[show abstract][hide abstract] ABSTRACT: The International Tamoxifen Pharmacogenomics Consortium (ITPC) was established to address the controversy over CYP2D6 status and clinical outcomes in tamoxifen therapy. We performed a meta-analysis on data from 4,973 tamoxifen treated patients (twelve globally-distributed sites). Using strict eligibility requirements (postmenopausal women with estrogen receptor (ER) positive breast cancer receiving 20 mg/day tamoxifen for 5 years, Criterion 1), CYP2D6 poor metabolizer status was associated with poorer Invasive Disease-Free Survival (IDFS: HR=1.25; 95% CI 1.06, 1.47; P=0.009). However, CYP2D6 was not statistically significant when tamoxifen duration, menopausal status, and annual follow-up were not specified (Criterion 2, n=2443; 49%; P=0.25) nor when no exclusions were applied (Criterion 3, n=4935; 99%; P=0.38). Although CYP2D6 is a strong predictor of IDFS using strict inclusion criteria, because the results are not robust to inclusion criteria (these were not defined a priori), prospective studies are necessary to fully establish the value of CYP2D6 genotyping in tamoxifen therapy.Clinical Pharmacology & Therapeutics (2013); accepted article preview online 23 September 2013 doi:10.1038/clpt.2013.186.
[show abstract][hide abstract] ABSTRACT: Aim: Because 5-fluorodeoxyuridine monophosphate (5-FdUMP), an anabolic active metabolite of 5-fluorouracil (5-FU), is a substrate of MRP8 (encoded by ABCC11), we investigated whether ABCC11 polymorphisms play a role in severe toxicity of 5-FU. Patients & methods: Genomic DNA from 672 cancer patients treated with 5-FU monotherapy and with documented toxicity according to WHO criteria was genotyped for 12 ABCC11 tag SNPs. Functional impact of polymorphisms was assessed in a Caucasian human liver cohort (n = 150) and by recombinant expression of MRP8 protein variants. Results: Univariate and multivariate analysis identified rs17822471 (G>A, T546M) as risk factor of severe leukopenia (p = 0.021, odds ratio [95%CI]: 3.31 [1.26-8.66]) but not of other toxicity types. MRP8 protein expression in human liver was 1.7-fold lower in carriers compared with wild-type (p = 0.02). Recombinant expression confirmed the effect of T546M on protein expression. Conclusion: Since MRP8 is expressed in bone marrow blasts and leukocytes, lower expression may lead to intracellular accumulation of 5-FdUMP and increased risk of leukopenia. Original submitted 25 April 2013; Revision submitted 17 July 2013.
[show abstract][hide abstract] ABSTRACT: Adjuvant tamoxifen is a valid treatment option for women with oestrogen receptor (ER)-positive breast cancer. However, up to 40% of patients experience distant or local recurrence or die. MicroRNAs have been suggested to be important prognosticators in breast cancer. This study aims to identify microRNAs with the potential to predict tamoxifen response.
We performed a global microRNA screen (1105 human microRNAs) in primary tumours of six matched pairs of postmenopausal, ER-positive breast cancer patients treated with tamoxifen, who were either recurrence free or had developed a recurrence (median follow up: 8.84years; range: 1.28-12.7years). Patients of this discovery set and the 81 patients of the validation set (median follow up: 8.64years; range: 0.21-19.85years) were treated at the Robert Bosch Hospital, Stuttgart, Germany, between 1986 and 2005.
Out of the top 20 deregulated microRNAs (12 up-regulated, eight down-regulated) miR-126 (Hazard Ratio (HR)=0.56, 95% confidence interval (CI): 0.38-0.83; Holm-adj. P=0.022) and miR-10a (HR=0.53, 95% CI: 0.33-0.85; Holm-adj. P=0.031) were identified as significant predictors of tamoxifen outcome by multivariate Cox regression analysis in the independent validation set of 81 postmenopausal, ER-positive patients. Kaplan-Meier survival analyses based on cut-offs determined by receiver operating characteristics curves confirmed that a higher expression of miR-126 and miR-10a in the patients tumour was associated with longer relapse-free time (log-rank P=0.037, P<0.0001, respectively).
Our data suggest that miR-126 and miR-10a are independent predictors for tumour relapse in early postmenopausal breast cancer patients treated with adjuvant tamoxifen.
European journal of cancer (Oxford, England: 1990) 08/2013; · 4.12 Impact Factor
[show abstract][hide abstract] ABSTRACT: The human olfactory bulb displays high morphologic dynamics changing its volume with olfactory function, which has been explained by active neurogenetic processes. Discussion continues whether the human olfactory bulb hosts a continuous turnover of neurons. We analyzed the transcriptome via RNA quantification of adult human olfactory bulbs and intersected the set of expressed transcriptomic genes with independently available proteomic expression data. To obtain a functional genomic perspective, this intersection was analyzed for higher-level organization of gene products into biological pathways established in the gene ontology database. We report that a fifth of genes expressed in adult human olfactory bulbs serve functions of nervous system or neuron development, half of them functionally converging to axonogenesis but no other non-neurogenetic biological processes. Other genes were expectedly involved in signal transmission and response to chemical stimuli. This provides a novel, functional genomics perspective supporting the existence of neurogenesis in the adult human olfactory bulb.
Brain Structure and Function 08/2013; · 7.84 Impact Factor
[show abstract][hide abstract] ABSTRACT: The monocarboxylate transporter 4 (MCT4) is a metabolic target in tumor biology since it mediates lactate transport across membranes resulting in anti-apoptotic effects. Cell experiments support the importance of MCT4 in clear cell renal cell carcinoma (ccRCC). In this study, we assessed the prognostic potential of MCT4 expression in ccRCC and its epigenetic regulation by DNA methylation as novel predictive marker for patient outcome using independent ccRCC-cohorts.
MCT4 protein expression was quantified in 207 ccRCC and corresponding non-tumor tissues. Data of an independent ccRCC-cohort from The Cancer Genome Atlas (TCGA) were analysed on MCT4 mRNA (n=482) and DNA methylation (n=283) level. The findings on MCT4 expression and DNA methylation in the SLC16A3 promoter were validated in a third cohort (n=64). Promoter activity assays were performed in four RCC cell lines.
MCT4 protein expression was upregulated (P<0.0001) in ccRCC and showed significant association with cancer-related death. Upregulation of MCT4 mRNA expression (P<0.00001) was confirmed in the TCGA-cohort. Single CpG-sites correlated inversely with mRNA expression and were associated with overall survival in Kaplan-Meier analyses (HR=0.39; 95%CI=0.24-0.64;P[log-rank]=1.23e(-04)). Promoter activity studies confirmed MCT4 regulation by DNA methylation. The significant correlation between MCT4 protein and gene expression or DNA methylation at single CpG-sites was validated in a third cohort. Again, higher methylation at individual CpG-sites was associated with prolonged survival (HR=0.05; 95%CI=0.01-0.40;P[log-rank]=6.91e(-05)).
We identified SLC16A3 promoter DNA methylation as a novel epigenetic mechanism for MCT4 regulation in ccRCC with first evidence of a biological rationale for prognosis and clinical outcome.
Clinical Cancer Research 07/2013; · 7.84 Impact Factor
[show abstract][hide abstract] ABSTRACT: Genes that are important for the detoxification of drugs and other xenobiotics show a high degree of interindividual variation due to regulation by diverse genetic, non-genetic, and epigenetic mechanisms including microRNAs. We selected a set of 56 microRNAs predicted to target the 3'-UTR of ADME (absorption, distribution, metabolism, excretion) genes to assess their hepatic expression levels and interindividual variability in a well-documented human liver tissue cohort (n=92), together with the well-known hepatic microRNAs miR-122, miR-21, miR-27b and miR-148a. Quantification by stem-loop real-time RT-PCR confirmed high expression for these microRNAs and revealed particularly strong variability of expression (>1000-fold) for miR-539, miR-200c, miR-31, miR-15a, and miR-22. Association analysis revealed a high degree of correlation among various miRNAs, suggesting coregulation. Statistical analysis considering liver donor meta-data including correction for multiple testing revealed strongly elevated levels of miR-21, miR-34a, miR-130b and miR-132 in cholestatic liver and of miR-21 and miR-130b during inflammation, as indicated by elevated C-reactive protein levels in serum. While none of the miRNAs was strongly associated with sex, several miRNAs including miR-34a and miR-200a/b were positively correlated with age. Association analysis with ADME gene expression profiles and with cytochrome P450 (CYP) gene expression phenotypes (mRNA, protein, enzymatic activity) revealed numerous significant correlations. In particular, negatively affected protein and/or activity levels were observed for CYP1A1/1A2 (miR-142-3p and miR-200a), CYP2C19 (miR-34a), CYP2D6 (let-7b) and CYP2E1 (miR-10a, let-7g). These data should be useful to further elucidate regulatory functions of microRNAs in liver pathophysiology and regulation of ADME gene expression.
Drug metabolism and disposition: the biological fate of chemicals 06/2013; · 3.74 Impact Factor
[show abstract][hide abstract] ABSTRACT: 17β-hydroxysteroid dehydrogenase type 1 (HSD17B1) plays an important role in the biosynthesis of 17β-estradiol. The current study aimed at confirming the reduced risk of breast cancer in carriers of the non-synonymous HSD17B1_937_A>G (rs605059) polymorphism who used any hormone replacement therapy (HRT) for 10 years or longer. We performed an independent association study using four breast cancer case-control studies from Australia, Germany, and Sweden. In all, 5,777 cases and 8,189 age-matched controls of European descent were genotyped by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and TaqMan. Risk estimates were calculated by interaction analysis and main effect analysis adjusted for age and study. Main effect analyses for women using any HRT for 10 years or longer (1,428 cases versus 1,724 controls) revealed a protective effect of the HSD17B1_937_G allele on breast cancer risk (OR 0.86, 95 % CI: 0.73-0.99; p = 0.048). Thus, our previous finding of a protective effect of the HSD17B1_937_G allele on HRT-associated breast cancer risk has now been confirmed both in independent large patient cohorts and a comprehensive pooled analysis supporting the hypothesis that a HSD17B1-mediated decreased conversion of estrone to the more potent 17β-estradiol may reduce the estrogenic effects, thereby reducing the risk of developing breast cancer during long-term HRT use.
Breast Cancer Research and Treatment 02/2013; · 4.47 Impact Factor
[show abstract][hide abstract] ABSTRACT: BACKGROUND: Organic anion transporting polypeptide (OATP) 1B1, OATP1B3, and OATP2B1 (encoded by SLCO1B1, SLCO1B3, SLCO2B1) mediate the hepatic uptake of endogenous compounds like bile acids and of drugs, e.g. the lipid-lowering atorvastatin, thereby influencing hepatobiliary elimination. Here we systematically elucidated the contribution of SLCO variants on expression of the three hepatic OATPs under consideration of additional important covariates. METHODS: Expression was quantified by RT-PCR and immunoblotting in 143 Caucasian liver samples. A total of 109 rare and common variants in the SLCO1B3-SLCO1B1 genomic region and the SLCO2B1 gene were genotyped by MALDI-TOF mass spectrometry and genome-wide SNP microarray technology. SLCO1B1 haplotypes affecting hepatic OATP1B1 expression were associated with pharmacokinetic data of the OATP1B1 substrate atorvastatin (n=82). RESULTS: Expression of OATP1B1, OATP1B3, and OATP2B1 on mRNA and protein level showed marked interindividual variability. All three OATPs were expressed in a coordinated fashion. By a multivariate regression analysis adjusted for non-genetic and transcription covariates increased OATP1B1 expression was associated with the coding SLCO1B1 variant c.388A>G (rs2306283) even after correction for multiple testing (P=0.00034). This held true for haplotypes harboring c.388A>G but not the functional variant c.521T>C (rs4149056) associated with statin-related myopathy. c.388A>G also significantly affected atorvastatin pharmacokinetics. SLCO variants, non-genetic, and regulatory covariates, accounted for 59% of variability of OATP1B1 expression. CONCLUSIONS: Our results showed that expression of OATP1B1, but not of OATP1B3 and OATP2B1, is significantly affected by genetic variants. The SLCO1B1 variant c.388A>G is the major determinant with additional consequences on atorvastatin plasma levels.
Genome Medicine 01/2013; 5(1):1. · 3.40 Impact Factor
[show abstract][hide abstract] ABSTRACT: Validation of an association between the UGT1A6_19_T>G (rs6759892) polymorphism and overall breast cancer risk. A pilot study included two population-based case-control studies from Germany (MARIE-GENICA). An independent validation study comprised four independent breast cancer case-control studies from Finland (KBCP, OBCS), Germany (BBCC), and Sweden (SASBAC). The pooled analysis included 7418 cases and 8720 controls from all six studies. Participants were of European descent. Genotyping was done by MALDI-TOF MS and statistical analysis was performed by logistic regression adjusted for age and study. The increased overall breast cancer risk for women with the UGT1A6_19_GG genotype which was observed in the pilot study was confirmed in the set of four independent study collections (OR 1.13, 95% CI 1.05-1.22; p = 0.001). The pooled study showed a similar effect (OR 1.09, 95% CI 1.04-1.14; p = 0.001). The risk effect on the basis of allele frequencies was highly significant, the pooled analysis showed an OR of 1.11 (95% CI 1.06-1.16; p = 5.8 × 10(-6)). We confirmed the association of UGT1A6_19_GG with increased overall breast cancer risk and conclude that our result from a well powered multi-stage study adds a novel candidate to the panel of validated breast cancer susceptibility loci.
[show abstract][hide abstract] ABSTRACT: Interindividual variability in cytochrome P450 3A4 (CYP3A4) is believed to be largely heritable; however, predictive genetic factors have remained scarce. Using a candidate-gene approach in a human liver bank, we identified single-nucleotide polymorphisms (SNPs) in the Ah-receptor nuclear translocator (ARNT), glucocorticoid receptor (GR), progesterone receptor membrane component 2 (PGRMC2), and peroxisome proliferator-activated receptor-α (PPARA) that are associated with CYP3A4 phenotype. Validation in atorvastatin-treated volunteers confirmed a decrease in atorvastatin-2-hydroxylation in carriers of PPARA SNP rs4253728. Homozygous carriers expressed significantly less PPAR-α protein in the liver. Moreover, shRNA-mediated PPARA gene knockdown in primary human hepatocytes decreased expression levels of the PPAR-α target ACOX1 and of CYP3A4 by more than 50%. In conclusion, this study identified novel genetic determinants of CYP3A4 that, together with nongenetic factors, explained 52, 55, and 33% of hepatic CYP3A4 mRNA, protein, and atorvastatin-2-hydroxylase activity, respectively. These findings have implications for variability in response to drug substrates of CYP3A4.
[show abstract][hide abstract] ABSTRACT: Delayed graft function (DGF) is an important complication in renal transplantation, contributing significantly to decrease in long-term allograft survival. In addition to donor- and recipient-related risk factors such as immunosuppression, altered renal excretion of xenobiotics by membrane transporters may influence DGF. Using DNA samples from recipients and donors, we assessed the impact on DGF of genetic variants in P-glycoprotein (ABCB1), multidrug resistance protein 2 (ABCC2), and the nuclear pregnane X receptor (PXR/NR1I2), which regulates the transcription of enzymes and transporters. In our local cohort of renal transplant recipients (n = 178), DGF occurred in 27.5%. The PXR 8055TT genotype of the donor only (not of the recipient) was significantly associated with an increased risk for DGF. This finding emerged from univariate as well as multivariate logistic regression analysis including 16 nongenetic factors and held true after correction for multiple testing. Our findings provide the first evidence that PXR may be associated with risk of DGF, independent of previously identified risk factors.
[show abstract][hide abstract] ABSTRACT: Artemisinins induce drug metabolism through the activation of the pregnane X receptor (PXR) in vitro. Here, we report the resequencing and genotyping of PXR variants in 75 Vietnamese individuals previously characterized for CYP3A enzyme activity after artemisinin exposure. We identified a total of 31 PXR variants, including 5 novel single nucleotide polymorphisms (SNPs), and we identified significantly different allele frequencies relative to other ethnic groups. A trend of significance was observed between the level of CYP3A4 induction by artemisinin and two PXR variants, the 8118C→T (Y328Y) and 10719A→G variants.
Antimicrobial Agents and Chemotherapy 01/2012; 56(4):2153-7. · 4.57 Impact Factor
[show abstract][hide abstract] ABSTRACT: Organic cation transporters (OCTs) determine not only physiological processes but are also involved in the cellular uptake of anticancer agents. Based on microarray analyses in hepatocellular carcinoma (HCC), SLC22A1/OCT1 mRNA seems to be downregulated, but systematic protein expression data are currently missing. Moreover, the underlying molecular mechanisms responsible for altered SLC22A1 expression in HCC are not fully understood. Therefore, we investigated the role of DNA methylation in the transcriptional regulation of the family members SLC22A1/OCT1, SLC22A2/OCT2 and SLC22A3/OCT3 in HCC.
Semiquantitative immunohistochemistry of SLC22A1 protein expression was performed in paired HCC and histological normal adjacent liver tissues (n = 71) using tissue microarray analyses, and the results were correlated with clinicopathological features. DNA methylation, quantified by MALDI-TOF mass spectrometry and gene expression of SLC22A1, SLC22A2 and SLC22A3 were investigated using fresh-frozen HCC (n = 22) and non-tumor adjacent liver tissues as well as histologically normal liver samples (n = 120) from a large-scale liverbank.
Based on tissue microarray analyses, we observed a significant downregulation of SLC22A1 protein expression in HCC compared to normal adjacent tissue (P < 0.0001). SLC22A1 expression was significantly inverse correlated with expression of the proliferation marker MIB1/Ki-67 (rs = -0.464, P < 0.0001). DNA methylation of SLC22A1 was significantly higher in HCC compared with non-tumor adjacent liver tissue and was lowest in histologically normal liver tissue. Methylation levels for SLC22A1 in combination with RASSF1A resulted in a specificity of > 90% and a sensitivity of 82% for discriminating HCC and tumor-free liver tissue.
DNA methylation of SLC22A1 is associated with downregulation of SLC22A1 in HCC and might be a new biomarker for HCC diagnosis and prognosis. Moreover, targeting SLC22A1 methylation by demethylating agents may offer a novel strategy for anticancer therapy of HCC.
Genome Medicine 12/2011; 3(12):82. · 3.40 Impact Factor
[show abstract][hide abstract] ABSTRACT: Expression quantitative trait loci (eQTL) analysis is a powerful approach toward identifying genetic loci associated with quantitative changes in gene expression. We applied genome-wide association analysis to a data set of >300 000 single-nucleotide polymorphisms and >48 000 mRNA expression phenotypes obtained by Illumina microarray profiling of 149 human surgical liver samples obtained from Caucasian donors with detailed medical documentation. Of 1226 significant associations, only 200 were validated when comparing with a previously published similar study. Potential explanations for low replication rate include differences in microarray platforms, statistical modeling, covariates considered and origin and collection procedures of tissues. Focused analysis revealed a subset of 95 associations related to absorption, distribution, metabolism and excretion of drugs. Of these, 21 were true replications and 74 were newly discovered associations in enzymes, transporters, transcriptional regulators and other genes. This study extends our knowledge about the genetics of inter-individual variability of gene expression with particular emphasis on pharmacogenomics.The Pharmacogenomics Journal advance online publication, 18 October 2011; doi:10.1038/tpj.2011.44.
The Pharmacogenomics Journal 10/2011; · 5.13 Impact Factor
[show abstract][hide abstract] ABSTRACT: Genetic polymorphisms of human ABC-transporter genes have been suggested to modulate breast cancer risk in the general population. In particular ABCC11 (MRP8), which is highly expressed in breast cancer tissue and involved in the efflux of conjugated estrogen metabolites such as estrone-3-sulfate and estradiol-17beta-glucuronide, has recently been proposed as a potential risk factor for female breast cancer. The wet earwax-associated G-allele of the c.538G>A polymorphism was associated with an increased risk for breast cancer in Japanese women. In contrast, no evidence for such an association could be observed in Caucasian women. We aimed to confirm/refute the association of the c.538G>A variant in ABCC11 with breast cancer risk and/or histo-pathological tumor characteristics in an independent population-based breast cancer case-control study from Germany comprising 1021 cases and 1015 age-matched controls. No association for allele and genotype frequencies of the 538G>A variant in ABCB11 with breast cancer risk was found. Our data suggest that the c.538G>A variation in ABCC11 does not contribute to breast carcinogenesis in women of European descent.
Breast Cancer Research and Treatment 06/2011; 129(3):993-9. · 4.47 Impact Factor
[show abstract][hide abstract] ABSTRACT: Tacrolimus (Tac) is effective in the treatment of steroid-refractory ulcerative colitis (UC); however, nonresponse and unpredictable side effects are major limitations. Because Tac response in patients who have undergone solid-organ transplantation has been associated with the presence of variants in CYP3A and ABCB1, we elucidated the contributions of CYP3A4*1B and CYP3A5*3 and of ABCB1 1236C>T, 2677G>T,A, and 3435C>T polymorphisms to Tac response in 89 patients with UC. Short-term remission and response were achieved in 61 and 14% of the patients, respectively, and were associated with colectomy-free survival. In a linear logistic regression model, patients with homozygous variants for one of the three ABCB1 alleles showed significantly higher short-term remission rates as compared with those of other genotypes. The effects held true after multivariate analysis including multiple comparisons and were more pronounced after correction for dose-adjusted Tac blood trough levels. We suggest that ABCB1, but not CYP3A5, may predict short-term remission of Tac in steroid-refractory UC.