correction for multiple comparisons (FDR). In the linear regression model adjusted for sex, body mass index (BMI), year of birth and month of blood sample rs7565264 (MLPH), rs10932949 (PAX3), and rs9328451 (BLOC1S5) showed a significant association with 25(OH)D. The combined impact on variation of 25(OH)D serum levels (coefficient of determination (R(2))) for the 11 SNPs was 1.6% and for the 3 SNPs after FDR 0.3%. In Cox Regression we identified rs2292881 (MLPH) as having a significant association (advantage) with overall survival. Kaplan-Meier analysis did not show any significant impact of individual SNPs on overall survival. In conclusion, these results shed new light on the role of sunlight, skin pigmentation and vitamin D for human evolution.

Natural and synthetic chemicals are often used in the fragrance industry. A toxicological profile of the synthetic fragrance booster, 4-cycloocten-1-carbaldehyde, was generated using a test program including the following methods: acute oral toxicity, acute dermal toxicity, acute skin and eye irritation, skin sensitization, subchronic toxicity, and mutagenicity. The substance was strongly irritating to the skin but only weakly irritating to the eye. It gave a clear indication of having skin-sensitizing properties. Based on the comprehensive data from a mutagenicity test battery, 4-cycloocten-1-carbaldehyde was assessed to be nonmutagenic. Although its acute toxicological profile shows no toxicity after oral or dermal application, 4-cycloocten-1-carbaldehyde displays a complex toxicological response after repeated dosing over 13 weeks. 4-Cycloocten-1-carbaldehyde or its metabolites show clear nephrotoxic properties focusing on tubular cells of the kidney. In view of these data no no-effect level can be derived from this study with 4-cycloocten-1-carbaldehyde. A broad interaction of the test substance with various tissue types and cell parameters together with severe and irreversible organic defects even at low doses leads to the conclusion that 4-cycloocten-1-carbaldehyde is unsuitable for the intended use in industrial fragrance formulations.

Emulsions of water-soluble cutting fluids (wsCF) are used in large quantities in the metal industry. In order to reduce the costs for use and disposal of these fluids, new technologies are being introduced. Minimist Lubricant Supply (MLS) uses only minimal amounts of cutting fluids. In contrast to conventional wsCF, which are complex multicompound mixtures, MLS cutting fluids are composed of one or two components only, like fatty alcohols and fatty acid esters. The aim of the study was to identify and compare the mutagenic potential of these cutting fluids as a first indicator of a possible hazard of systemic effects after inhalation or dermal absorption of the fluids at the workplace. The Salmonella typhimurium assay (Ames assay) was used as a screening method to detect mutagenic effects of cutting fluids. Conventional wsCF and MLS cutting fluids were tested in the strains TA 98, TA 100, TA 102 and TA 104, in the presence and absence of an external metabolizing enzyme system (rat liver S9-mix), using a preincubation (20 min) test protocol. For MLS fluids, no mutagenicity was found in a concentration range between 1 and 10 mg/plate in the Ames assay. Among five tested conventional wsCF, two were mutagenic in the Ames assay at concentration ranges between 2.5 and 15 mg/plate. In cooperation with the manufacturer, 18 defined components of cutting fluids were tested separately. The results revealed that formaldehyde generators, derivatives of oxazolidine and hexahydrotriazine used as biocides for preservation of the fluids, were mutagenic. Four components were nonmutagenic but cytotoxic, whereas the remaining components displayed no bacterial mutagenicity. The present results show the potential hazard of biocides for workers handling these fluids. An exposure via inhalation and/or dermal absorption could cause an additional risk due to mutagenic ingredients. Under aspects of workers’ safety, a further discussion about the use of specific components in cutting fluids is recommended.

A mendelian randomization study based on data from multiple cohorts conducted by Karani Santhanakrishnan Vimaleswaran and colleagues re-examines the causal nature of the relationship between vitamin D levels and obesity., Background Obesity is associated with vitamin D deficiency, and both are areas of active public health concern. We explored the causality and direction of the relationship between body mass index (BMI) and 25-hydroxyvitamin D [25(OH)D] using genetic markers as instrumental variables (IVs) in bi-directional Mendelian randomization (MR) analysis. Methods and Findings We used information from 21 adult cohorts (up to 42,024 participants) with 12 BMI-related SNPs (combined in an allelic score) to produce an instrument for BMI and four SNPs associated with 25(OH)D (combined in two allelic scores, separately for genes encoding its synthesis or metabolism) as an instrument for vitamin D. Regression estimates for the IVs (allele scores) were generated within-study and pooled by meta-analysis to generate summary effects. Associations between vitamin D scores and BMI were confirmed in the Genetic Investigation of Anthropometric Traits (GIANT) consortium (n = 123,864). Each 1 kg/m2 higher BMI was associated with 1.15% lower 25(OH)D (p = 6.52×10−27). The BMI allele score was associated both with BMI (p = 6.30×10−62) and 25(OH)D (−0.06% [95% CI −0.10 to −0.02], p = 0.004) in the cohorts that underwent meta-analysis. The two vitamin D allele scores were strongly associated with 25(OH)D (p≤8.07×10−57 for both scores) but not with BMI (synthesis score, p = 0.88; metabolism score, p = 0.08) in the meta-analysis. A 10% higher genetically instrumented BMI was associated with 4.2% lower 25(OH)D concentrations (IV ratio: −4.2 [95% CI −7.1 to −1.3], p = 0.005). No association was seen for genetically instrumented 25(OH)D with BMI, a finding that was confirmed using data from the GIANT consortium (p≥0.57 for both vitamin D scores). Conclusions On the basis of a bi-directional genetic approach that limits confounding, our study suggests that a higher BMI leads to lower 25(OH)D, while any effects of lower 25(OH)D increasing BMI are likely to be small. Population level interventions to reduce BMI are expected to decrease the prevalence of vitamin D deficiency. Please see later in the article for the Editors' Summary, Editors' Summary Background Obesity—having an unhealthy amount of body fat—is increasing worldwide. In the US, for example, a third of the adult population is now obese. Obesity is defined as having a body mass index (BMI, an indicator of body fat calculated by dividing a person's weight in kilograms by their height in meters squared) of more than 30.0 kg/m2. Although there is a genetic contribution to obesity, people generally become obese by consuming food and drink that contains more energy than they need for their daily activities. Thus, obesity can be prevented by having a healthy diet and exercising regularly. Compared to people with a healthy weight, obese individuals have an increased risk of developing diabetes, heart disease and stroke, and tend to die younger. They also have a higher risk of vitamin D deficiency, another increasingly common public health concern. Vitamin D, which is essential for healthy bones as well as other functions, is made in the skin after exposure to sunlight but can also be obtained through the diet and through supplements. Why Was This Study Done? Observational studies cannot prove that obesity causes vitamin D deficiency because obese individuals may share other characteristics that reduce their circulating 25-hydroxy vitamin D [25(OH)D] levels (referred to as confounding). Moreover, observational studies cannot indicate whether the larger vitamin D storage capacity of obese individuals (vitamin D is stored in fatty tissues) lowers their 25(OH)D levels or whether 25(OH)D levels influence fat accumulation (reverse causation). If obesity causes vitamin D deficiency, monitoring and treating vitamin D deficiency might alleviate some of the adverse health effects of obesity. Conversely, if low vitamin D levels cause obesity, encouraging people to take vitamin D supplements might help to control the obesity epidemic. Here, the researchers use bi-directional “Mendelian randomization” to examine the direction and causality of the relationship between BMI and 25(OH)D. In Mendelian randomization, causality is inferred from associations between genetic variants that mimic the influence of a modifiable environmental exposure and the outcome of interest. Because gene variants do not change over time and are inherited randomly, they are not prone to confounding and are free from reverse causation. Thus, if a lower vitamin D status leads to obesity, genetic variants associated with lower 25(OH)D concentrations should be associated with higher BMI, and if obesity leads to a lower vitamin D status, then genetic variants associated with higher BMI should be associated with lower 25(OH)D concentrations. What Did the Researchers Do and Find? The researchers created a “BMI allele score” based on 12 BMI-related gene variants and two “25(OH)D allele scores,” which are based on gene variants that affect either 25(OH)D synthesis or breakdown. Using information on up to 42,024 participants from 21 studies, the researchers showed that the BMI allele score was associated with both BMI and with 25(OH)D levels among the study participants. Based on this information, they calculated that each 10% increase in BMI will lead to a 4.2% decrease in 25(OH)D concentrations. By contrast, although both 25(OH)D allele scores were strongly associated with 25(OH)D levels, neither score was associated with BMI. This lack of an association between 25(OH)D allele scores and obesity was confirmed using data from more than 100,000 individuals involved in 46 studies that has been collected by the GIANT (Genetic Investigation of Anthropometric Traits) consortium. What Do These Findings Mean? These findings suggest that a higher BMI leads to a lower vitamin D status whereas any effects of low vitamin D status on BMI are likely to be small. That is, these findings provide evidence for obesity as a causal factor in the development of vitamin D deficiency but not for vitamin D deficiency as a causal factor in the development of obesity. These findings suggest that population-level interventions to reduce obesity should lead to a reduction in the prevalence of vitamin D deficiency and highlight the importance of monitoring and treating vitamin D deficiency as a means of alleviating the adverse influences of obesity on health. Additional Information Please access these Web sites via the online version of this summary at http://dx.doi.org/10.1371/journal.pmed.1001383. The US Centers for Disease Control and Prevention provides information on all aspects of overweight and obesity (in English and Spanish); a data brief provides information about the vitamin D status of the US population The World Health Organization provides information on obesity (in several languages) The UK National Health Service Choices website provides detailed information about obesity and a link to a personal story about losing weight; it also provides information about vitamin D The International Obesity Taskforce provides information about the global obesity epidemic The US Department of Agriculture's ChooseMyPlate.gov website provides a personal healthy eating plan; the Weight-control Information Network is an information service provided for the general public and health professionals by the US National Institute of Diabetes and Digestive and Kidney Diseases (in English and Spanish) The US Office of Dietary Supplements provides information about vitamin D (in English and Spanish) MedlinePlus has links to further information about obesity and about vitamin D (in English and Spanish) Wikipedia has a page on Mendelian randomization (note: Wikipedia is a free online encyclopedia that anyone can edit; available in several languages) Overview and details of the collaborative large-scale genetic association study (D-CarDia) provide information about vitamin D and the risk of cardiovascular disease, diabetes and related traits

Background— After age, sex is the most important risk factor for coronary artery disease (CAD). The mechanism through which women are protected from CAD is still largely unknown, but the observed sex difference suggests the involvement of the reproductive steroid hormone signaling system. Genetic association studies of the gene-encoding Estrogen Receptor α ( ESR1 ) have shown conflicting results, although only a limited range of variation in the gene has been investigated. Methods and Results— We exploited information made available by advanced new methods and resources in complex disease genetics to revisit the question of ESR1 's role in risk of CAD. We performed a meta-analysis of 14 genome-wide association studies (CARDIoGRAM discovery analysis, N=≈87 000) to search for population-wide and sex-specific associations between CAD risk and common genetic variants throughout the coding, noncoding, and flanking regions of ESR1 . In addition to samples from the MIGen (N=≈6000), WTCCC (N=≈7400), and Framingham (N=≈3700) studies, we extended this search to a larger number of common and uncommon variants by imputation into a panel of haplotypes constructed using data from the 1000 Genomes Project. Despite the widespread expression of ERα in vascular tissues, we found no evidence for involvement of common or low-frequency genetic variation throughout the ESR1 gene in modifying risk of CAD, either in the general population or as a function of sex. Conclusions— We suggest that future research on the genetic basis of sex-related differences in CAD risk should initially prioritize other genes in the reproductive steroid hormone biosynthesis system.

OBJECTIVE Glycated hemoglobin (HbA1c), used to monitor and diagnose diabetes, is influenced by average glycemia over a 2- to 3-month period. Genetic factors affecting expression, turnover, and abnormal glycation of hemoglobin could also be associated with increased levels of HbA1c. We aimed to identify such genetic factors and investigate the extent to which they influence diabetes classification based on HbA1c levels. RESEARCH DESIGN AND METHODS We studied associations with HbA1c in up to 46,368 nondiabetic adults of European descent from 23 genome-wide association studies (GWAS) and 8 cohorts with de novo genotyped single nucleotide polymorphisms (SNPs). We combined studies using inverse-variance meta-analysis and tested mediation by glycemia using conditional analyses. We estimated the global effect of HbA1c loci using a multilocus risk score, and used net reclassification to estimate genetic effects on diabetes screening. RESULTS Ten loci reached genome-wide significant association with HbA1c, including six new loci near FN3K (lead SNP/P value, rs1046896/P = 1.6 × 10−26), HFE (rs1800562/P = 2.6 × 10−20), TMPRSS6 (rs855791/P = 2.7 × 10−14), ANK1 (rs4737009/P = 6.1 × 10−12), SPTA1 (rs2779116/P = 2.8 × 10−9) and ATP11A/TUBGCP3 (rs7998202/P = 5.2 × 10−9), and four known HbA1c loci: HK1 (rs16926246/P = 3.1 × 10−54), MTNR1B (rs1387153/P = 4.0 × 10−11), GCK (rs1799884/P = 1.5 × 10−20) and G6PC2/ABCB11 (rs552976/P = 8.2 × 10−18). We show that associations with HbA1c are partly a function of hyperglycemia associated with 3 of the 10 loci (GCK, G6PC2 and MTNR1B). The seven nonglycemic loci accounted for a 0.19 (% HbA1c) difference between the extreme 10% tails of the risk score, and would reclassify ∼2% of a general white population screened for diabetes with HbA1c. CONCLUSIONS GWAS identified 10 genetic loci reproducibly associated with HbA1c. Six are novel and seven map to loci where rarer variants cause hereditary anemias and iron storage disorders. Common variants at these loci likely influence HbA1c levels via erythrocyte biology, and confer a small but detectable reclassification of diabetes diagnosis by HbA1c.

Extensive research has been conducted over the past decades to develop alternatives to the rabbit eye irritation test (Draize test) used in a regulatory context to assess eye irritation potentials. Although no single in vitro test has emerged as being completely acceptable for full replacement, various tests are considered to be suitable and are regularly used to assess certain aspects. Amongst these, the Hen's Egg Test Chorioallantoic Membrane (HET-CAM) has gained regulatory acceptance in various countries to classify severe eye irritants. In this retrospective study, historical eye irritation data (in vivo and in vitro) from 137 samples (approx. 75% non-irritants; 25% (severe) irritants) tested both in the HET-CAM and Draize eye test was compared with regard to the predicted eye irritation classes under the GHS and the traditional EU classification system (DSD).The overall concordance was in the range of 80-90%. A high specificity (96-98%, depending on the classification system and the chosen discrimination) but rather low sensitivity (48-65%) was observed. The study indicates that HET-CAM results are useful as part of weight-of-evidence assessments or in tiered approaches to assess eye irritation potentials rather than as stand-alone classification method.

Emulsions of water-soluble cutting fluids (wsCF) and non-water-soluble cutting fluids are used in large quantities for machining processes in the metal processing industry. From toxicological and epidemiological studies health risks resulting from handling of cutting fluids are well-known. Recently, the new technology of "minimist lubricant supply" (MLS) has been introduced which uses only minimal amounts of cutting fluids, in order to reduce the costs for use and disposal. This involves higher temperatures to which the fluids are subjected. Water-soluble cutting fluids supplemented with formaldehyde donors as biocides may lead to mutagenic potential. Current data show that attention should be given to generation of other genotoxic aldehydes from cutting fluids under condition of use, referring to both conventional and "minimist lubricant supply" technologies.

Statistical analysis plan for our study regarding anti-citrullinated protein antibodies in patients with coronary heart disease, without concomitant rheumatoid arthritis. Drafted and publishedbefore the start of data analysis. Abstract Background: Rheumatoid arthritis (RA) is a prevalent autoimmune disease leading to joint damage, disability, and increased mortality. Approximately 69% (95% confidence interval (CI) 39 to 94%) of patients with RA are positive for anti-citrullinated protein antibodies (ACPA). ACPA are very specific for RA (specificity of 97% (range 81–100%)) and are seen as markers for underlying autoimmune pathophysiology. ACPA is detected in approximately 1% of healthy individuals. Most risk factors known for RA, such as smoking, or the presence of HLA shared epitope alleles are exclusively associated with autoantibody-positive disease. ACPA-positive RA patients have increased cardiovascular mortality. ACPA have been described in 10% of patients with coronary heart disease (CHD) without concomitant RA, a much higher prevalence than would be expected. Furthermore, ACPA were independently associated with poor cardiovascular outcomes and significantly increased mortality in these patients. Objectives: To investigate the ACPA response in CHD by examining both the clinical phenotype of ACPA-positive CHD patients and the immunological characteristics of the ACPA in these patients. These findings will be compared to the existing data on the ACPA response in RA patients to gain more insight in the mechanisms underlying ACPA development and the specific ACPA features important in the pathophysiology of both diseases. Part 1 ACPA in coronary heart disease How many and which patients with CHD are ACPA-positive? Can we replicate the findings that around 10% of patients with CHD without concomitant RA is ACPA-positive? Part 2 Clinical phenotype, risk factors and disease outcomes Do ACPA-positive CHD patients differ from CHD-patients without ACPA regarding clinical phenotype and risk factors, such as female sex, smoking and low-grade inflammation? Do ACPA-positive CHD patients differ from CHD-patients without ACPA regarding disease outcomes, such as cardiovascular disease and all-cause mortality? Is ACPA-positivity in CHD associated with the same risk factors and clinical outcomes as ACPA-positivity in RA? Part 3 Immunological characteristics of ACPA What are the distinct immunological characteristics of ACPA in ACPA-positive CHD patients? We will focus on characteristics such as isotype usage, fine-specificity, avidity and glycosylation. Do the ACPA features in CHD differ from the ACPA features found in RA patients? Are certain ACPA features associated with clinical (CHD) outcomes, such as cardiovascular disease and all-cause mortality, in patients with and without RA? Methods:The specific patient populations and methods that will be used for each objective are stated in the table below. Stored samples and data from the following studies will be used: the clarithromycin for patients with stable coronary heart disease (CLARICOR) trial cohort containing CHD patients without RA ; the Leiden Early Arthritis Clinic (EAC) cohort containing RA patients ; and a cohort of healthy controls. The findings in the CLARICOR trial will be replicated in another CHD cohort: the LUdwigshafen Risk and Cardiovascular Health (LURIC) study . The flow of the analyses will be as following: First, we will determine the percentage of ACPA positive patients in the CLARICOR and LURIC studies. If the prevalence of ACPA is indeed increased in those patients compared to healthy donors, we will proceed with part 2 and 3. Table 2 Topic Populations Methods Part 1 Presence of ACPA in CHD (% of individuals positive) CLARICOR and LURIC Healthy population Chi-square Part 2A Risk factors associated with ACPA-positivity in CHD ACPA-positives in CLARICOR and LURIC ACPA-negatives in CLARICOR and LURIC Univariate and multivariate logistic regression Part 2B Disease outcome in ACPA-positive compared to ACPA-negative CHD patients ACPA-positives in CLARICOR (placebo group) and LURIC ACPA-negatives in CLARICOR (placebo group) and LURIC Univariate and multivariate (cause-specific) Cox regression Part 2C Compare risk factors/outcomes of ACPA-positivity in CHD and RA CLARICOR and LURIC Literature / EAC data Use results of 2A and 2B, compare with literature / EAC data Part 3A ACPA characteristics in CHD ACPA-positives in CLARICOR and LURIC Laboratory measurements Part 3B Compare ACPA features in CHD with ACPA features found in RA ACPA-positives in CLARICOR and LURIC ACPA-positive EAC RA patients T-test, Mann-Whitney U test, or chi-square Part 3C Association specific ACPA features and outcome in CHD and RA ACPA-positives in CLARICOR and LURIC ACPA-positive EAC RA patients Univariate and multivariate Cox regression Discussion:ACPA are considered specific for rheumatoid arthritis. The findings that ACPA can also be present in patients with coronary heart disease offers a unique opportunity to learn more about pathophysiology of both diseases. With this study we aim to perform an investigation of the ACPA response in coronary heart disease (CHD) by examining both the clinical phenotype, risk factors and disease outcomes of ACPA-positive CHD patients and the immunological characteristics of the ACPA. By investigating whether ACPA-positivity in CHD is associated with the same risk factors, clinical outcomes and ACPA characteristics as ACPA in RA patients, we hope to gain more insight in the mechanisms underlying ACPA development and the specific ACPA features important in the pathophysiology of both diseases.