Neural Stem Cell-Selective Drug Target for Small Molecule Therapy of Brain Tumors

描述（由申请人提供）尽管目前的积极治疗结合了手术、放疗、化疗和贝伐单抗（Avastin）（一种靶向VEGF-A的中和抗体），但从多形性胶质母细胞瘤（GBM）或IV期胶质瘤（脑癌的最恶性形式）诊断时起的中位生存期约为一年。需要新的疗法来控制这种侵袭性癌症。目前对GBM的机制和遗传学研究发现，许多特性与神经干细胞（NSC）相同，如通过CNS的迁移、细胞外基质降解和分化为多个谱系，这表明NSC增殖的机制可以为GBM的治疗提供线索。我们建议在这里确定和表征孤儿受体的第一个功能性配体，孤儿受体是神经干细胞增殖所需的，并选择性地表达在室管膜下区和齿状回，在那里发现NSC. Receptor过表达驱动胶质瘤形成的小鼠和人类患者中，胶质瘤表达升高与生存率低高度相关。与许多参与干细胞功能的转录因子不同，这种受体对于发现靶向GBM的药物具有双重优势。一是 主要在NSC中表达，并且似乎在控制增殖和分化的节点处起作用;第二，它具有受小分子配体调节的潜力。因此，该受体的配体具有阻断GBM增殖和抑制肿瘤生长的潜力。在目标1中，我们计划使用120，000种化合物的多样化小分子文库进行高通量筛选，该方案采用了一种新型的严格特异性方案，排除了大多数假阳性配体，并已在Orphagen得到验证。将从市售类似物中鉴定出更有效的配体（EC 50 < 1.0 mM），并且在目标2中，我们开发了一种生物化学测定法来检测配体诱导的受体与关键功能相互作用蛋白相互作用的变化，以便通过互补受体筛选技术确认命中。在目的3中，将测试化合物对正常NSC基因表达和增殖的调节。将在临床来源的人GBM细胞培养物中进一步检查活性化合物，以确定它们是否以受体特异性方式阻断肿瘤细胞生长。如果成功， SBIR第2阶段资助的项目将提出发现和测试抑制免疫受损小鼠中人GBM异种移植物生长的配体。我们的首要目标是商业化一类新的非细胞毒性药物治疗GBM。 公共卫生相关性：胶质母细胞瘤（GBM）是成人中最常见的原发性脑癌，占2010年美国预计诊断的22，000例恶性脑肿瘤病例的一半以上。治疗可以延缓癌症进展，但总体中位生存期仍然大约为一年，尽管在过去30年中努力引入改进的治疗方法。我们建议确定第一个配体的转录因子，调节神经干细胞（NSC）增殖。该因子的高水平表达与胶质瘤患者的不良预后密切相关。我们的目标是开发一种口服生物可利用的小分子药物，使这种转录因子失活，并抑制GBM的侵袭性临床行为。

DESCRIPTION (provided by applicant) Median survival from time of diagnosis of glioblastoma multiforme (GBM) or stage IV glioma, the most malignant form of brain cancer, is about one year, despite the current aggressive treatment that combines surgery, radiotherapy, chemotherapy, and bevacizumab (Avastin), a neutralizing antibody targeting VEGF-A. Novel therapies are needed for controlling this aggressive cancer. Ongoing mechanistic and genetic studies of GBM find many properties in common with neural stem cells (NSCs), such as migration through the CNS, extracellular matrix degradation, and differentiation into multiple lineages, suggesting that mechanisms of NSC proliferation can provide clues for treatment of GBM. We propose here to identify and characterize the first functional ligands to an orphan receptor that is required for neural stem cell proliferation and is selectively expressed in the subventricular zone and the dentate gyrus, where NSCs are found. Receptor overexpression drives glioma formation in mice and, in human patients, elevated expression in glioma is highly correlated with poor survival. Unlike many of the transcription factors involved in stem cell function, this receptor has a dual advantage for the discovery of drugs to target GBM. First, it is primarily expressed in NSCs and appears to act at a nodal point for control of proliferation and differentiation; and second, it has the potential to be regulated by small molecule ligands. Ligands to this receptor thus have the potential to block GBM proliferation and suppress tumor growth. In Aim 1, we plan high throughput screening with a diverse small molecule library of 120,000 compounds utilizing a novel, rigorous specificity protocol that excludes the majority of false positive ligands and has been validated at Orphagen. More potent ligands (EC50 < 1.0 mM) will be identified from commercially available analogues and, in Aim 2, we develop a biochemical assay to detect ligand-induced changes in receptor interaction with a key functional interacting protein in order to confirm hits by a complementary receptor screen technology. In Aim 3, compounds will be tested for regulation of normal NSC gene expression and proliferation. Active compounds will be further examined in clinically-derived human GBM cell cultures to determine if they block tumor cell growth in a receptor-specific manner. If successful, an SBIR Phase 2-funded project would propose to discover and test ligands that suppress human GBM xenotransplant growth in immunocompromised mice. Our overarching goal is to commercialize a novel class of non-cytotoxic drug for treatment of GBM. PUBLIC HEALTH RELEVANCE: Glioblastoma (GBM) is the most common primary brain cancer in adults, accounting for more than half of the 22,000 malignant brain tumor cases expected to be diagnosed in the US in 2010. Treatment can delay cancer progression, but overall median survival remains roughly one year, despite efforts to introduce improved therapies over the past 30 years. We propose to identify the first ligands to a transcription facto that regulates neural stem cell (NSC) proliferation. High level expression of this factor is closel linked to poor prognosis in glioma patients. Our goal is to develop an orally bioavailable small molecule drug that inactivates this transcription factor and inhibits the aggressive clinical behavior of GBM.