A tumor-specific gene therapy strategy targeting dysregulation of the VHL/HIF pathway in renal cell carcinomas
A tumor-specific gene therapy strategy targeting dysregulation of the VHL/HIF pathway in renal cell carcinomas
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DOI：
10.1111/j.1349-7006.2005.00044.x
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发表时间：
2005-05-01
期刊：
影响因子：
5.7
通讯作者：
Hiraoka, M
中科院分区：
文献类型：
--
作者：
Ogura, M;Shibata, T;Hiraoka, M
低氧诱导因子是低氧依赖基因表达的关键转录因子，在血管生成和肿瘤生长中发挥重要作用。VHL蛋白结合到(HIF-α)的a亚基上，进行氧依赖降解。VHL基因突变常见于散发性肾细胞癌。VHL的破坏导致HIF-α异常积聚，导致下游基因上调，如血管内皮生长因子基因。我们构建了由低氧反应元件驱动的荧光素酶报告载体(5HRE/Luc)和表达单纯疱疹病毒胸苷激酶基因的治疗性载体(5HRE/tk)。在VHL缺陷的786-O细胞的瞬时转染实验中，在有氧和低氧条件下都检测到了组成型荧光素酶的表达。相反，转染野生型VHL的786-O细胞显示出低氧诱导的荧光素酶活性。在体外MTS实验中，在有氧和缺氧条件下，0.2mgGCV对稳定表达5HRE/tk的786-O细胞的生长抑制率均达到50%。SCID小鼠移植的稳定克隆在每日注射GCV(50 mg/kg)10天后有明显的消退。综上所述，低氧反应载体可能对VHL突变的肾癌具有治疗潜力。
Hypoxia-inducible factors, key transcription factors for hypoxia-dependent gene expression, play important roles in angiogenesis and tumor growth. The VHL protein binds to the a subunit of (HIF-alpha) for its oxygen-dependent degradation. VHL mutations are found frequently in sporadic RCC. Disruption of VHL results in an abnormal accumulation of HIF-alpha, leading to the upregulation of downstream genes such as the vascular endothelial growth factor gene. We constructed a luciferase reporter vector driven by hypoxia-responsive elements (5HRE/luc) and a therapeutic vector expressing a herpes simplex virus thymidine kinase gene (5HRE/tk). In the transient transfection assay using VHL-deficient 786-O cells, constitutive luciferase expression was detected under both aerobic and hypoxic conditions. In contrast, 786-O cells transfected with a wild-type VHL showed hypoxia-inducible luciferase activity. In in vitro MTS assay, 50% of growth inhibition of 786-O cells stably transfected with 5HRE/tk was achieved with exposure to 0.2 mu g/mL of GCV under both aerobic and hypoxic conditions. Xenografts of the stable clone in SCID mice exhibited a marked regression on daily injections of GCV (50 mg/kg) for 10 days. In conclusion, a hypoxia-responsive vector may have therapeutic potential for RCC with VHL mutations.