The Role of PDGFR in Medulloblastoma Progression

描述（申请人提供）：髓母细胞瘤是儿童最常见的恶性脑肿瘤。为了解决有效治疗的问题，必须首先了解人髓母细胞瘤的进展。我们假设血小板衍生生长因子（PDGFR）介导的信号转导增强了肿瘤细胞的反应，促进了髓母细胞瘤的生长和转移扩散，并且已经证明(i) PDGFR在转移性髓母细胞瘤中显著过表达，（ii）髓母细胞瘤细胞PDGFR活性抑制剂降低下游信号靶标MAPK的磷酸化，改变基因表达。（iii）对髓母细胞瘤微阵列基因表达独立数据集中135个已知的促转移基因进行计算机分析，只有三个基因，包括DDGFR，在至少三分之一的肿瘤中显示可检测的mRNA表达，并且在每个数据集中转移性肿瘤中显着过表达。这些数据,结合初步数据显示,两个α(PDGFRA)和β受体的亚型(PDGFRB) 1) RNA和蛋白质水平的表达成神经管细胞瘤肿瘤和细胞,2)稀疏表达的大脑正常的,3)激活成神经管细胞瘤细胞自分泌和旁分泌的方式,和4)能够诱导细胞凋亡成神经管细胞瘤细胞的剂量依赖性的方式选择性抑制剂治疗后PDGFR酪氨酸激酶活性,提示PDGFR可能对成神经管细胞瘤的生长和进展至关重要的机制。因此，PDGFR是治疗干预的潜在新靶点。为了验证这一假设，我们将使用人类成神经管细胞瘤细胞系。我们将(i)进行全面的体外研究，以确定PDGFR信号级联及其对存活、增殖、粘附、迁移和侵袭的影响；（ii）确定PDGFR信号在这些细胞中诱导的关键基因表达变化；（iii）通过针对每种PDGFR特异性的诱导siRNA转染，培养PDGFR表达不足的成神经管细胞，并确定抑制表达和活性对存活的影响。体外的增殖和迁移以及体内的生长和转移，以及（iv）确定磷酸化PDGFR蛋白的表达模式及其与人髓母细胞瘤转移和结局的相关性，以此评估PDGFR抑制剂对该疾病的潜在临床应用价值

DESCRIPTION (provided by applicant): Medulloblastoma is the most common malignant brain tumor in children. To approach the problem of effective therapy, the progression of human medulloblastoma must first be understood. We hypothesize that platelet-derived growth factor (PDGFR)-mediated signal transduction enhances tumor cell responses that promote the growth and metastatic spread of medulloblastoma, and have shown that (i) PDGFR is significantly overexpressed by metastatic medulloblastomas, (ii) inhibitors of medulloblastoma cell PDGFR activity decrease phosphorylation of the downstream signaling target, MAPK, alter gene expression, and decrease cell migration and survival and (iii) in silico analysis of 135 known pro-metastatic genes within independent datasets of microarray gene expression of medulloblastomas, only three genes, including DDGFR, demonstrated detectable mRNA expression in at least one third of all tumors analyzed and significant overexpression by metastatic tumors in each dataset. These data, combined with the preliminary data showing that both alpha (PDGFRA) and beta (PDGFRB) subtypes of the receptor are 1) expressed on the RNA and protein level by medulloblastoma tumors and cells, 2) sparsely expressed by normal brain, 3) activated in an autocrine and paracrine fashion in medulloblastoma cells, and 4) capable of inducing apoptosis of medulloblastoma cells in a dose-dependent manner following treatment with a selective inhibitor of PDGFR tyrosine kinase activity, suggest a mechanism by which PDGFR may be vital for medulloblastoma growth and progression. Thus, PDGFR is a potential novel target for therapeutic intervention. To test this hypothesis we will employ human medulloblastoma cell lines. We will (i) conduct comprehensive in vitro studies to determine the PDGFR signaling cascade and its effects on survival, proliferation, adhesion, migration and invasion, (ii) determine the key gene expression changes induced by PDGFR signaling in these cells, (iii) develop medulloblastoma cells with deficient PDGFR expression by inducible siRNA transfection specific for each PDGFR and determine the effect of inhibited expression and activity on survival, proliferation and migration in vitro and growth and metastasis in vivo, and (iv) determine the expression pattern of phosphorylated PDGFR protein and its correlation with metastasis and outcome in human medulloblastomas as a way to assess the potential clinical utility of PDGFR inhibitors for this disease