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

BACKGROUND: 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 sile...

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Detalles Bibliográficos
Autores principales: Yang, Rui, Li, Wei-Wei, Hoang, Bang H, Kim, Hansoo, Banerjee, Debabrata, Kheradpour, Albert, Healey, John H, Meyers, Paul A, Bertino, Joseph R, Gorlick, Richard
Formato: Texto
Lenguaje:English
Publicado: BioMed Central 2008
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2387170/
https://www.ncbi.nlm.nih.gov/pubmed/18452618
http://dx.doi.org/10.1186/1471-2407-8-124
Descripción
Sumario:BACKGROUND: 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. METHODS: 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. RESULTS: A 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. CONCLUSION: 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.