Examination of ceramide signaling in the crosstalk between pancreatic cancer cells and the tumor microenvironment

检查胰腺癌细胞和肿瘤微环境之间的神经酰胺信号传导

• 批准号: 9548458
• 负责人: Audrey Marie Hendley
• 金额: $ 5.32万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-12-01 至 2019-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9548458
• 关键词: Address; Apoptosis; Attention; Biogenesis; Biological Assay; Cancer Etiology; Cancer cell line; Cell Aging; Cell Culture Techniques; Cell Cycle Arrest; Cell Cycle Regulation; Cell membrane; Cells; Ceramides; Cessation of life; Complex; Data; Development; Disease; Distant; Drug resistance; Event; Genes; Genetically Engineered Mouse; Growth; Health; Human; Immunodeficient Mouse; Implant; In Vitro; Incidence; Lipid Bilayers; Lipids; Location; Maintenance; Malignant Neoplasms; Malignant neoplasm of pancreas; Mediating; Metabolism; Mission; Modeling; Molecular; Mus; National Cancer Institute; Nature; Neoplasm Metastasis; Organ; Pancreas; Pancreatic Adenocarcinoma; Pancreatic Ductal Adenocarcinoma; Pathway interactions; Patients; Physiology; Play; Production; Regulation; Research; Research Design; Research Proposals; Role; Second Messenger Systems; Signal Transduction; Sphingolipids; Sphingomyelinase; Sphingomyelins; Stimulus; Stress; Structure; Testing; Tumor Burden; Tumor Expansion; United States; United States National Institutes of Health; base; cancer cell; cell growth regulation; design; effective therapy; exosome; experience; experimental study; gene function; implantation; interest; knock-down; loss of function; mouse model; neoplastic; new therapeutic target; novel therapeutics; overexpression; pancreatic cancer cells; pancreatic cancer model; pancreatic juice; pancreatic neoplasm; pancreatic tumorigenesis; prevent; response; senescence; small hairpin RNA; stable cell line; tumor growth; tumor microenvironment; tumor progression; tumorigenesis

Project Summary/Abstract Background: Deregulation of sphingolipid metabolism alters pancreatic cancer progression and has thus received appreciable attention as a potential target for development of novel therapeutics. Intracellular ceramide levels are imperative for regulation of cellular senescence, apoptosis, and cell cycle arrest in response to stress stimuli such as chemotherapeutics. Preliminary data from our lab indicate a prominent role for a gene that regulates ceramide synthesis and exosome biogenesis, in the maintenance of pancreatic tumor progression. Orthotopic implantation of murine pancreatic ductal adenocarcinoma (PDA) cell lines stably expressing shRNA targeting this gene into immunodeficient mice significantly reduced pancreatic tumor burden when compared to controls. In addition, our preliminary data also indicate that this gene regulates exosome secretion from pancreatic cancer cell lines. Based on our exciting preliminary data, I have designed a study to decipher the molecular mechanisms by which this gene modulates pancreatic tumorigenesis. Objective/Hypothesis: Given this gene’s complex function in regulation of both cancer cell exosome secretion and intracellular ceramide levels, I propose the following central hypothesis: The function of this gene in pancreatic cancer is multiplex, potentially playing distinct roles during initiation and progression. The overall objective of this study is to determine the mechanisms by which this gene controls pancreatic carcinogenesis. Specific Aims: The specific aims of this study are: Aim 1) Evaluate the function of this gene in normal murine pancreas and pancreatic neoplasia formation and progression Aim 2) Determine the mechanism(s) by which this gene regulates pancreatic tumor growth Aim 2A) Assess the ability of this gene to alter cellular activities downstream of ceramide signaling Aim 2B) Evaluate the ability of exosomes secreted via this gene’s pathway to modulate pancreatic tumor expansion. Study Design: The approach proposed herein rigorously determines the function of this gene during PDA formation and progression using loss of function studies in established mouse and cell culture models of PDA. I will ablate our gene of interest in a widely-used mouse model of pancreatic cancer to determine the ability of this gene to regulate pancreatic cancer formation and progression. In addition, I propose mechanistic assays using murine pancreatic cancer (MPDA) cell lines generated by our group which stably express constructs allowing knockdown and overexpression studies that will illuminate the effect of exosome secretion and regulation of intracellular ceramide levels on pancreatic carcinogenesis mediated by this gene. Impact: Pancreatic cancer is projected to become the 2nd leading cause of cancer- related death in the US by 2020. Given this notable projected increase in incidence, pancreatic cancer remains a major problem for overall human health. Consistent with the overall mission of the NIH National Cancer Institute, the proposed studies may identify new therapeutic targets critical for development of new strategies to prevent, treat, and cure patients.

项目概要/摘要 背景：鞘脂代谢失调会改变胰腺癌的进展，因此 作为开发新型疗法的潜在目标，受到了相当大的关注。细胞内 神经酰胺水平对于调节细胞衰老、细胞凋亡和细胞周期停滞至关重要 对化疗等应激刺激的反应。我们实验室的初步数据显示了显着的作用 调节神经酰胺合成和外泌体生物发生的基因，在胰腺肿瘤的维持中 进展。小鼠胰腺导管腺癌（PDA）细胞系原位稳定植入 在免疫缺陷小鼠中表达靶向该基因的shRNA可显着减少胰腺肿瘤负荷 与对照相比。此外，我们的初步数据还表明该基因调控外泌体 胰腺癌细胞系的分泌。根据我们令人兴奋的初步数据，我设计了一项研究 破译该基因调节胰腺肿瘤发生的分子机制。 目的/假设：鉴于该基因在调节癌细胞外泌体分泌中的复杂功能 和细胞内神经酰胺水平，我提出以下中心假设：该基因在 胰腺癌是多发性的，在发生和进展过程中可能发挥不同的作用。整体 本研究的目的是确定该基因控制胰腺癌发生的机制。 具体目的：本研究的具体目的是： 目的 1) 评估该基因在正常小鼠中的功能 胰腺和胰腺肿瘤的形成和进展 目标 2) 确定胰腺和胰腺肿瘤形成和进展的机制 该基因调节胰腺肿瘤生长 目标 2A) 评估该基因改变细胞活动的能力 神经酰胺信号下游 目标 2B) 评估通过该基因途径分泌外泌体的能力 调节胰腺肿瘤的扩张。研究设计：本文提出的方法严格确定 使用已建立的功能丧失研究来确定该基因在 PDA 形成和进展过程中的功能 PDA 的小鼠和细胞培养模型。我将在广泛使用的小鼠模型中消除我们感兴趣的基因 胰腺癌，以确定该基因调节胰腺癌形成和进展的能力。 此外，我建议使用我们的小鼠胰腺癌（MPDA）细胞系进行机械测定。 稳定表达构建体的小组允许敲低和过度表达研究，这将阐明 外泌体分泌和细胞内神经酰胺水平调节对胰腺癌发生的影响 由该基因介导。影响：胰腺癌预计将成为第二大癌症原因 到 2020 年，美国的相关死亡人数将增加。鉴于预计发病率显着增加，胰腺癌仍然 人类整体健康的一个主要问题。与 NIH 国家癌症中心的总体使命一致 研究所，拟议的研究可能会确定对于制定新策略至关重要的新治疗靶点 预防、治疗和治愈患者。