Protein kinase C and lung carcinogenesis

蛋白激酶C与肺癌发生

• 批准号: 10733467
• 负责人: David Feldser
• 金额: $ 67.44万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-06 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10733467
• 关键词: Actins; Adenocarcinoma Cell; Affect; Animals; Benzo(a)pyrene; Biology; Biosensor; CRISPR/Cas technology; Cancer Etiology; Cancer Model; Carcinogens; Carcinoma; Cell Separation; Cells; Cellular biology; Cessation of life; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Competence; Cytoskeletal Modeling; Data; Defect; Development; Dimensions; Disease; Environmental Carcinogens; Epithelial Cells; Epithelium; Event; Exposure to; Extracellular Matrix; Family; Fluorescence; Fluorescence Resonance Energy Transfer; Gene Expression; Genetic; Genetically Engineered Mouse; Goals; Grant; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate; Hematopoietic; Human; In Vitro; Invaded; Joints; KRAS oncogenesis; KRAS2 gene; KRASG12D; Knock-out; Link; Lipids; LoxP-flanked allele; Lung; Lung Adenocarcinoma; Lung Neoplasms; Malignant Neoplasms; Malignant neoplasm of lung; Malignant neoplasm of prostate; Mass Spectrum Analysis; Mediating; Membrane; Mesenchyme; Modeling; Molecular; Monomeric GTP-Binding Proteins; Mus; Mutation; Neoplasm Metastasis; Non-Small-Cell Lung Carcinoma; Nude Mice; Oncogenic; Outcome; Patients; Peptide Hydrolases; Phenotype; Phorbol Esters; Phosphorylation; Phosphotransferases; Play; Positioning Attribute; Production; Protein Analysis; Protein Deficiency; Protein Kinase C; Publishing; RAS driven cancer; Rationalization; Receptor Protein-Tyrosine Kinases; Recurrence; Role; Second Messenger Systems; Series; Signal Pathway; Signal Transduction; Stromal Cells; Techniques; Testing; Tumor Promoters; Work; adenoma; cancer cell; cancer initiation; candidate identification; cell motility; cell type; experience; in vivo; innovation; insight; lung cancer cell; lung carcinogenesis; lung metastatic; lung tumorigenesis; malignant breast neoplasm; member; mouse model; mutant; neoplastic cell; novel; permissiveness; protein activation; protein kinase C epsilon; protein kinase C kinase; rac1 GTP-Binding Protein; receptor; screening; targeted treatment; tumor; tumor initiation; tumor microenvironment; tumorigenesis

ABSTRACT Lung cancer is the leading cause of cancer-related deaths in the U.S., with ~236,700 new cases and ~130,200 deaths estimated for 2022. A joint effort by both PIs in this grant led to the identification of the oncogenic kinase protein kinase C epsilon (PKCe) as a key player in lung carcinogenesis. PKCe is aberrantly up-regulated in lung adenocarcinoma and is associated with poor outcome in patients specifically harboring KRAS mutations. Using genetically engineered mouse (GEM) models, we demonstrated that PKCe is required for both carcinogen- and Kras-driven lung tumorigenesis. In addition to this role in cancer initiation, our work established novel roles for PKCe in cellular events associated with late cancer stages. Indeed PKCe is a major player in lung cancer cell motility and invasion via activation of the small G-protein Rac1, linking this kinase to metastatic dissemination. That PKCe plays critical permissive roles first during adenoma initiation and then again later during the acquisition of metastatic competency highlights the functional complexities of the PKCe signaling events in lung cancer. Interestingly, CRISPR-mediated inactivation of PKCe in the initiating cell-of-origin does not significantly affect Kras- G12D-induced tumor development, leading us to hypothesize that PKCe does not strictly act in a tumor cell autonomous manner to permit oncogenic KRAS-mediated tumorigenesis. To test this hypothesis, in Aim 1 we will generate and characterize a series of GEM models to restrict genetic deletion of PKCe to either oncogenic Kras- expressing cancer cells or to diverse stromal cell types present in the tumor microenvironment, including non- cancerous epithelial, mesenchyme and hematopoietic cells. Gene expression studies on isolated cells using fluorescence-based lineage tracing techniques will provide significant mechanistic insights. In Aim 2, we will thoroughly dissect the mechanistic basis of motility/invasive signaling activation by PKCe. Our hypothesis is that PKCe activates Rac1-mediated formation of cell ruffles and motility in KRAS mutant lung cancer cells via Rac Guanine nucleotide Exchange Factors (Rac-GEFs). We will identify and characterize candidate Rac-GEFs as PKCe effectors responsible for this phenotype. In Aim 3, we will establish the involvement of PKCe and its effector Rac-GEFs for the development of metastatic lung cancer. We will use combined in vitro and in vivo approaches to pin down mechanistic defects in the metastatic cascade upon CRISPR-mediated deletion of PKCe and Rac-GEFs in lung adenocarcinoma cells. We will establish GEM models and use lentiviral CRISPR-based approaches to determine the permissive contribution of PKCe and its GEF effectors to lung adenocarcinoma metastasis, and identify relevant gene expression and signaling signatures contributing to this phenotype. Thus, by using innovative, state-of-the-art genetic and molecular approaches, our studies should reveal the vast multidimensional complexity of PKCe signaling in lung cancer development as well as underscore unappreciated mechanisms of carcinogen- and KRAS-mediated lung oncogenesis.

摘要 在美国，肺癌是癌症相关死亡的主要原因，约有236,700例新病例和约130,200例 据估计，2022年的死亡人数。在这笔赠款中，两个私人投资者的共同努力导致了致癌激酶的鉴定。 蛋白激酶C epsilon(PKCE)在肺癌发生中的关键作用。PKCE在肺中异常上调 腺癌，在特别携带KRAS突变的患者中与不良预后有关。vbl.使用 在基因工程小鼠(GEM)模型中，我们证明了PKCE是致癌物质和 KRAS驱动的肺肿瘤发生。除了在癌症启动中的这一角色外，我们的工作还为 PKCE在与晚期癌症相关的细胞事件中的作用。事实上，PKCE是肺癌细胞的主要参与者 运动性和侵袭性通过激活小G蛋白rac1，将该激酶与转移扩散联系起来。那 PKCE首先在腺瘤形成过程中发挥关键的许可作用，然后在随后的获得过程中再次发挥作用。 转移能力突出了肺癌中PKCE信号事件的功能复杂性。 有趣的是，CRISPR介导的起始细胞中PKCE的失活并不显著地影响Kras- G12D诱导的肿瘤发展，使我们假设PKCE并不严格地作用于肿瘤细胞 自主方式允许致癌的KRAS介导的肿瘤发生。为了验证这一假设，在目标1中，我们将 建立并表征了一系列GEM模型，以限制PKCE的基因缺失 表达癌细胞或肿瘤微环境中存在的不同类型的间质细胞，包括非 癌上皮、间充质和造血细胞。分离细胞的基因表达研究 基于荧光的血统追踪技术将提供重要的机械性见解。在目标2中，我们将 深入剖析PKCE激活运动性/侵袭性信号的机制基础。我们的假设是 PKCE通过Rac激活Rac介导的Rac1介导的KRAS突变肺癌细胞皱褶的形成和运动 鸟嘌呤核苷酸交换因子(RAC-GEF)。我们将确定候选RAC-GEF并将其描述为 PKCE效应器对此表型负责。在目标3中，我们将确定PKCE及其效应者的参与 RAC-GEF在转移性肺癌发展中的作用我们将使用体外和体内相结合的方法来 CRISPR介导的PKCE和RAC-GEF缺失后转移级联中的机械性缺陷 在肺腺癌细胞中。我们将建立GEM模型并使用基于慢病毒CRISPR的方法来 确定PKCE及其全球环境基金效应因子对肺腺癌转移的允许贡献，以及 确定与此表型相关的基因表达和信号特征。因此，通过使用 创新的、最先进的遗传和分子方法，我们的研究应该揭示广阔的多维 PKCE信号在肺癌发生发展中的复杂性以及强调未被认识的 致癌物和KRAS介导的肺肿瘤发生。