Overcoming apoptotic resistance in glioblastoma by CP-d/n-ATF5, a novel tumor spe

描述(申请人提供)：多形性胶质母细胞瘤(GBM)是最常见的原发脑肿瘤，目前无治疗方法，进展迅速，复发，导致12-18个月内死亡。转录因子ATF5是激活转录因子(ATF)/cAMP反应元件结合蛋白(CREB)家族的成员之一，与非肿瘤性星形胶质细胞和神经元相比，在GBM中高表达。在这项提议中，ATF5是一种新设计的多肽药物，称为CP-d/n-ATF5(细胞穿透显性负性ATF5)。初步研究表明，CP-d/n-ATF5在体内外均能有效杀伤GBM细胞。我们将确定CP-d/n-ATF5产生抗胶质瘤作用的机制，并测试其在体内外联合药物治疗中的适用性。在我们的初步数据中，我们已经表明，CP-d/n-ATF5介导了p73、DR5(促凋亡配体TRAIL的受体)和PUMA(促凋亡的Bcl-2家族蛋白)蛋白水平的显著增加。P73是肿瘤抑制基因P53的同系物，与TP53不同，P53在GBM中不常见突变，但具有相似的下游靶点，如DR5和PUMA，它们促进细胞凋亡(程序性细胞死亡)并促进细胞周期停滞。因此，诱导p73的试剂是非常有价值的肿瘤治疗药物。我们将检验这一假设，即CP-d/n-ATF5通过上调p73而杀死GBM细胞，而p73又通过PUMA依赖的凋亡诱导死亡。我们还将评估CP-d/n-ATF5诱导DR5依赖于p73的假设，以及升高的DR5将允许CP-d/n-ATF5与DR5配体TRAIL的GBM治疗相结合。我们与TRAIL/CP-d/n-ATF5组合的初步数据显示，与单一治疗相比，TRAIL/CP-d/n-ATF5联合治疗对GBM细胞具有协同杀伤作用。这些作用是由CP-d/n-ATF5诱导的DR5上调所介导的，并与p73蛋白水平的升高相一致。SiRNA对DR5的特异性抑制可减轻CP-d/n-ATF5/TRAIL介导的细胞死亡。这项研究计划将在劳埃德·格林博士(哥伦比亚大学病理学和细胞生物学系)的指导下进行，他在培训内科科学家方面有着重要的记录，并开创了许多成功和有成就的生物医学研究人员的职业生涯。AS联合导师将服务于Peter Canoll博士(哥伦比亚大学病理和细胞生物系)和Jeffrey Bruce博士(神经外科)。卡诺尔博士和布鲁斯博士是哥伦比亚大学脑瘤中心的负责人，他们都很有成就。这两家公司都有NIH资助的与胶质母细胞瘤动物模型相关的活跃研究项目，这些动物模型具有复杂的药物输送系统。这个拟议的项目，连同它著名和有成就的导师、课程工作、培训机会和职业发展计划，将培训申请者与胶质母细胞瘤相关的研究，重点是临床前药物开发，并帮助他成为一名独立的研究员。

DESCRIPTION (provided by applicant): Glioblastoma multiform WHO IV (GBM) is the most common primary brain tumor with no current curative treatment, rapid progression and recurrence, leading to death within 12-18 month. The transcription factor ATF5, a member of the activating transcription factor (ATF)/cAMP response-element binding protein (CREB) family, is over-expressed in GBMs compared to non-neoplastic astrocytes and neurons. In this proposal ATF5 is targeted by a novel designed peptide drug, called CP-d/n-ATF5 (cell penetrating dominant-negative ATF5). Preliminary data show that CP-d/n-ATF5 effectively kills GBM cells in vitro and in vivo. We will determine the mechanism by which CP-d/n-ATF5 elicits its anti-glioma effects and test its suitability for drug combination therapy in vitro and in vivo. In our preliminry data, we have shown that CP-d/n-ATF5 mediates striking increases in p73, DR5 (a pro-apoptotic receptor for death ligand TRAIL) and PUMA (a pro-apoptotic Bcl-2 family protein) protein levels. P73 is a homolog of the tumor suppressor p53, which, in contrast to TP53 is not commonly mutated in GBM, but which shares similar downstream targets, e.g. DR5 and PUMA, that enhance apoptosis (programmed cell death) and promote cell cycle arrest. Therefore, reagents that induce p73 are highly valuable tumor therapeutics. We will test the hypothesis that CP-d/n-ATF5 kills GBM cells by up- regulating p73 that in turn induces death via PUMA-dependent apoptosis. We will also evaluate the hypotheses that induction of DR5 by CP-d/n-ATF5 is dependent on p73 and that elevated DR5 will permit a combination GBM therapy of CP-d/n-ATF5 with the DR5 ligand TRAIL. Our preliminary data with the TRAIL/CP-d/n-ATF5 combination demonstrate synergistic killing of GBM cells as compared to single treatments. These effects were mediated by CP-d/n-ATF5-induced up-regulation of DR5, and coincided with an increase of p73 protein levels. Specific suppression of DR5 by siRNA mitigates CP-d/n-ATF5/TRAIL mediated cell death. The research program will be conducted under the guidance of Dr. Lloyd Greene (Department of Pathology and Cell Biology at Columbia University) who has a significant track record of training physician-scientists and who has launched many careers of successful and accomplished biomedical researchers. As Co-Mentors will serve Drs. Peter Canoll (Department of Pathology and Cell Biology at Columbia University) and Jeffrey Bruce (Department of Neurosurgery). Dr. Canoll and Dr. Bruce are leading the Brain Tumor Center at Columbia and are highly accomplished. Both have active NIH-funded research projects related to glioblastoma animal models with sophisticated drug delivery systems. This proposed project along with its renowned and accomplished mentors, course work, training opportunities and career development plan will train the applicant in glioblastoma-related research with an emphasis on preclinical drug development and assist him to become an independent investigator.