活性氧介导的p38MAPK/mTOR通路通过调控细胞自噬与凋亡影响舌鳞癌顺铂耐药的分子机制研究

Tongue squamous cell carcinoma (TSCC) is the most common malignant head and neck tumor. Cisplatin-based chemotherapy is the leading therapy for late stage TSCC. However, the consequent chemoresistance severely impairs the therapy effect. Cellular reactive oxygen species (ROS) mediated-p38MAPK/mTOR signaling pathway plays a key role in the process of cisplatin resistance in TSCC cells; however, the exact molecular mechanisms are not yet clear. Recent studies from other groups indicated that ROS could induce continuous activation of p38MAPK/mTOR signaling pathway. Our previous studies showed that the autophagy level was increased greatly while the phosphor-level of p38MAPK and mTOR proteins were enhanced significantly in cisplatin-resistant TSCC cell line CAL27 after the treatment of cisplatin. However, in the chemosensitive TSCC cell line SCC25, the apoptosis level was increased significantly while the phosphor-level of p38MAPK and mTOR proteins were decreased greatly. Moreover, the level of autophagy and apoptosis were enhanced significantly when CAL27 and SCC25 cells were both treated with the combination of cisplatin and plumbagin which is a ROS inducing agent. Based on our previous studies and studies from other groups, we hypothesize that ROS-mediated activation of p38MAPK/mTOR signaling pathway plays an important role in cisplatin resistance of TSCC via the modulation of autophagy and apoptosis. This research project will be carried out in chemo-resistant/sensitive cell lines and nude mouse model; we will use specific activators or inhibitors to regulate the cellular ROS level. Moreover, we will apply the techniques of siRNA and lentiviral transfection to modulate the expression of key proteins in p38MAPK/mTOR signaling pathway. Finally the chemosensitivity of cisplatin will be examined in TSCC cells and nude mice. The project aims to further reveal the role of ROS-mediated p38MAPK/mTOR pathway in cisplatin resistance of TSCC and lay the foundation for developing potent drugs to reverse cisplatin resistance in TSCC.

活性氧介导的p38MAPK/mTOR通路在舌鳞癌顺铂耐药过程中起关键作用，近年来研究发现活性氧可诱导该通路持续活化，但在化疗耐药中其分子机制尚不明确。我们前期研究发现：①舌鳞癌顺铂耐药细胞经顺铂处理后自噬水平明显上升且p38MAPK和mTOR蛋白磷酸化水平显著升高，而化疗敏感细胞经顺铂处理后凋亡水平明显升高且p38MAPK和mTOR磷酸化水平降低；②顺铂联合活性氧生成药物白花丹素作用后，耐药及敏感细胞中自噬及凋亡水平均显著升高，据此我们提出假设：活性氧介导的该通路活化通过调控细胞自噬与凋亡影响舌鳞癌顺铂耐药。本研究拟采用顺铂耐药细胞模型和裸鼠移植瘤模型、应用特异性诱导剂和抑制剂调控活性氧水平、应用siRNA及慢病毒转染技术调控通路水平后检测舌鳞癌细胞对顺铂化疗的敏感性，揭示活性氧介导的p38MAPK/mTOR通路在舌鳞癌细胞顺铂耐药中的作用及分子机制，为探索研发相关化疗增敏剂奠定理论基础。

舌鳞状细胞癌是口腔颌面部最常见的恶性肿瘤，其发病率和死亡率高。以顺铂为基础的传统化疗仍是晚期舌鳞癌患者的首选治疗方案，但常常产生化疗耐药而导致治疗效果不佳。我们前期研究发现，p38MAPK和mTOR信号通路与舌鳞癌的自噬凋亡水平及顺铂耐药密切相关，且顺铂联合活性氧生成药物白花丹素作用后，舌鳞癌细胞中自噬、凋亡水平均显著升高。据此我们提出以下假设：活性氧介导的p38MAPK和mTOR信号通路通过调控细胞自噬与凋亡影响舌鳞癌顺铂耐药。本研究拟采用顺铂耐药细胞和裸鼠移植瘤模型、应用特异性诱导剂和抑制剂调控细胞内活性氧水平，探索低水平活性氧是否可以通过调控p38MAPK和mTOR信号通路参与舌鳞癌细胞自噬和凋亡；探索低水平活性氧与舌鳞癌细胞顺铂耐药的关系及ROS生成药物白花丹素逆转该耐药的过程及其机制；并进一步通过体内实验验证活性氧生成药物白花丹素逆转该耐药的作用。我们发现：1.低水平活性氧能够通过调控p38MAPK和mTOR信号通路参与舌鳞癌细胞的自噬与凋亡；2.低水平活性氧能够增加舌鳞癌对顺铂的耐药性，且白花丹素能够逆转该耐药过程；3.体内实验验证白花丹素增加舌鳞癌对顺铂的敏感性。该发现为寻找舌鳞癌化疗新靶点提供方向；为今后临床逆转舌鳞癌化疗耐药提供了新的思路与理论基础。