GDF15调控GRP75/Nanog抑制CSCs样特性在三氧化二砷增加肝癌顺铂化疗敏感性中的作用

Improving the efficiency of chemotherapy on hepatocellular carcinoma (HCC) plays a key role in enhancing the overall therapeutic effect and the improvement of the prognosis of HCC patients. The elevation of cancer stem cells (CSCs)-like properties, is the root cause of HCC chemotherapy resistance. Here, based on our previous studies and on the researches worldwide, we suggest a new mechanism underlying the effects of arsenic trioxide (ATO) on the improvement of the efficiency of chemotherapy on HCC. Tumor tissues release a lot of cytokine when cisplatin causes tumor necrosis. Of all, GDF15 plays a key role in the induction of chemotherapy resistance. That is GDF15 improves the CSCs-like properties via up-regulating the GRP75/Nanog signal pathway. However, ATO can inhibit the expression of GDF15 by directly binding to this protein, which causes the ubiquitination of GDF15 and then blocks the GRP75/Nanog signal pathway. This process leads to an attenuation of CSCs-like properties, which finally improves the efficiency of chemotherapy on HCC. This project intends to perform the work in the cell model, animal model, as well as in the population. Meanwhile, we will also clarify the significance of the serum GDF15 and GRP75 in the HCC chemotherapy resistance and prognosis. By understanding a new mechanism by which ATO improves the efficiency of chemotherapy on HCC, this project will expand our knowledge in the discovering of the new ways to increase the sensitive of HCC chemotherapy.

化疗增敏对于提升肝癌总体治疗效果、改善预后具有十分重要的意义。肿瘤干细胞（CSCs）样特性增强是肝癌产生化疗抵抗的关键因素。本课题根据前期积累，结合国内外报道，提出三氧化二砷（ATO）增加肝癌化疗敏感性的分子机制假说如下：当顺铂诱导肿瘤坏死过程中，肿瘤组织释放多种细胞因子，其中GDF15作为关键信号分子之一，通过上调GRP75继而激活Nanog，诱导肝癌细胞CSCs样特性显著增强，进而产生化疗抵抗；而ATO则可通过直接与GDF15的半胱氨酸残基结合，诱导其发生泛素化降解，进而阻滞其下游GRP75/Nanog信号通路，抑制肝癌细胞CSCs样特性，最终增加肝癌对顺铂化疗的敏感性。本项目拟从细胞、动物及人群水平系统探讨我们提出的科学假说，同时阐明血清GDF15和GRP75在肝癌化疗产生抵抗以及化疗预后评估中的意义，为寻找有效的肝癌化疗增敏途径提供新思路。

深入研究肝癌化疗抵抗机制，寻找有效的化疗增敏或逆转化疗抵抗途径以提高肝癌化疗有效率，对于提高肝癌总体治疗效果并改善患者预后具有十分重要的意义。本项目从细胞、动物及人群水平系统探讨了我们提出的科学假说，初步阐明GDF15调控GRP75/Nanog抑制CSCs样特性在三氧化二砷增加肝癌顺铂化疗敏感性中的作用。主要研究结果如下：.1. 顺铂单纯处理可显著上调细胞的CSCs特性；而ATO单纯处理则能够显著抑制CSCs特性；应用ATO联合顺铂处理可显著抑制顺铂诱导的CSCs特性。.2. ATO能够显著增强低剂量顺铂对肝癌细胞杀伤作用；进一步构建肝癌多药耐药细胞株，发现ATO处理能够显著逆转细胞对5-FU、奥沙利铂和多柔比星的耐药性。上述结果在动物模型中亦得到验证。.3. 敲除GDF15或GRP75均可显著抑制肝癌细胞的CSCs特性，而过表达GRP75则呈现相反的效应。进一步研究发现：ATO可能通过与GDF15靶向结合，继而诱导GDF15泛素化降解，进而抑制其下游GRP75的表达水平。.4. 通过Meta分析发现：高表达Mortalin与包括肝癌在内多种肿瘤预后不良相关。基于TCGA数据库：Mortalin在肝癌人群组织中的表达水平显著高于癌旁组织，且随着肿瘤进展表达水平进一步升高；高表达Mortalin与肝癌人群总体生存（总人群）和亚洲人群无复发生存呈负相关。本课题组收集在肝癌人群组织中（术后复发，接受TACE治疗），Mortalin高表达组的生存情况显著差于Mortalin低表达组。.5. 原创性发现了GRP75下游重要信号分子14-3-3及其在炎症、CSCs特性中的作用，ATO亦可与14-3-3靶向结合继而诱导14-3-3泛素化降解，抑制炎症微环境和CSCs样特性，逆转肝癌的多药耐药性（研究拓展）。.本研究初步阐明了ATO逆转肝癌MDR的部分机制，为寻找有效的逆转化疗抵抗途径以提高肝癌化疗的有效率提供了新的理论和实践依据。