Epigenetic Silencing of Death Receptor 4 Mediates Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand Resistance in Gliomas
Epigenetic Silencing of Death Receptor 4 Mediates Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand Resistance in Gliomas
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DOI：
10.1158/1078-0432.ccr-09-1125
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发表时间：
2009-09-01
影响因子：
11.5
通讯作者：
Mueller, Wolf
中科院分区：
文献类型：
--
作者：
Elias, Agnes;Siegelin, Markus D.;Mueller, Wolf
目的：鉴定和表征表观遗传调控基因，这些基因能够预测胶质瘤治疗中对目前测试的化疗药物的敏感性或耐药性。实验设计：我们使用甲基化敏感的BeadArray技术来鉴定与细胞凋亡和胶质瘤治疗中潜在治疗靶点相关的新型表观遗传调控基因。为了阐明目标死亡受体4 （DR4）启动子甲基化的功能后果，我们在DR4缺失的胶质瘤细胞系（U373和A172）和DR4强表达的胶质瘤细胞系（LN18）中研究了肿瘤坏死因子相关凋亡诱导配体（TRAIL）介导和抗DR4介导的细胞凋亡。结果：在人类星形细胞肿瘤中，我们在WHO 2级弥漫性星形细胞瘤中检测到60% （n = 5）的DR4启动子高甲基化，在WHO 3级间变性星形细胞瘤中检测到75% （n = 8）的DR4启动子高甲基化，在WHO 4级胶质母细胞瘤中检测到70% （n = 33）的DR4启动子高甲基化。DR4是一种局限于胶质瘤细胞的细胞表面蛋白，是TRAIL的靶标。胶质瘤细胞系U373和A172含有重度甲基化的DR4启动子，5-aza-2-脱氧胞苷介导的去甲基化重组了这些细胞系中DR4的表达。DR4特异性小干扰RNA在trail敏感胶质瘤细胞系LN18中功能性敲低DR4可显著减轻拮抗DR4抗体诱导的细胞凋亡。5- aza -2-脱氧胞苷介导的去甲基化导致trail抗性胶质瘤细胞系U373细胞表面DR4的功能重构，并使U373对trail介导的凋亡敏感。在去甲基化的U373中，通过小干扰RNA抑制DR4成功地重建了U373的trail抗性表型。结论：DR4启动子甲基化在人类星形细胞胶质瘤中很常见，DR4的表观遗传沉默介导了对基于TRAIL/DR4的胶质瘤治疗的耐药性。[临床癌症杂志，2009;15(17):5457-65]
Purpose: To identify and characterize epigenetically regulated genes able to predict sensitivity or resistance to currently tested chemotherapeutic agents in glioma therapy.Experimental Design: We used methylation-sensitive BeadArray technology to identify novel epigenetically regulated genes associated with apoptosis and with potential therapeutic targets in glioma therapy. To elucidate the functional consequences of promoter methylation in the identified target death receptor 4 (DR4), we investigated tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated and anti-DR4-mediated apoptosis in glioma cell lines (U373 and A172) with loss of DR4 and one glioma cell line (LN18) with robust DR4 expression.Results: In human astrocytic tumors, we detected DR4 promoter hypermethylation in 60% (n = 5) of diffuse astrocytomas WHO grade 2, in 75% (n = 8) of anaplastic astrocytomas WHO grade 3, and in 70% of glioblastomas WHO grade 4 (n = 33). DR4 is a cell surface protein restricted to glioma cells and is targeted by TRAIL. Glioma cell lines U373 and A172 harbored heavily methylated DR4 promoters, and 5-aza-2-deoxycytidine-mediated demethylation reconstituted DR4 expression in these cell lines. Functional knockdown of DR4 by DR4-specific small interfering RNA in TRAIL-sensitive glioma cell line LN18 significantly mitigated apoptosis induced by an agonistic anti-DR4 antibody. 5-Aza-2-deoxycytidine-mediated demethylation resulted in a functional reconstitution of DR4 on the cell surface of TRAIL-resistant glioma cell line U373 and sensitized U373 to TRAIL-mediated apoptosis. Suppression of DR4 by small interfering RNA in demethylated U373 successfully reestablished the TRAIL-resistant phenotype of U373.Conclusions: DR4 promoter methylation is frequent in human astrocytic gliomas, and epigenetic silencing of DR4 mediates resistance to TRAIL/DR4-based glioma therapies. (Clin Cancer Res 2009;15(17):5457-65)