Defining the role of BRAF in glioma initiation and maintenance

描述（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，但这些肿瘤发生发展的分子机制还远未完全了解。鉴定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的激活突变已经在许多人类癌症中被发现，最近的一些研究已经在不同级别的儿童和成人胶质瘤中发现了这些病变；然而，BRAF激活在胶质瘤发生、进展和维持中的作用尚未得到证实。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。一旦确定了基因靶点或途径，下一步就是证明它在疾病发生、维持或两者中起关键作用。我们对活化的BRAF在体内胶质瘤发育中的作用进行了初步评估。我们发现，虽然BRAFV600E本身不具有致瘤性，但与周期蛋白依赖性激酶抑制剂2a （Cdkn2a; Ink4a/Arf）缺失或AKT激活合作，可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具有这种改变的儿童和成人胶质瘤的相关靶点。小鼠反向遗传学已经成为在哺乳动物生理自然环境中解码基因功能和靶标验证的有力方法，我们将使用这种方法结合四环素调节的表达系统来确定Ink4a/ arf缺乏的情况下，BRAFV600E是否需要维持胶质瘤。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促进胶质瘤形成。通过表达谱分析，将生成分子特征来进一步确定突变BRAF是定义了GBM的新分子亚型还是具有已知胶质瘤亚型的簇。基于信号通路差异激活的GBM亚型不仅可以帮助预测治疗反应，还可以在靶向药物的临床试验中对患者进行合理分层。BRAF或相关下游信号介质是否能有效靶向治疗干预该疾病尚不清楚。由于该模型也可用于测试治疗剂，因此我们将使用具有临床重要性的药物抑制剂，在体外和体内评估药物抑制BRAFV600E在胶质瘤中的有效性。更好地了解这些胶质瘤的生物学将指导新疗法的发展，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.