皂苷类单体抑制自噬体－溶酶体融合过程在食管癌治疗中的作用与分子机制

Seeking new molecular target and developing novel treatment has become an urgent need for esophageal cancer therapy. Since autophagy is closely related to the tumorigenesis, development and drug-resistance of tumor, regulating autophagy provides a new idea for cancer therapy. In the early research work: ① we have screened and identified new saponins suppressing autophagosome-lysosome fusion; ② 5-Fu induced a protective autophagy; ③ saponins enhanced the sensitivity of esophageal cancer cells to 5-Fu or induced autophagy-related cell deaths. Based on the hypothesis that saponins can enhance chemosensitivity of esophageal cancer or induce cell death through inhibiting autophagic flux, the present project intends to: ① use cell biology and cancer pharmacology means to confirm that though ROS, endoplasmic reticulum stress and other signals, saponins suppress autophagosome-lysosome fusion and thereby antagonize the protective effect of autophagy, reverse multidrug resistance and cause cell death; ② use chemical inhibitors and other tools to explore the crosstalk of DNA damage repair and autophagy signaling pathway in saponins-induced death; ③ explore the role of autophagy in esophageal cancer treatment in vivo by evaluating the antitumor effects of saponins alone or in combination with chemotherapeutic agents in nude mice. This work will clarify the mechanism of asaponins-mediated esophageal cancer cell death by inhibiting autophagosome-lysosome fusion, and provide a new strategy of esophageal cancer chemotherapy.

寻找和发现新的分子靶点和治疗手段已成为当前食管癌治疗的迫切需要。细胞自噬与肿瘤的发生发展以及耐药密切相关，调控自噬为肿瘤治疗提供了新思路。申请者在前期研究工作中：①筛选得到新的抑制自噬体－溶酶体融合的皂苷类单体；②5-Fu诱导保护作用的自噬；③皂苷类单体增强食管癌细胞对5-Fu的敏感性及诱导自噬相关的死亡。本项目拟以皂苷类单体抑制自噬流增强食管癌的化疗敏感性和诱导细胞死亡为切入点，①应用细胞生物学、肿瘤药理学手段，证实皂苷单体通过ROS，内质网应激等信号抑制自噬体－溶酶体融合，拮抗自噬的保护作用，从而逆转多药耐药及引起细胞死亡；②利用工具药等探究DNA损伤修复与自噬信号通路的串话在皂苷类单体诱导的死亡中的作用；③在裸鼠体内评价皂苷类单体单独或联合用药，探讨细胞自噬在治疗食管癌中的作用。本课题将为阐明皂苷单体抑制细胞自噬在食管癌细胞死亡的机制提供科学依据，同时提供一种全新的食管癌化疗策略。

寻找和发现新的分子靶点和治疗手段已成为当前食管癌治疗的迫切需要。细胞自噬与肿瘤的发生发展以及耐药密切相关，调控自噬为肿瘤治疗提供了新思路。在前期工作中，申请人筛选得到多种新的具有自噬调控作用的皂苷单体（如人参皂苷Ro），多项实验结果表明自噬通过选择性的调控周期蛋白的降解，促进DNA损伤下肿瘤细胞周期的运转和耐药的形成。而皂苷通过抑制细胞自噬，调控细胞周期-DNA损伤修复通路增强食管癌化疗敏感性，揭示了细胞自噬在肿瘤耐药中的潜在重要意义。在本项目中，我们着重探讨了绞股蓝皂苷Gyp-L对溶酶体调控的作用机制，明确皂苷Gyp-L通过脂筏介导的内吞作用，激活ROS-NOX2信号轴，进而促进转录因子TFEB入核，促进异常溶酶体的生成和碱化，最终诱导食管癌及肝癌细胞的死亡。此外，我们证明皂苷Gyp-L通过ROS激活 MAPK/NF-kB信号通路，导致细胞周期阻滞和衰老相关炎症因子的释放，诱导食管癌细胞和肝癌细胞发生衰老，皂苷Gyp-L还能增强肿瘤细胞对化疗药物的敏感性。最后，我们还筛选得到具有自噬调控作用的异硫氰酸盐萝卜硫素和莱菔素，其通过抑制自噬诱导食管癌细胞衰老。本课题将为皂苷Gyp-L作为先导化合物用于食管癌治疗提供药理基础，为调控自噬-溶酶体途径与衰老互作在肿瘤细胞死亡与耐药中决择的机理提供依据，同时也为食管癌治疗提供新的化疗策略。