Reduced expression of p27 is a novel mechanism of docetaxel resistance in breast cancer cells

Abstract

Introduction: Docetaxel is one of the most effective chemotherapeutic agents in the treatment of breast cancer. Breast cancers can have an inherent or acquired resistance to docetaxel but the causes of this resistance remain unclear. However, apoptosis and cell cycle regulation are key mechanisms by which most chemotherapeutic agents exert their cytotoxic effects.

Methods: We created two docetaxel-resistant human breast cancer cell lines (MCF-7 and MDA-MB-231) and performed cDNA microarray analysis to identify candidate genes associated with docetaxel resistance. Gene expression changes were validated at the RNA and protein levels by reverse transcription PCR and western analysis, respectively.

Results: Gene expression cDNA microarray analysis demonstrated reduced p27 expression in docetaxel-resistant breast cancer cells. Although p27 mRNA expression was found to be reduced only in MCF-7 docetaxel-resistant sublines (2.47-fold), reduced expression of p27 protein was noted in both MCF-7 and MDA-MB-231 docetaxel-resistant breast cancer cells (2.83-fold and 3.80-fold, respectively).

Conclusions: This study demonstrates that reduced expression of p27 is associated with acquired resistance to docetaxel in breast cancer cells. An understanding of the genes that are involved in resistance to chemotherapy may allow further development in modulating drug resistance, and may permit selection of those patients who are most likely to benefit from such therapies.

Figure 1

Modulation of p27 mRNA expression in docetaxel-resistant breast cancer cells. Total RNA extracted from each cell line was amplified using specific oligonucleotide primer sequences and separated by agarose gel electrophoresis. The p27 and 18S internal control amplified PCR products produced fragment sizes of 253 bp and 489 bp, respectively. Lane 1, DNA size standards; lane 2, MCF-7 docetaxel-resistant subline; lane 3, MCF-7 breast cancer cells; lane 4, MDA-MB-231 breast cancer cells; lane 5, MDA-MB-231 docetaxel-resistant subline.

Figure 2

Modulation of p27 protein expression in docetaxel-resistant breast cancer cells. Protein extracted from each cell line was separated by PAGE, transferred to nitrocellulose membranes and probed with anti-human p27 and β-actin antibodies. Lane 1, MCF-7 breast cancer cells; lane 2, MCF-7 docetaxel-resistant subline; lane 3, MDA-MB-231 breast cancer cells; lane 4, MDA-MB-231 docetaxel-resistant subline.