SBP1表达下调/缺失在肺鳞癌发生发展中的作用与分子机制

In our previous study,we found that selenium binding protein 1(SBP1) was progressively downregulated in human bronchial epithelial carcinogenic process.SBP1 expression being knockdown could increase the susceptibility of human bronchial epithelial cell(16HBE) transformation induced by B[a]P.To investigate the role and its molecular mechanism of SBP1 during the onset and progression of lung squamous cell carcinoma(LSCC),we will construct NCI-H520 cell line upregulating SBP1 expression using gene transfection,and 16HBE cell line downregulating SBP1 expression using gene knocked down by siRNA technology,and the model of malignant transformation of bronchial epithelial cell and the xenograft model of LSCC.Effect of SBP1 expression change on the occurrence and development of LSCC in vivo and in vitro will be studied,and on oxidative stress,DNA damage,and P53 DNA repair pathway activity will be explored.Then,we will identify the proteins interacting with SBP1 using targeted proteomics technology, and build the protein regulatory networks of SBP1 during the onset and progression of LSCC,and screen the important target proteins regulated by SBP1,and find the key signaling pathway related to the occurrence and development of LSCC by bioinformatics analysis.Next,we will detect SBP1 expression in various stage tissues of bronchial epithelial carcinogenesis by Immunohistochemistry,and evaluate the clinical significance of SBP1 as a early diagnosis and prognosis marker.This study not only helps us to elucidate the role and molecular mechanism of SBP1 during the onset and progression of LSCC,but also can provide scientific basis for chemoprevention and targeted treatment of LSCC.

我们前期研究发现：硒结合蛋白1(SBP1)在支气管上皮细胞癌变过程中逐步下调；在永生化的人支气管上皮细胞(16HBE)中，沉默SBP1的表达可增强其对苯并芘诱癌的敏感性。为探讨SBP1在肺鳞癌发生发展中的作用及其机制，本项目拟建立SBP1表达上调的肺鳞癌细胞系及SBP1表达下调的16HBE细胞系、支气管上皮细胞恶性转化模型和裸鼠移植瘤模型，体外和体内研究SBP1表达变化对肺鳞癌发生发展的影响；分析SBP1表达变化对细胞氧化应激、DNA损伤修复以及p53 DNA修复通路活性的影响，采用靶向蛋白质组学技术鉴定SBP1的相互作用蛋白，挖掘SBP1涉及的信号通路；采用免疫组化技术检测支气管上皮癌变各阶段组织中SBP1的表达，分析SBP1在肺鳞癌早期诊断和预后评估中的价值。本研究不仅有助于揭示SBP1表达下调/缺失在肺鳞癌发生发展中的作用及分子机制，而且能为肺鳞癌的化学预防和靶向治疗提供科学依据。

我们前期研究发现：硒结合蛋白1(SBP1)在支气管上皮细胞癌变过程中逐步下调；在永生化的人支气管上皮细胞(16HBE)中，沉默SBP1的表达可增强其对苯并芘诱癌的敏感性。为探讨SBP1在肺鳞癌发生发展中的作用及其机制，本项目首先建立了SBP1表达上调的肺鳞癌细胞系NCI-H520-SBP1及SBP1表达下调的16HBE-shSBP1细胞系，然后建立了支气管上皮细胞恶性转化模型，在体外研究SBP1表达变化对肺鳞癌发生发展的影响；分析SBP1表达变化对细胞氧化应激、DNA损伤修复以及p53 DNA修复通路活性的影响，采用免疫组织化学技术检测了SBP1在支气管上皮癌变各阶段组织中的表达水平，研究了SBP1的表达水平与临床病理特征、复发及生存期之间的关系，分析了SBP1作为肺鳞癌复发与预后标志物的临床价值。通过研究，我们证实SBP1表达下调可促进B[a]P诱导16HBE恶性转化和16HBE细胞的生长，增强细胞的侵袭能力，减少细胞凋亡，降低细胞G0/G1期百分率，增加S期的百分率。SBP1表达下调可降低SOD、GSH/GSSG、MnSOD在16HBE细胞中的表达水平，增加ROS、MDA 、8-OhdG在16HBE细胞中的表达水平，使细胞增殖加快，细胞凋亡减少，说明SBP1表达下调降低细胞抗氧化能力，导致DNA氧化损伤。SBP1表达下调不仅可抑制P53的表达水平，降低P53的生物活性，也能显著抑制P53、Gadd45a、PCNA和APE1/Ref-1的表达，从而抑制P53的下游基因Gadd45a与 PCNA及APE1/Ref-1的相互作用，进而抑制依赖P53的碱基切除修复(base-excision repair, BER)，促进肺鳞癌的发生。SBP1的表达水平与病人的性别、年龄、吸烟状况和肿瘤大小无关，而与肿瘤的TNM分期、淋巴结转移、远处转移、复发率和生存率密切相关。SBP1表达越低，其复发的可能性越大，生存率越低，SBP1低表达是肺鳞癌病人独立的预后因子。SBP1可与TNM分期、淋巴结转移、远处转移一起作为肺鳞癌预后因子，SBP1有望成为肺鳞癌的预后标志物。至今，已发表或接收标注项目资助的SCI论文2篇，CSCD论文3篇。