Transcriptional profile of breast cancer cell lines after treatment with docetaxel.

Among microtubule-targeting agents, docetaxel has received recent interest owing to its good therapeutic index. Clinical trials have underlined its potential for the treatment of advanced breast cancer, although little is known about its molecular mode of action in this context. We characterized the molecular changes induced by docetaxel in two well-known human breast carcinoma cell lines. Two mechanisms of action according to drug concentration were suggested by a biphasic sensitivity curve, and were further validated by cell morphology, cell cycle and cell death changes. Two to four nanomolar docetaxel induced aberrant mitosis followed by late necrosis, and 100 nM docetaxel induced mitotic arrest followed by apoptosis. Passing through mitosis phase was a requirement for hypodiploidy to occur, as shown by functional studies in synchronized cells and by combining docetaxel with the proteasome inhibitor MG132. Transcriptional profiling showed differences according to cell line and docetaxel concentration, with cell cycle, cell death and structural genes commonly regulated in both cell lines. Although p53 targets were mainly induced with low concentration of drug in MCF7 cells, its relevance in the dual mechanism of docetaxel cytotoxicity was ruled out by using an isogenic shp53 cell line. Many of the genes shown in this study may contribute to the dual mechanism by which docetaxel inhibits the growth of breast cancer cells at different concentrations. These findings provide a basis for rationally enhancing docetaxel therapy, considering lower concentrations, and better drug combinations. Keywords: Dose-response analysis MCF7 and MDA-MB-231 breast cancer cells lines were treated with low (2-4 nM) and high (100 nM) concentrations of docetaxel. After 8, 24, and 48 hours of exposure total RNA was extracted, amplified and hybridized onto CNIO Oncochip cDNA microarrays. Untreated control cells were used for normalization of duplicate arrays.

在微管靶向药物（microtubule-targeting agents）中，多西他赛（docetaxel）凭借优异的治疗指数近年来受到广泛关注。临床试验已证实其在晚期乳腺癌治疗中的潜力，但目前对其在此场景下的分子作用机制仍知之甚少。
本研究对两种经典人类乳腺癌细胞系中多西他赛诱导的分子变化进行了表征。双相敏感性曲线提示存在两种依赖药物浓度的作用机制，并通过细胞形态、细胞周期及细胞死亡变化得到进一步验证。
2~4纳摩尔浓度的多西他赛可诱导异常有丝分裂，随后引发迟发性坏死；而100纳摩尔浓度的多西他赛则会导致有丝分裂阻滞，继而触发细胞凋亡。
同步化细胞功能实验以及多西他赛与蛋白酶体抑制剂MG132联合处理的结果均证实，进入有丝分裂期是发生亚二倍体的必要条件。
转录谱分析结果显示，不同细胞系及多西他赛浓度下的基因表达存在差异，但细胞周期、细胞死亡及结构相关基因在两种细胞系中均存在共同调控现象。
尽管在MCF7细胞中，p53靶基因主要在低浓度药物处理时被诱导，但通过使用同基因shp53细胞系的实验排除了p53在多西他赛双重细胞毒性机制中的核心作用。
本研究鉴定的诸多基因或可解释多西他赛在不同浓度下抑制乳腺癌细胞增殖的双重机制。上述发现为优化多西他赛治疗策略提供了理论依据，包括采用更低给药浓度及优化联合用药方案。
关键词：剂量反应分析；将MCF7与MDA-MB-231乳腺癌细胞系分别以低浓度（2~4 nM）和高浓度（100 nM）多西他赛处理。于给药后8、24及48小时提取总RNA，经扩增后与CNIO Oncochip cDNA微阵列进行杂交。以未处理的对照细胞用于重复阵列的归一化分析。