Functional contributions of glycogen metabolism to ovarian cancer metastasis

糖原代谢对卵巢癌转移的功能贡献

• 批准号: 9974038
• 负责人: Ernst Lengyel
• 金额: $ 44.05万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/9974038
• 关键词: Adhesions; Biological Assay; Biology; Cancer Biology; Cancer cell line; Cell Proliferation; Cells; ChIP-seq; Citric Acid Cycle; Coculture Techniques; Complement; Coupled; Cultured Cells; DNA; DNA Methylation; Data; Dioxygenases; Enzymes; Epigenetic Process; Epithelial; Epithelial Cells; Epithelium; Event; Fibroblasts; Gene Expression; Genes; Glucose-6-Phosphate; Glycogen; Glycogen Storage Disease; Glycolysis; Goals; Greater sac of peritoneum; Growth; Hematogenous; Human; In Vitro; Investigation; Label; Libraries; Link; MAP Kinase Gene; Malignant Neoplasms; Malignant neoplasm of ovary; Mammalian Oviducts; Mediating; Metabolic; Neoplasm Metastasis; Omentum; Organ; Peritoneum; Phenotype; Phosphorylation; Phosphotyrosine; Protein Glycosylation; Proteins; RNA Interference; Regulation; Role; Seeds; Serous; Signal Pathway; Signal Transduction; Site; alpha ketoglutarate; base; cancer cell; cell type; clinically relevant; cofactor; epigenetic regulation; experimental study; genome-wide; glucose 1 phosphate; glycogen metabolism; glycogenolysis; glycosylation; glycosyltransferase; histone methylation; in vivo; knock-down; metabolomics; mouse model; nano; neoplastic cell; novel; ovarian neoplasm; overexpression; phosphoproteomics; promoter; response; seal; three-dimensional modeling; tumor; tumor growth; tumor microenvironment; tumor progression

PROJECT SUMMARY/ABSTRACT The biology of high grade serous ovarian cancer (OvCa) is distinct from that of most epithelial tumors, in that hematogenous metastases are rare. Ovarian tumors remain confined to the peritoneal cavity and primarily seed to the omentum and peritoneum. There, cancer cells interact with cancer associated fibroblasts (CAFs) which promote cancer cell proliferation, invasion, and metastasis. To study bidirectional signaling between the two cell types, we co-cultured them and used SILAC coupled with quantitative, label-free phosphoproteomics to identify phospho-tyrosine signaling events in both OvCa cells and fibroblasts. We identified activation of phosphoglucomutase 1 (PGM1) in the cancer cells, pointing towards regulation of glycogen metabolism in OvCa cells by CAFs. Further, preliminary experiments showed that glycogenolysis provides energy to cancer cells and regulates protein glycosylation and histone methylation. Based on these data, the primary hypothesis underlying this application is that CAF-mediated glycogenolysis promotes metastasis through several different mechanisms, including glycolysis, protein glycosylation, and epigenetics. In Aim I, we will explore the metabolic consequences of PGM1 activation or inhibition in both OvCa cells and normal fallopian tube epithelial cells using untargeted metabolomics and metabolic flux studies. We will use PGM knockdown cells and investigate the effects of PGM1 inhibition on adhesion/invasion/proliferation using a 3D model and a syngeneic mouse model of metastasis. We will then utilize a high-throughput RNAi library with the goal of identifying the upstream signaling pathways regulating glycogenolysis in response to CAFs. In Aim II, we will systematically study glycogen-mediated glycosylation events using human protein glycosylation arrays to identify which proteins are specifically glycosylated by CAF-mediated glycogenolysis. This investigation will be complemented by studies determining how glycosyltransferase enzymes regulate CAF- mediated glycosylation. We will then study the role of glycosyltransferases on in vivo metastasis assays using primary CAF/OvCa cells and a syngeneic mouse model of OvCa metastasis. Using targeted metabolomics data and flux analysis of CAFs co-cultured with OvCa cells, we discovered that glycogenolysis induced α- ketoglutarate (KG). We therefore propose, in Aim III, to study how glycogenolysis alters epigenetic changes in cancer cells. We will characterize genome-wide changes in histone methylation (ChIP-seq) and DNA hydroxymethylation (Nano-hmC-Seal) to identify genes epigenetically regulated by glycogenolysis. The proposed experiments aim to define the link between glycogenolysis and epigenetics to identify functional regulators of OvCa metastasis. By understanding glycogen metabolism in the tumor organ, we may be able to elucidate novel metabolic mechanisms important for metastasis, which could result in the identification of a new and clinically relevant approach to the treatment of metastatic ovarian cancer.

项目摘要/摘要 高级浆液卵巢癌（OVCA）的生物学与大多数上皮肿瘤的生物学不同， 在该血源转移中很少见。卵巢肿瘤仍然局限于腹膜腔， 首先将种子变成大脑和腹膜。在那里，癌细胞与癌症相关的成纤维细胞相互作用 （CAF）促进癌细胞增殖，侵袭和转移。研究双向信号 在两种细胞类型之间，我们共同培养它们，并使用Silac与定量，无标签 鉴定OVCA细胞和成纤维细胞中的磷酸蛋白质组学鉴定磷酸酪氨酸信号传导事件。我们 鉴定出癌细胞中磷酸葡萄糖酶1（PGM1）的激活，指向调节 CAFS中OVCA细胞中的糖原代谢。此外，初步实验表明糖原分解 为癌细胞提供能量，并调节蛋白质糖基化和Hisstone甲基化。基于这些 数据，该应用的主要假设是CAF介导的糖原分解促进 通过几种不同的机制转移，包括糖酵解，蛋白质糖基化和表观遗传学。 在目标I中，我们将探索两个OVCA细胞中PGM1激活或抑制作用的代谢后果 正常的输卵管上皮细胞使用非靶向的代谢组学和代谢通量研究。我们将使用 PGM敲除细胞并研究PGM1抑制对广告/入侵/增殖的影响 3D模型和转移的合成小鼠模型。然后，我们将使用一个高通量RNAi库 确定对CAF响应糖原分解的上游信号通路的目的。目标 II，我们将使用人蛋白糖基化系统地研究糖原介导的糖基化事件 通过CAF介导的糖原分解来识别哪些蛋白质是特异性糖基化的阵列。这 研究将通过确定糖基转移酶如何调节CAF-的研究来完成研究 介导的糖基化。然后，我们将研究糖基转移酶在使用体内转移测定中的作用 原代CAF/OVCA细胞和OVCA转移的同性小鼠模型。使用靶向代谢组学 与OVCA细胞共培养的CAF的数据和通量分析，我们发现糖原分解诱导α- 酮谷酸酯（kg）。因此，我们建议在AIM III中研究糖原分解如何改变表观遗传的变化 癌细胞。我们将表征整个基因组的组蛋白甲基化（CHIP-SEQ）和DNA的变化 羟甲基化（纳米-HMC-丝）以鉴定受糖基解体表观遗传调节的基因。这 提出的实验旨在定义糖原分解与表观遗传学之间的联系，以鉴定功能 OVCA转移的调节剂。通过了解肿瘤器官中的糖原代谢，我们可能能够 阐明新型代谢机制对转移很重要，这可能导致鉴定 用于治疗转移性卵巢癌的新的和临床相关的方法。