Muc4 Involvement in Mammary Tumor Progression

描述（由申请方提供）：拟定研究的总体目的是阐明膜粘蛋白Muc4在促进乳腺肿瘤发生和进展中的作用。当在培养的细胞中表达时，Muc4具有有效的抗粘附和生长信号传导活性，其原则上可以在健康上皮细胞向早期增生表型的初始转化和向高度侵袭性和转移性状态的后期转化中发挥作用。与此一致，据报道，与患者匹配的正常组织样品相比，Muc4在高达20%的局部乳腺肿瘤中过表达和错误定位，并且相对于匹配的原发性肿瘤，在高达40%的淋巴结转移中过表达和错误定位。该项目将通过分析Muc4对体外和体内逐渐更具侵略性状态发展的贡献来整合这些观察结果。驱动假设是乳腺肿瘤中Muc4蛋白的异常表达促进了其向恶性肿瘤的进展。目的1将分析一系列来自MCF10A细胞的同基因培养乳腺上皮细胞系中诱导型Muc4过表达的结果，这些细胞模拟正常、增生和恶性乳腺组织，以及模拟ErbB2阳性、ER阳性和三阴性疾病的乳腺肿瘤细胞系。当细胞在2D和3D培养物中生长时，将进行细胞生长特性和信号传导通路使用的分析。全长Muc4与缺乏其VNTR结构域的肿瘤相关剪接变体的比较将揭示Muc4抗粘附活性在调节细胞生长特性中的作用。在Ai 2中，Muc4将在转基因小鼠的乳腺中诱导表达，并且这些将被杂交到乳腺癌小鼠模型中。在肿瘤发展的两个不同点诱导Muc4表达将允许在疾病的不同阶段体内评估Muc4对肿瘤进展的贡献。还将分析Muc4敲除对小鼠乳腺肿瘤发生和进展的影响。目的1和2的研究将共同确定在患者肿瘤中观察到的Muc4过表达是否有助于乳腺癌进展。目的3将检查患者样本中Muc4失调的机制，并将开始评估一种新的内质网定位的、TGF β调节的蛋白质降解途径在控制细胞Muc4水平中的作用。总的来说，这些研究将验证Muc4作为乳腺癌进展的新贡献者，并将开始解开可能最终用于治疗干预的生化途径。

DESCRIPTION (provided by applicant): The overall purpose of the proposed studies is to elucidate the role of the membrane mucin Muc4 in promoting breast tumor initiation and progression. Muc4 harbors potent anti-adhesive and growth signaling activities when expressed in cultured cells, which in principle could play roles in both the initial transformation of health epithelia to early hyperplastic phenotypes and in later transitions to highly invasive and metastatic states. Consistent with this, it has been reported that Muc4 is overexpressed and mislocalized in up to 20% of local breast tumors compared to patient-matched normal tissue samples, and up to 40% of lymph node metastases relative to matched primary tumor. This project will integrate these observations by providing an analysis of Muc4 contribution to the development of progressively more aggressive states in vitro and in vivo. The driving hypothesis is that the aberrant expression of Muc4 protein in breast tumors facilitates their progression to malignancy. Aim 1 will analyze the outcome of inducible Muc4 overexpression in a series of isogenic cultured breast epithelial cell lines derived from MCF10A cells that model normal, hyperplastic, and malignant breast tissue, as well as in breast tumor cell lines that model ErbB2-positive, ER-positive and triple-negative disease. Analysis of cellular growth properties and signaling pathway usage will be carried out when cells are grown in 2D and 3D culture. Comparison of full-length Muc4 with a tumor-associated splice variant lacking its VNTR domain will reveal the role of Muc4 anti-adhesive activity in regulating cellular growth properties. In Ai 2, Muc4 will be inducibly expressed in the mammary glands of transgenic mice, and these will be crossed into mouse models of breast cancer. Induction of Muc4 expression at two different points in tumor development will permit the in vivo assessment of Muc4 contribution to tumor progression at different stages of disease. The impact of Muc4 knockout on mouse mammary tumor development and progression will also be analyzed. Together the studies of Aims 1 and 2 will determine whether the Muc4 overexpression observed in patient tumors contributes to breast cancer progression. Aim 3 will examine the mechanisms of Muc4 dysregulation in patient samples, and will begin to assess the role of a novel endoplasmic reticulum-localized, TGFß-regulated protein degradation pathway in governing cellular Muc4 levels. Overall, these studies will validate Muc4 as a novel contributor to breast cancer progression, and will begin to unravel biochemical pathways that might ultimately be exploited for therapeutic intervention.