In Vivo Drug Testing of Pediatric CNS Tumors Using Patient Derived Orthotopic Xenograft Models

 描述(由申请人提供)：本申请是为了响应资助机会而准备的：儿科临床前试验联盟研究计划(U01)，RFA-CA-14-018。具体地说，我们提出了B型：中枢神经系统肿瘤(CNS)活体测试的研究计划。脑瘤是儿童癌症相关死亡的主要原因。临床药物开发的挑战之一是如何有效地对候选药物进行优先排序，以确保癌症患者的临床成功。由于儿科候选药物的数量比患者数量少得多，因此有必要进行全面的临床前测试，以确定最有可能在临床上有效的研究药物。然而，由于缺乏临床上相关的和分子上准确的模型系统，这种努力多年来一直受阻。幸运的是，我们已经通过将患者肿瘤标本直接注射到SCID小鼠的大脑中，建立了一大组(&gt；70)儿童脑肿瘤患者来源的原位异种移植(PDOX)模型。这些PDOX模型被证明复制了原始原发肿瘤的组织病理学特征、侵袭表型和主要遗传异常(基因表达、DNA拷贝数和基因突变)，即使在小鼠脑内连续亚移植的过程中也是如此。异种移植的肿瘤细胞也可以被冷冻保存，以供持续和按需供应致瘤的PDOX细胞。这种能力与我们优化的手术程序相结合，每天可以植入多达260只小鼠，使我们能够每年为每种肿瘤类型测试多种(例如，6-10种)药物。因此，我们的目标是利用这组独特的PDOX模型来检验新药物的治疗效果，并分析高级别胶质瘤、髓母细胞瘤和室管膜瘤的作用机制和治疗阻力。我们的假设是，这些患者特异性的PDOX肿瘤对抗癌治疗的反应与相应的人类原发肿瘤相似，通过该系统确定的有效药物将有更好的临床成功机会。为了验证这一假说，我们将进行一系列体外和体内试验，以达到下列目标：1)通过数据挖掘我们的小鼠模型分子特征数据库，寻找基因上准确的候选PDOX新药的治疗靶点(S)；2)通过体外功能筛选选择反应最灵敏的模型，以确定时间和剂量效应；3)在多个靶向携带PDOX的模型中验证新药的治疗效果；4)详细分析体内外细胞杀伤的细胞和分子机制以及产生治疗耐药的原因。我们的新型PDOX小鼠模型小组代表了儿童中枢神经系统肿瘤的广泛遗传异常。所有的分析都是在我们的实验室中建立并常规进行的；我们处于独特的地位，可以在体内完成拟议的药物研究。我们的发现应该提供强有力的临床前证据来支持临床试验的启动。

 DESCRIPTION (provided by applicant): This application is prepared in response to the funding opportunity: Pediatric Preclinical Testing Consortium Research Programs (U01), RFA-CA-14-018. Specifically, we propose for Type B: Research Program for tumors of central nervous system (CNS) in vivo testing. Brain tumors are the leading cause of cancer-related death in children. One of the challenges in clinical drug development is how to effectively prioritize drug candidates to ensure clinical success in cancer patients. As pediatric drug candidates are much more numerous than the small number of patients, it is essential to perform comprehensive pre-clinical testing to identify the investigational agents that are most likely to be effective in the clinic. However, such effort has been blocked for many years due to the lack of clinically relevant and molecularly accurate model systems. Fortunately, we have established a large panel (>70) of patient derived orthotopic xenograft (PDOX) models of pediatric brain tumors through direct injection of patient tumor specimens into the brains of SCID mice. These PDOX models are shown to have replicated the histopathological features, invasive phenotypes and major genetic abnormalities (gene expression, DNA copy number and gene mutations) of the original primary tumors even during serial sub-transplantations in vivo in mouse brains. The xenograft tumor cells can also be cryopreserved for sustained and on-demand supply of tumorigenic PDOX cells. This capacity combined with our optimized surgical procedure, with which we can implant up to 260 mice per day, makes it possible for us to test multiple (e.g., 6-10) drugs per year for every tumor type. Our objective is therefore to make use of this unique panel of PDOX models to examine therapeutic efficacy of new agents and to analyze mechanisms of action and therapy resistance in high grade glioma, medulloblastoma and ependymoma. Our hypothesis is that these patient-specific PDOX tumors will respond to anti-cancer therapies similarly to the corresponding human primary tumors, and the effective agents identified through this system would have better chances of clinical success. To test this hypothesis, we will perform a series of in vitro and in vivo assays to achieve the following aims: 1) to identify genetically accurate candidate PDOX models that bear the therapeutic target(s) of new investigational drugs through data mining of our mouse model molecular characterization databases; 2) to select the most responsive models through functional in vitro screening to determine time- and dose- responses; 3) to demonstrate therapeutic efficacy of new investigational drugs in multiple target-bearing PDOX models; and 4) to perform detailed analysis of cellular and molecular mechanisms of cell killing as well as the causes of therapy resistance both in vitro and in vivo. Our novel panel of PDOX mouse models represents a broad spectrum of genetic abnormalities of pediatric CNS tumors. All the assays are well established and routinely performed in our laboratory; we are uniquely positioned to accomplish the proposed drug studies in vivo. Our findings should provide strong preclinical evidence to support the initiation of clinical trials.