EphA2介导的细胞自噬在鼻咽癌紫杉醇耐药中的作用及分子机制研究

Autophagy has been a hot spot in drug resistance of cancer chemotherapy. Our previous study has clearly demonstrated that EphA2 upregulation, on the one hand, led to drug resistance of Paclitaxel in human poor differentiated nasopharyngeal carinoma cell line 5-8F. On the other hand, EphA2 overexpression induced the increased formation of autophagosome and upregulation of autophagy-associated biomarker protein LC3-Ⅱ, indicating the potential associatin between EphA2 and autophagy phenomenon. Therefore, combined with the close association between autophagy and drug resistance of chemotherapy, we hypothesize that EphA2 may lead to drug resistance of chemotherapy in nasopharyngeal carcinoma (NPC) via modulating the process of autophagy. In order to test the hypothesis, we shall utilize lentivirus-mediated EphA2 overexpression and silencing vector to change EphA2 expression level in NPC cell lines. Autophagy process and its down signalling pathway and the drug sensitivity of NPC to paclitaxel are observed. Morever, autophagy process is impeded or enhanced to observe whether the changed response to paclitaxel and cisplatin mediated by EphA2 will be reversed. In summary, we clarify that EphA2 lead to drug resistance of chemotherapy in NPC via regulating autophagy process, which will provide novel theory and experimental data in the field of drug resistance in cancer chemotherapy.

自噬作用为目前肿瘤化疗耐药研究中的热点。前期研究表明：EphA2过表达后，①鼻咽部低分化鳞癌细胞5-8F对化疗药物紫杉醇的敏感性显著降低；②能引起自噬小体增加和自噬相关标记物LC3-Ⅱ蛋白表达升高，提示EphA2过表达能在鼻咽癌细胞中诱导出自噬现象。结合目前研究表明自噬作用与化疗耐药关系密切，我们设想EphA2可能通过介导自噬作用调控鼻咽癌细胞对紫杉醇的敏感性。为此我们利用慢病毒介导的EphA2过表达和沉默载体，在体内外实验中，改变鼻咽癌细胞EphA2的表达，观察改变后自噬及其通路蛋白的改变和对紫杉醇敏感性的影响；再通过阻断或诱导自噬作用，观察EphA2 改变所致的紫杉醇敏感性改变能否逆转。从而阐明EphA2 介导的自噬作用在鼻咽癌细胞紫杉醇化疗敏感性中的重要作用，为肿瘤紫杉醇耐药研究提供新的理论依据和实验基础。

紫杉醇作为鼻咽癌（NPC）的一线临床用药，其耐药性的产生严重影响着鼻咽癌患者的临床治疗效果、生存质量及预后。因此，探讨紫杉醇耐药的产生机理有助于提高其临床效率。本研究首先发现EphA2蛋白在NPC组织及细胞中的表达较鼻咽部非肿瘤组织中的表达明显升高，且其表达水平与NPC的T分级、临床分期和颈部淋巴结转移密切相关。再通过慢病毒介导EphA2过表达和沉默载体，在多株鼻咽癌细胞中证实EphA2对紫杉醇的药物敏感性具有调控作用。进一步机制研究表明，该调控作用的产生与三个方面因素相关：①自噬对NPC化疗药物敏感性具有调控作用，EphA2的表达改变能影响NPC细胞自噬的发生。②EphA2通过PI3-K/Akt/GSK-3β介导的信号通路影响细胞周期调控蛋白的表达，进而调控NPC细胞对紫杉醇的敏感性。③EphA2可通过PI3-K/Akt通路介导多药耐药P-gp蛋白的表达，进而调控NPC细胞紫杉醇敏感性。综上所述，该项目基本按照研究计划进行，达到预期研究目的，初步阐明EphA2介导的自噬作用和非自噬途径在鼻咽癌紫杉醇化疗敏感性中的作用，为肿瘤紫杉醇耐药研究提供了新的实验数据和理论基础。