Molecular Mechanisms of Fructose-induced Colorectal Cancer Cell Survival

果糖诱导结直肠癌细胞存活的分子机制

基本信息

批准号：
10548829
负责人：
Marcus DaSilva Goncalves
金额：
$ 57.3万
依托单位：
WEILL MEDICAL COLL OF CORNELL UNIV
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-01-10 至 2026-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10548829
关键词：
Adult Binding Biochemical Biological Assay Cell Count Cell Density Cell Hypoxia Cell Survival Cells Cellular Metabolic Process Chemicals Clinical Research Colon Colorectal Cancer Consumption Data Development Dietary Factors Dose Drug Design Drug Targeting Eating Ensure Enzyme Kinetics Enzymes Exposure to Food Fructose Future Gene Activation Generations Genes Genetic Genetic Models Glucose Goals Growth Human Hypoxia Incidence Ingestion Intake Intestines Isoenzymes Isotopes Ketohexokinase Kinetics Link Malignant Neoplasms Mass Spectrum Analysis Measures Mediator Metabolic Metabolic syndrome Metabolism Modality Modeling Molecular Mus Mutation Nutrient Obesity Oral Organ Organoids Oxygen Patients Physiology Placebos Play Process Prospective, cohort study Protein Biochemistry Protein Isoforms Proteins Public Health Publications Pyruvate Kinase Recombinant Proteins Recombinants Role Science Structure Sucrose Testing Time Tracer Transactivation Tumor Promotion Variant Woman cancer cell clinical development colon cancer cell line colorectal cancer progression colorectal cancer risk combat cost dietary experimental study fructose-1-phosphate hypoxia inducible factor 1 improved inhibitor intestinal adenoma men metabolomics microbiota middle age mortality mouse model mutant new therapeutic target novel therapeutics pharmacologic pre-clinical programs small molecule sugar tumor tumor growth tumor metabolism

项目摘要

Project Summary Clear associations have been established between the food we eat and the development and progression of colorectal cancer (CRC). For example, the consumption of fructose increases the risk for CRC development and CRC-specific mortality. However, the mechanism underlying this association is unknown. We have shown that moderate daily exposure to oral high fructose corn syrup (HFCS, a mix of fructose and glucose) leads to larger and more aggressive intestinal adenomas in mice. These effects were absent in mice with genetic deficiency of ketohexokinase (KHK), the enzyme that converts fructose to fructose 1-phosphate (F1P). A metabolomic analysis of these tumors showed that F1P is highly abundant following HFCS exposure, and this increase correlates with a reduction in pyruvate kinase (PK) activity. Therefore, we hypothesize that F1P, the product of KHK, enhances tumor growth by acting as an allosteric inhibitor of PK to promote anabolic metabolism and cell survival. We will test this hypothesis using mouse physiology and organ metabolism, cell and human organoid culture, and recombinant protein biochemistry. In Aim 1, we will genetically and pharmacologically manipulate the M2 isozyme of PK (PKM2) in mice to interrogate its role as a mediator of HFCS-induced tumor growth. In Aim 2, we will define the mechanistic linkage between fructose exposure and cancer cell survival. We have found that cells in culture do not grow faster when exposed to fructose, however we observed a significant improvement in cell viability, especially under conditions of high cell density and hypoxia with fructose in the media. Therefore, we hypothesize that F1P inhibits PKM2 to promote hypoxic cell survival. We will test this hypothesis using cell and organoid culture models exposed to fructose and hypoxia. We will genetically and pharmacologically manipulate KHK and PKM2 expression and activity in these models to determine the specific effects of these proteins on cell metabolism and survival. In Aim 3, we will assess the effects of F1P on recombinant PK isoforms with a particular focus on PKM2. We hypothesize that fructose- derived F1P binds to and inhibits PKM2. We will perform biochemical activity and structural assays to determine the kinetic parameters and oligomeric state of PK isoforms in the presence of F1P. These experiments will reveal the molecular mechanisms of how F1P binds and inhibits PKM2. Together, these aims will change our fundamental understanding of how fructose alters tumor cell metabolism, define the fructose/F1P/PKM2 axis as a metabolic vulnerability of CRC, and provide pre-clinical evidence for PKM2 activators as a novel therapeutic modality to combat CRC.

项目概要我们所吃的食物与身体的发育和进展之间已经建立了明确的联系结直肠癌（CRC）。例如，摄入果糖会增加结直肠癌发生的风险和 CRC 特异性死亡率。然而，这种关联背后的机制尚不清楚。我们已经展示了每天适度接触口服高果糖玉米糖浆（HFCS，果糖和葡萄糖的混合物）会导致小鼠肠道腺瘤更大、更具侵袭性。这些效应在具有遗传基因的小鼠中不存在酮己糖激酶 (KHK) 缺乏，这种酶将果糖转化为果糖 1-磷酸 (F1P)。一个这些肿瘤的代谢组学分析表明，HFCS 暴露后 F1P 含量很高，这增加与丙酮酸激酶（PK）活性的降低相关。因此，我们假设 F1P KHK 的产品，通过作为 PK 的变构抑制剂促进合成代谢来促进肿瘤生长新陈代谢和细胞存活。我们将利用小鼠生理学和器官代谢、细胞和人类类器官培养，以及重组蛋白生物化学。在目标 1 中，我们将通过遗传和在小鼠中药理学操纵 PK (PKM2) 的 M2 同工酶以探究其作为介导的作用 HFCS 诱导的肿瘤生长。在目标 2 中，我们将定义果糖暴露与癌细胞存活。我们发现，当暴露于果糖时，培养中的细胞不会生长得更快，但是我们观察到细胞活力显着提高，特别是在高细胞密度和介质中存在果糖缺氧。因此，我们推测F1P抑制PKM2促进细胞缺氧生存。我们将使用暴露于果糖和缺氧的细胞和类器官培养模型来检验这一假设。我们将从遗传和药理学角度操纵这些模型中的 KHK 和 PKM2 表达和活性确定这些蛋白质对细胞代谢和存活的具体影响。在目标 3 中，我们将评估 F1P 对重组 PK 同工型的影响，特别关注 PKM2。我们假设果糖- 衍生的 F1P 结合并抑制 PKM2。我们将进行生化活性和结构测定确定 F1P 存在下 PK 亚型的动力学参数和寡聚状态。这些实验将揭示 F1P 如何结合和抑制 PKM2 的分子机制。这些目标共同实现将改变我们对果糖如何改变肿瘤细胞代谢的基本理解，定义果糖/F1P/PKM2轴作为CRC的代谢脆弱性，并为PKM2提供临床前证据激活剂作为对抗结直肠癌的新型治疗方式。