MiR-133a调控神经胶质瘤中TRAIL耐受性及分子机理研究

Aim The aim of this project is to investigate the role of miRNA-133a in mediating TRAIL-inducing apoptotic resistance in vitro and in vivo so as to identify its clinical potential as a promising therapeutic target for TRAIL resistant glioblastoma. Project Background Glioblastoma multiforme (GBM) is the most aggressive form of brain tumor, and despite multimodal treatment approaches, the patients have poor prognoses and the average length of survival is only about 12-15 months. Recombinant tumor necrosis factor (TNF)-related apoptosis inducing ligand (TRAIL) is currently being tested in phase II/III clinical studies as a novel cancer therapy; however, a big obstacle to its widely clinical application is TRAIL resistance in many tumors. Therefore, sensitizing cancer cells to TRAIL-induced apoptosis may constitute an effective therapeutic strategy to fight TRAIL-resistant glioblastoma. Brief Project Description In this proposal, we are exploring an alternative approach to reduce TRAIL resistance through activating its specific death receptors DR4 and DR5. Although many cancer cells frequently express both receptors, DR5 expression was significantly higher than DR4 in glioblastoma. Therefore, activating DR5 might be a more effective way to lower TRAIL-resistance. In our preliminary studies, microRNA-133a was found to correlate positively with TRAIL-resistance by microarray and bioinformatics analyses. In addition, DR5 expression was suppressed by miR-133a mimics and activated by anti-miR-133a. We further investigated the effect of miR-133a on TRAIL-induced apoptosis and found that miR-133a may be required to maintain the TRAIL-resistant phenotype in glioma. And we also found that caspase-dependent apoptotic pathway and alternative NF-κB signaling may be involved in this process. Therefore, these data support our hypothesis that miR-133a mediates TRAIL-resistance in glioma through suppressing DR5 expression directly and regulating caspase-dependent apoptotic pathway and NF-κB signaling. Significance of the Project In the present proposal, we will: (1) characterize the effects of miR-133a on TRAIL resistance in vitro and in vivo; (2) elucidate the underlying mechanism and (3) investigate its therapeutic potential in glioma specimens. The information generated from this study will improve our understanding to the pathology of TRAIL resistance and explore the potential of miR-133a as a promising therapeutic target for TRAIL resistant glioblastoma.

TRAIL是一种新型蛋白药物，目前在美国正进行II/III临床研究。研究表明，TRAIL主要是通过与死亡受体DR4/5结合来诱导肿瘤细胞凋亡，但一半以上的肿瘤对TRAIL产生耐受性。到目前为止，关于肿瘤产生TRAIL耐受性的原因还不清楚。在神经胶质瘤细胞中，TRAIL的耐受性主要与DR5的分布有关。本项目组在前期研究中发现，miR-133a调控神经胶质瘤细胞TRAIL的耐受性，更为重要的是miR-133a直接抑制DR5的表达。由此激活的相关信号通路可能是TRAIL耐受性的分子机制。本项目以miR-133a/DR5为切入点，拟从细胞水平、分子水平、蛋白水平、动物水平及组织水平探讨miR-133a对神经胶质瘤细胞中TRAIL耐受性的影响，并深入阐明其分子机理及相关信号通路，为miR-133a作为一个神经胶质瘤治疗的新型分子靶标提供更充分的科学依据。

神经胶质瘤是脑肿瘤中最富挑战性的一种恶性肿瘤，居颅脑肿瘤发病率的首位。因耐药性以及高度迁移性使得神经胶质瘤成为了最致命的肿瘤之一。虽然多模式治疗方法已取得了很大的进展，但病人预后较差，现在的平均存活时间仅为12-15个月。而且近年来，中国、台湾及香港的年青人中神经胶质瘤的发病率呈大幅度的上升趋势。因此，开发新的治疗方法具有重要的临床意义和研究价值。..肿瘤坏死因子相关凋亡诱导配体(TRAIL)是TNF超家族新成员。因为它能诱导多种肿瘤细胞凋亡同时对正常细胞没有明显影响使其成为了肿瘤治疗的一个具有巨大潜力的生物制剂，目前已经在美国进行II/III临床研究。TRAIL诱导细胞凋亡主要是通过与靶细胞表面特异性的受体结合来实现的。其中最重要的表面受体是被称死亡受体的DR4和DR5，TRAIL与他们结合后，其死亡信号传导至胞质，激活半胱天冬氨酸蛋白酶(caspase)系统，最终引起细胞凋亡。而在正常的组织细胞中中另外存在着一种诱骗受体DcR1和DcR2。DcR1没有死亡结构域，而是通过一个糖磷酯酰肌醇的尾部锚链到细胞膜上；而DcR2有一个截短的、无功能的死亡结构域，二者都不能传递TRAIL介导的死亡信号，因此不具有诱导细胞内凋亡信号传导的功能，只能竞争性的与配体结合，起到保护作用。许多研究表明诱骗受体仅存在于正常组织，肿瘤细胞没有分布。因此，TRAIL能选择性诱导肿瘤细胞凋亡，而对正常细胞无明显毒性作用，故作为一种新型的肿瘤治疗生物制剂，具有良好的临床应用前景。..microRNA（miRNA）是一类内源性的非编码的微小RNA，它通过调控靶标基因转录后水平的表达，从而广泛的参与生命活动的整个过程。到目前为止，已经发现有一些miRNAs能调控肿瘤细胞的TRAIL耐受性，如miR-221/222，miR-130等。在本项目的研究中，我们发现DR5的表达与神经胶质瘤TRAIL的耐受程度呈负向关系，生物信息学预测和生物学结果发现miR-133a直接靶标DR5，并且体内体外实验证明miR-133a能直接TRAIL在神经胶质瘤中的耐受性，并且激活NF-kB信号通路。因此，miR-133a可以调控TRAIL在肿瘤中的耐受性，为神经胶质瘤治疗提供一个新的思路。