ING5通过促进肺癌细胞糖代谢调控激酶PDHK1酪氨酸Y163磷酸化逆转Warburg效应的研究

Warburg effect is a hallmark of cancer metabolism which contributes to cancer invasion and metastasis. This effect is characterized by a high rate of aerobic glycolysis instead of oxidative phosphorylation (OXPHOS). Thus, promoting the switch from glycolysis to OXPHOS may reverse Warburg effect. Pyruvate is the glucose metabolite intermediate which could be catalyzed by pyruvate dehydrogenase (PDH) in mitochondria and enter OXPHOS pathway, leading to decreased glycolysis in cytoplasm. The enzyme activity of PDH is negatively phosphor-regulated by PDH kinase PDHK1. Therefore, PDHK1 is the key protein whose kinase activity determines the metabolite choice of cancer cells. In our previous work, we have found that the Inhibitor of Growth (ING) family member ING5 could significantly inhibit lung cancer cell invasion and metastasis. We also found that ING5 could upregulate phosphorylation of PDHK1 at tyrosine (Y) 163 by 14 fold. Furthermore, ING5 overexpression could reverse Warburg effect. Whether ING5 reverses Warburg effect and inhibits cancer invasion by regulating PDHK1 Y163 phosphorylation and the underlying mechanisms are unknown. In current project, we are going to perform experiments including in vitro kinase assay, gene knockdown and rescue, protein kinase siRNA library and et al. on lung cancer cells, to investigate increased phosphorylation of PDHK1 at Y163 by ING5 and its role on lung cancer cell metabolism, and to explore the mechanisms by which ING5 upregulates PDHK1 Y163 phosphorylation. Our study would not only reveal the mechanisms of how ING5 regulates lung cancer cell metabolism and reverses Warburg effect, but also provide evidence for developing anti-cancer strategies targeting key regulators in cancer metabolism.

Warburg瓦博格效应是恶性肿瘤最显著的代谢特征，是促进肿瘤侵袭转移的重要机制，其核心是有氧糖酵解取代氧化磷酸化。因此促进肿瘤细胞糖酵解转向氧化磷酸化可逆转该效应。丙酮酸脱氢酶可促进糖代谢中间产物丙酮酸进入线粒体氧化磷酸化而减少胞浆糖酵解，其活性受丙酮酸脱氢酶激酶PDHK1磷酸化负调控，故PDHK1是肿瘤代谢途径选择的关键开关分子。我们前期发现生长抑制因子ING5可显著抑制肺癌细胞侵袭转移，上调PDHK1酪氨酸Y163磷酸化14倍，且逆转瓦博格效应即增加细胞氧耗量、减少乳酸分泌。ING5是否通过调控PDHK1磷酸化逆转瓦博格效应而抑制肿瘤侵袭？机制如何？本课题拟以常见肿瘤细胞为研究对象，以基因敲减回补和激酶试验等研究ING5促进PDHK1Y163磷酸化及其对肿瘤细胞代谢选择的影响，揭示ING5在翻译后修饰水平调控肿瘤糖代谢而逆转瓦博格效应的机制，为靶向肿瘤糖代谢调控分子治疗策略提供依据。

侵袭和转移是导致肿瘤患者死亡的重要原因，瓦氏效应是恶性肿瘤最显著的代谢特征，是促进肿瘤侵袭转移的重要机制，其核心是有氧糖酵解取代氧化磷酸化。因此促进肿瘤细胞糖酵解转向氧化磷酸化可逆转该效应。丙酮酸脱氢酶可促进糖代谢中间产物丙酮酸进入线粒体氧化磷酸化而减少胞浆糖酵解，其活性受丙酮酸脱氢酶激酶PDK1磷酸化负调控，故PDK1是肿瘤代谢途径选择的关键开关分子。我们前期发现生长抑制因子（Inhibitor of Growth）家族成员ING5可显著抑制肺癌细胞侵袭转移，修饰蛋白质组学研究发现ING5过表达上调PDK1酪氨酸Y163磷酸化高达14倍，这是我们新发现的一个磷酸化位点。本课题中我们对PDK1Y163磷酸化是否介导ING5抑癌作用及其调控机制进行了研究，有如下新的发现：1. ING5过表达促进PDK1Y163磷酸化，抑制其磷酸激酶活性，进而使其下游丙酮酸脱氢酶核心酶PDHA1S293脱磷酸活化，促进氧化磷酸化作用；2. ING5过表达逆转瓦氏效应，表现为氧耗量增加，乳酸减少；3. PDK1Y163磷酸化介导ING5逆转瓦氏效应和对肺癌细胞增殖侵袭的抑制作用；4. PDK1Y163磷酸化介导ING5对裸鼠移植瘤的生长和转移作用；5. PDK1Y163磷酸化在临床肺癌肿瘤组织表达明显低于癌旁正常组织，PDK1Y163磷酸化程度与预后正相关；6. TIE1为磷酸化PDK1Y163 的特异性上游激酶，其表达受ING5正调控，ING5对TIE1 的调控作用和机制值得进一步研究。本课题研究揭示了抑癌蛋白ING5作用的新机制，即通过翻译后修饰水平调控肺癌细胞糖代谢过程酶的活性抑制瓦氏效应而发挥抑制肺癌侵袭转移的作用，本课题还发现了PDK1Y163磷酸化的细胞生物学和临床意义以及其上游调控机制，这些新的发现为靶向肺癌细胞糖代谢通路关键分子的抗肺癌转移治疗提供了方向和靶点。