STUDY ON THE DIAGNOSIS AND THERAPY OF BRAIN TUMORS IN TERMS OF GENETIC ENGINEERING

在本研究中，我们研究了肿瘤抑制基因p53的表达改变和基因突变，以阐明p53基因突变是否确实参与了人类胶质性肿瘤的发生和发展。作为第一步，我们采用免疫细胞化学和PCR-SSCP分析方法检测了7种不同的人胶质瘤细胞系。7个细胞系均显示p53蛋白表达异常，这种基因表达异常与胶质瘤细胞p53基因发生点突变有关。第二步，采用PCR-SSCP方法对50例开颅脑肿瘤标本进行分析。8例星形细胞瘤中有3例（38%）和16例胶质母细胞瘤中有5例（31%）分别显示p53基因的点突变或缺失，尽管突变或缺失的位置在个别病例中有所不同。提示p53基因在低级别和高级别胶质瘤中的突变率相似，p53基因突变直接参与了至少1 / 3的人类胶质瘤的发生。因此，p53似乎在人类脑肿瘤，特别是神经胶质肿瘤的DNA诊断中非常有用。另一方面，获得GFAP启动子的中枢神经系统特异性表达载体系统是建立恶性胶质瘤基因治疗的关键。在本研究中，GFAP启动子区域增强子的详细序列已经确定，并且由2个或4个GFAP增强子组成的串联结构显示转录率提高了4 ~ 5倍，表明其在未来中枢神经系统基因治疗中的潜在应用。

In the present study, we have investigated the altered expression and gene mutation of a tumor suppressor gene, p53, in order to clarify whether p53 gene mutation is actually involved in tumorigenesis and development of human glial tumors. As the first step, 7 different human glioma cell lines were examined by immunocytochemistry and PCR-SSCP analysis. All of the 7 cell llines examined showed abnormal expression of p53 protein and this altered gene expression was ascribed to the point mutations occurred in p53 gene of glioma cells. As the second step, 50 brain tumor specimens obtained at craniotomy were examined by PCR-SSCP analysis. Three cases (38%) out of 8 astrocytomas and 5 (31%) out of 16 glioblastomas revealed point mutations or deletions of p53 gene, respectively, though the sites of the mutations or deletions were different in indivedual cases. This results indicate that the mutation rate of p53 gene is similar between low grade and high grade gliomas, and that p53 gene mutations are directly invovled in the tumorigenesis of al lest one third of human glial tumors. Thus, p53 seems to be very useful in the DNA diagnosis of human brain tumors, especially glial tumors. On the other hand, it is essential to obtain the CNS specific expression vector sytem in terms of GFAP promoter in establishing the gene therapy for malignant gliomas. In the present study, the detailed sequence of enhancer in the GFAP promoter region has been determined and furthermore, the tandem constructs consisting of 2 or 4 enhancers of GFAP revealed 4 to 5 times increase in transcription rate, indicating its potential use in CNS gene therapy in the fulure.