Quantitative correlation between promoter methylation and messenger RNA levels of the reduced folate carrier

• Main Authors: Kheradpour Albert, Banerjee Debabrata, Kim Hansoo, Hoang Bang H, Li Wei-Wei, Yang Rui, Healey John H, Meyers Paul A, Bertino Joseph R, Gorlick Richard
• Format: Article
• Language: English
• Published: BMC 2008-05-01
• Series: BMC Cancer
• Online Access: http://www.biomedcentral.com/1471-2407/8/124

<p>Abstract</p> <p>Background</p> <p>Methotrexate (MTX) uptake is mediated by the reduced folate carrier (RFC). Defective drug uptake in association with decreased RFC expression is a common mechanism of MTX resistance in many tumor types. Heavy promoter methylation was previously identified as a basis for the complete silencing of RFC in MDA-MB-231 breast cancer cells, its role and prevalence in RFC transcription regulation are, however, not widely studied.</p> <p>Methods</p> <p>In the current study, RFC promoter methylation was assessed using methylation specific PCR in a panel of malignant cell lines (n = 8), including MDA-MB-231, and M805, a MTX resistant cell line directly established from the specimen of a patient with malignant fibrohistocytoma, whom received multiple doses of MTX. A quantitative approach of real-time PCR for measuring the extent of RFC promoter methylation was developed, and was validated by direct bisulfite genomic sequencing. RFC mRNA levels were determined by quantitative real-time RT-PCR and were related to the extent of promoter methylation in these cell lines.</p> <p>Results</p> <p>A partial promoter methylation and RFC mRNA down-regulation were observed in M805. Using the quantitative approach, a reverse correlation (correlation coefficient = -0.59, <it>p </it>< 0.05) was identified between the promoter methylation and RFC mRNA levels in this a panel of malignant cell lines.</p> <p>Conclusion</p> <p>This study further suggests that promoter methylation is a potential basis for MTX resistance. The quantitative correlation identified in this study implies that promoter methylation is possibly a mechanism involved in the fine regulation of RFC transcription.</p>