Progressive increases in de novo methylation of CPG islands in bladder cancer

We conducted a quantitative analysis of the extent of de novo methylation o f four CpG islands in human urinary transitional cell carcinomas of differe nt stages and grades to determine how frequently these CpG islands became m ethylated in transition cell carcinomas during progression. The CpG islands included exon 5 of PAX6, exon 2 of p16, the 5' end of the deleted in bladd er cancer gene, and the 5' end of transmembrane protein containing epiderma l growth factor and follistatin domains. These sequences were not methylate d in normal urothelial tissues; however, 48 of the 54 tumors examined (89%) showed methylation levels in excess of 20% for at least one of the markers . The number of markers concurrently methylated in individual tumors increa sed with the stage of the tumor, with several of the more aggressive invasi ve cancers showing hypermethylation of all four markers compared with the L ess aggressive invasive cancers. However, considerable methylation defects were present in superficial, preinvasive, papillary tumors. These data demo nstrate that 89% of bladder cancers have increased methylation of CpG islan ds relative to their normal counterparts and suggest the occurrence of a hy permethylator phenotype in which multiple independent CpG islands become co ncurrently methylated in individual tumors in a process associated with tum or progression.