Promoter CpG Hypermethylation Research Articles

Promoter hypermethylation of MGMT, CDH1, RAR-beta and SYK tumour suppressor genes in granulosa cell tumours (GCTs) of ovarian origin.

Ovarian carcinoma (OC) is a leading cause of death among women throughout the world. A number of cancer-associated genes have been shown to be inactivated by hypermethylation of CpG islands during tumorigenesis. We tested the hypothesis that methylation status of MGMT, CDH1, RAR-β and SYK could be important in the ovarian tumorigenic process and can lead to the gene(s) inactivation. Therefore, we assessed the promoter hypermethylation of MGMT, CDH1, RAR-β and SYK in 43 ovarian granulosa cell tumours (GCTs) (adult type) using methylation-specific PCR. These tumours are relatively rare, accounting for approximately 3% of all ovarian cancers. Hypermethylation of MGMT (in 14 tumours), CDH1 (in nine tumours), RAR-β (in eight tumours) and SYK (in seven tumours) have been found. Selective loss of RAR-β and RAR-β2 mRNA has been found in seven patients, while that of MGMT and SYK in three patients who also show aberrant methylation in promoter region of RAR-β in addition to MGMT, SYK and CDH1 genes. Promoter CpG hypermethylation may be an alternative to mutation(s) to inactivate tumour suppressor genes such as MGMT, CDH1, RAR-β and SYK, and this can also be an early event in the pathogenesis of OCs. Moreover, hypermethylation of the MGMT and CDH1, MGMT and RAR-β and CDH1 and RAR-β promoters occurred concordantly (P< 0.001, 0.0421 and 0.0005 respectively; Fischer's exact test). In addition to this, monosomy 22 and trisomy 14 have also been found in 10 tumours. It is clear from the results that hypermethylation of the promoter region of these tumour suppressor genes, monosomy 22 and trisomy 14, may be critical steps in the tumorigenesis, which consequently play a permissive role for tumour aggressiveness. All these events might play an important role in the early clinical diagnosis of the disease. Our results, therefore, suggest a potential role for epigenetic modification of these critical tumour suppressor genes in pathways relevant to the transformation and differentiation of rare type of ovarian cancer (GCTs).

Bioinformatics for Oncogenomic Target Identification

Activation of proto-oncogenes and inactivation of tumor suppressor genes (TSGs) occur during multistage carcinogenesis. Proto-oncogenes are activated by gene amplification, point mutation and chromosomal translocation, while TSGs are inactivated by promoter CpG hypermethylation, point mutation, chromosomal translocation, and deletion. Array CGH combined with mRNA microarray analysis is applied for genome-wide screening of proto-oncogenes as well as TSGs [2]. Microarray analysis for lase-captured microdissection (LCM) sample is applied for screening of novel tumor markers specifically expressed in tumor cells [1]. Because huge amounts of data concerning genome sequences, expression profile, copy-number changes of human chromosomal regions in tumors are available in the post-genomic era, we applied bioinformatics for oncogenomic target identification.

Inactivation of retinoic acid receptor β by promoter CpG hypermethylation in gastric cancer

Inactivation of nuclear retinoic acid receptor β (RARβ) expression is implicated in tumorigenesis. We hypothesized that loss of RARβ in gastric cancer cells may occur as a result of multiple factors, including epigenetic modifications which alter RARβ promoter chromatin structure. We examined hypermethylation of CpG islands present in the RARβ promoter by methylation-specific PCR and the expression of RARβ in gastric cancer cell lines and tissues. Three (MKN-28, -45 and -74) out of eight gastric cancer cell lines had a loss of RAR expression associated with promoter methylation. RARβ expression was retrieved in these cell lines by treatment with 5-azacytidine or by the histone deacetylase inhibitor trichostatin A. Promoter hypermethylation was detected in 64 % (7/11) of gastric carcinoma tissues with reduced expression of RARβ, whereas it was detected in 22 % (2/9) of tumors with retained RARβ expression. To investigate the functions of exogenous RARβ in gastric cancer cells, we transfected a retroviral RARβ expression vector (LNSβ) into MKN-28 cells that have hypermethylation of the RARβ promoter. Overexpression of RAR in MKN-28 cells appeared to regulate the expression of DNA methyltransferase and DNA demethylase and the acetylation of hitone H4. These results suggest that the transcriptional inactivation of the RARβ by promoter CpG hypermethylation is frequently associated with gastric carcinoma. Our data also suggests that DNA methylation plays a pivotal role in establishing and maintaining an inactive state of RARβ by rendering the chromatin structure inaccessible to the transcription machinery.

Molecular characteristics of eight gastric cancer cell lines established in Japan.

Molecular characterization of eight gastric cancer cell lines established in Japan are summarized according to the genetic and epigenetic alterations and growth factor status. TMK-1 poorly differentiated adenocarcinoma cell line harbors mutant p53 tumor suppressor gene and rearrangement of p15MTS2. MKN-1 adenosquamous carcinoma line with mutant p53 reveals silencing of E-cadherin by promoter CpG hypermethylation. MKN-7 well-differentiated adenocarcinoma cell line has amplification of c-erbB2 oncogene and cyclin E gene. MKN-28 well-differentiated adenocarcinoma cell line reveals mutations in p53 and APC tumor suppressor genes and silencing of CD44. The MKN-45 poorly differentiated adenocarcinoma cell line with wild-type p53 is characterized by homozygous deletion of p16CDKN2/MTS1/INK4A and p15MTS2, amplification of c-met oncogene and promoter mutation of E-cadherin. MKN-74 derived from moderately differentiated tubular adenocarcinoma has wild-type p53. KATO-III signet ring cell carcinoma line has genomic deletion of p53, amplification of K-sam and c-met oncogene and mutation of E-cadherin. HSC-39 signet ring cell carcinoma cell line harboring p53 missense mutation has homozygous deletion of p16CDKN2/MTS1/INK4A and p15MTS2, amplifications of c-myc, c-met, K-sam and CD44 gene and mutation in beta-catenin gene.