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

描述（由申请人提供）：WHO IV型多形性胶质母细胞瘤（GBM）是最常见的原发性脑肿瘤，目前没有治愈治疗，进展和复发迅速，可在12-18个月内导致死亡。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，与非肿瘤性星形胶质细胞和神经元相比，在GBMs中过度表达。在这个提议中，ATF5被一种新的肽药物靶向，称为CP-d/n-ATF5（细胞穿透显性阴性ATF5）。初步数据显示，CP-d/n-ATF5在体外和体内均能有效杀伤GBM细胞。我们将确定CP-d/n-ATF5引发其抗胶质瘤作用的机制，并在体外和体内测试其药物联合治疗的适用性。在我们的初步数据中，我们已经表明，CP-d/n-ATF5介导p73、DR5（死亡配体TRAIL的促凋亡受体）和PUMA（促凋亡Bcl-2家族蛋白）蛋白水平的显著增加。P73是肿瘤抑制因子p53的同源物，与TP53不同，p53在GBM中不常见突变，但它们具有相似的下游靶点，如DR5和PUMA，可增强细胞凋亡（程序性细胞死亡）并促进细胞周期阻滞。因此，诱导p73的试剂是非常有价值的肿瘤治疗药物。我们将验证CP-d/n-ATF5通过上调p73杀死GBM细胞，进而通过puma依赖性凋亡诱导死亡的假设。我们还将评估CP-d/n-ATF5对DR5的诱导依赖于p73的假设，以及升高的DR5将允许CP-d/n-ATF5与DR5配体TRAIL联合治疗GBM。我们的初步数据显示，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上调介导的，并与p73蛋白水平的升高相一致。siRNA特异性抑制DR5可减轻CP-d/n-ATF5/TRAIL介导的细胞死亡。该研究项目将在Lloyd Greene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。作为共同导师，将为博士提供服务。Peter Canoll（哥伦比亚大学病理与细胞生物学系）和Jeffrey Bruce（神经外科学系）。卡诺尔博士和布鲁斯博士是哥伦比亚大学脑肿瘤中心的负责人，他们成就斐然。他们都有美国国立卫生研究院资助的研究项目，与胶质母细胞瘤动物模型和复杂的药物输送系统有关。该项目将提供知名导师、课程、培训机会和职业发展计划，培养申请人在胶质母细胞瘤相关研究方面的能力，重点是临床前药物开发，并帮助他成为一名独立研究者。

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.