Suppression of VEGF transcription in renal cell carcinoma cells by pyrrole-imidazole hairpin polyamides targeting the hypoxia responsive element
Suppression of VEGF transcription in renal cell carcinoma cells by pyrrole-imidazole hairpin polyamides targeting the hypoxia responsive element
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DOI：
10.1080/02841860500486648
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发表时间：
2006-04-01
期刊：
影响因子：
3.1
通讯作者：
Kihara, K
中科院分区：
文献类型：
--
作者：
Kageyama, Y;Sugiyama, H;Kihara, K
低氧诱导因子(HIF)是细胞在低氧条件下存活的关键基因的主要调节因子，已知与肾癌的发生和发展有关。N-甲基吡咯(Py)-N-甲基咪唑(Im)发夹状聚酰胺是一种人工合成的有机化合物，能够识别并结合特定DNA序列的细小凹槽。我们合成了三个针对血管内皮生长因子基因启动子区域缺氧反应元件侧翼序列(HRE；HIF结合部位)的Py-Im发夹状聚酰胺。用含有血管内皮生长因子启动子的报告质粒进行荧光素酶分析，评价了聚酰胺对HIF诱导转录的影响。采用实时定量逆转录聚合酶链式反应和酶联免疫吸附试验检测聚酰胺对具有von Hippel-Lindau基因移码突变的A498肾癌细胞中血管内皮生长因子转录和分泌的影响。在293细胞的报告分析中，三种Py-Im发夹多酰胺的组合抑制了HIF诱导的转录，并成功地抑制了A498细胞中血管内皮生长因子基因的转录和翻译。HIF-HRE相互作用的抑制通过凝胶迁移率改变分析得到证实。用Py-Im发夹状聚酰胺治疗肾癌可能是一种新的治疗策略。
Hypoxia inducible factor (HIF), a master regulator of critical genes for cell survival under hypoxic conditions, is known to be related to tumorigenesis and progression of renal cell carcinoma. N-methylpyrrole (Py)-N-methylimidazole (Im) hairpin polyamides are synthetic organic compounds that recognize and bind to the minor grooves of specific DNA sequences. We synthesized three Py-Im hairpin polyamides targeting the flanking sequences of hypoxia responsive element (HRE; a binding site of HIF) in the promoter region of the vascular endothelial growth factor ( VEGF) gene. The effects of the polyamides on HIF-induced transcription were evaluated by a luciferase assay using a reporter plasmid containing a VEGF promoter. Real time reverse-transcriptase polymerase chain reaction and enzyme-linked immunosorbent assay were performed to examine the effects of the polyamides on the transcription and secretion of VEGF in A498 renal cell carcinoma cells, which have a frame-shift mutation in the von Hippel-Lindau gene. A combination of three Py-Im hairpin polyamides suppressed HIF-induced transcription in reporter assays using 293 cells and successfully suppressed transcription and translation of the VEGF gene in A498 cells. Inhibition of the HIF-HRE interaction was confirmed by an electrophoresis mobility shift assay. An approach using Py-Im hairpin polyamides may be a new strategy for the treatment of renal cell carcinoma.