Functional analysis of malignant brain tumor related gene <h-neu> and development of a novel molecular tageting therapy based on the h-neu gene function

我们对一个新基因&lt；h-neu&gt；的产物进行了功能分析。我们还尝试了几个实验来研究该基因如何在恶性胶质瘤细胞中发挥作用。首先，我们自行研制了一种识别h-neu蛋白表位的抗体。由于我们可以证实这种抗体在免疫组织化学中起作用，我们试图对每一级胶质瘤的肿瘤样本进行染色。但是，我们没有发现染色敏感性的显著差异，因此，我们得出结论，h-neu在胶质瘤中的表达水平并不是很高。接下来，我们对每个级别的胶质瘤样本进行了实时荧光定量聚合酶链式反应。H-neu基因在恶性胶质瘤中的表达低于低级别胶质瘤。为了研究h-neu产物的减少对恶性胶质瘤生长速度的影响，我们制备了h-neu的siRNA，并将其转染到几种胶质瘤培养细胞中。在胶质瘤细胞系中，h-neu siRNA的表达对细胞生长速度有不同程度的抑制作用。我们认为，脑胶质瘤细胞系生长减少率的差异主要是由于h-neu在每个细胞系中的天然表达，因此，我们进行了Nothernb印迹分析，并测定了h-neu在每个细胞系中的表达水平。而h-neu的表达与siRNA对胶质瘤细胞生长的抑制作用无相关性。我们认为应该尝试研究h-neu蛋白的功能，我们已经开始使用蛋白质组学程序对h-neu蛋白进行功能分析。我们现在正在准备h-neu过表达的胶质瘤细胞系。此外，我们仍在计划几个实验来研究h-neu功能。我们想知道h-neu的正常功能，然后我们想要知道h-neu功能在胶质瘤细胞系中的意义3。

We have been investigated about the analysis of the function in the product of a novel gene < h-neu>. We also have tried several experiments to investigate how this gene prduct functions in the malignant glioma cells. First, we made a new antibody to recognize the epitope of the h-neu protein by ourselves. Since we could confirm that this antibody worked in immunhistochemistry, we tried to stain the tumor samples in every grade of gliomas. However, we could not find out the significant differences in the sensitivity of the staining, therefore, we concluded that the expression level of h-neu in glioma was not so high. Next, we performed a Real-Time PCR in every grade of glioma samples. The expression of the mRNA of h-neu in malignant glioma was low compared as those of low grade gliomg. To investigate how the reduction of the product of h-neu affect the growth rate of malignant gliomas, we made a siRNA of h-neu and performed the transfection of it to several glioma culture cells. The reduction of the growth rate due to the transfection of h-neu siRNA was various in glioma cell lines. We considered that the variety of the reduction rate of growth in glioma cell lines was basically due to the native expression of h-neu in each cell lines, therefore, we performed the Nothern blot analysis and measured the expression level of h-neu in each cell lines. However the there was no relation between the expression of h-neu and the reduction rate of the growth due to siRNA in glioma cell lines. We considered that we should attempt to investigate the function of h-neu protein, we have started the functional analysis of h-neu using the proteomix procedure. We are now preparing the glioma cell line in which h-neu was overexpressed. Furthermore, we are still planning several experiments to investigate the h-neu function. We would like to know the normal function of h-neu and after that we would like to try to know the meaning 3 of h-neu function in glioma cell lines.