Cyclooxygenase-2 induces genomic instability, BCL2 expression, doxorubicin resistance, and altered cancer-initiating cell phenotype in MCF7 breast cancer cells
Cyclooxygenase-2 induces genomic instability, BCL2 expression, doxorubicin resistance, and altered cancer-initiating cell phenotype in MCF7 breast cancer cells
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DOI：
10.1016/j.jss.2008.02.026
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发表时间：
2008-06-15
影响因子：
2.2
通讯作者：
Lucci, Anthony
中科院分区：
文献类型：
--
作者：
Singh, Balraj;Cook, Kendra R.;Lucci, Anthony
导论.原发性乳腺癌中环氧合酶（COX 2）表达与预后不良相关。我们以前报道过，COX 2在MCF 10A乳腺上皮细胞的基底亚型表达诱导基因组不稳定性。为了了解COX 2在雌激素受体阳性（非基底）乳腺癌中的作用，我们用COX 2转染MCF 7细胞系，并分析其染色体特征、BCL 2蛋白表达和对阿霉素的耐药性。我们还分析了作为乳腺球生长的细胞培养物，以确定COX 2表达是否影响MCF 7细胞中的癌症起始（“干”）细胞表型。用pTRE 2 pur-COX 2或pTRE 2 pur-COX 2-GFP稳定转染MCF 7泰特On细胞（获自Clontech）以分别产生COX 2或COX 2-GFP蛋白。Western blotting检测BCL 2蛋白表达。MTT法检测细胞对药物的敏感性。使用卡方检验比较各组。我们通过Giemsa染色后对对照和COX 2转染的中期阻滞MCF 7细胞进行染色体分析来分析基因组不稳定表型。我们用克隆形成试验评估了作为乳腺球生长的细胞的致瘤潜力。早期传代COX 2转染的MCF 7细胞的细胞遗传学分析表明，与亲本MCF 7细胞相比，具有显著的基因组不稳定性。COX 2过表达与染色体畸变（染色单体断裂、染色体融合、C后期）的显著增加相关。COX 2转染的MCF 7细胞产生的抗凋亡蛋白BCL 2的水平显着高于亲本细胞。在一项功能测定中，我们发现COX 2表达与阿霉素耐药性增加相关。在确定细胞致瘤潜力的补充方法中，我们发现COX 2增加了MCF 7细胞在培养物中作为乳腺球生长的能力，这与克隆形成效率的增加相关。我们发现MCF 7乳腺癌细胞中COX 2的表达诱导了基因组不稳定性、BCL 2表达和阿霉素耐药，从而使其显著更具致瘤性。这些数据表明，考克斯-2可能是乳腺癌治疗的一个重要靶点。(C)2008年爱思唯尔公司All rights reserved.
Introduction. Cyclooxygenase (COX2) expression in primary breast cancer correlates with a worse prognosis. We reported previously that COX2 expression in MCF10A breast epithelial cells of basal subtype induces genomic instability. To understand the role of COX2 in estrogen receptor-positive (non-basal) breast cancer, we transfected the MCF7 cell line with COX2 and analyzed its chromosomal profile, BCL2 protein expression, and resistance to doxorubicin. We also analyzed cell cultures grown as mammospheres to determine whether COX2 expression affects the cancer-initiating ("stem") cell phenotype in MCF7 cells.Methods. MCF7 Tet On cells (obtained from Clontech) were stably transfected with pTRE2pur-COX2 or pTRE2pur-COX2-GFP to produce COX2 or COX2-GFP protein, respectively. BCL2 protein was detected by Western blotting. Sensitivity of cells to drug treatment was analyzed by MTT assay. Groups were compared using Chi-Square test. We analyzed the genomic instability phenotype by chromosome analysis of control and COX2 transfected metaphase-arrested MCF7 cells after Giemsa staining. We assessed the tumorigenic potential of cells grown as mammospheres with a clonogenic assay.Results. Cytogenetic analysis of early passage COX2 transfected MCF7 cells demonstrated significant genomic instability as compared to parental MCF7 cells. COX2 overexpression was associated with a significant increase in chromosomal aberrations (chromatid breaks, chromosome fusions, C anaphase). COX2 transfected MCF7 cells produced a significantly higher level of the anti-apoptotic protein BCL2 than the parental cells. In a functional assay, we found that COX2 expression correlated with increased resistance to doxorubicin. In a complimentary approach to determine tumorigenic potential of cells, we found that COX2 increased the ability of MCF7 cells to grow as mammospheres in culture, which correlated with an increase in clonogenic efficiency.Conclusions. We found that COX2 expression in MCF7 breast cancer cells induced genomic instability, BCL2 expression, and doxorubicin resistance, thus making them significantly more tumorigenic. This data suggests that COX-2 may be an important target for breast cancer treatment. (C) 2008 Elsevier Inc. All rights reserved.