缺氧及辐射双敏感性启动子增强抑瘤素M表达治疗肺癌的实验研究

Oncostatin M gene therapy in mice bearing lung adenocarcinoma xenograft using a hypoxia/radiation dual-sensitive promoter

目的 构建缺氧及辐射双敏感性启动子 ,增强缺氧条件下放射诱导的抑瘤素M (OSM)表达水平 ,提高肺癌放射 基因治疗效果。方法 利用基因重组构建缺氧及辐射双敏感性缺氧反应元件早期生长反应基因 (HRE Egr)启动子及其调控的OSM表达载体 ;以脂质体转染肺癌A5 4 9细胞 ,给予照射 ( 6Gy)和 (或 )缺氧 ( 1%氧浓度 )处理 ,观察其OSM表达水平及细胞存活率的变化。利用肺癌移植瘤观察不同处理后的抑瘤效应。结果 HRE Egr启动子具有辐射及缺氧双重敏感性 ,它可使照射后的肺癌细胞OSM表达水平在缺氧条件下显著提高 ,为常氧条件下的 3倍。在缺氧条件下 ,重组质粒 pHEO转染合并照射处理后细胞存活率 [( 9 5± 2 8) % ]显著低于常氧组 [( 34 8± 3 6 ) % ;χ2 =11 375 ,P =0 0 0 3]。HRE Egr启动子转染合并照射可明显抑制肺癌移植瘤 ,并可使 6 0 %的移植瘤完全消退。结论 用HRE Egr启动子提高OSM表达水平 ,可增强肺癌移植瘤放射

Objective To improve the efficacy of radiogenetic therapy for lung cancer, a hypoxia/radiation dual sensitive promoter was constructed to enhance the expression of oncostatin M (OSM) in transfected cells exposed to radiation under hypoxia Methods The chimeric promoter HRE Egr was generated by insertion of hypoxia response elements(HREs) upstream of the Early growth response gene 1(Egr 1) promoter OSM expression vector was constructed by cloning HRE Egr promoter upstream of OSM gene, which was transfected into A549 cells The expression of OSM in transfected cells exposed to irradiation and(or) hypoxia was analyzed, and the relative survival rate of transfected cells exposed to the above conditions was tested To examine the efficacy of this HRE Egr OSM gene therapy in vivo, the tumor suppression effects were investigated in 40 nude mice bearing lung adenocarcinoma xenograft Results Expression of OSM gene in transfected cells exposed 6 Gy irradiation was markedly increased under hypoxia A gene therapy experiment in vitro showed that the survival rate of transfected cells exposed to radiation under hypoxia was obviously decreased with comparison of cells under normoxia HRE Egr promoter transfected tumors regressed significantly after a combination therapy of irradiation and HRE Egr transfection in all mice ( n =10), and six tumors disappeared in 3 weeks without any side effects Conclusion The data indicate that tumor targeted expression of OSM gene under the control of a hypoxia/radiation dual sensitive promoter represents a novel strategy for safe and effective gene therapy of lung carcinoma and might have clinical application in the future