2017-08-012024-05-14https://scholars.lib.ntu.edu.tw/handle/123456789/658685摘要：結直腸癌在癌症死亡率中排名第二，臨床上常見癌細胞產生抗藥性成為治療上棘手問題。過去文獻記載結直腸癌病人檢體有異常表現葡萄糖轉運蛋白，一般推論此現象與癌細胞代謝異常及腫瘤生長所需能量有關。正常細胞代謝過程中，葡萄糖藉由無氧醣解作用轉變為丙酮酸，在有氧環境下丙酮酸進入粒線體的電子傳遞鏈產生能量(ATP)。然而，癌細胞因快速生長之壓力，致使代謝異常只進行醣解作用，而無粒線體的有氧呼吸反應，稱之為Warburg’s effect。最近有研究指出葡萄糖供給異常可能是癌細胞基因突變和抗藥性的導因之一。我們初步研究顯示葡萄糖攝取可增加人類結直腸癌細胞株在基因毒性藥物5-Fluorouracil (5-FU) 處理下的活性。時至今日，葡萄糖代謝參與腸癌細胞抗藥性產生之詳盡機制（如阻抗死亡、調節增生、誘發幹細胞特性）仍不清楚。本實驗室過去研究顛覆了傳統認知ATP 為細胞主要保護因子的概念，發現葡萄糖可阻抗腸道上皮和腸癌細胞因缺氧壓力造成之死亡，而其阻抗死亡機制是藉由丙酮酸的自由基清除, 並非透過ATP 補給 (Huang CY et al., 2013 Cell Death Dis; HuangCY et al., 2016 J Physiol)。因此，本計畫實驗假設是葡萄糖個別代謝物(如丙酮酸、ATP)可經由獨立之機轉參與癌細胞各面向的存活增生反應，導致異質性腸癌細胞的抗藥性。本計畫研究目的是利用人類結直腸癌細胞株、體外癌球狀體培養、致變劑誘發結直腸癌和異種移植腸癌之小鼠模式，探討葡萄糖促進癌細胞抗藥性之分子機轉，期望能提供新的抗癌策略。本計畫目標和策略：目標＃1：採用致變劑誘發腸癌小鼠模式探討葡萄糖運輸器表現量在活體腫瘤上的改變，並利用體外培養的初代小鼠腸癌球狀體和人類腸癌株探究葡萄糖對基因毒性藥物的抑制作用。目標＃2：利用人類腸癌株和初代小鼠腸癌球狀體之離體模式來釐清葡萄糖及其代謝物參與抗藥性之詳盡機轉，如阻抗死亡、調節增生、誘發幹細胞特性。目標＃3：藉由基質膠包覆腸癌球狀體作異種移植小鼠模式佐證葡萄糖、丙酮酸、ATP 對腫瘤生長和腸癌抗藥性之影響。目標＃4： 探索長期重複性葡萄糖對癌細胞在基因毒性藥物下的援救是否會促發腸癌幹細胞特性增加和基因表現變異，導致頑強抗藥性。<br> Abstract: Colorectal cancer (CRC) is the second leading cause of cancer mortality, of whichchemoresistance to genotoxic agents is commonly reported. Glucose transporters areabnormally expressed in human CRC specimens, which are suspected to be linked to metabolicshift and energy demand for tumor growth. In normal cells, glucose converts to pyruvatethrough anaerobic glycolytic pathways, followed by entry to mitochondrial electron transportchain for ATP synthesis. Owing to the pressure of rapid tumor growth, cancer cells showedaltered metabolism with mainly glycolysis, termed Warburg’s effect. Recent studies indicatethat aberrant glucose uptake and metabolism may induce gene mutation and drug resistancein CRC. Our preliminary data show that high glucose uptake increases the viability of humanCRC cell lines under treatment of genotoxic agent 5-Fluorouracil (5-FU). To date, detailedmechanisms of glucose-mediated chemoresistance remain elusive. The effect of glucosemetabolism on multiple aspects in chemoresistance (including death resistance, regulationof proliferation, and stemness promotion) are poorly understood.Our previous findings challenged the traditional view of energy being the maincytoprotective factor. We showed that glucose prevented cell death in intestinal epitheliumand cancer cells under hypoxic stress via free radical scavenging by glycolytic pyruvate in anATP-independent manner (Huang et al. 2013 Cell Death Dis; Huang et al., 2016 J Physiol). The workinghypothesis of the current project is that glucose provides genotoxic protection toheterogeneous cancer cells through divergent mechanisms conferred by distinct glucosemetabolites such as pyruvate and ATP.The overall goal is to investigate the roles of distinct glucose metabolites in novelchemoresistance mechanisms by using human CRC cell lines, primary mouse tumorspheroids, as well as mutagen-induced and xenograft and orthotopic transplant models ofmurine CRC. The study will shed light to the development of glucose transporter- andmetabolism-targeting strategies for clinical management of refractory colon cancers.Project Objective and Aims:Objective #1 To characterize glucose transport abnormality in mutagen-induced mouse CRC,and to verify glucose-dependent cell survival against genotoxic agents in primary mousetumor spheroids and human CRC cell lines.Objective #2 To identify the role of glucose metabolism in distinct mechanisms ofchemoresistance, including death resistance, regulation of proliferation, and cancerstemness, using in vitro human CRC cell lines and mouse tumor spheroids.Objective #3 To evaluate in vivo tumor growth and chemoresistance conferred by glucose,pyruvate, and ATP using mutagen-induced and matrigel-based transplant models ofmouse CRC.Objective #4 To explore long-term glucose modulation on stemness and genetic signatures inCRC cells by repeated rescue against genotoxicity.結直腸癌抗藥性葡萄糖代謝細胞死亡細胞增生幹細胞特性癌球狀體小鼠模式colorectal carcinomachemoresistanceglucose metabolismcell deathcell proliferationstemnesstumor spheroidsmouse model