INDUCTION OF ANTIESTROGEN RESISTANCE IN HUMAN BREAST-CANCER CELLS BY RANDOM INSERTIONAL MUTAGENESIS USING DEFECTIVE RETROVIRUSES - IDENTIFICATION OF BCAR-1, A COMMON INTEGRATION SITE
INDUCTION OF ANTIESTROGEN RESISTANCE IN HUMAN BREAST-CANCER CELLS BY RANDOM INSERTIONAL MUTAGENESIS USING DEFECTIVE RETROVIRUSES - IDENTIFICATION OF BCAR-1, A COMMON INTEGRATION SITE
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DOI：
10.1210/me.7.7.870
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发表时间：
1993-07-01
影响因子：
--
通讯作者：
KOK, EM
中科院分区：
文献类型：
--
作者：
DORSSERS, LCJ;VANAGTHOVEN, T;KOK, EM
由于抗雌激素耐药肿瘤的发展，转移性乳腺癌对抗雌激素治疗的反应持续时间有限。所涉及的机制尚不清楚，但可能源于肿瘤细胞内的（表观）遗传改变。我们利用复制缺陷型逆转录病毒进行体外随机插入诱变，以确定那些在人类乳腺癌细胞抗雌激素耐药性发展中起关键作用的基因。从 7 x 10(8) 雌激素依赖性 ZR-75-1 细胞中分离出 80 个抗雌激素抗性细胞克隆，用缺陷逆转录病毒大量感染并进行 4-OH-他莫昔芬选择。整合位点特异性 DNA 探针通过反向聚合酶链式反应技术制备，用于寻找常见的整合位点。在单个基因座中以相同方向鉴定了六个细胞克隆，该基因组被逆转录病毒基因组整合，指定为乳腺癌抗雌激素耐药基因座-1 (bcar-1)。这些bcar-1细胞克隆失去了雌激素受体表达，并且变得不依赖雌激素。我们的结果强烈表明，bcar-1 位点的改变是人乳腺癌细胞在体外产生抗雌激素耐药性的原因。此外，我们还表明，使用有缺陷的逆转录病毒进行体外插入诱变可应用于人类细胞中的基因标记。
Duration of response to antiestrogen therapy in metastatic breast cancer is limited due to the development of antiestrogen-resistant tumors. The mechanisms involved are not understood but could originate from (epi)genetic alterations within the tumor cells. We have applied in vitro random insertional mutagenesis with replication defective retroviruses to identify those genes playing a key role in development of antiestrogen resistance in human breast cancer cells. Eighty antiestrogen-resistant cell clones were isolated f rom 7 x 10(8) estrogen-dependent ZR-75-1 cells, mass-infected with defective retroviruses and subjected to 4-OH-tamoxifen selection. Integration site-specific DNA probes were made by inverse polymerase chain reaction techniques and used to search for common integration sites. Six cell clones were identified with retroviral genome integrations in the same orientation in a single locus, designated breast cancer antiestrogen resistance locus-1 (bcar-1). These bcar-1 cell clones had lost estrogen receptor expression and had become estrogen independent. Our results strongly suggest that alteration of the bcar-1 locus is responsible for development of antiestrogen resistance in human breast cancer cells in vitro. In addition, we have shown that in vitro insertional mutagenesis using defective retroviruses can be applied for gene tagging in human cells.