AGK调节p53蛋白稳定性促进结肠癌细胞衰老的研究

Cellular senescence is an important way of cell death and plays a novel role in the progression of colon cancer. p53 is a key molecule to regulate cell senescence. The abnormal regulation of p53 protein stability is a major cause for dysregulation of p53 function. We found that AGK (Acylglycerol Kinase) is involved in the regulation of colon cancer cell senescence. AGK may bind with p53 to increase the phosphorylation of p53, thus the ubiquitination and degradation of p53 is inhibited to sustain the stability of p53 protein, then p21 is upregulated to promote cell senescence. In this way, AGK can improve the colon cancer cell senescence and inhibit the progression of colon cancer. This study aims to investigate the roles of AGK in the progression of CRC and cellular senescence with clinical CRC samples, to demostrate the mechanism how AGK-mediated phosphorylation of p53 regulates its stability and the p53/p21 pathway through knockdown /knockout /overexpression, transgenic mice model, protein mass spectrometry, RNA-Seq, and point mutation. In addition, this project try to explore the possibilities of individualized colon cancer treatment based on the AGK/p53/p21 pathway. This study is expected to further clarify the regulation of p53 signal pathway, the functions of AGK in cellular senescence, and provide a new target for the treatment of CRC.

细胞衰老是细胞重要的死亡方式，在结肠癌发生发展中发挥重要作用。p53是细胞衰老调控的关键分子。p53蛋白稳定性的调节异常是导致p53失调的重要原因。前期研究发现：甘油三酯酰基激酶AGK参与对结肠癌细胞衰老的调节，其机制可能是通过结合p53，介导p53磷酸化以促进p53蛋白稳定性，进而上调p21的表达，促进细胞衰老。本项目将通过临床样本分析，明确AGK在结肠癌发生发展和结肠癌细胞衰老中的作用；通过基因敲减/敲除/高表达，转基因小鼠模型等明确AGK磷酸化p53，进而增强其稳定性的信号转导机制；通过蛋白质质谱、RNA-Seq、点突变等技术阐述AGK介导的p53磷酸化调节p53蛋白稳定性和p53/p21通路活性的分子机制；最后探讨基于AGK-p53-p21信号轴对结肠癌实施个体化治疗的可能性。项目的研究可望丰富p53信号通路调控理论，阐释AGK在细胞衰老中的作用机理，并为结肠癌治疗提供新的靶点。

AGK是一种脂质激酶，不仅参与调节细胞脂质代谢，而且与线粒体蛋白转运，糖酵解密切相关。AGK基因突变可导致常染色体隐性遗传性疾病Sengers综合征。AGK功能异常与多种肿瘤发生发展有关，包括前列腺癌，乳腺癌，宫颈鳞状细胞癌等，参与调节肿瘤细胞生长、侵袭、转移和耐药性。本项目前期研究表明，在结肠癌细胞系中，AGK可通过磷酸化p53, 抑制p53与MDM2的结合，从而抑制p53泛素化及其降解，并持续激活p21，发挥促进抑制结肠癌细胞衰老的作用。在研究不同肿瘤类型中AGK的具体作用时，我们发现，在弥漫性大B细胞淋巴瘤(DLBCLs)中，一种BCL-2小分子抑制剂，Venetoclax，其杀伤效应与AGK的表达水平呈显著负相关。在AGK表达水平高的DLBCLs细胞系中，Venetoclax对细胞的杀伤能力显著下降。同时，在AGK表达水平低的DLBCLs细胞系中，Venetoclax的杀伤能力显著增强。进一步的体内外研究均表明，AGK敲低可抑制PTEN，AKT及FOXO1磷酸化，促进FOXO1入核及BCL-2转录，进而增强DLBCL细胞对Venetoclax的敏感性；且在DLBCL临床肿瘤组织中，AGK表达与FOXO1入核及BCL-2表达负相关。总之，我们的研究初步阐明，AGK可作为重要的生物学标志物预测DLBCL患者对Venetoclax的敏感性，共靶向AGK与BCL-2可能提高对DLBCL的临床治疗效果。