title: Quantitative genome-wide methylation analysis of high-grade non-muscle invasive bladder cancer.
abstract: High-grade non-muscle invasive bladder cancer (HG-NMIBC) is a clinically unpredictable disease with greater risks of recurrence and progression relative to their low-intermediate-grade counterparts. The molecular events, including those affecting the epigenome, that characterize this disease entity in the context of tumor development, recurrence, and progression, are incompletely understood. We therefore interrogated genome-wide DNA methylation using HumanMethylation450 BeadChip arrays in 21 primary HG-NMIBC tumors relative to normal bladder controls. Using strict inclusion-exclusion criteria we identified 1,057 hypermethylated CpGs within gene promoter-associated CpG islands, representing 256 genes. We validated the array data by bisulphite pyrosequencing and examined 25 array-identified candidate genes in an independent cohort of 30 HG-NMIBC and 18 low-intermediate-grade NMIBC. These analyses revealed significantly higher methylation frequencies in high-grade tumors relative to low-intermediate-grade tumors for the ATP5G2, IRX1 and VAX2 genes (P<0.05), and similarly significant increases in mean levels of methylation in high-grade tumors for the ATP5G2, VAX2, INSRR, PRDM14, VSX1, TFAP2b, PRRX1, and HIST1H4F genes (P<0.05). Although inappropriate promoter methylation was not invariantly associated with reduced transcript expression, a significant association was apparent for the ARHGEF4, PON3, STAT5a, and VAX2 gene transcripts (P<0.05). Herein, we present the first genome-wide DNA methylation analysis in a unique HG-NMIBC cohort, showing extensive and discrete methylation changes relative to normal bladder and low-intermediate-grade tumors. The genes we identified hold significant potential as targets for novel therapeutic intervention either alone, or in combination, with more conventional therapeutic options in the treatment of this clinically unpredictable disease.
authors.affiliation: a Institute for Science and Technology in Medicine, Keele University , UK.|b Urology Department , University Hospitals of North Midlands NHS Trust , UK.
authors.forename: Mark O
authors.initials: MO
authors.lastname: Kitchen
authors.affiliation: c Institute of Cancer and Genomic Sciences, University of Birmingham , UK.
authors.forename: Richard T
authors.initials: RT
authors.lastname: Bryan
authors.affiliation: d Advanced Data Analysis Center, University of Nottingham , UK.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Emes
authors.affiliation: a Institute for Science and Technology in Medicine, Keele University , UK.
authors.forename: John R
authors.initials: JR
authors.lastname: Glossop
authors.affiliation: b Urology Department , University Hospitals of North Midlands NHS Trust , UK.
authors.forename: Christopher
authors.initials: C
authors.lastname: Luscombe
authors.affiliation: c Institute of Cancer and Genomic Sciences, University of Birmingham , UK.
authors.forename: K K
authors.initials: KK
authors.lastname: Cheng
authors.affiliation: c Institute of Cancer and Genomic Sciences, University of Birmingham , UK.|e Department of Complex Genetics , Maastricht University Medical Center , The Netherlands.|f NUTRIM School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center , The Netherlands.|g CAPHRI School for Public Health and Primary Care, Maastricht University Medical Center , The Netherlands.
authors.forename: Maurice P
authors.initials: MP
authors.lastname: Zeegers
authors.affiliation: h Cancer Research Unit, University of Warwick , UK.
authors.forename: Nicholas D
authors.initials: ND
authors.lastname: James
authors.affiliation: c Institute of Cancer and Genomic Sciences, University of Birmingham , UK.
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Devall
authors.affiliation: i The Genome Center, Barts and the London School of Medicine and Dentistry , London , UK.
authors.forename: Charles A
authors.initials: CA
authors.lastname: Mein
authors.affiliation: b Urology Department , University Hospitals of North Midlands NHS Trust , UK.
authors.forename: Lyndon
authors.initials: L
authors.lastname: Gommersall
authors.affiliation: a Institute for Science and Technology in Medicine, Keele University , UK.
authors.forename: Anthony A
authors.initials: AA
authors.lastname: Fryer
authors.affiliation: a Institute for Science and Technology in Medicine, Keele University , UK.
authors.forename: William E
authors.initials: WE
authors.lastname: Farrell
mesh_terms: D043303:Aryldialkylphosphatase / Q000235:genetics*; D014408:Biomarkers, Tumor / Q000235:genetics; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation / Q000235:genetics*; D005260:Female; D015894:Genome, Human; D055106:Genome-Wide Association Study; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D008297:Male; D009361:Neoplasm Invasiveness / Q000235:genetics; D009363:Neoplasm Proteins / Q000235:genetics; D009364:Neoplasm Recurrence, Local / Q000235:genetics / Q000473:pathology; D011401:Promoter Regions, Genetic / Q000235:genetics; D064067:Rho Guanine Nucleotide Exchange Factors / Q000235:genetics; D050799:STAT5 Transcription Factor / Q000235:genetics*; D025521:Tumor Suppressor Proteins / Q000235:genetics*; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000473:pathology
chemical_list: C576602:ARHGEF4 protein, human; D014408:Biomarkers, Tumor; D018398:Homeodomain Proteins; D009363:Neoplasm Proteins; D064067:Rho Guanine Nucleotide Exchange Factors; D050799:STAT5 Transcription Factor; C494092:STAT5A protein, human; D025521:Tumor Suppressor Proteins; C492362:VAX2 protein, human; D043303:Aryldialkylphosphatase; C113656:PON3 protein, human
keywords: Epigenetics; HumanMethylation450 BeadChip Array; gene expression; high-grade non-muscle invasive bladder cancer; methylation