Ron Receptor Tyrosine Kinase Signaling in Breast Cancer

描述（由申请人提供）： 罗恩受体酪氨酸激酶最近已被证明是过度表达和激活的一组人类癌症，最引人注目的数据发现在乳腺癌。罗恩在大约50%的人类乳腺癌中过表达，并且已经被证明是患有这种疾病的女性中转移和预后不良的独立预测因子。虽然罗恩过表达似乎是人类乳腺癌和转移的一个重要因素，但我们对罗恩在乳腺肿瘤中激活的信号通路以及这些通路对整体肿瘤生长和转移性扩散的重要性的认识存在重大差距。我们实验室的研究表明，罗恩在乳腺上皮中的选择性过表达导致雌性小鼠的乳腺肿瘤发生率为100%，并且与乳腺癌向肝脏和肺部的高度转移相关。在该模型中，乳腺肿瘤发生与罗恩的表达升高、受体磷酸化增加和罗恩相关酪氨酸激酶活性增加相关。然而，在这个模型中，罗恩配体，肝细胞生长因子样蛋白（HGFL）的依赖程度仍然未知。对这些小鼠的原发性乳腺肿瘤的生化分析表明，罗恩过表达与酪氨酸磷酸化2-连环蛋白水平升高和下游2-连环蛋白靶基因上调相关。与罗恩相似，2-连环蛋白表达也与乳腺癌患者的不良预后相关。此外，我们的初步数据显示，在人类乳腺癌标本中高罗恩表达和高2-连环蛋白水平之间存在关联。我们还发现，人乳腺癌细胞系中HGFL诱导的罗恩激活诱导2-连环蛋白的快速积累和2-连环蛋白的酪氨酸磷酸化，并转移到细胞核中。基于我们的初步和已发表的数据，我们假设配体介导的罗恩激活增强了2-连环蛋白信号传导，导致乳腺癌生长、生存和转移扩散增加。为了验证这一假设，提出了三个具体的目的：（i）描述罗恩配体HGFL在体内乳腺癌细胞生长、存活和转移扩散中的作用;（ii）检测2-连环蛋白表达在Ron介导的乳腺肿瘤发生和转移中的体内生物学意义，和（iii）确定HGFL诱导的，乳腺癌细胞中ron依赖的2-catenin酪氨酸磷酸化。总之，本提案中概述的研究将是第一个直接测试肿瘤发生过程中配体诱导的罗恩受体激活的意义以及定义乳腺癌中罗恩和2-连环蛋白信号通路之间的分子串扰的研究。我们将利用最先进的体内方法，包括在罗恩癌基因过表达的情况下，2-连环蛋白的条件性丢失，或罗恩配体的基因靶向丢失，来分析临床相关小鼠模型中自发性乳腺肿瘤形成和转移。

DESCRIPTION (provided by applicant): The Ron receptor tyrosine kinase has recently been shown to be overexpressed and activated in a cohort of human cancers, with the most compelling data found in breast cancer. Ron is overexpressed in approximately 50% of human breast cancers, and has been shown to be an independent predictor of both metastases and poor prognosis in women with this disease. While Ron overexpression appears to be an important factor in human breast cancer and metastasis, a significant gap exists in our knowledge about the signaling pathways that Ron activates in breast tumors, and about the importance of these pathways with respect to overall tumor growth and metastatic dissemination. Studies from our laboratory have shown that Ron overexpression selectively in the mammary epithelium leads to mammary tumorigenesis in female mice with 100% incidence and is associated with a high degree of breast cancer metastasis to the liver and lungs. In this model, mammary tumorigenesis is associated with elevated expression of Ron, increased receptor phosphorylation and increased Ron-associated tyrosine kinase activity. However, the degree of dependency of the Ron ligand, hepatocyte growth factor-like protein (HGFL), in this model remains unknown. Biochemical analyses of primary mammary tumors from these mice demonstrate that Ron overexpression is associated with elevated levels of tyrosine phosphorylated 2-catenin and with the upregulation of downstream 2-catenin target genes. Similar to Ron, 2-catenin expression is also associated with poor prognosis in breast cancer patients. Moreover, our preliminary data show an association between high Ron expression and high2-catenin levels in human breast cancer specimens. We also show that HGFL-induced Ron activation in human breast cancer cell lines induces rapid 2-catenin accumulation and tyrosine phosphorylation of2-catenin that is translocated to the nucleus. Based on our preliminary and published data we hypothesize that ligand-mediated activation of Ron augments 2-catenin signaling, leading to increased breast cancer growth, survival, and metastatic dissemination. To test this hypothesis three Specific Aims are proposed: (i) to delineate the role of the Ron ligand, HGFL, in breast cancer cell growth, survival and metastatic dissemination in vivo; (ii) to examine the in vivo biological significance of2-catenin expression in Ron-mediated mammary tumorigenesis and metastasis, and (iii) to determine the association, mechanism, and biological relevance of HGFL-induced, Ron-dependent 2-catenin tyrosine phosphorylation in breast cancer cells. In total, the studies outlined in this proposal will be the first to directly test the significance of ligand-induced Ron receptor activation during tumorigenesis as well as to define the molecular cross-talk between the Ron and 2-catenin signaling pathways in breast cancer. We will utilize cutting edge in vivo approaches involving the conditional loss of2-catenin, or a gene targeted loss of the Ron ligand, in the presence of Ron oncogene overexpression to analyze spontaneous breast tumor formation and metastasis in a clinically relevant murine model.