Department of Urology, Keio University School of Medicine, Tokyo, Japan.
Urology. 2010 Apr;75(4):805-12. doi: 10.1016/j.urology.2009.11.048. Epub 2010 Feb 13.
To explore the potential therapeutic effects of the nuclear factor-kappaB (NF-kappaB) inhibitor dehydroxymethylepoxyquinomicin (DHMEQ). KU-19-19 cells, originally derived from a patient with invasive bladder cancer who exhibited marked leukocytosis, produce multiple cytokines. This model of clinically advanced bladder cancer, in which NF-kappaB is constitutively activated, was used in this study.
Expression of p65 protein in fractionated KU-19-19 cells was determined by Western blotting analysis. DNA-binding activity of NF-kappaB was detected by electrophoretic mobility shift assay. The cytotoxic effects and induction of apoptosis by DHMEQ were analyzed, and cytokines in the supernatant of KU-19-19 cells cultured with or without DHMEQ were measured by enzyme-linked immunosorbent assay (ELISA). Athymic nude mice bearing KU-19-19 subcutaneous tumors were subjected to intraperitoneal administration of 2 mg/kg/d DHMEQ for 3 weeks. Tumor growth was monitored and microvessel density, vascular endothelial growth factor expression, and the apoptotic index of tumors were evaluated by tissue immunohistochemistry.
NF-kappaB was constitutively activated in KU-19-19 cells. DHMEQ reversibly inhibited the DNA-binding activity of NF-kappaB by blocking its nuclear translocation. Both cell viability and production of cytokines were significantly and dose-dependently suppressed by DHMEQ, and significant apoptosis was also induced. In in vivo studies, the mean tumor volume in mice treated with DHMEQ was significantly smaller than in controls. Immunohistochemical analysis of tumors revealed marked reduction in microvessel density, vascular endothelial growth factor expression, and induction of apoptosis.
Blockade of NF-kappaB function by DHMEQ may be a useful new molecular targeting treatment for highly aggressive bladder cancer.
探讨核因子-κB（NF-κB）抑制剂去甲氧基环氧小檗碱（DHMEQ）的潜在治疗作用。KU-19-19 细胞最初源自一名患有浸润性膀胱癌的患者，该患者表现出明显的白细胞增多症，并产生多种细胞因子。本研究使用了这种临床上进展期膀胱癌的模型，其中 NF-κB 持续激活。
通过 Western 印迹分析确定分馏的 KU-19-19 细胞中 p65 蛋白的表达。通过电泳迁移率变动分析检测 NF-κB 的 DNA 结合活性。分析 DHMEQ 的细胞毒性作用和诱导细胞凋亡的作用，并通过酶联免疫吸附试验（ELISA）测量培养有或没有 DHMEQ 的 KU-19-19 细胞上清液中的细胞因子。将荷有 KU-19-19 皮下肿瘤的裸鼠进行腹腔内注射 2mg/kg/d 的 DHMEQ 治疗 3 周。通过组织免疫组织化学评估肿瘤生长、微血管密度、血管内皮生长因子表达和肿瘤的凋亡指数。
NF-κB 在 KU-19-19 细胞中持续激活。DHMEQ 通过阻断其核易位可逆地抑制 NF-κB 的 DNA 结合活性。DHMEQ 显著且剂量依赖性地抑制细胞活力和细胞因子的产生，并诱导显著的细胞凋亡。在体内研究中，用 DHMEQ 治疗的小鼠的平均肿瘤体积明显小于对照组。肿瘤的免疫组织化学分析显示微血管密度、血管内皮生长因子表达和诱导的细胞凋亡明显减少。
DHMEQ 阻断 NF-κB 功能可能是一种针对高度侵袭性膀胱癌的有用的新型分子靶向治疗方法。
