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BMC Cancer

, 8:124

First Online: 01 May 2008Received: 10 December 2007Accepted: 01 May 2008DOI: 10.1186-1471-2407-8-124

Cite this article as: Yang, R., Li, WW., Hoang, B.H. et al. BMC Cancer 2008 8: 124. doi:10.1186-1471-2407-8-124

Abstract

BackgroundMethotrexate 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.

MethodsIn 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.

ResultsA partial promoter methylation and RFC mRNA down-regulation were observed in M805. Using the quantitative approach, a reverse correlation correlation coefficient = -0.59, p < 0.05 was identified between the promoter methylation and RFC mRNA levels in this a panel of malignant cell lines.

ConclusionThis 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.

AbbreviationsRFCreduced folate carrier

MTXmethotrexate

DHFRdihydrofolate reductase

MSPmethylation specific PCR.

Electronic supplementary materialThe online version of this article doi:10.1186-1471-2407-8-124 contains supplementary material, which is available to authorized users.

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Autor: Rui Yang - Wei-Wei Li - Bang H Hoang - Hansoo Kim - Debabrata Banerjee - Albert Kheradpour - John H Healey - Paul A Mey

Fuente: https://link.springer.com/







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