title: Aberrant methylation of the CDH13 (H-cadherin) promoter region in colorectal cancers and adenomas.
abstract: Expression of the cadherin family member CDH13 (H-cadherin) is reduced in several human tumors, and it has been hypothesized that this gene functions as a tumor suppressor gene. Previously, we reported that the 5' region of CDH13 is frequently methylated in breast and lung cancers. Here we confirmed the promoter activity of 5' region of CDH13 by luciferase assay and examined its aberrant methylation in colorectal cancers, cell lines, and adenomas. Methylation status was investigated by methylation-specific PCR (MSP) and by bisulfite DNA sequencing of cloned DNA of PCR amplicons. In cell lines, we examined the correlation between methylation status and mRNA expression by reverse transcription-PCR. Aberrant methylation of CDH13 was present in 7 of 13 (54%) cell lines, and expression was absent in 6 of 13 (46%) cell lines. CDH13 expression was present in six cell lines that showed only the unmethylated form by MSP and in one cell line that showed both the methylated and unmethylated forms. Treatment with 5-aza-2'-deoxycytidine restored CDH13 expression in methylated cell lines. In surgically resected samples, 17 of 35 (49%) cases of primary colorectal cancer, 2 of 33 (6%) cases of corresponding nonmalignant colorectal mucosa, and 8 of 19 (42%) adenomas were methylated. Sequence data after bisulfite treatment indicated that primary cancers and two cell lines with loss of expression were highly methylated compared with nonmalignant colorectal epithelial cells, especially at the attachment sites of primers for MSP, although there was heterogeneity in methylation status. Our results suggest that CDH13 expression is frequently silenced by aberrant methylation in colorectal cancers and adenomas and that methylation of CDH13 commences at an early stage of multistep colorectal tumorigenesis.
authors.affiliation: Hamon Center for Therapeutic Oncology Research, University of Texas Southwestern Medical Center, Dallas 75390, USA.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Toyooka
authors.forename: Kiyomi O
authors.initials: KO
authors.lastname: Toyooka
authors.forename: Kenichi
authors.initials: K
authors.lastname: Harada
authors.forename: Kuniharu
authors.initials: K
authors.lastname: Miyajima
authors.forename: Prakash
authors.initials: P
authors.lastname: Makarla
authors.forename: Ubaradka G
authors.initials: UG
authors.lastname: Sathyanarayana
authors.forename: Jing
authors.initials: J
authors.lastname: Yin
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Sato
authors.forename: Narayan
authors.initials: N
authors.lastname: Shivapurkar
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Meltzer
authors.forename: Adi F
authors.initials: AF
authors.lastname: Gazdar
mesh_terms: D000236:Adenoma / Q000235:genetics* / Q000378:metabolism; D015820:Cadherins / Q000096:biosynthesis / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics* / Q000378:metabolism; D019175:DNA Methylation; D006801:Humans; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D017422:Sequence Analysis, DNA; D013447:Sulfites / Q000737:chemistry; D014407:Tumor Cells, Cultured
chemical_list: D015820:Cadherins; C099972:H-cadherin; D013447:Sulfites; D008156:Luciferases; C042345:hydrogen sulfite