Association of family structure with type 1 diabetes management and outcomes in adolescents: A population-based cross-sectional survey.

BACKGROUND: Diabetes therapies have enormously changed during past decades, but only few studies have analyzed the association between family structure and diabetes management and outcomes. OBJECTIVE: To analyze cross-sectionally the associations of family structure with type 1 diabetes (T1D) management and various diabetes outcomes. METHODS: A total of 1635 11- to 17-year-old participants and their parents completed one of three baseline surveys as part of a nationwide, population-based cohort study on early-onset, long-standing T1D. Associations between family structure and outcome variables were analyzed by multivariable linear/logistic regression. RESULTS: Compared to adolescents living with both parents (reference), HbA1c was 0.48% (95% confidence interval 0.24; 0.71) / 5.2 (2.6; 7.8) mmol/mol higher in adolescents living with one parent and 0.34% (0.08; 0.59) / 3.7 (0.9; 6.5) mmol/mol higher in those living with one parent and her/his partner. The blood glucose self-monitoring (SMBG) frequency was lower (single parent: -0.6 (-1.1; -0.2), parent and partner:-0.5 (-1.0; 0.0)) and parents reported more long-term consequences related to school or work (ORsingle-parent 1.52 (0.90; 2.57), ORparent + partner 1.50 (0.86; 2.60)). While living with one parent was associated with increased odds of insulin injection vs. insulin pump therapy (OR 1.61 [1.13; 2.29]), the odds of low hypoglycemia awareness (OR 1.75 [1.00; 3.08]) and diabetes complications (1.32 [0.78; 2.22]) were higher in people living with a parent and her/his partner. CONCLUSIONS: Living with only one parent with or without a new partner was associated with less SMBG and pump use and poor diabetes outcomes. Future studies to explore the underlying mechanisms are required.

Genetic determinants of steatosis and fibrosis progression in paediatric non-alcoholic fatty liver disease.

BACKGROUND AND AIMS: Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in children and adolescents today. In comparison with adult disease, paediatric NAFLD may show a periportal localization, which is associated with advanced fibrosis. This study aimed to assess the role of genetic risk variants for histological disease pattern and severity in childhood NAFLD. METHODS: We studied 14 single nucleotide polymorphisms (SNP) in a cohort of 70 adolescents with biopsy-proven NAFLD. Genotype was compared to an adult control cohort (n = 200) and analysed in relation to histological disease severity and liver tissue proteomics. RESULTS: Three of the 14 SNPs were significantly associated with paediatric NAFLD after FDR adjustment, rs738409 (PNPLA3, P = 2.80 × 10-06 ), rs1044498 (ENPP1, P = 0.0091) and rs780094 (GCKR, P = 0.0281). The severity of steatosis was critically associated with rs738409 (OR=3.25; 95% CI: 1.72-6.52, FDR-adjusted P = 0.0070). The strongest variants associated with severity of fibrosis were rs1260326, rs780094 (both GCKR) and rs659366 (UCP2). PNPLA3 was associated with a portal pattern of steatosis, inflammation and fibrosis. Proteome profiling revealed decreasing levels of GCKR protein with increasing carriage of the rs1260326/rs780094 minor alleles and downregulation of the retinol pathway in rs738409 G/G carriers. Computational metabolic modelling highlighted functional relevance of PNPLA3, GCKR and UCP2 for NAFLD development. CONCLUSIONS: This study provides evidence for the role of PNPLA3 as a determinant of portal NAFLD localization and severity of portal fibrosis in children and adolescents, the risk variant being associated with an impaired hepatic retinol metabolism.

Identification of interactions of binary variables associated with survival time using survivalFS.

Many medical studies aim to identify factors associated with a time to an event such as survival time or time to relapse. Often, in particular, when binary variables are considered in such studies, interactions of these variables might be the actual relevant factors for predicting, e.g., the time to recurrence of a disease. Testing all possible interactions is often not possible, so that procedures such as logic regression are required that avoid such an exhaustive search. In this article, we present an ensemble method based on logic regression that can cope with the instability of the regression models generated by logic regression. This procedure called survivalFS also provides measures for quantifying the importance of the interactions forming the logic regression models on the time to an event and for the assessment of the individual variables that take the multivariate data structure into account. In this context, we introduce a new performance measure, which is an adaptation of Harrel's concordance index. The performance of survivalFS and the proposed importance measures is evaluated in a simulation study as well as in an application to genotype data from a urinary bladder cancer study. Furthermore, we compare the performance of survivalFS and its importance measures for the individual variables with the variable importance measure used in random survival forests, a popular procedure for the analysis of survival data. These applications show that survivalFS is able to identify interactions associated with time to an event and to outperform random survival forests.

Identification of a novel susceptibility locus at 13q34 and refinement of the 20p12.2 region as a multi-signal locus associated with bladder cancer risk in individuals of European ancestry.

Candidate gene and genome-wide association studies (GWAS) have identified 15 independent genomic regions associated with bladder cancer risk. In search for additional susceptibility variants, we followed up on four promising single-nucleotide polymorphisms (SNPs) that had not achieved genome-wide significance in 6911 cases and 11 814 controls (rs6104690, rs4510656, rs5003154 and rs4907479, P < 1 × 10(-6)), using additional data from existing GWAS datasets and targeted genotyping for studies that did not have GWAS data. In a combined analysis, which included data on up to 15 058 cases and 286 270 controls, two SNPs achieved genome-wide statistical significance: rs6104690 in a gene desert at 20p12.2 (P = 2.19 × 10(-11)) and rs4907479 within the MCF2L gene at 13q34 (P = 3.3 × 10(-10)). Imputation and fine-mapping analyses were performed in these two regions for a subset of 5551 bladder cancer cases and 10 242 controls. Analyses at the 13q34 region suggest a single signal marked by rs4907479. In contrast, we detected two signals in the 20p12.2 region-the first signal is marked by rs6104690, and the second signal is marked by two moderately correlated SNPs (r(2) = 0.53), rs6108803 and the previously reported rs62185668. The second 20p12.2 signal is more strongly associated with the risk of muscle-invasive (T2-T4 stage) compared with non-muscle-invasive (Ta, T1 stage) bladder cancer (case-case P ≤ 0.02 for both rs62185668 and rs6108803). Functional analyses are needed to explore the biological mechanisms underlying these novel genetic associations with risk for bladder cancer.

Identification and replication of the interplay of four genetic high-risk variants for urinary bladder cancer.

Little is known whether genetic variants identified in genome-wide association studies interact to increase bladder cancer risk. Recently, we identified two- and three-variant combinations associated with a particular increase of bladder cancer risk in a urinary bladder cancer case-control series (Leibniz Research Centre for Working Environment and Human Factors at TU Dortmund (IfADo), 1501 cases, 1565 controls). In an independent case-control series (Nijmegen Bladder Cancer Study, NBCS, 1468 cases, 1720 controls) we confirmed these two- and three-variant combinations. Pooled analysis of the two studies as discovery group (IfADo-NBCS) resulted in sufficient statistical power to test up to four-variant combinations by a logistic regression approach. The New England and Spanish Bladder Cancer Studies (2080 cases and 2167 controls) were used as a replication series. Twelve previously identified risk variants were considered. The strongest four-variant combination was obtained in never smokers. The combination of rs1014971[AA] near apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3A (APOBEC3A) and chromobox homolog 6 (CBX6), solute carrier family 1s4 (urea transporter), member 1 (Kidd blood group) (SLC14A1) exon single nucleotide polymorphism (SNP) rs1058396[AG, GG], UDP glucuronosyltransferase 1 family, polypeptide A complex locus (UGT1A) intron SNP rs11892031[AA] and rs8102137[CC, CT] near cyclin E1 (CCNE1) resulted in an unadjusted odds ratio (OR) of 2.59 (95% CI = 1.93-3.47; P = 1.87 × 10-10), while the individual variant ORs ranged only between 1.11 and 1.30. The combination replicated in the New England and Spanish Bladder Cancer Studies (ORunadjusted = 1.60, 95% CI = 1.10-2.33; P = 0.013). The four-variant combination is relatively frequent, with 25% in never smoking cases and 11% in never smoking controls (total study group: 19% cases, 14% controls). In conclusion, we show that four high-risk variants can statistically interact to confer increased bladder cancer risk particularly in never smokers.

Occupational bladder cancer: Polymorphisms of xenobiotic metabolizing enzymes, exposures, and prognosis.

Approximately 7% of all bladder cancer cases in males are associated with occupation. The question arises whether the use of genome-wide association studies was able to identify bladder cancer risk factors that may modulate occupational bladder cancer risk and prognosis. One hundred and forty-three bladder cancer cases with suspected occupational bladder cancer and 337 controls were genotyped for the following polymorphisms: N-acetyltransferase 2 (NAT2), glutathione S-transferase M1 (GSTM1), glutathione S-transferase T1 (GSTT1), UDP-glucuronyltransferase 1A rs11892031 (UGT1A), rs9642880 (close to c-MYC), and rs710521 (close to TP63). The most relevant polymorphisms for occupational bladder cancer risk were GSTM1 and UGT1A, especially when co-occurring (GSTM1 negative and rs11892031[A/A]: 48% cases vs. 38% controls, OR 1.47, 95% CI 0.99-2.20). The effect was more pronounced in smokers. GSTM1 negative genotype occurred more frequently in cancer cases exposed to aromatic amines, carbolineum, and in painters and varnishers. UGT1A (rs11892031[A/A]) was found frequently in cases exposed to carbolineum, crack test spray, PAH, and in painters and varnishers. All investigated polymorphisms except rs710521 (TP63) seemed to exert an impact on recurrence risk. Relapse-free times were shorter for NAT2 slow and ultra-slow, GSTT1 positive and GSTM1 negative cases. Occupational bladder cancer cases with a number of risk variants displayed significantly shorter relapse-free times compared to cases with few, less relevant risk alleles as evidenced by median difference 8 months. In conclusion, in the present, suspected occupational bladder cancer cases phase II polymorphisms involved in bladder carcinogen metabolism modulate bladder cancer recurrence. Most relevant for bladder cancer risk were GSTM1 and UGT1A but not NAT2.

Urinary bladder cancer risk factors in an area of former coal, iron, and steel industries in Germany.

This study was performed to investigate the frequency of bladder cancer in patients with an occupational history such as underground hard coal mining and/or painting after the structural change in the local industry. A total of 206 patients with bladder cancer and 207 controls were enlisted regarding occupational and nonoccupational bladder cancer risk factors by questionnaire. The phase II enzymes N-acetyltransferase 2 (NAT2), glutathione S-transferases M1 (GSTM1), and T1 (GSTT1) and the single nucleotide polymorphism (SNP) rs11892031[A/C] reported to be associated with bladder cancer in genome-wide association studies were genotyped. The bladder cancer risk in varnishers and underground hard coal miners was increased as previously shown in a study in this area performed in the 1980s. The occupation of a car mechanic was associated with a significantly elevated bladder cancer risk and higher in the case of underground hard coal miners even though the mine was closed in 1987. The frequency of GSTM1 negative genotype was comparable in cases and controls (53% versus 54%). In the case of NAT2, the slow NAT2 genotype was more frequent (62% versus 58%) and ultra-slow NAT2 genotype (NAT2*6A and/or *7B alleles only) was 23% versus 15%. An occupational history of a varnisher or an underground hard coal miner remains a risk factor for bladder cancer occurrence. Data indicate that in the case of bladder cancer, GSTM1 is a susceptibility factor related to environmental and/or occupational exposure.

Polymorphisms of xenobiotic metabolizing enzymes in bladder cancer patients of the Semmelweis University Budapest, Hungary.

Polymorphic xenobiotic metabolizing enzymes such as N-acetyltransferase 2 (NAT2) or glutathione S-transferase M1 (GSTM1) are known to modulate bladder cancer risk. As no apparent data were available from Hungary, a former member of the eastern European economic organization, a study was performed in Budapest. In total, 182 bladder cancer cases and 78 cancer-free controls were investigated by questionnaire. Genotypes of NAT2, GSTM1, GSTT1, rs1058396 and rs17674580 were determined by standard methods. Current smokers' crude odds ratio (OR) (3.43) and former smokers crude OR (2.36) displayed a significantly increased bladder cancer risk. The risk rose by a factor of 1.56 per 10 pack years. Exposure to fumes was associated with an elevated bladder cancer risk (23% cases, 13% controls). Sixty-four % of the cases and 59% of controls were slow NAT2 acetylators. It was not possible to establish a particular impact of NAT2*6A and *7B genotypes (15 cases, 8%, 5 controls, 7%). GSTT1 exerted no marked influence on bladder cancer (negative 21% cases vs. 22% controls). The portion of GSTM1 negative bladder cancer patients was increased (63% cases vs. 54% controls). The SLC14A1 SNPs rs1058396[AG/GG] and the nearby rs17674580[CT/TT] occurred more frequently in cases (79% and 68%) than controls (77% and 55%). The portion of GSTM1 negative bladder cancer patients is comparable with portions reported from other industrialized areas like Lutherstadt Wittenberg/Germany (58%), Dortmund/Germany (70%), Brescia/Italy (66%) or an occupational case-control series in Germany (56%). Data indicate that GSTM1 is a susceptibility factor for environmentally triggered bladder cancer rather than for smoking-mediated bladder cancer.

N-acetyltransferase 1*10 genotype in bladder cancer patients.

In a large bladder cancer study in the greater Berlin area with 425 cases and 343 controls, the haplotype N-acetyltransferase 1*10 (NAT1*10) was associated with a decreased bladder cancer risk. In a recently published meta-analysis, results of the studies were found to be inconclusive. Therefore, the aim of this study was to investigate the frequency of NAT1*10 in bladder cancer patients and controls recruited in an area without industries reported to be associated with increased bladder cancer risk. Rs1057126 (1088 T > A) and rs15561 (1095 C > A) were determined in 412 bladder cancer patients and 415 controls without a known history of malignancies. With these two single-nucleotide polymorphisms (SNP), it was possible to distinguish between NAT1*4 (wild type), NAT1*3 (1095 C > A), and NAT1*10 (1088 T > A, 1095C > A). The frequencies of the determined NAT1 haplotypes did not differ markedly between cases and controls: NAT1*4: 74%, NAT1*3: 6%, NAT1*10: 20%. Bladder cancer risk was not significantly modulated by NAT1*10/*10 (OR 1.03, 95% CI 0.71-1.48) but was higher for NAT1*3/*3 genotypes (OR 2.05, 95% CI 1.32-3.21). In contrast to the Berlin study from 2001, data in present study demonstrated that NAT1*10 haplotype was not associated with a significantly decreased bladder cancer risk. This may be due to local effects in the greater Berlin area, particularly at the time of investigation. The findings of the present study are in agreement with observations of a recently published meta-analysis which also showed no relevant impact of NAT1*10 haplotype on bladder cancer risk. The impact of the rare NAT1*3/*3 genotype was significant but this may be attributed to rarity without major practical relevance.

Urinary cadmium levels in active and retired coal miners.

A meta-analysis, based upon 24 publications, showed a significantly elevated risk for urinary bladder cancer amongst miners. In European underground hard coal mining areas, an increased risk for urinary bladder cancer development was noted among hard coal miners, in particular in three investigations in the greater Dortmund area. However, the cause remains unclear. As cadmium (Cd), which was reported to be a bladder carcinogen in humans and is a constituent of coal, the aim of this study was to determine urinary Cd levels in active and retired hard coal miners and assess whether hard coal miners demonstrated elevated metal levels. In total, 103 retired and 25 active hard coal miners as well as 18 controls without any history of hard coal mining were investigated for urinary Cd levels. Urinary Cd concentrations, in addition to other elements, were analyzed in spot urines by ICP-MS-based multi-element analysis in a Department for Forensic and Clinical Toxicology. Limit of detection (LOD) for Cd was 0.5 µg/L. Reference value for occupationally non-exposed working age population was 0.8 µg/L. In total, 49% of all underground coal miners were exposed to coal dust, 12% to grinded rock, and 39% to both. Urinary Cd levels in retired as well as active coal miners and controls were clearly below the Biological Exposure Index. Urinary Cd concentration is a suitable biomarker to evaluate the metallic load of the body, as the half-life is > than 10 years. The detected urinary Cd levels in retired and active coal miners indicated underground hard coal miners were not apparently exposed to Cd to a occupationally-relevant concentration.

Occupational risk factors for prostate cancer in an area of former coal, iron, and steel industries in Germany. Part 2: results from a study performed in the 1990s.

Currently, there is no established occupational risk factor for prostate cancer. However, in the 1980s, a hospital-based case-control study in the greater Dortmund area showed an elevated risk for hard coal miners and, based on few cases, for painters and varnishers. Therefore, approximately 10 yr later, a similar study regarding prostate cancer was performed in this area. In total, 292 patients with prostate cancer who underwent radical prostatectomy and 313 controls who underwent transurethral resection of a benign prostatic hyperplasia were investigated by questionnaire. All of them were operated on between 1995 and 1999. This study showed a decreased risk for prostate cancer in hard coal miners (odds ratio [OR] = 0.67, 95% confidence interval [CI] = 0.44-1.03). Occupational exposures related to an elevated risk for prostate cancer were exposures to combustion products (20% cases vs. 11% controls), colorants and dyes (19 vs. 13%), and cutting fluids (8 vs. 6%). The different prostate cancer risks for underground coal miners in two studies with a time interval of approximately 10 yr are striking. Factors to be discussed are the introduction of prostate-specific antigen (PSA) screening for prostate cancer and investigation of cases that underwent radical prostatectomy, where the disease in general is locally confined. Working conditions in the local underground coal mines improved over time but did not change markedly in the period of interest. In essence, the present study does not corroborate an elevated prostate cancer risk in former underground hard coal miners from the greater Dortmund area.

Occupational risk factors for relapse-free survival in bladder cancer patients.

The influence of occupational risk factors on bladder cancer development is well investigated. However, studies on the influence on bladder cancer prognosis are rare. Therefore, it was of interest to investigate the time to first relapse in the follow-ups of three case-control series from Dortmund, Neuss, and Lutherstadt Wittenberg, Germany. Relapse-free survival of in total 794 urinary bladder cancer patients (Dortmund 174, Neuss 407, Lutherstadt Wittenberg 213) was derived from medical records. Cox regression models were used to determine the impact of profession and exposure to bladder carcinogens if the risk factor was present in at least four cases. One or several relapses were observed in 416 cases (52%). Median time to first relapse was 0.94 yr. Ten professions were observed in at least 4 patients. No significant associations were found. However, workers in the leather industry (n = 4), printing industry (n = 4), transportation (n = 43), and chemical industry (n = 40) and locksmiths/mechanics (n = 44) showed shorter relapse-free times. No trend to shorter relapse-free time was observed for miners (n = 42), agriculturists (n = 18), painters/lacquerers (n = 21), colorant production and processing workers (n = 7), foundry workers (n = 5), and persons exposed to aromatic amines (n = 45). Although the follow-up comprised nearly 800 cases, data on occupations and exposures of interest were not sufficient to obtain significant results. However, first results indicated potential associations that are worth further investigations.

Prostate Specific Antigen (PSA) as Predicting Marker for Clinical Outcome and Evaluation of Early Toxicity Rate after High-Dose Rate Brachytherapy (HDR-BT) in Combination with Additional External Beam Radiation Therapy (EBRT) for High Risk Prostate Cancer.

High-dose-rate brachytherapy (HDR-BT) with external beam radiation therapy (EBRT) is a common treatment option for locally advanced prostate cancer (PCa). Seventy-nine male patients (median age 71 years, range 50 to 79) with high-risk PCa underwent HDR-BT following EBRT between December 2009 and January 2016 with a median follow-up of 21 months. HDR-BT was administered in two treatment sessions (one week interval) with 9 Gy per fraction using a planning system and the Ir192 treatment unit GammaMed Plus iX. EBRT was performed with CT-based 3D-conformal treatment planning with a total dose administration of 50.4 Gy with 1.8 Gy per fraction and five fractions per week. Follow-up for all patients was organized one, three, and five years after radiation therapy to evaluate early and late toxicity side effects, metastases, local recurrence, and prostate-specific antigen (PSA) value measured in ng/mL. The evaluated data included age, PSA at time of diagnosis, PSA density, BMI (body mass index), Gleason score, D'Amico risk classification for PCa, digital rectal examination (DRE), PSA value after one/three/five year(s) follow-up (FU), time of follow-up, TNM classification, prostate volume, and early toxicity rates. Early toxicity rates were 8.86% for gastrointestinal, and 6.33% for genitourinary side effects. Of all treated patients, 84.81% had no side effects. All reported complications in early toxicity were grade 1. PSA density at time of diagnosis (p = 0.009), PSA on date of first HDR-BT (p = 0.033), and PSA on date of first follow-up after one year (p = 0.025) have statistical significance on a higher risk to get a local recurrence during follow-up. HDR-BT in combination with additional EBRT in the presented design for high-risk PCa results in high biochemical control rates with minimal side-effects. PSA is a negative predictive biomarker for local recurrence during follow-up. A longer follow-up is needed to assess long-term outcome and toxicities.

The ultra-slow NAT2*6A haplotype is associated with reduced higher cognitive functions in an elderly study group.

N-Acetyltransferase 2 (NAT2) genotype is associated with age-related declines in basic sensory hearing functions. However, the possible modulatory role of NAT2 for higher cognitive functions has not yet been studied. We tested auditory goal-directed behavior and attentional control in 120 NAT2 genotyped subjects (63-88 years), using an auditory distraction paradigm in which participants responded to the duration of long and short tone stimuli. We studied involuntary shifts in attention to task-irrelevant deviant stimuli and applied event-related potentials (ERPs) to examine which cognitive subprocesses are affected by NAT2 status on a neurophysiological level. Relative to the standard stimuli, deviant stimuli decreased performance in the recently described ultra-slow acetylators (NAT2*6A and *7B): The increase in error-corrected reaction times (a combined measure of response speed and accuracy) in ultra-slow acetylators (254 ms increase) was more than twice as high as in the rapid acetylator reference group (111 ms increase; p < 0.01). The increase was still higher than in the other slow acetylators (149 ms increase, p < 0.05). In addition, clear differences were found in the ERP results: Ultra-slow acetylators showed deficits specifically in the automatic detection of changes in the acoustic environment as evidenced by reduced mismatch negativity (MMN, p < 0.005 compared to rapid acetylators). Refocussing of attention after a distracting event was also impaired in the ultra-slow acetylators as evidenced by a reduced re-orienting negativity (RON, p < 0.01 compared to rapid acetylators). In conclusion, the ultra-slow acetylation status was associated with reduced higher cognitive functions.

Glycerophospholipid profile in oncogene-induced senescence.

Alterations in lipid metabolism and in the lipid composition of cellular membranes are linked to the pathology of numerous diseases including cancer. However, the influence of oncogene expression on cellular lipid profile is currently unknown. In this work we analyzed changes in lipid profiles that are induced in the course of ERBB2-expression mediated premature senescence. As a model system we used MCF-7 breast cancer cells with doxycycline-inducible expression of NeuT, an oncogenic ERBB2 variant. Affymetrix gene array data showed NeuT-induced alterations in the transcription of many enzymes involved in lipid metabolism, several of which (ACSL3, CHPT1, PLD1, LIPG, MGLL, LDL and NPC1) could be confirmed by quantitative realtime PCR. A study of the glycerophospholipid and lyso-glycerophospholipid profiles, obtained by high performance liquid chromatography coupled to Fourier-transform ion cyclotron resonance-mass spectrometry revealed senescence-associated changes in numerous lipid species, including mitochondrial lipids. The most prominent changes were found in PG(34:1), PG(36:1) (increased) and LPE(18:1), PG(40:7) and PI(36:1) (decreased). Statistical analysis revealed a general trend towards shortened phospholipid acyl chains in senescence and a significant trend to more saturated acyl chains in the class of phosphatidylglycerol. Additionally, the cellular cholesterol content was elevated and accumulated in vacuoles in senescent cells. These changes were accompanied by increased membrane fluidity. In mitochondria, loss of membrane potential along with altered intracellular distribution was observed. In conclusion, we present a comprehensive overview of altered cholesterol and glycerophospholipid patterns in senescence, showing that predominantly mitochondrial lipids are affected and lipid species less susceptible to peroxidation are increased.

European genome-wide association study identifies SLC14A1 as a new urinary bladder cancer susceptibility gene.

Three genome-wide association studies in Europe and the USA have reported eight urinary bladder cancer (UBC) susceptibility loci. Using extended case and control series and 1000 Genomes imputations of 5 340 737 single-nucleotide polymorphisms (SNPs), we searched for additional loci in the European GWAS. The discovery sample set consisted of 1631 cases and 3822 controls from the Netherlands and 603 cases and 37 781 controls from Iceland. For follow-up, we used 3790 cases and 7507 controls from 13 sample sets of European and Iranian ancestry. Based on the discovery analysis, we followed up signals in the urea transporter (UT) gene SLC14A. The strongest signal at this locus was represented by a SNP in intron 3, rs17674580, that reached genome-wide significance in the overall analysis of the discovery and follow-up groups: odds ratio = 1.17, P = 7.6 × 10(-11). SLC14A1 codes for UTs that define the Kidd blood group and are crucial for the maintenance of a constant urea concentration gradient in the renal medulla and, through this, the kidney's ability to concentrate urine. It is speculated that rs17674580, or other sequence variants in LD with it, indirectly modifies UBC risk by affecting urine production. If confirmed, this would support the 'urogenous contact hypothesis' that urine production and voiding frequency modify the risk of UBC.

Refinement of the prediction of N-acetyltransferase 2 (NAT2) phenotypes with respect to enzyme activity and urinary bladder cancer risk.

Polymorphisms of N-acetyltransferase 2 (NAT2) are well known to modify urinary bladder cancer risk as well as efficacy and toxicity of pharmaceuticals via reduction in the enzyme's acetylation capacity. Nevertheless, the discussion about optimal NAT2 phenotype prediction, particularly differentiation between different degrees of slow acetylation, is still controversial. Therefore, we investigated the impact of single nucleotide polymorphisms and their haplotypes on slow acetylation in vivo and on bladder cancer risk. For this purpose, we used a study cohort of 1,712 bladder cancer cases and 2,020 controls genotyped for NAT2 by RFLP-PCR and for the tagSNP rs1495741 by TaqMan(®) assay. A subgroup of 344 individuals was phenotyped by the caffeine test in vivo. We identified an 'ultra-slow' acetylator phenotype based on combined *6A/*6A, *6A/*7B and *7B/*7B genotypes containing the homozygous minor alleles of C282T (rs1041983, *6A, *7B) and G590A (rs1799930, *6A). 'Ultra-slow' acetylators have significantly about 32 and 46 % lower activities of caffeine metabolism compared with other slow acetylators and with the *5B/*5B genotypes, respectively (P < 0.01, both). The 'ultra-slow' genotype showed an association with bladder cancer risk in the univariate analysis (OR = 1.31, P = 0.012) and a trend adjusted for age, gender and smoking habits (OR = 1.22, P = 0.082). In contrast, slow acetylators in general were not associated with bladder cancer risk, neither in the univariate (OR = 1.02, P = 0.78) nor in the adjusted (OR = 0.98, P = 0.77) analysis. In conclusion, this study suggests that NAT2 phenotype prediction should be refined by consideration of an 'ultra-slow' acetylation genotype.

Pharmacokinetics explain in vivo/in vitro discrepancies of carcinogen-induced gene expression alterations in rat liver and cultivated hepatocytes.

Cultivated hepatocytes represent a well-established in vitro system. However, the applicability of hepatocytes in toxicogenomics is still controversially discussed. Recently, an in vivo/in vitro discrepancy has been described, whereby the non-genotoxic rat liver carcinogen methapyrilene alters the expression of the metabolizing genes SULT1A1 and ABAT, as well as the DNA damage response gene GADD34 in vitro, but not in vivo. If the collagen sandwich cultures of hepatocytes really produce false-positive data, this would compromise its application in toxicogenomics. To revisit the putative in vivo/in vitro discrepancy, we first analyzed and modeled methapyrilene concentrations in the portal vein of rats. The relatively short half-life of 2.8 h implies a rapid decrease in orally administered methapyrilene in vivo below concentrations that can cause gene expression alterations. This corresponded to the time-dependent alteration levels of GADD34, ABAT and SULT1A1 RNA in the liver: RNA levels are altered 1, 6 and 12 h after methapyrilene administration, but return to control levels after 24 and 72 h. In contrast, methapyrilene concentrations in the culture medium supernatant of primary rat hepatocyte cultures decreased slowly. This explains why GADD34, ABAT and SULT1A1 were still deregulated after 24 h exposure in vitro, but not in vivo. It should also be considered that the earliest analyzed time point in the previous in vivo studies was 24 h after methapyrilene administration. In conclusion, previously observed in vitro/in vivo discrepancy can be explained by different pharmacokinetics present in vitro and in vivo. When the in vivo half-life is short, levels of some initially altered genes may have returned to control levels already 24 h after administration.

Bladder cancer course, four genetic high-risk variants, and histopathological findings.

Urinary bladder cancer, a smoking and occupation related disease, was subject of several genome-wide association studies (GWAS). However, studies on the course of the disease based on GWAS findings differentiating between muscle invasive bladder cancer (MIBC) and non-muscle invasive bladder cancer (NMIBC) are rare. Thus we investigated 4 single nucleotide polymorphisms (SNPs) detected in GWAS, related to the genes coding for TACC3 (transforming, acidic coiled-coil containing protein 3), for FGFR3 (fibroblast growth factor receptor 3), for PSCA (prostate stem cell antigen) and the genes coding for CBX6 (chromobox homolog 6) and APOBEC3A (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3A). This study is based on 712 bladder cancer patients and 875 controls from 3 different case control studies in Germany. The 4 SNPs of interest (PSCA rs2294008 and rs2978974, FGFR3-TACC3 rs798766, and CBX6-APOBEC3A rs1014971) were determined by real-time polymerase chain reaction. The distribution of the 4 SNPs does not vary significantly between cases and controls in the entire study group and in the 3 local subgroups, including two former highly industrialized areas and a region without such history. Also, no significant differences in the bladder cancer subgroups of MIBC and NMIBC were observed. The 4 investigated SNPs do not noticeably contribute differently to the bladder cancer risk for the bladder cancer subgroups of MIBC and NMIBC.

Genome-wide Association Study of Bladder Cancer Reveals New Biological and Translational Insights.

BACKGROUND: Genomic regions identified by genome-wide association studies (GWAS) for bladder cancer risk provide new insights into etiology. OBJECTIVE: To identify new susceptibility variants for bladder cancer in a meta-analysis of new and existing genome-wide genotype data. DESIGN, SETTING, AND PARTICIPANTS: Data from 32 studies that includes 13,790 bladder cancer cases and 343,502 controls of European ancestry were used for meta-analysis. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSES: Log-additive associations of genetic variants were assessed using logistic regression models. A fixed-effects model was used for meta-analysis of the results. Stratified analyses were conducted to evaluate effect modification by sex and smoking status. A polygenic risk score (PRS) was generated on the basis of known and novel susceptibility variants and tested for interaction with smoking. RESULTS AND LIMITATIONS: Multiple novel bladder cancer susceptibility loci (6p.22.3, 7q36.3, 8q21.13, 9p21.3, 10q22.1, 19q13.33) as well as improved signals in three known regions (4p16.3, 5p15.33, 11p15.5) were identified, bringing the number of independent markers at genome-wide significance (p < 5 × 10-8) to 24. The 4p16.3 (FGFR3/TACC3) locus was associated with a stronger risk for women than for men (p-interaction = 0.002). Bladder cancer risk was increased by interactions between smoking status and genetic variants at 8p22 (NAT2; multiplicative p value for interaction [pM-I] = 0.004), 8q21.13 (PAG1; pM-I = 0.01), and 9p21.3 (LOC107987026/MTAP/CDKN2A; pM-I = 0.02). The PRS based on the 24 independent GWAS markers (odds ratio per standard deviation increase 1.49, 95% confidence interval 1.44-1.53), which also showed comparable results in two prospective cohorts (UK Biobank, PLCO trial), revealed an approximately fourfold difference in the lifetime risk of bladder cancer according to the PRS (e.g., 1st vs 10th decile) for both smokers and nonsmokers. CONCLUSIONS: We report novel loci associated with risk of bladder cancer that provide clues to its biological underpinnings. Using 24 independent markers, we constructed a PRS to stratify lifetime risk. The PRS combined with smoking history, and other established risk factors, has the potential to inform future screening efforts for bladder cancer. PATIENT SUMMARY: We identified new genetic markers that provide biological insights into the genetic causes of bladder cancer. These genetic risk factors combined with lifestyle risk factors, such as smoking, may inform future preventive and screening strategies for bladder cancer.