Messenger RNA expression and methylation of candidate tumor-suppressor genes and risk of ovarian cancer-a case-control analysis

Abstract

To investigate the association of expression and promoter methylation of tumor-suppressor genes with risk of ovarian cancer, we conducted a case-control study of 102 patients with serous epithelial ovarian cancer and 100 patients without ovarian cancers. We measured mRNA expression levels (by real-time reverse transcription polymerase chain reaction) and methylation status (by methylation-specific polymerase chain reaction) of five candidate genes (BRCA1, BRCA2, hMLH1, MGMT, and DNMT3B) in tumors from the cases and normal ovaries from the controls. We found that mRNA expression levels of the five genes were decreased in tumors than in normal ovaries with 0.39-fold for BRCA1, 0.25-fold for BRCA2, 0.42-fold for hMLH1, 0.45-fold for MGMT, and 0.87-fold for DNMT3B, calculated by the 2(-ΔΔCT) method. Ovarian cancer risk (odds ratios, ORs) was associated with low expression of all genes (2.95 [95% confidence interval (CI), 1.51 - 5.78] for BRCA1, 3.65 (95% CI, 1.82 - 7.30) for BRCA2, 5.25 (95% CI, 2.52 - 10.96) for hMLH1, and 4.72 (95% CI, 2.32 - 9.62) for MGMT) but not DNMT3B. However, methylation status was not associated with gene expression levels in the tumors, except for hMLH1 whose mean (± SD) gene expression was significantly lower in methylated (13.0 ± 7.6) than in unmethylated (31.2 ± 44.8) tumors (P < 0.001). We concluded that low mRNA expression of these tumor-suppressor genes, likely due to molecular mechanisms in addition to the promoter methylation in some instances, may be a biomarker for ovarian cancer risk in this study population. Larger studies are needed to validate our findings.

Figure 1

A, Methylation-specific PCR analysis of the methylation status in BRCA1, hMLHl, and MGMT. Representative PCR products of the promoter region of these genes were amplified by the MSP method. P, positive control (CpGnome Universal Methylated DNA); Ca, ovarian cancer tissues; Cn, normal ovarian tissues; N, negative control (water blank); M, methylated; U, unmethylated. B, Relative mRNA expression levels of methylated and unmethylated BRCA1, hMLHl, and MGMT in the cases.