Stathmin 1经p53、PI3K/AKT信号通路在口腔鳞癌诱导化疗中的分子机制研究

Although induction chemotherapy agents could be used to treat malignancies in head and neck cancers, there are still debates on the clinical application of induction chemotherapy in oral squmaous cell carcinoma (OSCC). The keypoint of improving the entire prognosis is to screen the proper patients who might benefit from induction chemotherapy before deciding the treatment protocol. Stathmin 1 is a microtubule-destabilizing phosphoprotein, ubiquitously expressed and involved in regulating the global dynamics of mitotic and interphase microtubules; however, the molecular role of Stathmin 1 in induction chemotherapy is still unclear in OSCC. Based on our previously established in vitro cellular carcinogenesis model of OSCC and proteomic analysis, we found that Stathmin 1 played an important role in progression of OSCC, it was possibly regulated by mutant p53 genes and correlated with the cellular apoptosis in OSCC; detection of stathmin 1 in biopsy samples from 256 OSCC patients in a clincial trial focusing on TPF induction chemotherpay shew that the patients with low Stathmin 1 expression could significantly benefit from TPF indcuction chemotherpay on suvival, the patients with high Stathmin 1 expression could not benefit from TPF induction chemotherapy. However, in the literatures, there are few studies reporting the molecular mechanism of Stathmin 1 expression on cellular apoptosis through mutant 53 and PI3K/AKT pathways, there are either few studies reporting the mechanism of treatment resistance in patients with Stathmin 1 overexpression against TPF induction chemotherapy in OSCC; therefore, there are still a lot of unknown mechanisms to be further investigated. Our present study will use the combination of basic machanism investigations and clincial trials to reveal the molecular mechanism of Stathmin 1 on cellular apoptosis through mutant p53 and PI3K/AKT pathways and the molecular mechanism of treatment resistance of Stathmin 1 overexpression against TPF induction chemotherapy in OSCC; a new clinical trial of Stathmin 1 expression as a predictive biomarker for choosing TPF induction chemotherpay will also be designed and excuted based on the retrospective results of Stathmin 1 anslysis in OSCC patients from the previous TPF trial. The aim of this study is to clarify the molecular mechanism of Stathmin 1 in cellular apoptosis through mutant 53 and PI3K/AKT pathways, and a personalized treatment strategy based on predictive biomarker in OSCC patients.

临床上诱导化疗在口腔鳞癌中还存在一定争议，如何筛选获益人群是整体提高疗效的关键。微管解聚蛋白Stathmin 1参与调解微管稳定性的动态平衡。课题组在建立口腔黏膜上皮细胞体外癌变模型和蛋白质组学研究的基础上，发现Stathmin 1在口腔鳞癌中起重要作用，并可能受p53基因突变调控；通过检测口腔鳞癌TPF诱导化疗临床试验患者组织中Stathmin 1表达，发现低表达Stathmin 1的患者可从TPF诱导化疗中生存获益，而高表达Stathmin 1的患者没有生存获益。本研究旨在结合分子机制研究和临床试验，重点围绕Stathmin 1与p53突变、PI3K/AKT信号通路，以及Stathmin 1在口腔鳞癌抵抗TPF诱导化疗中的作用，阐明Stathmin 1在口腔鳞癌发展和TPF诱导化疗中的分子机制，探索以Stathmin 1作为预测性生物标志物指导口腔鳞癌个体化治疗选择的肿瘤诊治新策略。

本项目旨在研究Stathmin 1在p53与PI3K/AKT信号通路调控口腔鳞癌发展的作用机制，及其在口腔鳞癌细胞抵抗化疗药物多西他赛、顺铂和5氟尿嘧啶（TPF）的分子机制。探索以Stathmin 1作为预测性生物标志物指导口腔鳞癌个体化治疗选择的肿瘤诊治新策略。.采用CCK-8细胞增殖实验、药敏实验、流式细胞术、免疫荧光和裸鼠皮下移植瘤体内实验，检测Stathmin 1在口腔鳞癌中的生物学功能。利用基因转染技术，干预Stathmin 1、野生型/突变型p53的表达，采用免疫沉淀实验进一步验证p53对Stathmin 1的转录激活作用。采用免疫组织化学方法分析Stathmin 1与p53表达以及患者预后的相关性。根据Stathmin 1的表达高低分组，采用Kaplan-Meier法和log-rank检验的统计学方法，进行亚组生存分析。使用PI3K-AKT-mTOR信号通路抑制剂联合TPF化疗药物处理口腔鳞癌细胞和裸鼠皮下移植瘤，评价联合用药的疗效。.发现Stathmin 1在口腔鳞癌细胞和患者肿瘤组织中过表达。敲减Stathmin 1的表达可抑制细胞增殖、集落形成和迁移、移植瘤的生长，促进细胞凋亡，并提高细胞对于TPF化疗药物的敏感性。突变型p53在口腔鳞癌中的转录激活Stathmin 1的表达，而野生型p53不能激活Stathmin 1的表达。在170例局部晚期口腔鳞癌患者中，Stathmin 1低表达43例，高表达127例。低表达Stathmin 1的患者可以从TPF诱导化疗中生存获益，而高表达Stathmin 1的患者不能生存获益。不同靶向位点的PI3K-AKT-mTOR信号通路抑制剂处理后，口腔鳞癌细胞中Stathmin 1的表达和磷酸化水平降低；联合应用TPF化疗药物和抑制剂，可以显著抑制口腔鳞癌细胞增殖、细胞周期、诱导细胞凋亡，有效抑制移植瘤的生长。.综上所述，突变型p53激活Stathmin 1的高表达，Stathmin 1可以作为预测局部晚期口腔鳞癌患者TPF诱导化疗疗效的生物标志物。联合应用PI3K-AKT-mTOR信号通路抑制剂和TPF化疗药物，可以有效抑制口腔鳞癌细胞和移植瘤的生长。