van Agthoven Ton, Dorssers Lambert C J, Lehmann Ulrich, Kreipe Hans, Looijenga Leendert H J, Christgen Matthias
Department of Pathology, Erasmus MC Cancer Institute, Rotterdam, the Netherlands.
Institute of Pathology, Hannover Medical School, Hannover, Germany.
PLoS One. 2015 Aug 28;10(8):e0136845. doi: 10.1371/journal.pone.0136845. eCollection 2015.
Most breast cancers depend on estrogenic growth stimulation. Functional genetic screenings in in vitro cell models have identified genes, which override growth suppression induced by anti-estrogenic drugs like tamoxifen. Using that approach, we have previously identified Breast Cancer Anti-Estrogen Resistance 4 (BCAR4) as a mediator of cell proliferation and tamoxifen-resistance. Here, we show high level of expression and function of BCAR4 in human breast cancer.
BCAR4 mRNA expression was evaluated by (q)RT-PCR in a panel of human normal tissues, primary breast cancers and cell lines. A new antibody raised against C78-I97 of the putative BCAR4 protein and used for western blot and immunoprecipitation assays. Furthermore, siRNA-mediated gene silencing was implemented to study the function of BCAR4 and its downstream targets ERBB2/3.
Except for placenta, all human normal tissues tested were BCAR4-negative. In primary breast cancers, BCAR4 expression was comparatively rare (10%), but associated with enhanced proliferation. Relative high BCAR4 mRNA expression was identified in IPH-926, a cell line derived from an endocrine-resistant lobular breast cancer. Moderate BCAR4 expression was evident in MDA-MB-134 and MDA-MB-453 breast cancer cells. BCAR4 protein was detected in breast cancer cells with ectopic (ZR-75-1-BCAR4) and endogenous (IPH-926, MDA-MB-453) BCAR4 mRNA expression. Knockdown of BCAR4 inhibited cell proliferation. A similar effect was observed upon knockdown of ERBB2/3 and exposure to lapatinib, implying that BCAR4 acts in an ERBB2/3-dependent manner.
BCAR4 encodes a functional protein, which drives proliferation of endocrine-resistant breast cancer cells. Lapatinib, a clinically approved EGFR/ERBB2 inhibitor, counteracts BCAR4-driven tumor cell growth, a clinical relevant observation.
大多数乳腺癌依赖雌激素刺激生长。体外细胞模型中的功能基因筛选已鉴定出一些基因，这些基因可克服他莫昔芬等抗雌激素药物诱导的生长抑制。通过该方法，我们之前已鉴定出乳腺癌抗雌激素耐药4（BCAR4）是细胞增殖和他莫昔芬耐药的介质。在此，我们展示了BCAR4在人类乳腺癌中的高表达水平及功能。
通过（定量）逆转录聚合酶链反应（qRT-PCR）评估BCAR4 mRNA在一组人类正常组织、原发性乳腺癌及细胞系中的表达。制备了一种针对假定的BCAR4蛋白C78-I97区域的新抗体，并用于蛋白质印迹和免疫沉淀分析。此外，采用小干扰RNA（siRNA）介导的基因沉默来研究BCAR4及其下游靶点ERBB2/3的功能。
除胎盘外，所有检测的人类正常组织均为BCAR4阴性。在原发性乳腺癌中，BCAR4表达相对少见（10%），但与增殖增强相关。在源自内分泌耐药性小叶乳腺癌的IPH-926细胞系中鉴定出相对较高的BCAR4 mRNA表达。在MDA-MB-134和MDA-MB-453乳腺癌细胞中可见中等水平的BCAR4表达。在具有异位（ZR-75-1-BCAR4）和内源性（IPH-926、MDA-MB-453）BCAR4 mRNA表达的乳腺癌细胞中检测到了BCAR4蛋白。敲低BCAR4可抑制细胞增殖。敲低ERBB2/3并暴露于拉帕替尼后观察到类似效果，这表明BCAR4以ERBB2/3依赖的方式发挥作用。
BCAR4编码一种功能性蛋白，其驱动内分泌耐药性乳腺癌细胞的增殖。临床上已获批的表皮生长因子受体（EGFR）/ERBB2抑制剂拉帕替尼可对抗BCAR4驱动的肿瘤细胞生长，这是一项具有临床相关性的观察结果。
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