Defining the role of BRAF in glioma initiation and maintenance

描述（由申请人提供）：胶质瘤是最常见的原发性中枢神经系统肿瘤，也是儿童癌症相关死亡的主要原因，但这些肿瘤发生和进展的分子机制远未完全了解。识别和验证致病基因是药物发现和开发过程中必不可少的第一步。BRAF激活突变已在大量人类癌症中被鉴定，最近的几项研究在不同级别的儿童和成人胶质瘤中检测到这些病变;然而，BRAF激活在胶质瘤发生、进展和维持中的作用尚未得到验证。随着体内模型的有效使用，昂贵的药物发现实验可以集中在最有可能导致有效治疗的药物靶点上。一旦确定了一个基因靶点或途径，下一步就是证明它在疾病的发生、维持或两者中起着关键作用。我们对激活的BRAF在体内胶质瘤发展中的作用进行了初步评估。我们发现，虽然BRAFV 600 E本身不是致瘤性的，但与细胞周期蛋白依赖性激酶抑制剂2a（Cdkn 2a; Ink 4a/Arf）的丢失或AKT活化的合作导致恶性胶质瘤的形成，这是环境依赖性的。基于这些数据，我们假设BRAF或下游信号介质可能是治疗儿童和成人胶质瘤的相关靶点。小鼠反向遗传学已成为一种强大的方法，用于破译基因功能和目标验证在哺乳动物生理学的自然环境中，我们将使用这种方法与四环素调控的表达系统，以确定是否BRAFV 600 E是必要的，在体内的背景下，Ink 4a/Arf缺陷的胶质瘤的维护。我们还将进行机制研究，以确定BRAF激活是否与其他遗传改变合作，以促进胶质瘤的发生。使用表达谱分析，将产生分子特征以进一步确定突变BRAF是否定义GBM的新分子亚型或具有已知胶质瘤亚型的簇。基于信号通路的差异激活的GBM亚型不仅可以帮助预测对治疗的反应，而且还可以允许在靶向药物的临床试验中对患者进行合理分层。BRAF或相关的下游信号传导介质是否可以有效地靶向用于这种疾病的治疗干预尚未确定。由于该模型也可用于测试治疗剂，我们将使用具有临床重要性的药理学抑制剂在体外和体内评价BRAFV 600 E在胶质瘤中的药理学抑制的有效性。更好地了解这些胶质瘤的生物学将指导新疗法的开发，以提高这些患者的生存率和降低发病率。

DESCRIPTION (provided by applicant): Gliomas are the most common primary central nervous system tumor, and the leading cause of cancer-related death in children, but the molecular mechanisms responsible for the development and progression of these tumors are far from being completely understood. The identification and validation of genes that cause disease is an essential first step in the drug discovery and development process. Activating mutations of BRAF have been identified in a significant number of human cancers and several recent studies have detected these lesions in both pediatric and adult gliomas of varying grades; however, the role of BRAF activation in glioma initiation, progression, and maintenance has not been validated. With the effective use of in vivo models, expensive drug-discovery experiments can be focused on the drug targets that are most likely to lead to effective therapeutics. Once a gene target or pathway is identified, the next step is to demonstrate that it plays a critical role in disease initiation, maintenance, or both. We performed a preliminary assessment of the role of activated BRAF in glioma development in vivo. We found that while BRAFV600E was not tumorigenic on its own, cooperation with cyclin-dependent kinase inhibitor 2a (Cdkn2a; Ink4a/Arf) loss or AKT activation leads to the formation of malignant gliomas that is context dependent. Based on these data, we hypothesize that BRAF or downstream signaling mediators may be relevant targets for therapy in both pediatric and adult gliomas with this alteration. Mouse reverse genetics has become a powerful approach for deciphering gene function and target validation in the natural environment of mammalian physiology and we will use this approach in combination with the tetracycline-regulated expression system to determine if BRAFV600E is required for the maintenance of gliomas in vivo in the context of Ink4a/Arf-deficiency. We will also perform mechanistic studies to determine if BRAF activation cooperates with other genetic alterations to promote gliomagenesis. Using expression profiling, a molecular signature will be generated to further define whether mutant BRAF defines a novel molecular subtype of GBM or clusters with a known glioma subtype. GBM subtypes based on differential activation of signaling pathways may not only assist with predicting response to therapy but may also allow for rational stratification of patients in clinical trials of targeted agents. Whether BRAF or relevant downstream signaling mediators can be productively targeted for therapeutic intervention in this disease has yet to be determined. Since this model can also be used to test therapeutic agents, we will evaluate the effectiveness of pharmacological inhibition of BRAFV600E in gliomas both in vitro and in vivo using pharmacological inhibitors of clinical importance. A better understanding of the biology of these gliomas will guide the development of new therapies to improve survival and reduce morbidity in these patients.