Identification of predictive factors of adjuvant chemotherapy for breast cancer

背景：紫杉烷类药物是治疗乳腺癌的重要药物。这些药物与微管蛋白结合，抑制纺锤体微管动力学，导致细胞周期停滞于G2/M期，继而发生细胞凋亡。然而，对紫杉烷治疗的抗药性阻碍了一些患者从这些药物中受益。紫杉烷耐药的几种机制已被描述，包括微管相关蛋白tau(MAPT)的参与。MAPT与微管中的紫杉烷结合在同一个口袋上，并阻碍药物的功能。雌激素受体(ER)是一种转录因子，在乳腺癌的发生发展中起重要作用。然而，ER和MAPT在乳腺癌中的关系尚不完全清楚。在这项研究中，我们研究了MAPT的表达与人乳腺癌细胞对紫杉烷类药物敏感性的相关性，以及ER和MAPT在这些细胞中的蛋白水平的关系。我们还检查了组合Thera…更多的是激素药物和紫杉烷。方法：采用实时定量聚合酶链式反应、免疫印迹分析和MTS法检测12株人乳腺癌细胞株中MAPT的表达与紫杉烷类药物敏感性的关系。用小干扰RNA(SiRNA)抑制MAPT的表达后，用MTS法、流式细胞仪和免疫荧光法检测细胞对紫杉烷类药物敏感性的变化。为了研究ER和MAPT之间的关系，用小干扰RNA抑制MAPT和ER阳性细胞(分别为MCF7和ZR75.1)的ER表达或用17-β-雌二醇刺激ER的表达。用激素药物(他莫昔芬和ICI182,780)处理细胞，检测MAPT蛋白表达的变化。结果：6个细胞株MAPT mRNA高表达，4个细胞株MAPT蛋白高表达，即MAPT在mRNA水平和蛋白质水平的表达并不总是相关。MAPT基因表达与紫杉烷耐药性无关，但70 kDa以下MAPT蛋白亚型的表达与紫杉烷耐药性相关。MAPT的下调增加了对紫杉烷的敏感性。ER下调MAPT蛋白表达，17-β-雌二醇刺激MAPT蛋白表达增加。他莫昔芬可使MAPT蛋白水平升高，而IC1182,780可使MAPT蛋白水平降低。紫杉烷类化合物与ICI182,780联合作用显示出较强的协同作用，而与他莫昔芬类似的处理对两种细胞株均有拮抗作用。结论：70 kDa以下MAPT蛋白亚型的表达与乳腺癌细胞对紫杉烷的耐药性有关。MAPT在乳腺癌中的表达受ER的影响，ICI182,780是一种选择性ER抑制剂，可以逆转MAPT和ER阳性乳腺癌对紫杉烷的耐药性。较少

Background : Taxanes are important drugs in treatment of breast cancer. These drugs bind to tubulin and suppress spindle microtubule dynamics, which leads to cell cycle arrest in the G2/M phase followed by apoptosis. However, resistance to taxane therapy prevents some patients from benefiting from these drugs. Several mechanisms of taxane resistance have been described, including the involvement of microtubule-associated protein-tau (MAPT). MAPT binds to the same pocket as taxanes in microtubules and obstructs the function of the drug. Estrogen receptors (ER) are transcriptional factors that play an important role in the development and progression of breast cancer. However, the relationship between ER and MAPT in breast cancer is not entirely clear. In this study we examined the correlation between MAPT expression and the sensitivity of human breast cancer cells to taxanes, and the relationship between ER and MAPT at the protein level in these cells. We also examined combination thera … More py with hormone drugs and taxanes. Methods : The correlation between MAPT expression and sensitivity to taxanes was examined in 12 human breast cancer cell lines using real time PCR, western blotting analysis and an MTS assay. Following small interfering RNA (siRNA) knockdown of MAPT expression, the alteration of cellular sensitivity to taxanes was examined by MTS assay, flow cytometry and immunofluorescence. To examine the relationship between ER and MAPT, ER expression was knocked down with siRNA or stimulated with 17-β-estradiol in MAPT- and ER-positive cell lines (MCF-7 and ZR75.1, respectively). The cells were also treated with hormone drugs (tamoxifen and ICI182,780) and changes in MAPT protein expression were examined. Results : Six cell lines showed high MAPT mRNA expression and four showed high MAPT protein expression ; that is, expression at the mRNA level did not always correlate with that at the protein level. MAPT mRNA expression did not correlate with taxane resistance, but expression of MAPT protein isoforms under 70kDa correlated with taxane resistance. Downregulation of MAPT increased sensitivity to taxanes. MAPT protein expression was decreased by ER knockdown and increased by 17-β-estradiol stimulation. The MAPT protein level was also increased by tamoxifen, but decreased by IC1182,780. Combination treatment of taxanes with ICI182,780 showed a strong synergistic effect, but similar treatment with tamoxifen had an antagonistic effect in both cell lines. Conclusions : Expression of MAPT protein isoforms under 70kDa correlates with taxane resistance in breast cancer cells. MAPT expression is influenced by ER in breast cancer and ICI182,780, a selective ER inhibitor, can reverse the resistance to taxanes in both MAPT- and ER-positive breast cancer. Less