Elucidating the Role of Akt and Keratins in Autophagy and Tumorigenesis

阐明 Akt 和角蛋白在自噬和肿瘤发生中的作用

基本信息

批准号：
8541786
负责人：
Richard C Wang
金额：
$ 13.5万
依托单位：
UT SOUTHWESTERN MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-09-10 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8541786
关键词：
Address Affect Agar Autophagocytosis Award Binding Binding Proteins Biological Assay Cancer Biology Cancer cell line Cell Proliferation Cell Size Cell Survival Cells Chemicals Clinical Trials Complex Cytoskeletal Proteins Defect Deletion Mutation Dermatology Epithelial Foundations Funding Future Genetic Genetic Induction Goals Human In Vitro Intermediate Filament Proteins Intermediate Filaments K-18 conjugate K-Series Research Career Programs Keratin Malignant Neoplasms Malignant Squamous Cell Neoplasm Mediating Medical center Mentors Mentorship Methods Modeling Mus Mutation Oncogenes PTEN gene Pathway interactions Patient Care Phenotype Phosphorylation Phosphotransferases Physicians Protein-Serine-Threonine Kinases Reading Reagent Regulation Research Research Personnel Research Project Grants Role Scientist Signal Transduction Skin Skin Carcinoma Structure Testing Texas Therapeutic United States National Institutes of Health Universities Vimentin cancer cell cancer therapy career cell growth human FRAP1 protein improved in vitro Assay in vivo inhibition of autophagy mouse model mutant novel prevent research study tumorigenesis

项目摘要

DESCRIPTION (provided by applicant): The NIH K08 mentored career development award provides the necessary foundation for me to fulfill my long-term career goals of being an independently-funded translational investigator focusing on the therapeutic manipulation of autophagy in the treatment of cancer. The mentored award will also help me achieve my immediate goals of obtaining expertise both in autophagy research and in the management of non- melanoma skin cancer. Under the auspices of the University of Texas Southwestern Medical Center at Dallas, I hope to attain these objectives through the following: 1) career mentorship from internationally recognized physicians and scientists who are experts in autophagy, dermatology, and cancer biology, 2) specific guidance in the form of structured reading and courses, which I will apply to experiments and patient care. My mentored research project centers on a novel model for the regulation of autophagy by Akt and keratin intermediate filaments and tests this model for its effects on tumorigenesis; our model challenges the current paradigm that the regulation of autophagy occurs primarily through mTOR. Addressing the question of how autophagy is regulated will be essential not only for interpreting the results of ongoing clinical trials but also for improving the way autophagy is manipulated in future cancer therapies. Aim 1 tests our hypothesis that Akt and phosphorylation of Beclin 1 directly regulate autophagy. First, we will test our hypothesis that Akt can regulate autophagy independently of mTOR through phosphorylation of Beclin 1. Next, we will determine how the status of PI3K-Akt activation in cancer cells affects the phosphorylation of Beclin 1. Finally, we will address what effects the phosphorylation of Beclin 1 has on autophagy and cell proliferation in cancer cell lines. Aim 2 will test whether the phosphorylation of epithelial keratins regulates autophagy in vitro and whether known pathogenic keratin mutations affect autophagy and cell proliferation. First, mutant intermediate filament (IF) proteins and cells deficient for specific IFs cells will b tested for their effects on autophagy in established in vitro assays. Next, we will determine whether this inhibition of autophagy results in excess cell proliferation in vitro. Finally, we wil use mouse models with keratin defects (both deletions and mutations) to test whether impaired autophagy contributes to keratinopathies and tumorigenesis. Aim 3 will test what effects the disruption of the predicted complex has on tumorigenesis in in vitro assays and in vivo mouse models. The effects of IF mutations on tumorigenesis will be determined in vitro. Next, we will test whether the chemical induction of autophagy or genetic induction of autophagy through disruption of proposed regulatory complex can prevent tumorigenesis in vivo. These studies will employ an established model of non-melanoma skin cancer formation in skin with an inducible deletion of PTEN. These studies will transform our current understanding of how autophagy is regulated and impact how autophagy is manipulated to treat cancer in future clinical trials.

描述（由申请人提供）：NIH K08 指导的职业发展奖为我实现长期职业目标提供了必要的基础，即成为一名独立资助的转化研究者，专注于癌症治疗中自噬的治疗操作。该指导奖项还将帮助我实现近期目标，即获得自噬研究和非黑色素瘤皮肤癌治疗方面的专业知识。在达拉斯德克萨斯大学西南医学中心的支持下，我希望通过以下方式实现这些目标：1）来自国际公认的自噬、皮肤病学和癌症生物学专家的医生和科学家的职业指导，2）具体指导以结构化阅读和课程的形式，我将应用于实验和患者护理。我指导的研究项目以 Akt 和角蛋白中间丝调节自噬的新模型为中心，并测试该模型对肿瘤发生的影响；我们的模型挑战了当前自噬调节主要通过 mTOR 发生的范式。解决自噬如何调控的问题不仅对于解释正在进行的临床试验的结果至关重要，而且对于改进未来癌症治疗中自噬的操纵方式也至关重要。目标 1 检验我们的假设，即 Akt 和 Beclin 1 磷酸化直接调节自噬。首先，我们将检验我们的假设，即 Akt 可以通过 Beclin 1 的磷酸化独立于 mTOR 调节自噬。接下来，我们将确定癌细胞中 PI3K-Akt 激活的状态如何影响 Beclin 1 的磷酸化。最后，我们将讨论什么Beclin 1 磷酸化对癌细胞系自噬和细胞增殖的影响。目标 2 将测试上皮角蛋白的磷酸化是否在体外调节自噬，以及已知的致病性角蛋白突变是否影响自噬和细胞增殖。首先，将在已建立的体外测定中测试突变中间丝 (IF) 蛋白和缺乏特定 IF 细胞的细胞对自噬的影响。接下来，我们将确定这种自噬抑制是否会导致体外细胞过度增殖。最后，我们将使用具有角蛋白缺陷（缺失和突变）的小鼠模型来测试受损的自噬是否会导致角蛋白病和肿瘤发生。目标 3 将在体外试验和体内小鼠模型中测试预测复合物的破坏对肿瘤发生的影响。 IF 突变对肿瘤发生的影响将在体外确定。接下来，我们将测试化学诱导自噬或通过破坏所提出的调节复合物遗传诱导自噬是否可以预防体内肿瘤发生。这些研究将采用一个已建立的模型，在具有 PTEN 诱导缺失的皮肤中形成非黑色素瘤皮肤癌。这些研究将改变我们目前对自噬如何调节的理解，并影响未来临床试验中如何操纵自噬来治疗癌症。