Role of Cdc25A in breast cancer

描述（由申请人提供）：Cdc 25 A是一种双特异性蛋白磷酸酶，在乳腺癌中经常过表达，并与生存率低相关。该计划的长期目标是了解Cdc 25 A在乳腺癌中的确切作用，并为针对该蛋白质的抗癌疗法建立科学基础。Cdc 25 A通过在G1至S期间激活细胞周期蛋白E（A）/Cdk 2以及在G2/M边界激活细胞周期蛋白B（A）/Cdk 1来促进细胞周期进程。Cdc 25 A的异位表达缩短了通过G1的通道，并与H-ras在转化小鼠胚胎成纤维细胞中合作，而Cdc 25 A的沉默导致在G1和G2的停滞。我们已经表明，转化生长因子-β（TGF β），这是重要的形态发生和肿瘤抑制的乳腺，下调Cdc 25 A在转录和翻译后的方式。这些观察结果表明，Cdc 25 A可能是乳腺癌中的一个关键癌基因，虽然它还没有被证明在体内使用动物模型。该建议基于我们的以下初步数据：（i）我们产生的Cdc 25 A杂合敲除小鼠显示出对MMTV-ras诱导的乳腺肿瘤发生的显著抗性;（ii）MMTV-Cdc 25 A转基因与MMTV-ras转基因在鼠肿瘤发生中协同合作;（iv）TGF-β促进Smad 3-受体中Cdc 25 A蛋白降解。依赖的方式，该途径可能在MCF 7和其他人乳腺癌细胞系中存在缺陷;（iii）Cdc 25 A过表达抑制凋亡信号调节激酶-1（ASK 1）并降低细胞对氧化应激的敏感性。诱导凋亡。本提案中评估的中心假设是Cdc 25 A蛋白的缺陷性降解对乳腺癌的发生和/或进展具有限速作用，导致细胞周期进展和细胞凋亡的失控。具体目标是：（2）使用具有Cdc 25 A siRNA和四环素诱导表达的Cdc 25 A突变体的MCF-10a细胞，确定Cdc 25 A表达降低或Cdc 25 A稳定性改变如何影响细胞周期进程、细胞对氧化应激的敏感性以及响应ras和neu的恶性转化;（3）使用我们的新型Cdc 25 A基因敲除小鼠和MMTV-Cdc 25 A转基因小鼠系，确定Cdc 25 A表达改变如何影响MMTV-neu、MMTV-myc或MMTV-Wnt-1转基因小鼠的乳腺肿瘤发生。我们期望这些研究将为我们提供对Cdc 25 A稳定在乳腺癌发展中的作用的重要见解，并为开发针对癌基因的有效疗法提供分子手段。

DESCRIPTION (provided by applicant): Cdc25A, a dual-specific protein phosphatase, is frequently overexpressed in breast cancer, and associated with poor survival. The long-term goal of this program is to understand the exact role of Cdc25A in breast cancer and establish a scientific basis for anti-cancer therapies targeted on this protein. Cdc25A promotes cell cycle progression by activating cyclin E (A)/Cdk2 during G1 through S, and also cyclin B(A)/Cdk1 at the G2/M boundary. Ectopic expression of Cdc25A shortens the passage through G1 and cooperates with H-ras in transforming mouse embryonic fibroblasts, while silencing of Cdc25A results in arrest at both G1 and G2. We have shown that transforming growth factor-beta (TGFbeta), which is important for morphogenesis and tumor suppression of the mammary gland, downregulates Cdc25A in both transcriptional and posttranslational manners. These observations suggest that Cdc25A could be a critical oncogene in breast cancer, although it has not been demonstrated in vivo using animal models. This proposal is based on our following preliminary data: (i) Cdc25A-heterozygous knockout mice we generated show marked resistance to MMTV-ras induced mammary tumorigenesis; (ii) An MMTV-Cdc25A transgene synergistically cooperates with the MMTV-ras transgene in murine tumorigenesis; (iv) TGF-beta promotes Cdc25A protein degradation in a Smad3-dependent manner and this pathway could be defective in MCF7 and other human breast cancer cell lines; (iii) Cdc25A overexpression inhibits apoptosis signal-regulating kinase-1 (ASK1) and diminishes cellular sensitivity to oxidative stress-induced apoptosis. The central hypothesis evaluated in this proposal is that defective degradation of Cdc25A protein is rate-limiting for initiation and/or progression of breast cancer, causing deregulated control of cell cycle progression and apoptosis. The specific aims are: (1) Determine whether deregulated TGF-beta/Smad signaling is involved in Cdc25A stabilization in human breast cancer cell lines; (2) Determine how reduced Cdc25A expression or altered stability ofCdc25A affects cell cycle progression, cellular sensitivity to oxidative stress, and malignant transformation in response to ras and neu, using MCF-10a cells with Cdc25A siRNA and tetracycline-inducible expression ofCdc25A mutants; (3) Determine how altered Cdc25A expression affects breast tumorigenesis in mice with MMTV-neu, MMTV-myc or MMTV-Wnt-1 transgene, using our novel Cdc25A-knockout and MMTV-Cdc25A transgenic mouse lines. We expect that these studies will provide us with significant insight into the role of Cdc25A stabilization in breast cancer development and a molecular handle to develop effective therapies to target the oncogene.