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Coronavirus disease-2019 (COVID-19) is associated with increased thromboembolic complications. Long-term alteration in the coagulation system after acute COVID-19 infection is still a subject of research. Furthermore, the effect of sera from convalescent subjects on platelets is not known. In this study, we investigated platelet phenotype, coagulation, and fibrinolysis in COVID-19 convalescent plasma (CCP) donors and analyzed convalescent sera-induced effects on platelets. We investigated CCP donors who had a history of mild COVID-19 infection and donors who did not have COVID-19 were used as controls. We analyzed phosphatidylserine (PS) externalization, CD62p expression, and glycoprotein VI (GPVI) shedding both in platelet-rich plasma (PRP) and after incubation of washed healthy platelets with donors' sera using flow cytometry. Coagulation and fibrinolysis systems were assessed with thromboelastometry. Forty-seven CCP donors (22 males, 25 females; mean age (±SD): 41.4 ± 13.7 years) with a history of mild COVID-19 infection were included. Median duration after acute COVID-19 infection was 97 days (range, 34–401). We did not find an increased PS externalization, CD62p expression, or GPVI shedding in platelets from CCP donors. Sera from CCP donors did not induce PS externalization or GPVI shedding in healthy platelets. Sera-induced CD62p expression was slightly, albeit statistically significantly, lower in CCP donors than in plasma donors without a history of COVID-19. One patient showed increased maximum clot firmness and prolonged lysis time in thromboelastometry. Our findings suggest that procoagulant platelet phenotype is not present after mild COVID-19. Furthermore, CCP sera do not affect the activation status of platelets.

Background The calcium sensing receptor (CASR) is a G protein-coupled receptor that is responsible for assessing extracellular Ca2+ levels and thus plays a crucial role in calcium homeostasis. Hypercalcemia is a metabolic risk factor for pancreatitis and rare CASR variants have been described in patients with chronic pancreatitis. At the carboxy-terminal tail of CASR, there is a cluster of three common polymorphisms, p.A986S (rs1801725), p.R990G (rs1042636) and p.Q1011E (rs1801726), which have been associated with chronic pancreatitis in various studies, but with conflicting results. Methods We examined 542 German and 339 French patients with chronic pancreatitis as well as 1025 German controls for the 3 common CASR polymorphism by melting curve analysis. For comparison, we used genotype data from 583 French controls from a previous study. In addition, we functionally analyzed the three variants by NFAT and SRE luciferase reporter systems as well as Western blotting and verified cell surface expression by ELISA. Results In both cohorts, neither the genotype nor the allele frequencies differed significantly between patients and controls. In both luciferase assays, p.R990G showed a significant leftward shift, indicating an increased responsiveness of the receptor. p.A986S showed a leftward shift in the SRE but not in the NFAT reporter assay, while the responsiveness of p.Q1011E did not differ from the wild-type. These functional studies therefore do not support the contributions of variant CASR to increasing the risk of pancreatitis. Conclusions The three frequent CASR polymorphisms are unlikely to increase the risk for chronic pancreatitis.

Contains fulltext : 229360.pdf (Publisher’s version ) (Closed access) BACKGROUND: The G-protein-coupled receptor Class C Group 6 Member A (GPRC6A) is activated by multiple ligands and is important for the regulation of calcium homeostasis. Extracellular calcium is capable to increase NLRP3 inflammasome activity of the innate immune system and deletion of this proinflammatory pathway mitigated pancreatitis severity in vivo. As such this pathway and the GPRC6A receptor is a reasonable candidate gene for pancreatitis. Here we investigated the prevalence of sequence variants in the GPRC6A locus in different pancreatitis aetiologies. METHODS: We selected 6 tagging SNPs with the SNPinfo LD TAG SNP Selection tool and the functional relevant SNP rs6907580 for genotyping. Cohorts from Germany, further European countries and China with up to 1,124 patients and 1,999 controls were screened for single SNPs with melting curve analysis. RESULTS: We identified an association of rs1606365(G) with alcoholic chronic pancreatitis in a German (odds ratio (OR) 0.76, 95% confidence interval (CI) 0.65-0.89, p = 8 × 10(-5)) and a Chinese cohort (OR 0.78, 95% CI 0.64-0.96, p = 0.02). However, this association was not replicated in a combined cohort of European patients (OR 1.18, 95% CI 0.99-1.41, p = 0.07). Finally, no association was found with acute and non-alcoholic chronic pancreatitis. CONCLUSIONS: Our results support a potential role of calcium sensing receptors and inflammasome activation in alcoholic chronic pancreatitis development. As the functional consequence of the associated variant is unclear, further investigations might elucidate the relevant mechanisms.

Background and purpose The vasomotor role of K2P potassium channels during early postnatal development has never been investigated. We tested the hypothesis that TASK-1 channel (K2P family member) contribution to arterial vascular tone and BP is higher in the early postnatal period than in adulthood. Experimental approach We studied 10- to 15-day-old ("young") and 2- to 3-month-old ("adult") male rats performing digital PCR (dPCR) (using endothelium-intact saphenous arteries), isometric myography, sharp microelectrode technique, quantitative PCR (qPCR) and Western blotting (using endothelium-denuded saphenous arteries), and arterial pressure measurements under urethane anaesthesia. Key results We found mRNA of Kcnk1-Kcnk7, Kcnk12, and Kcnk13 genes to be expressed in rat saphenous artery, and Kcnk3 (TASK-1) and Kcnk6 (TWIK-2) were most abundant in both age groups. The TASK-1 channel blocker AVE1231 (1 μmol·L-1 ) prominently depolarized arterial smooth muscle and increased basal tone level and contractile responses to methoxamine of arteries from young rats but had almost no effect in adult rats. The level of TASK-1 mRNA and protein expression was higher in arteries from young compared with adult rats. Importantly, intravenous administration of AVE1231 (4 mg·kg-1 ) had no effect on mean arterial pressure in adult rats but prominently raised it in young rats. Conclusion and implications We showed that TASK-1 channels are important for negative feedback regulation of vasocontraction in young but not adult rats. The influence of TASK-1 channels most likely contributes to low BP level at perinatal age.

OBJECTIVES. Premature activation of the digestive protease trypsin within the pancreatic parenchyma is a critical factor in the pathogenesis of pancreatitis. Alterations in genes that affect intra-pancreatic trypsin activity are associated with chronic pancreatitis (CP). Recently, carboxyl ester lipase (CEL) emerged as a trypsin-independent risk gene. Here, we evaluated PNLIP encoding pancreatic lipase as a potential novel susceptibility gene for CP. METHODS. We analyzed all 13 PNLIP exons in 429 German non-alcoholic CP patients and in 600 German control subjects, in 632 patients and 957 controls from France, and in 223 patients and 1070 controls from Japan by DNA sequencing. Additionally, we analyzed selected exons in further 545 CP patients and 1849 controls originating from Germany, USA and India. We assessed the cellular secretion, lipase activity and proteolytic stability of recombinant PNLIP variants. RESULTS. In the German discovery cohort, 8/429 (1.9%) patients and 2/600 (0.3%) controls carried a PNLIP missense variant (P=0.02, OR=5.7, 95% CI=1.1–38.9). Variants detected in patients were prone to proteolytic degradation by trypsin and chymotrypsin. In the French replication cohort, protease-sensitive variants were also enriched in patients with early-onset CP (5/632 [0.8%]) versus controls (1/957 [0.1%]) (P=0.04, OR=7.6, 95% CI=0.9–172.9). In contrast, we detected no protease-sensitive variants in the non-European populations. In the combined European data, protease-sensitive variants were found in 13/1163 cases (1.1%) and in 3/3000 controls (0.1%) (OR=11.3, 95% CI=3.0–49.9, P

Background and Objectives: Non-invasive assays for predicting foetal blood group status in pregnancy serve as valuable clinical tools in the management of pregnancies at risk of detrimental consequences due to blood group antigen incompatibility. To secure clinical applicability, assays for non-invasive prenatal testing of foetal blood groups need to follow strict rules for validation and quality assurance. Here, we present a multi-national position paper with specific recommendations for validation and quality assurance for such assays and discuss their risk classification according to EU regulations. Materials and Methods: We reviewed the literature covering validation for in-vitro diagnostic (IVD) assays in general and for non-invasive foetal RHD genotyping in particular. Recommendations were based on the result of discussions between co-authors. Results: In relation to Annex VIII of the In-Vitro-Diagnostic Medical Device Regulation 2017/746 of the European Parliament and the Council, assays for non-invasive prenatal testing of foetal blood groups are risk class D devices. In our opinion, screening for targeted anti-D prophylaxis for non-immunized RhD negative women should be placed under risk class C. To ensure high quality of non-invasive foetal blood group assays within and beyond the European Union, we present specific recommendations for validation and quality assurance in terms of analytical detection limit, range and linearity, precision, robustness, pre-analytics and use of controls in routine testing. With respect to immunized women, different requirements for validation and IVD risk classification are discussed. Conclusion: These recommendations should be followed to ensure appropriate assay performance and applicability for clinical use of both commercial and in-house assays. (Less)

In systemic mastocytosis (SM), qualitative and serial quantitative assessment of the KIT D816V mutation is of diagnostic and prognostic relevance. We investigated peripheral blood and bone marrow samples of 161 patients (indolent SM (ISM), n = 40, advanced SM, AdvSM, n = 121) at referral and during follow-up for the KIT D816V variant allele frequency (VAF) at the DNA-level and the KIT D816V expressed allele burden (EAB) at the RNA-level. A round robin test with four participating laboratories revealed an excellent correlation (r &gt, 0.99, R2 &gt, 0.98) between three different DNA-assays. VAF and EAB strongly correlated in ISM (r = 0.91, coefficient of determination, R2 = 0.84) but only to a lesser extent in AdvSM (r = 0.71, R2 = 0.5). However, as compared to an EAB/VAF ratio ≤2 (cohort A, 77/121 patients, 64%) receiver operating characteristic (ROC) analysis identified an EAB/VAF ratio of &gt, 2 (cohort B, 44/121 patients, 36%) as predictive for an advanced phenotype and a significantly inferior median survival (3.3 vs. 11.7 years, p = 0.005). In terms of overall survival, Cox-regression analysis was only significant for the EAB/VAF ratio &gt, 2 (p = 0.006) but not for VAF or EAB individually. This study demonstrates for the first time that the transcriptional activity of KIT D816V may play an important role in the pathophysiology of SM.

A pro-coagulative state is related to increased risk of cardiovascular diseases but also certain cancers. Since experimental and smaller human studies suggest that diet, physical activity, and body weight may all affect coagulation, we evaluated associations between these lifestyle factors and hemostatic biomarkers in a population-based study. Cross-sectional baseline data from 2267 randomly selected participants of EPIC-Heidelberg (age range 35&ndash, 65 years) was used. Fibrinogen, glycoprotein IIb/IIIa, P-selectin, thrombomodulin (TM), and thrombopoietin (TPO) were measured in baseline plasma samples. A score reflecting adherence to the World Cancer Research Fund/American Institute for Cancer Research (WCRF/AICR) recommendations for cancer prevention was created. Associations between the WCRF/AICR score as well as its individual components and hemostatic biomarkers were analyzed by linear regression models. Multivariable-adjusted geometric means (95% confidence intervals) of TM and TPO were higher with greater adherence to the WCRF/AICR recommendations (TM, lowest vs. highest score category: 2.90 (2.7,3.1) vs. 3.10 (2.9,3.3) ng/mL, plinear trend = 0.0001, TPO: 328 (302,356) vs. 348 (321,378) pg/mL, plinear trend = 0.0007). These associations were driven by lower alcohol and meat consumption among persons with higher WCRF/AICR scores. Our results indicate that lifestyle factors favorably affect TM and TPO, two hemostatic factors implicated in chronic disease development.

Background Early diagnosis of Alzheimer's disease (AD) is challenging, and easily accessible biomarkers are an unmet need. Blood platelets frequently serve as peripheral model for studying AD pathogenesis and might represent a reasonable biomarker source. Objective In the present study, we investigated the potential to differentiate AD patients from healthy controls (HC) based on blood count, platelet morphology, and function as well as molecular markers at the time of first clinical diagnosis. Methods Blood samples from 40 AD patients and 29 age-matched HC were included for determination of 78 parameter by blood counting, platelet morphometry, aggregometry, flow cytometry (CD62P, CD63, activated fibrinogen receptor), protein quantification of nicotinic acetylcholine receptor α7 (nAChRα7) and caveolin-1 (CAV-1), and miRNA quantification (miR-26b, miR-199a, miR-335). Group comparison between patients and controls was performed in univariate and multivariate statistical analyses. Results AD patients showed significantly lower aggregation response to ADP and arachidonic acid and significantly decreased CD62P and CD63 surface expression induced by ADP and U46619 compared to HC. Relative nAChRα7 and CAV-1 expression was significantly higher AD platelets than in HC. Multivariate analysis of 63 parameter revealed significant differences between AD patients and healthy controls. The best performing feature model revealed a sensitivity of 96.6%, a specificity of 80.0%, and a positive predictive value of 89.3%. No grouping could be achieved by using single parameter groups. Conclusion Significant differences between platelet characteristics from AD patients and HC at the time of first clinical diagnosis were observed. The best performing parameter can be used as a blood-based biomarker for AD diagnosis in a multivariate model in addition to the standardized mental tests.

/Objectives: AGE and their receptors like RAGE and Galectin-3 can activate inflammatory pathways and have been associated with chronic inflammatory diseases. Several studies investigated the role of AGE, Galectin-3 and sRAGE in pancreatic diseases, whereas no comprehensive data for chronic pancreatitis (CP) are available.Serum samples from CP patients without an active inflammatory process (85 ACP; 26 NACP patients) and 40 healthy controls were collected. Levels of AGE, sRAGE and Galectin-3 were measured by ELISA. To exclude potential influences of previously described RAGE SNPs on detected serum levels, we analyzed variants rs207128, rs207060, rs1800625, and rs1800624 by melting curve technique in 378 CP patients and 338 controls.AGE and Galectin-3 serum levels were significantly elevated in both ACP and NACP patients compared to controls (AGE: 56.61 ± 3.043 vs. 31.71 ± 2.308 ng/mL; p 0.001; Galectin-3: 16.63 ± 0.6297 vs. 10.81 ± 0.4835 ng/mL; p 0.001). In contrast, mean serum sRAGE levels were significantly reduced in CP patients compared to controls (sRAGE: 829.7 ± 37.10 vs. 1135 ± 55.74 ng/mL; p 0.001). All results were consistent after correction for gender, age and diabetes mellitus. No genetic association with CP was found.Our extensive analysis demonstrated the importance of aging related pathways in the pathogenesis of CP. As the results were consistent in ACP and NACP, both entities most likely share common pathomechanisms. Most probably the involved pathways are a general hallmark of an inflammatory state in CP that is even present in symptom-free intervals.

Chronic hemorrhagic diathesis in patients showing normal levels of plasmatic clotting factors strongly suggests for congenital platelet disorders. We report on a pediatric patient (male, 3 years, D1) with mild bleeding. A sibling (D2), his mother (D3) and father (D4) were included for laboratory investigation. Platelet counts in D1, D2 and D4 indicated mild thrombocytopenia (100 Gpt/L). D1 and D3 platelets showed significantly diminished aggregation response on arachidonic acid and U46619 stimulation. Immunostaining for platelet proteins on blood smears of D1 and D2 indicated defects in ß1-tubulin. Exon sequencing of

Contains fulltext : 215401.pdf (Publisher’s version ) (Open Access) INTRODUCTION: Chronic pancreatitis (CP) may be caused by oxidative stress. An important source of reactive oxygen species (ROS) is the methylglyoxal-derived formation of advanced glycation endproducts (AGE). Methylglyoxal is detoxified by Glyoxalase I (GLO1). A reduction in GLO1 activity results in increased ROS. Single nucleotide polymorphisms (SNPs) of GLO1 have been linked to various inflammatory diseases. Here, we analyzed whether common GLO1 variants are associated with alcoholic (ACP) and non-alcoholic CP (NACP). METHODS: Using melting curve analysis, we genotyped a screening cohort of 223 ACP, 218 NACP patients, and 328 controls for 11 tagging SNPs defined by the SNPinfo LD TAG SNP Selection tool and the functionally relevant variant rs4746. For selected variants the cohorts were extended to up to 1,441 patient samples. RESULTS: In the ACP cohort, comparison of genotypes for rs1937780 between patients and controls displayed an ambiguous result in the screening cohort (p = 0.08). However, in the extended cohort of 1,441 patients no statistically significant association was found for the comparison of genotypes (p = 0.11), nor in logistic regression analysis (p = 0.214, OR 1.072, 95% CI 0.961-1.196). In the NACP screening cohort SNPs rs937662, rs1699012, and rs4746 displayed an ambiguous result when patients were compared to controls in the recessive or dominant model (p = 0.08, 0.08, and 0.07, respectively). Again, these associations were not confirmed in the extended cohorts (rs937662, dominant model: p = 0.07, logistic regression: p = 0.07, OR 1.207, 95% CI 0.985-1.480) or in the replication cohorts for rs4746 (Germany, p = 0.42, OR 1.080, 95% CI 0.673-1.124; France, p = 0.19, OR 0.90, 95% CI 0.76-1.06; China, p = 0.24, OR 1.18, 95% CI 0.90-1.54) and rs1699012 (Germany, Munich; p = 0.279, OR 0.903, 95% CI 0.750-1.087). CONCLUSIONS: Common GLO1 variants do not increase chronic pancreatitis risk.

Background & Aims Changes in pancreatic calcium levels affect secretion and might be involved in development of chronic pancreatitis (CP). We investigated the association of CP with the transient receptor potential cation channel subfamily V member 6 gene (TRPV6), which encodes a Ca2+-selective ion channel, in an international cohort of patients and in mice. Methods We performed whole-exome DNA sequencing from a patient with idiopathic CP and from his parents, who did not have CP. We validated our findings by sequencing DNA from 300 patients with CP (not associated with alcohol consumption) and 1070 persons from the general population in Japan (control individuals). In replication studies, we sequenced DNA from patients with early-onset CP (20 years or younger) not associated with alcohol consumption from France (n = 470) and Germany (n = 410). We expressed TRPV6 variants in HEK293 cells and measured their activity using Ca2+ imaging assays. CP was induced by repeated injections of cerulein in TRPV6mut/mut mice. Results We identified the variants c.629C>T (p.A210V) and c.970G>A (p.D324N) in TRPV6 in the index patient. Variants that affected function of the TRPV6 product were found in 13 of 300 patients (4.3%) and 1 of 1070 control individuals (0.1%) from Japan (odds ratio [OR], 48.4; 95% confidence interval [CI], 6.3–371.7; P = 2.4 × 10–8). Twelve of 124 patients (9.7%) with early-onset CP had such variants. In the replication set from Europe, 18 patients with CP (2.0%) carried variants that affected the function of the TRPV6 product compared with 0 control individuals (P = 6.2 × 10–8). Variants that did not affect the function of the TRPV6 product (p.I223T and p.D324N) were overrepresented in Japanese patients vs control individuals (OR, 10.9; 95% CI, 4.5–25.9; P = 7.4 × 10–9 for p.I223T and P = .01 for p.D324N), whereas the p.L299Q was overrepresented in European patients vs control individuals (OR, 3.0; 95% CI, 1.9–4.8; P = 1.2 × 10–5). TRPV6mut/mut mice given cerulein developed more severe pancreatitis than control mice, as shown by increased levels of pancreatic enzymes, histologic alterations, and pancreatic fibrosis. Conclusions We found that patients with early-onset CP not associated with alcohol consumption carry variants in TRPV6 that affect the function of its product, perhaps by altering Ca2+ balance in pancreatic cells. TRPV6 regulates Ca2+ homeostasis and pancreatic inflammation.

Little is known about circulating biomarkers of vascular injury in relation to cardiovascular disease risk. Thus, we evaluated associations between six novel markers (E-Selectin, P-Selectin, thrombomodulin, thrombopoietin, intercellular adhesion molecule 3 and GPIIb/IIIa) and established cardiovascular risk factors as well as the risk of myocardial infarction (MI) in a population-based study. Biomarkers were measured in pre-diagnostic plasma samples of a case-cohort subset of EPIC-Heidelberg (incident MI cases: n = 369, random sub-cohort: n = 2,418). Generalized Linear models were used to analyse cross-sectional associations between biomarkers and cardiovascular risk factors. Multivariable Cox Regression analyses were carried out to obtain Hazard Ratios (HRs) of MI across quartiles of biomarkers levels. Cross-sectional analyses showed that sex, smoking, alcohol consumption, diabetes and exogenous hormone use were associated with biomarker levels. However, while fibrinogen was associated with MI risk (HR per standard deviation: 2.97 [95% confidence interval: 1.61, 5.46]), none of the six novel biomarkers was associated with MI risk after multivariable adjustment. In a population-based cohort, biomarkers of vascular injury were associated with established cardiovascular risk factors, but not MI risk. The tested biomarkers may reflect pathophysiological alterations in cardiovascular disease development rather than constituting independent MI risk factors.

DNA methylation changes in peripheral blood DNA have been shown to be associated with solid tumors. We sought to identify methylation alterations in whole blood DNA that are associated with breast cancer (BC). Epigenome-wide DNA methylation profiling on blood DNA from BC cases and healthy controls was performed by applying Infinium HumanMethylation450K BeadChips. Promising CpG sites were selected and validated in three independent larger sample cohorts via MassARRAY EpiTyper assays. CpG sites located in three genes (cg06418238 in RPTOR, cg00736299 in MGRN1 and cg27466532 in RAPSN), which showed significant hypomethylation in BC patients compared to healthy controls in the discovery cohort (p < 1.00 x 10-6) were selected and successfully validated in three independent cohorts (validation I, n =211; validation II, n=378; validation III, n=520). The observed methylation differences are likely not cell-type specific, as the differences were only seen in whole blood, but not in specific sub cell-types of leucocytes. Moreover, we observed in quartile analysis that women in the lower methylation quartiles of these three loci had higher ORs than women in the higher quartiles. The combined AUC of three loci was 0.79 (95%CI 0.73-0.85) in validation cohort I, and was 0.60 (95%CI 0.54-0.66) and 0.62 (95%CI 0.57-0.67) in validation cohort II and III, respectively. Our study suggests that hypomethylation of CpG sites in RPTOR, MGRN1 and RAPSN in blood is associated with BC and might serve as blood-based marker supplements for BC if these could be verified in prospective studies.

Background: While enhanced platelet activation and a procoagulant state may drive lung cancer progression and metastases, less is known about their role in earlier phases of cancer development. Thus, we evaluated whether prediagnostic biomarkers of platelet activation and coagulation are related to the risk of lung cancer in the prospective EPIC-Heidelberg Study using a case–cohort design. Methods: Levels of fibrinogen, soluble glycoprotein (sGP) IIb/IIIa, soluble P-selectin (sP-selectin), soluble thrombomodulin (sTM), and thrombopoietin (TPO) were measured in baseline plasma samples of a random subcohort (n = 2,480) and incident cases of lung cancer (n = 190). Multivariable-adjusted Cox proportional hazards regression analyses were used to obtain HRs of lung cancer across quartiles of biomarker levels. Results: Fibrinogen [HR highest vs. lowest quartile: 1.91 (95% confidence interval: 1.09–3.34)] and sP-Selectin [HR: 2.51 (1.39–4.52)] were significantly associated with lung cancer risk in multivariable adjusted Cox regression models. Adding both biomarkers to the established PLCOm2012 algorithm, which alone showed a C-statistic of 0.788, led to a slight increment in lung cancer risk prediction, with a C-statistic of 0.814. Conclusion: Our findings indicate that enhanced platelet activation and a procoagulative state contribute to lung carcinogenesis. Impact: The current prospective study supports the hypothesis of increased coagulation being a possible driver of lung carcinogenesis, as strong positive associations were found between two procoagulative markers, sP-Selectin and fibrinogen, with lung cancer risk. Both biomarkers could improve lung cancer risk prediction, but external validation of the results is needed.

Platelets are thought to play an important role in metastasis formation, although the mechanisms involved remain incompletely understood. Here we studied the influence of platelet numbers on organ-specific metastasis to the lungs and lymph nodes using Tpo deficient mice that have low platelet counts. After tail vein injection of 4T1 breast cancer cells, the number of lung metastases was significantly lower in Tpo

While enhanced platelet activation may drive cancer progression and metastases, less is known about its role in early cancer development. Thus, we evaluated whether pre-diagnostic biomarkers of platelet activation and coagulation are related to the risks of common cancers in the prospective EPIC-Heidelberg Study using a case-cohort design. Levels of fibrinogen, soluble glycoprotein (sGP) IIb/IIIa, soluble P-selectin (sP-selectin), soluble thrombomodulin (sTM), and thrombopoietin (TPO) were measured in baseline plasma samples of a random subcohort (n = 2,480) and incident cases of breast (n = 605), prostate (n = 543), and colorectal cancer (n = 249). Multivariable Cox regression models revealed no statistically significant associations between biomarker concentrations and any of the cancer endpoints. Subgroup analyses showed a significant inverse relationship between TPO and colorectal cancer among men, with a hazard ratio (HR, highest vs. lowest quartile) of 0.60 (95% confidence interval: 0.37,0.95), whereas no significant association was observed among women. With regard to fibrinogen levels and breast cancer risk, there was a significant positive association among nulliparous women (HR: 2.53 [95% CI: 1.21, 5.30]), but not among parous women. Overall, our data suggest that enhanced platelet activation and a pro-coagulative state may not be related to increased risks of common cancers, although studies on other potential biomarkers of platelet activation and further cancer types are needed. Findings from our subgroup analyses require further investigation, as potential underlying mechanisms are not known.

In our previous investigations we have shown that platelets and their precursors express nicotinic α7 acetylcholine receptors (nAChRα7) that are involved in platelet function and in vitro differentiation of the megakaryoblastic cell line MEG-01. In this study, we were interested in the expression analysis of additional nAChR and the effects of nicotine in an ex vivo model using megakaryocytic cells differentiated from cord blood derived CD34(+) cells (CBMK) and an in vivo model using blood samples from smokers. CBMK were differentiated with thrombopoietin (TPO) for up to 17 days. Quantitative real-time PCR (QRT-PCR), Western blot analysis and flow cytometry were used to investigate nAChR expression (nAChRα7, nAChRα4, nAChRβ2) and nicotine effects. In blood samples of 15 nonsmokers and 16 smokers platelet parameters (count, mean platelet volume--MPV and platelet distribution width--PDW) were determined as indicators for changes of in vivo megakaryopoiesis. Platelet function was determined by the use of whole blood aggregometry and flow cytometry. The functional role of nAChR was evaluated using specific antagonists in aggregometry. CHRNA7, CHRNA4 and CHRNB2 gene transcripts and the corresponding proteins could be identified in CBMK during all stages of differentiation. Platelets contain nAChRα7 and nAChRβ2 but not nAChRα4. Nicotine had no effect on TPO-induced differentiation of CBMK. There was no significant difference in all platelet parameters of the smokers compared to the nonsmokers. In line with this, cholinergic gene transcripts as well as the encoded proteins were equally expressed in both the study groups. Despite our observation of nAChR expression in megakaryopoiesis and platelets, we were not able to detect effects of nicotine in our ex vivo and in vivo models. Thus, the functional role of the nAChR in these cells remains open.

White blood cell-associated antibodies can lead to transfusion-related acute lung injury (TRALI). Female donors with a history of pregnancies have been identified as a main cause for these antibodies. Male or female donors without a history of pregnancy are considered as safe donors.Following the identification of two TRALI cases associated with blood products from male donors, we investigated the frequency of granulocyte-specific and human leukocyte antigen (HLA) antibodies in the entire blood donor population using a high throughput automated flow-cytometry-based granulocyte immunofluorescence test (Flow-GIFT). We investigated sera from 14,343 whole blood donors (female, n = 6974, 48.7%; male, n = 7369, 51.3%) using automated Flow-GIFT. Of the female blood donors, 60.4% had a history of pregnancy. Positive sera were retested by the standard granulocyte immunofluorescence test and granulocyte agglutination test. For the detection of HLA Class I and II immunoglobulin G antibodies, we used a commercial screening enzyme-linked immunosorbent assay.We detected in 924 (21.9%) of the 4212 females with a history of pregnancy antibodies against granulocyte antigens (n = 62, 1.5%), HLA Class I and/or II antigens (n = 864, 20.5%). Notably, in 3.5% (n = 96) of 2762 females without a history of pregnancy and in 2.1% (n = 154) of 7369 males antibodies against granulocyte antigens (n = 13, 0.47% and n = 45, 0.6%), HLA Class I and/or II (n = 83, 3% and n = 109, 1.4%, respectively), were also detected.Human neutrophil antigen antibodies are rare in male and females without a history of pregnancy compared to females with a history of pregnancy, but their relevance is not negligible.

Alcohol-related pancreatitis is associated with a disproportionately large number of hospitalisations among GI disorders. Despite its clinical importance, genetic susceptibility to alcoholic chronic pancreatitis (CP) is poorly characterised. To identify risk genes for alcoholic CP and to evaluate their relevance in non-alcoholic CP, we performed a genome-wide association study and functional characterisation of a new pancreatitis locus.1959 European alcoholic CP patients and population-based controls from the KORA, LIFE and INCIPE studies (n=4708) as well as chronic alcoholics from the GESGA consortium (n=1332) were screened with Illumina technology. For replication, three European cohorts comprising 1650 patients with non-alcoholic CP and 6695 controls originating from the same countries were used.We replicated previously reported risk lociAn inversion in the

Background Enhanced platelet activation has been implicated in several pathophysiological processes. Here, we evaluated the biological reproducibility of circulating P-Selectin, Thrombomodulin (TM), Thrombopoietin (TPO), and Glycoprotein IIb/IIIa (GPIIb/IIIa) to assess whether these analytes can be used as reliable biomarkers of platelet activation in epidemiological studies. Methods We measured circulating P-Selectin, TM, TPO and GPIIb/IIIa by immunoassays in two blood samples of 78 participants of the EPIC Heidelberg study (47–80 years, 50% female) that were collected one year apart. Biological reproducibility of biomarker levels over time and associations with routine biochemistry parameters were assessed by Spearman's correlation coefficients. Results Statistical analyses revealed good reproducibility over one year for two of the analyzed markers, with Spearman coefficients of ρ = 0.80 (P-Selectin) and ρ = 0.73 (TPO) and reasonable reproducibility for TM (ρ = 0.63) and GPIIb/IIIa (ρ = 0.51). Levels of P-Selectin, TM, TPO and GPIIb/IIIa were not significantly associated with routine biochemistry parameters, such as glucose, HbA1c, LDL, HDL, Triglycerides and CRP. Conclusions Our findings suggest that a single assessment of P-Selectin, TM, TPO and GPIIb/IIIa at baseline in prospective epidemiological studies is appropriate to investigate associations between platelet activation and risks of chronic diseases.

Background/aim Previous genetic and morphologic characterisation of mucinous tubular and spindle cell carcinoma (MTSCC) have yielded controversial results. The aim of this study was to explain the phenotypic heterogeneity of MTSCC diagnosed by genetic means. Materials and methods We analyzed 7 MTSCC by array CGH and microsatellite allelotyping and by histology for morphological variation. We worked-up two entire kidneys with MTSCC to find microscopic alterations. Results We confirmed the diagnosis of MTSCC by detecting copy number changes at chromosomes 1, 4, 6, 8, 13, 14, 15, 18 and 22. We detected 13 small, microscopic precursor lesions in the two kidneys and found similar histological structures in precursor lesions and MTSCC. Conclusion MTSCC develops from embryonal rest-like lesions of impaired differentiation which may explain its morphological variations. Until diagnosis of a "malignant" MTSCC" is not confirmed by genetic means, it should not be called carcinoma.

Colorectal cancer (CRC) is one of the most common cancer worldwide. However, a large number of genetic risk factors involved in CRC have not been understood. Copy number variations (CNVs) might partly contribute to the 'missing heritability' of CRC. An increased overall burden of CNV has been identified in several complex diseases, whereas the association between the overall CNV burden and CRC risk is largely unknown. We performed a genome-wide investigation of CNVs on genomic DNA from 384 familial CRC cases and 1285 healthy controls by the Affymetrix 6.0 array. An increase of overall CNV burden was observed in familial CRC patients compared with healthy controls, especially for CNVs larger than 50kb (case/control ratio = 1.66, P = 0.025). In addition, we discovered for the first time a novel structural variation at 12p12.3 and determined the breakpoints by strategic PCR and sequencing. This 12p12.3 structural variation was found in four of 2862 CRC cases but not in 6243 healthy controls (P = 0.0098). RERGL gene (RERG/RAS-like), the only gene influenced by the 12p12.3 structural variation, sharing most of the conserved regions with its close family member RERG tumor suppressor gene (RAS-like, estrogen-regulated, growth inhibitor), might be a novel CRC-related gene. In conclusion, this is the first study to reveal the contribution of the overall burden of CNVs to familial CRC risk and identify a novel rare structural variation at 12p12.3 containing RERGL gene to be associated with CRC.

Aims Oxidative stress may contribute to the development of chronic pancreatitis (CP). The enzymes manganese superoxide dismutase 2 (MnSOD, SOD2) and catalase (CAT) counteract free radical activity within the mitochondria and the cytosol. Moreover, CAT activity contributes to the transformation of ethanol to acetaldehyde, a toxic intermediate product of ethanol metabolism, which has been associated with pancreatic damage. Common functional polymorphisms have been described in the MnSOD gene [rs4880, NM_000636.3:c.47 T > C, alanine (ALA) to valine (Val)] and in the CAT promoter region [rs1001179, NG_013339.1:g.4760 C > T]. We investigated whether these polymorphisms are associated with alcoholic CP. Methods We genotyped 470 patients with alcoholic CP for these MnSOD and CAT polymorphisms. We also analysed these variants in 357 healthy control subjects, and in an additional control group of 113 individuals with non-alcoholic CP. We used the age at onset of CP as marker of disease severity and investigated whether different genotypes are associated with different ages at onset. In patients with alcoholic CP, we investigated whether an interaction exists between smoking behaviour and genotypes by comparing genotype distributions in smokers and non-smokers. Results We did not observe significant differences of genotype frequencies between patient groups and controls. In patient groups, we did not find significant differences in the ages at onset between different genotypes. We did not observe an interaction between these polymorphisms. We did not find an association of these variants with smoking behaviour. Conclusions The investigated MnSOD and CAT polymorphisms do not predispose to the development of alcoholic CP. Short summary Patients with alcoholic pancreatitis and controls were genotyped for polymorphisms in oxidative stress genes. There were no significant differences of genotype frequencies between patients and controls, and no association with the age at onset of disease was observed. The polymorphisms are not associated with the development of alcoholic pancreatitis.

International audience; For maintaining energy homeostasis, creatine kinase (CK) is present at elevated levels in tissueswith high and/or fluctuating energy requirements such as muscle, brain, and epithelia, while there is very fewCK, if any, in peripheral blood cells. However, an ectopic expression of brain-type creatine kinase (BCK) has been reported for platelets and leukocytes in an autosomal dominant inherited anomaly named CKBE. Here we investigated CK inerythrocytes of CKBE individuals from eight unrelated families. The data revealed a varying but significant increase of CK activity in CKBE individuals as compared to controls, reaching an almost 800-fold increase in two CKBE individuals which also had increased erythrocyte creatine. Immunoblotting with highly specific antibodies confirmed that the expressed CK isoform is BCK. Cell fractionation evidenced soluble BCK, suggesting cytosolic and not membrane localization of erythrocyte CK as reported earlier. These results are discussed in the context of putative CK energy buffering and transfer functions in red blood cells.

We performed a multistage genome-wide association study including 7,683 individuals with pancreatic cancer and 14,397 controls of European descent. Four new loci reached genome-wide significance: rs6971499 at 7q32.3 (LINC-PINT, per-allele odds ratio (OR) = 0.79, 95% confidence interval (CI) 0.74-0.84, P = 3.0 x 10(-12)), rs7190458 at 16q23.1 (BCAR1/CTRB1/CTRB2, OR = 1.46, 95% CI 1.30-1.65, P = 1.1 x 10(-10)), rs9581943 at 13q12.2 (PDX1, OR = 1.15, 95% CI 1.10-1.20, P = 2.4 x 10(-9)) and rs16986825 at 22q12.1 (ZNRF3, OR = 1.18, 95% CI 1.12-1.25, P = 1.2 x 10(-8)). We identified an independent signal in exon 2 of TERT at the established region 5p15.33 (rs2736098, OR = 0.80, 95% CI 0.76-0.85, P = 9.8 x 10(-14)). We also identified a locus at 8q24.21 (rs1561927, P = 1.3 x 10(-7)) that approached genome-wide significance located 455 kb telomeric of PVT1. Our study identified multiple new susceptibility alleles for pancreatic cancer that are worthy of follow-up studies.

Breast cancer (BC) is the leading cancer in women worldwide. Changes in DNA methylation in peripheral blood could be associated with malignant diseases. Making use of screening results by llumina 27K Methylation Assay, we validated demethylation of five CpG sites of S100P gene in blood cell DNA of BC patients by three independent retrospective studies with subjects from different centers (Validation I: 235 familial BC case and 206 controls, odds ratio per -1% methylation1.03, and P6.00 × 10-8 for all five CpG sites; Validation II: 189 sporadic BC case and 189 controls, odds ratio per -1% methylation1.03, P8.0 × 10-5 for four CpG sites; Validation III: 156 sporadic BC case and 151 controls, odds ratio per -1% methylation1.03, P6.0 × 10-4 for four CpG sites). In addition, the blood-based S100P methylation pattern was similar among BC patients with differential clinical characteristics regardless of stage, receptor status and menopause status. The observed BC-associated decreased S100P methylation in blood mainly originates from the leucocytes subpopulations but not B cells. The methylation levels of most S100P CpG sites were inversely correlated with the expression of S100P in leucocytes (P1.2 × 10-4) and in tissue (P1.1 × 10-4). This study reveals significant association between blood-based decreased S100P methylation and BC, and provides another proof for the application of altered DNA methylation signatures from blood cells as potential markers for the detection of BC, especially for the early stage.

Pancreatic neuroendocrine tumors (PNETs) are heterogeneous neoplasms which represent only 2% of all pancreatic neoplasms by incidence, but 10% by prevalence. Genetic risk factors could have an important role in the disease aetiology, however only a small number of case control studies have been performed yet. To further our knowledge, we genotyped 13 SNPs belonging to the pleiotropic CDKN2A/B gene region in 320 PNET cases and 4436 controls, the largest study on the disease so far. We observed a statistically significant association between the homozygotes for the minor allele of the rs2518719 SNP and an increased risk of developing PNET (ORhom = 2.08, 95% CI 1.05–4.11, p = 0.035). This SNP is in linkage disequilibrium with another polymorphic variant associated with increased risk of several cancer types. In silico analysis suggested that the SNP could alter the sequence recognized by the Neuron-Restrictive Silencer Factor (NRSF), whose deregulation has been associated with the development of several tumors. The mechanistic link between the allele and the disease has not been completely clarified yet but the epidemiologic evidences that link the DNA region to increased cancer risk are convincing. In conclusion, our results suggest rs2518719 as a pleiotropic CDKN2A variant associated with the risk of developing PNETs.

Introduction A single nucleotide polymorphism of the factor VII activating protease (FSAP), FSAP Marburg I (rs7080536) has been identified as a risk factor for venous thrombosis, but its clinical role has so far been controversial in part due to small cohort sizes. The aim of the present case-control study was to elucidate the impact of the FSAP Marburg I polymorphism (FSAP-MI) on the development of venous thromboembolic disease (VTE) with other known sequence variations, including Factor V Leiden (rs6025) and Factor II G20210A (rs1799963). Materials and Methods The study included 891 patients (312 male and 579 female) with a history of deep venous thrombosis (DVT) and/or pulmonary embolism (PE) and 1283 healthy blood donors with no history of venous thromboembolic disease. Results We found that besides to the well-established aforementioned sequence variations of FV and Prothrombin, the FSAP Marburg I (FSAP-MI) polymorphism was significantly associated with the development of DVTs (1.65 (1.16-2.34) OR (95% CI)) and recurrent thromboembolic events (DVT and PE) (2.13 (1.35-3.36) OR (95% CI)). Comparing patients displaying one or more events FSAP-MI was still associated with the development of recurrent thromboembolic events (1.64 (1- 2.69) OR (95% CI)). Conclusions We conclude that FSAP Marburg-I genotyping may be used to determine the risk for thromboembolic disorders in patients with suspected thrombophilia and known DVT or PE.

In this study, we analyzed a "variant of uncertain significance" (VUS) located in exon 23 of the BRCA2 gene exhibited by six members of five distinct families with hereditary breast cancer (BC). The variant was identified by DNA sequencing, and cDNA analysis revealed its co-expression with wild-type mRNA. We analyzed co-occurrence with other pathological mutations in BRCA1/2, performed a case-control study, looked for evolutionary data and used in-silico analyses to predict its potential clinical significance. Sequencing revealed an in frame deletion of 126 nucleotides in exon 23, leading to a deletion of 42 amino acids (c.9203_9328del126, p.Pro2992_Thr3033del). All of the VUS-carriers suffered from either BC or ovarian/pancreatic cancer. No other definite pathologic mutation of BRCA genes was found in the five families. The identified deletion could not be observed in a control cohort of 2,652 healthy individuals, but in 5 out of 916 (0.5%) tested BC families without a bona fide pathogenic BRCA1/2 mutation (P = 0.0011). According to these results, the in frame deletion c.9203_9328del126 is a rare mutation strongly associated with familial BC. In summary, our investigations indicate that this BRCA2 deletion is pathogenic.

The ectopic expression in peripheral blood cells of the brain-type creatine kinase (CKB) is an autosomal dominant inherited anomaly named CKBE (MIM ID 123270). Here, we characterized the CK activity in serum, platelets (PLT) and leukocytes (WBC) of 22 probands (from 8 unrelated families) and 10 controls. CK activity was measured by standard UV-photometry. Expression of the CKB gene was analyzed by real-time PCR and Western blotting. DNA sequencing including bisulfite treatment was used for molecular analysis of the CKB gene. Serum CK levels were comparable between probands and controls. CKBE probands revealed significantly higher CK activity in PLT (3.7 +/- 2.7 versus 179.2 +/- 83.0 U/10(12) PLT; p

The P-Selectin Gene Polymorphism Val168Met. Aims: Ventricular fibrillation (VF) in the setting of acute myocardial infarction (AMI) is the leading cause of sudden cardiac death (SCD). Family history of SCD is described as risk factor for primary VF during acute AMI. Genetic factors may be associated with primary VF. We examined polymorphisms in genes related to the activation and adhesion of blood platelets in patients with and without VF in the setting of AMI and among healthy controls. Methods: Two hundred and forty patients with a history of AMI and 475 healthy controls were studied. Seventy-three patients (30%) had primary VF during AMI. By using PCR techniques with sequence-specific primers (PCR-SSP), we genotyped 5 single nucleotide polymorphisms (SNPs) in P-selectin (SELP) (V168M, S290N, N592D, V599L, T715P), 2 SNPs (M62I, S273F) in P-selectin glycoprotein ligand-1 (SELPLG), 5 SNPs in CD40LG (−3459A>G, −122A>C, −123A>C, 148T>C, intr4–13T>C), the H558R SNP in SCN5A, and rs2106261 in ZFHX3. In addition, length polymorphisms in SELPLG (36bp-tandem repeat) and CD40LG (CA-repeat) were genotyped by PCR methods. Results were evaluated by 2-sided t-tests, chi-square tests, and logistic regression analysis. Results: None of the gene polymorphisms showed significant differences between AMI patients and healthy controls. Among patients with a history of VF, however, the SELP 168M variant showed a significantly higher prevalence (14/73 patients; 19.2%) as compared with patients without VF (13/167 patients; 7.8%; P < 0.01). This association remained significant in a logistic regression analysis after adjustment for age and gender (P = 0.013; odds ratio 2.8; 95% confidence interval 1.2–6.3). Conclusions: This is the first description of an association of the SELP gene variant 168M with primary VF during acute MI. This variant may be a candidate polymorphism for evaluating the susceptibility for VF in the setting of acute MI. (J Cardiovasc Electrophysiol, Vol. 21, pp. 1260-1265, November 2010)

MicroRNAs (miRNAs) regulate pathways involved in cell differentiation, proliferation, development, and apoptosis by degradation of target mRNAs and/or repression of their translation. Although the single nucleotide polymorphisms (SNPs) in miRNAs target sites have been studied, the effects of SNPs in miRNAs are largely unknown. In our study, we first systematically sequenced miRNA genes reported to be involved in breast cancer to identify/verify SNPs. We analyzed four SNPs, one located in the pre-miRNA and the other three located in miRNA flanking regions, for a putative association with breast cancer risk. The SNP rs895819, located in the terminal loop of pre-miRNA-27a, showed a protective effect. In a large familial breast cancer study cohort, the rare [G] allele of rs895819 was found to be less frequent in the cases than in the controls, indicating a reduced familial breast cancer risk ([G] vs. [A]: OR = 0.88, 95% CI 0.78–0.99, P = 0.0287). Furthermore, age stratification revealed that the protective effect was mainly observed in the age group < 50 years of age ([G] vs. [A]: OR = 0.83, 95% CI 0.70–0.98, P = 0.0314), whereas no significant effect was observed in the age group ≥ 50 years of age, indicating a possible hormone-related effect. It has been shown that artificial mutations in the terminal loop of miR-27a can block the maturation process of the miRNA. We hypothesize that the G-variant of rs895819 might impair the maturation of the oncogenic miR-27a and thus, is associated with familial breast cancer risk.

Aspirin-like defect (ALD) is a rare, mostly autosomal dominant inherited dysfunction of the intraplatelet arachidonic acid (AA) pathway leading to impaired thromboxane A2 signalling. We aimed to establish diagnostic criteria for ALD diagnosis and present clinical and laboratory phenotypes of 52 individuals from 17 unrelated families. Platelet in vitro function was determined on the basis of platelet aggregation response (PAR) to AA, adenosine diphosphate, collagen and ristocetin as well as PFA-100 closure times (CT). Using impaired PAR to AA (< or =10%) as the mandatory diagnostic criterion, ALD could be confirmed in 17 patients. Subsequently, family members were investigated and among 35 individuals an additional 13 ALD patients as well as 4 individuals with mild ALD (PAR to AA: 19-32%) were identified. At least one bleeding symptom was reported by 25 (74%) ALD patients and prolonged CT was detected in 24 (71%) of the cases, both significantly correlated with impaired PAR to AA (P = 0.001 and P = 0.002, respectively). An estimated 0.6% prevalence was determined for ALD in our paediatric patients with suspected coagulation disorders. Due to the mild bleeding symptoms, ALD is probably underdiagnosed. If ALD is suspected, PAR to AA is suitable for the identification of individuals at risk of increased haemorrhage.

MicroRNAs (miRNAs) negatively regulate expression of target transcripts by hybridization to complementary sites of their messenger RNA targets. Chen et al. have described several putative functional single nucleotide polymorphisms (SNPs) in miRNA target sites. Here, we selected 11 miRNA target site SNPs located in 3' untranslated regions of genes involved in cancer and breast cancer to analyze their impact on breast cancer risk using a large familial study population. Whereas no association was observed for 10 SNPs, a significant association was revealed for the variant affecting a miRNA target site in the estrogen receptor (ESR) 1. Age stratification showed that the association was stronger in premenopausal women [C versus T: odds ratio (OR) = 0.60, confidence interval (CI) = 0.41-0.89, P = 0.010]. Furthermore, the effect was stronger in high-risk familial cases (C versus T: OR = 0.42, CI = 0.25-0.71, P = 0.0009). Clinical studies have shown that elimination of ESR1 significantly reduces breast cancer risk. Thus, therapies that inhibit ESR1 are used for breast cancer treatment. According to in silico analysis, ESR1_rs2747648 affects the binding capacity of miR-453, which is stronger when the C allele is present. In contrast, the T allele attenuates the binding of miR-453, which might lead to a reduced miRNA-mediated ESR1 repression, in consequence higher ESR1 protein levels and an increased breast cancer risk. Thus, the breast cancer protective effect observed for the C allele in premenopausal women is biologically reasonable. The analysis of large study populations in multicentre collaboration will be needed to verify the association and answer questions regarding the possible impact of this variant on therapeutic and clinical outcome.

Data from several studies have suggested that polymorphisms in A-kinase anchoring proteins (AKAPs), which are key components of signal transduction, contribute to carcinogenesis. To evaluate the impact of AKAP variants on breast cancer risk, we genotyped six nonsynonymous single-nucleotide polymorphisms that were predicted to be deleterious and found two (M463I, 1389G>T and N2792S, 8375A>G) to be associated with an allele dose-dependent increase in risk of familial breast cancer in a German population. We extended the analysis of AKAP9 M463I, which is in strong linkage disequilibrium with AKAP9 N2792S, to 9523 breast cancer patients and 13770 healthy control subjects from seven independent European and Australian breast cancer studies. All statistical tests were two-sided. The collaborative analysis confirmed the association of M463I with increased breast cancer risk. Among all breast cancer patients, the combined adjusted odds ratio (OR) of breast cancer for individuals homozygous for the rare allele TT (frequency = 0.19) compared with GG homozygotes was 1.17 (95% confidence interval [CI] = 1.08 to 1.27, P = .0003), and the OR for TT homozygotes plus GT heterozygotes compared with GG homozygotes was 1.10 (95% CI = 1.04 to 1.17, P = .001). Among the combined subset of 2795 familial breast cancer patients, the respective ORs were 1.27 (95% CI = 1.12 to 1.45, P = .0003) and 1.16 (95% CI = 1.06 to 1.27, P = .001).

Copy number variants (CNVs), insertions, deletions and duplications, contribute considerably to human genetic variation and disease development. A recent study has characterized 100 CNVs including a deletion in the mitochondrial tumor suppressor gene 1 (MTUS1) lacking the coding exon 4. MTUS1 maps to chromosome 8p, a region frequently deleted and associated with disease progression in human cancers, including breast cancer (BC). To investigate the effect of the MTUS1 CNV on familial BC risk, we analyzed 593 BC patients and 732 control individuals using a case-control study design. We found a significant association of the deletion variant with a decreased risk for both familial and high-risk familial BC (odds ratio (OR) = 0.58, 95% confidence interval (CI) = 0.37-0.90, P = 0.01 and OR = 0.41, 95% CI = 0.23-0.74, P = 0.003), supporting its role in human cancer. To our knowledge, the present study is the first to determine the impact of a CNV in a tumor suppressor gene on cancer risk.

Individuals with lupus anticoagulants (LA) are at risk for thromboembolism (TE). Chronic inflammation is an important characteristic in LA patients which may dispose for TE. Platelets play a key role in inflammation and TE. We therefore investigated gene polymorphisms as well as plasma levels of platelet receptors as predictors of TE in 107 LA patients. We compared 74 patients with a history of thromboembolic disease (TE+), 56 with venous thrombosis (VT), 12 with arterial thrombosis (AT), and six patients who had both, with 33 LA patients without previous thrombosis (TE-). The P-selectin Pro715 allele was slightly more frequent in VT (OR = 3.167,95% CI 0.955-10.503; p = 0.0594), but no patient with AT had this allele (OR = 0.099, 95 % CI 0.001-0.790; p = 0.0238) which therefore may protect from AT. Plasma levels of P-selectin, collected a median of 35 months (range 2-329 months) after the last thrombotic event, were higher in patients withVT (p = 0.0096) than in TE-, but not with AT (p = 0.4713). These high P-selectin levels were not explained by the P-selectin polymorphism. The CA repeat polymorphism in the 3'-noncoding region of CD154 was significantly associated with the development of AT (OR = 4.035,95 % CI 1.329-12.249; p = 0.0138). Plasma levels of CD154 were not significantly different among the subgroups. Thus, the Thr715Pro polymorphism of P-selectin and CA repeats of CD154 are differentiating between the risk for VT and AT. Further, soluble P-selectin is elevated in LA patients with previous VT, but its role to predict VT needs to be evaluated in prospective studies.

Background: A recent association study identified a common variant (rs9790517) at 4q24 to be associated with breast cancer risk. Independent association signals and potential functional variants in this locus have not been explored. Methods: We conducted a fine-mapping analysis in 55,540 breast cancer cases and 51,168 controls from the Breast Cancer Association Consortium. Results: Conditional analyses identified two independent association signals among women of European ancestry, represented by rs9790517 [conditional P = 2.51 × 10−4; OR, 1.04; 95% confidence interval (CI), 1.02–1.07] and rs77928427 (P = 1.86 × 10−4; OR, 1.04; 95% CI, 1.02–1.07). Functional annotation using data from the Encyclopedia of DNA Elements (ENCODE) project revealed two putative functional variants, rs62331150 and rs73838678 in linkage disequilibrium (LD) with rs9790517 (r2 ≥ 0.90) residing in the active promoter or enhancer, respectively, of the nearest gene, TET2. Both variants are located in DNase I hypersensitivity and transcription factor–binding sites. Using data from both The Cancer Genome Atlas (TCGA) and Molecular Taxonomy of Breast Cancer International Consortium (METABRIC), we showed that rs62331150 was associated with level of expression of TET2 in breast normal and tumor tissue. Conclusion: Our study identified two independent association signals at 4q24 in relation to breast cancer risk and suggested that observed association in this locus may be mediated through the regulation of TET2. Impact: Fine-mapping study with large sample size warranted for identification of independent loci for breast cancer risk. Cancer Epidemiol Biomarkers Prev; 24(11); 1680–91. ©2015 AACR.

Infectious hepatitis C virus (HCV) particles bind to host lipoproteins such as low-density lipoproteins (LDLs). Low-density lipoprotein receptors (LDLR) have been termed candidate receptors for HCV-LDL complexes. Functional host genetic single nucleotide polymorphisms (SNPs) in the apolipoprotein E (APOE) gene encoding apolipoprotein E (apoE) - a major structural LDL component and natural ligand of LDLR - likely influence the course of HCV infection. We investigated the prevalence of APOE SNPs in two large and independent cohorts of patients with chronic HCV infection compared to respective controls.We genotyped 996 chronically HCV-infected patients; 179 patients with spontaneous HCV clearance; 283 individuals with non-HCV-associated liver disease; and 2 234 healthy controls.APOE genotype proportions in patients with persistent HCV infection significantly differed from healthy controls (P = 0.007) primarily because of a substantial under-representation of APOE4 alleles in chronically HCV-infected patients (10.2%) compared to 13.0% in healthy controls (P = 0.001). The distribution of APOE4 allele positive genotypes (?2?4, ?3?4, ?4?4) also significantly differed between chronically HCV-infected patients and healthy controls (1.4%, 17%, 1% vs. 2.4%, 20.5%, 1.7%; P = 0.001), suggesting a protective effect of the APOE4 allele in HCV infection. This was confirmed by a significant over-representation of the APOE4 allele in patients with spontaneous HCV clearance (17.6%; P = 0.00008). The APOE4 allele distribution in patients with non-HCV-associated liver disease (14.0%) was very similar to healthy controls and also differed from chronically HCV-infected patients (P = 0.012), suggesting HCV specificity.Our findings suggest that the APOE4 allele may confer a protective effect in the course of HCV infection.

Carboxyl ester lipase is a digestive pancreatic enzyme encoded by the CEL gene. Mutations in CEL cause maturity-onset diabetes of the young as well as pancreatic exocrine dysfunction. Here we describe a hybrid allele (CEL-HYB) originating from a crossover between CEL and its neighboring pseudogene, CELP. In a discovery series of familial chronic pancreatitis cases, we observed CEL-HYB in 14.1% (10/71) of cases compared to 1.0% (5/478) of controls (odds ratio (OR) = 15.5; 95% confidence interval (CI) = 5.1-46.9; P = 1.3 × 10 -6 by two-tailed Fisher's exact test). In three replication studies of nonalcoholic chronic pancreatitis, we identified CEL-HYB in a total of 3.7% (42/1,122) cases and 0.7% (30/4,152) controls (OR = 5.2; 95% CI = 3.2-8.5; P = 1.2 × 10 -11; formal meta-analysis). The allele was also enriched in alcoholic chronic pancreatitis. Expression of CEL-HYB in cellular models showed reduced lipolytic activity, impaired secretion, prominent intracellular accumulation and induced autophagy. These findings implicate a new pathway distinct from the protease-antiprotease system of pancreatic acinar cells in chronic pancreatitis., SCOPUS: ar.j, info:eu-repo/semantics/published

The chromosomal passenger complex (CPC) plays a pivotal role in the regulation of cell division. Therefore, inherited CPC variability could influence tumor development. The present candidate gene approach investigates the relationship between single nucleotide polymorphisms (SNPs) in genes encoding key CPC components and breast cancer risk. Fifteen SNPs in four CPC genes (INCENP, AURKB, BIRC5 and CDCA8) were genotyped in 88 911 European women from 39 case-control studies of the Breast Cancer Association Consortium. Possible associations were investigated in fixedeffects meta-analyses. The synonymous SNP rs1675126 in exon 7 of INCENP was associated with overall breast cancer risk per A allele odds ratio (OR) 0.95, 95% confidence interval (CI) 0.92-0.98, P = 0.007 and particularly with estrogen receptor (ER)-negative breast tumors (per A allele OR 0.89, 95% CI 0.83-0.95, P = 0.0005). SNPs not directly genotyped were imputed based on 1000 Genomes. The SNPs rs1047739 in the 3' untranslated region and rs144045115 downstream of INCENP showed the strongest association signals for overall (per T allele OR 1.03, 95% CI 1.00-1.06, P = 0.0009) and ER-negative breast cancer risk (per A allele OR 1.06, 95% CI 1.02-1.10, P = 0.0002). Two genotyped SNPs in BIRC5 were associated with familial breast cancer risk (top SNP rs2071214: per G allele OR 1.12, 95% CI 1.04-1.21, P = 0.002). The data suggest that INCENP in the CPC pathway contributes to ER-negative breast cancer susceptibility in the European population. In spite of a modest contribution of CPC-inherited variants to the total burden of sporadic and familial breast cancer, their potential as novel targets for breast cancer treatment should be further investigated. © The Author 2015.

The nuclear receptor coactivator 3 (NCOA3, also known as AIB1) is a coactivator of nuclear receptors like the estrogen receptor. NCOA3 is overexpressed in ∼60% of primary human breast tumors, and high levels of NCOA3 expression are associated with tamoxifen resistance and worse survival rate. In contrast, NCOA3 deficiency suppresses v-Ha-ras–induced breast cancer initiation and progression in mice. Here, we analyzed the influence of NCOA3 coding single nucleotide polymorphisms on breast cancer risk by performing a case-control study using a German and a Polish study population and identified an association between NCOA3 polymorphisms and breast cancer. A joint analysis of the German and the Polish study population revealed a significant protective effect for the 1758G>C (Q586H) and 2880A>G (T960T) variants. In addition, haplotype analysis showed a protective effect of the 1758C-2880A and 1758G-2880G haplotypes (odds ratio 0.79; 95% confidence interval, 0.67-0.93; P = 0.004). Because of the impact of NCOA3 in antiestrogen therapy resistance, these polymorphisms might also influence therapy outcome in breast cancer.

c-MYC is a multifaceted protein that regulates cell proliferation, differentiation and apoptosis. Its crucial role in diverse cancers has been demonstrated in several studies. Here, we analysed the influence of the rare c-MYC Asn11Ser polymorphism on familial breast cancer risk by performing a case-control study with a Polish (cases n = 349; controls n = 441) and a German (cases n = 356; controls n = 655) study population. All cases have been tested negative for mutations in the BRCA1 and BRCA2 genes. A joint analysis of the Polish and the German study population revealed a 54% increased risk for breast cancer associated with the heterozygous Asn11Ser variant (OR = 1.54, 95% CI 1.05–2.26, p = 0.028). The breast cancer risk associated with this genotype increases above the age of 50 years (OR = 2.24, 95% CI 1.20–4.21, p = 0.012). The wild-type amino acid Asn of this polymorphism is located in the N-terminal MYC transactivation domain and is highly conserved not only among most diverse species but also in the N-MYC homologue. Due to the pivotal role of c-MYC in diverse tumours, this variant might affect the genetic susceptibility of other cancers as well. © 2005 Wiley-Liss, Inc.