Cripto Antagonism of Activin and TGF-Beta Signalling

激活素和 TGF-β 信号传导的 Cripto 拮抗作用

• 批准号: 6898291
• 负责人: WYLIE W. VALE
• 金额: $ 30.54万
• 依托单位: SALK INSTITUTE FOR BIOLOGICAL STUDIES
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-06-01 至 2009-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6898291
• 关键词: MCF7 cell; activins; biological signal transduction; breast neoplasms; cell proliferation; growth factor receptors; hormone inhibitor; laboratory rabbit; molecular oncology; neutralizing antibody; oncoproteins; phosphorylation; protein binding; protein structure function; transcription factor; transforming growth factors

DESCRIPTION (provided by applicant): TGF-B and activin are functionally related members of the TGF-Beta superfamily of growth and differentiation factors. TGF-Beta and activin activate Smad2 and Smad3 and they both regulate tissue homeostasis by potently inhibiting proliferation of multiple cell types, including epithelial cells. Disruption of the antiproliferative response to activin and/or TGF-Beta is associated with dysregulated cellular proliferation, oncogenesis and progression toward the malignant phenotype. Cripto is a developmental oncoprotein that is overexpressed in multiple human tumors including approximately 80% of human breast tumors. Cripto overexpression transforms mammary epithelial ceils and activates the growth promoting Erk/MAPK and PI3K signaling pathways. Cripto and related EGF-CFC proteins also play essential signaling roles during embryonic development and Cripto is required for mesoderm induction and cardiogenesis. It has been shown that Cripto acts as a coreceptor to facilitate receptor assembly and signaling via activin type II/I receptors by the TGF-Beta superfamily members nodal, Vgl and GDF1. This led us to test whether Cripto may regulate the ability of activin and possibly other TGF-Beta ligands to assemble functional receptor complexes and we showed that Cripto binds activin in the presence of type II activin receptors, competes with ALK4 for binding to activin and antagonizes activin signaling. We now have evidence showing that Cripto also binds TGF-Beta in the presence of TBRII, competes with ALK5 for TGF-Beta binding and antagonizes TGF-Beta signaling; we propose, therefore, that Cripto may be generally capable of blocking antiproliferative Smad2/3 signals. Under Aim I of this proposal we will identify the molecular determinants on Cripto that mediate Cripto binding to activin/TFG-Beta and antagonism of activin/TGF-Beta signaling. We also propose to develop reagents including anti-Cripto neutralizing antibodies designed to prevent the ability of Cripto to antagonize activin and TGF-Beta signaling. Aim II proposes to characterize the ability of Cripto to block type I receptor recruitment to ligand-type II receptor complexes and block type I receptor phosphorylation in breast cancer cells in vitro in response to activin/TGF-Beta and the ability of Cripto to block phosphorylation of Smad2/3 following activin/TGF-beta treatment of breast cancer cells in vitro. Under Aim III we will characterize the effects of Cripto on activin/TGF-Beta-induced growth inhibition in three mammary epithelial-derived cell lines: MCF-7; MCF-10A; and CID-9 cells. Together, these studies will lead to an improved understanding of the mechanisms of Cripto action and will help illuminate therapeutic targets for the management of neoplastic disease.

描述(申请人提供)：转化生长因子-β和激活素是生长和分化因子转化生长因子-β超家族中功能相关的成员。转化生长因子-β和激活素激活Smad2和Smad3，它们都通过有效地抑制包括上皮细胞在内的多种细胞的增殖来调节组织的动态平衡。对激活素和/或转化生长因子-β的抗增殖反应的破坏与细胞增殖、肿瘤发生和向恶性表型的进展有关。Cripto是一种发育性癌蛋白，在多种人类肿瘤中过表达，包括大约80%的人类乳腺肿瘤。CRIPTO过表达可转化乳腺上皮细胞，激活促进生长的ERK/MAPK和PI3K信号通路。Cripto和相关的EGF-CFC蛋白在胚胎发育过程中也发挥着重要的信号作用，并且Cripto是中胚层诱导和心脏发生所必需的。已有研究表明，在转化生长因子-β超家族成员Node1、VGL和GDF1中，CRIPTO作为辅助受体，通过激活素II/I受体促进受体组装和信号传递。这使得我们测试了Cripto是否可以调节激活素以及可能的其他转化生长因子-β配体组装功能性受体复合体的能力，我们发现Cripto在存在II型激活素受体的情况下与激活素结合，与ALK4竞争结合激活素，并拮抗激活素信号转导。我们现在有证据表明，在TBRII存在的情况下，Cripto也能结合转化生长因子-β，与Alk5竞争转化生长因子-β结合，并拮抗转化生长因子-β信号；因此，我们认为，Cripto通常可能能够阻断抗增殖的Smad2/3信号。根据这项建议的目标I，我们将确定CRIPTO上介导CRIPTO与激活素/TFG-β结合和激活素/转化生长因子-β信号拮抗的分子决定因素。我们还建议开发包括抗CRIPTO中和抗体在内的试剂，旨在阻止CRIPTO对抗激活素和转化生长因子-β信号转导的能力。目的研究CRIPTO对激活素/转化生长因子-β(激活素/转化生长因子-β)诱导的乳腺癌细胞I型受体聚集和I型受体磷酸化的阻断作用，以及CRIPTO对激活素/转化生长因子-β诱导的乳腺癌细胞Smad2/3磷酸化的阻断作用。在AIM III中，我们将研究Cripto对激活素/转化生长因子-β诱导的三种乳腺上皮源性细胞系：MCF-7、MCF-10A和CID-9细胞生长抑制的影响。总之，这些研究将有助于更好地理解CRIPTO的作用机制，并将有助于阐明肿瘤疾病管理的治疗目标。