Purpose: Epigenetic alterations are common and can now be addressed in a parallel fashion. We investigated the methylation in bladder cancer with respect to location in genome, consistency, variation in metachronous tumors, impact on transcripts, chromosomal location, and usefulness as urinary markers.Experimental Design: A microarray assay was utilized to analyze methylation in 56 samples. Independent validation was conducted in 63 samples by a PCR-based method and bisulfite sequencing. The methylation levels in 174 urine specimens were quantified. Transcript levels were analyzed using expression microarrays and pathways were analyzed using dedicated software.Results: Global methylation patterns were established within and outside CpG islands. We validated methylation of the eight tumor markers genes ZNF154 (P < 0.0001), HOXA9 (P < 0.0001), POU4F2 (P < 0.0001), EOMES (P ¼ 0.0005), ACOT11 (P ¼ 0.0001), PCDHGA12 (P ¼ 0.0001), CA3 (P ¼ 0.0002), and PTGDR (P ¼ 0.0110), the candidate marker of disease progression TBX4 (P < 0.04), and other genes with stage-specific methylation. The methylation of metachronous tumors was stable and targeted to certain pathways. The correlation to expression was not stringent. Chromosome 21 showed most differential methylation (P < 0.0001) and specifically hypomethylation of keratins, which together with keratin-like proteins were epigenetically regulated. In DNA from voided urine, we detected differential methylation of ZNF154 (P < 0.0001), POU4F2 (P < 0.0001), HOXA9 (P < 0.0001), and EOMES (P < 0.0001), achieving 84% sensitivity and 96% specificity.Conclusions: We initiated a detailed mapping of the methylome in metachronous bladder cancer. Novel genes with tumor, chromosome, as well as pathway-specific differential methylation in bladder cancer were identified. The methylated genes were promising cancer markers for early detection of bladder cancer. Clin Cancer Res; 17(17); 5582-92. Ó2011 AACR.

Background: Blue collar men have both the lowest average life and longest patient delay, and currently no research has been conducted with mobile health applications as health promotion interventions for this target group. This study wants to contribute with knowledge about how mobile health applications affect the level of physical activity amongst blue collar men as well as the frequency of how often they think and do something for their own physical health.Methods: Controlled clinical trial, men, aged 19-62 years, working in industrial companies, who are assigned to an intervention group (n=35) or a control group (n=35) are allocated based on mobile phone type/model. Baseline and effect measurement is carried out in both groups. Intervention period of 6 months in which the intervention group uses mobile applications that record the number of steps, and they receive a health-promoting text message every other week, and, every 4 weeks, the men send the recorded number of steps as well as visual analogue scale (VAS) thinking and doing in a selected week.Results: During the intervention period, the following parameters have improved significantly in the intervention group: muscle mass, from 67.9 to 68.8 kg (P=0.02); oxygen absorption in the fitness test, from 3.09 to 3.22 L/min (P=0.03); VAS, how often the men do something to improve their own physical health, from 6.00 to 7.00 (P=0.0051). There are improved parameters in the intervention group for resting heart rate (RHR), fitness test, fat percentage and VAS thinking. For blood pressure (BP), there is a surprising increase in the intervention group: systolic blood pressure (SBP) from 134 to 136 mm/Hg (P=0.09) and diastolic 83 to 86 mm/Hg (P=0.03). Conclusions:The use of mobile health applications, health promoting text messages and step-bystep control increases the physical activity level of blue-collar men as well as how often they think and do something to improve their physical health. Significant improvement is seen in their muscle mass and oxygen uptake as well as in relation to how often they do something to improve their own health.

Background: Human TOX3 (TOX high mobility group box family member 3) regulates Ca 2+ dependent transcription in neurons and has been associated with breast cancer susceptibility. Aim of the study was to investigate the expression of TOX3 in bladder cancer tissue samples and to identify genes and pathways altered upon TOX3 dysregulation using a cell line model. Methods:We performed microarray transcript profiling of biopsies and validated the data with RT-qPCR. We used cell line models for Overexpression and siRNA mediated knockdown of TOX3. Pathway analysis was applied for target gene identification and immunoprecipitation studies were used for DNA binding studies.Results: Microarray transcript profiling of 89 bladder biopsies showed a significant up-regulation of TOX3 (p<10 -4 ) in non-muscle invasive (Ta-T1) bladder tumors compared to muscle-invasive (T2-T4) bladder tumors and normal urothelium. Microarray expression profiling of human bladder cancer cells over expressing TOX3 followed by Pathway analysis showed that TOX3 Overexpression mainly affected the Interferon Signaling Pathway. TOX3 up regulation induced the expression of several genes with a gamma interferon activation site (GAS), e.g. STAT1.In vitro functional studies showed that TOX3 was able to bind to the GAS-sequence located at the STAT1 promoter. siRNA mediated knockdown of TOX3 in RT4 bladder cancer cells decreased STAT1 expression suggesting a direct impact of TOX3 on STAT1. Immunoprecipitation of TOX3 over expressing cell extracts with an artificial "GAS"-DNA element resulted in an enrichment of the GAS containing DNA-sequence, providing evidence for a potential interaction of TOX3 with the GAS-sequence of STAT1.Conclusions: These results provide evidence for an alternative activation of the downstream interferon targets, independent of the initial interferon-receptor interaction, and consequently a biological role for TOX3.

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