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The Role of Rb phosphorylation in proliferation and apoptosis of breast cancer ce

Rb磷酸化在乳腺癌细胞增殖和凋亡中的作用

基本信息

批准号：
8625004
负责人：
Nancy A Krucher
金额：
$ 36.82万
依托单位：
PACE UNIVERSITY NEW YORK
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-03-01 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8625004
关键词：
Alanine Amino Acids Apoptosis Apoptotic Binding Binding Proteins Biological Models Breast Cancer Cell Breast Cancer Model Breast Cancer Treatment Breast Epithelial Cells Cell Culture Techniques Cell Proliferation Cell physiology Cells Cleaved cell Clinic Complex Cyclin-Dependent Kinases Cyclins Development Exhibits Future Glutamic Acid Human Individual Investigation Length Malignant Neoplasms Methods Modeling Mutation Pathway interactions Phosphoric Monoester Hydrolases Phosphorylation Phosphorylation Site Physiological Proliferating Protein Dephosphorylation Proteins Regulation Retinoblastoma Retinoblastoma Protein Role Site Site-Directed Mutagenesis Tumor Suppressor Proteins cancer cell cancer type clinically relevant design in vivo inhibitor/antagonist malignant breast neoplasm novel pro-apoptotic protein public health relevance retinoblastoma tumor suppressor three-dimensional modeling tumor

项目摘要

DESCRIPTION (provided by applicant): The molecules that control Retinoblastoma (Rb) phosphorylation called "the Rb pathway" consisting of D-type cyclins, cyclin dependent kinases (cdks) and cdk inhibitors (cdkis) as well as Rb itself are regularly disrupted in most cancer types including breast cancer leading to excessive phosphorylation (hyperphosphorylation) of the Rb protein. Hyperphosphorylation of Rb is known to promote proliferation and block apoptosis. Treatments that inhibit cdk activity have begun to exhibit promise in the clinic in several cancer types. Although Rb can be phosphorylated on 15 amino acid residues, very little is known about the contribution any individual phosphorylated amino acid has on the function of Rb in proliferation and apoptosis. In this project we propose to undertake the first investigation of Rb phosphorylation in proliferation and apoptosis utilizing a model of breast epithelial cells using three-dimensional cultures. This model system is employed to recapitulate the physiological context in which breast epithelial cells regulate cellular processes. We propose to target Rb hyper- phosphorylation by using a novel method of activating Rb-specific phosphatase activity in breast epithelial cells grown in 3D cultures. In this way we can determine the effect of dephosphorylation of Rb on proliferation and apoptosis in these cells. In addition, we will perform site directed mutagenesis of each Rb phosphorylation site to either alanine or glutamic acid to block phosphorylation or mimic phosphorylation at each site and evaluate the effects on proliferation and apoptosis in breast epithelial cells grown in the 3D culture model. Finally, we will clarify the functional significance of complex formation between Rb and the pro-apoptotic protein Bak that is regulated by Rb dephosphorylation in apoptosis. Thus, the proposed studies will elucidate the role of specific Rb phosphorylation sites involved in the regulation of proliferation and apoptosis in breast cancer and will yield useful information that could inform the development of future therapies that target Rb phosphorylation in cancer.

描述（由申请人提供）：控制视网膜母细胞瘤（Rb）磷酸化的分子（称为“Rb途径”），由D型细胞周期蛋白、细胞周期蛋白依赖性激酶（cdks）和cdk抑制剂（cdkis）组成，以及Rb本身在大多数癌症类型中经常被破坏包括导致Rb蛋白过度磷酸化（过度磷酸化）的乳腺癌。已知Rb的过度磷酸化促进增殖并阻断凋亡。抑制cdk活性的治疗方法已经开始在临床上对几种癌症类型显示出希望。虽然Rb可以在15个氨基酸残基上磷酸化，但是关于任何单个磷酸化氨基酸对Rb在增殖和凋亡中的功能的贡献知之甚少。在这个项目中，我们建议进行第一次调查的Rb磷酸化的增殖和凋亡利用模型的乳腺上皮细胞，使用三维培养。该模型系统被用来概括乳腺上皮细胞调节细胞过程的生理背景。我们建议通过使用一种新的方法来激活在3D培养中生长的乳腺上皮细胞中Rb特异性磷酸酶活性，从而靶向Rb过度磷酸化。通过这种方式，我们可以确定Rb去磷酸化对这些细胞增殖和凋亡的影响。此外，我们将对每个Rb磷酸化位点进行丙氨酸或谷氨酸的定点诱变，以阻断每个位点的磷酸化或模拟磷酸化，并评估对3D培养模型中生长的乳腺上皮细胞增殖和凋亡的影响。最后，我们将阐明Rb和凋亡前蛋白巴克之间形成复合物的功能意义，该蛋白在凋亡中受Rb去磷酸化的调节。因此，拟议的研究将阐明参与乳腺癌增殖和凋亡调控的特定Rb磷酸化位点的作用，并将产生有用的信息，可以为未来癌症中靶向Rb磷酸化的治疗方法的发展提供信息。