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基因突变是否真的参与了人类胶质瘤的发生和发展。作为第一步，对7种不同的人脑胶质瘤细胞系进行免疫细胞化学和PCR-SSCP分析。7株胶质瘤细胞系均显示p53蛋白异常表达，这种基因表达的改变与胶质瘤细胞p53基因点突变有关。第二步，对50例开颅手术切除的脑肿瘤标本进行PCR-SSCP分析。8例星形细胞瘤和16例胶质母细胞瘤中分别有3例（38%）和5例（31%）存在p53基因点突变或缺失，但突变或缺失的位点在不同的病例中不同。结果表明，p53基因突变率在低级别胶质瘤和高级别胶质瘤中相似，p53基因突变直接参与了近1/3的胶质瘤的发生。因此，p53似乎在人类脑肿瘤（尤其是神经胶质瘤）的DNA诊断中非常有用。另一方面，获得胶质纤维酸性蛋白启动子的中枢神经系统特异性表达载体系统，对于建立恶性胶质瘤的基因治疗是至关重要的。本研究对GFAP基因启动子区增强子序列进行了详细的测定，并进一步研究了由2个或4个增强子组成的串联构建体，其转录速率提高了4 ~ 5倍，表明其在中枢神经系统基因治疗中具有潜在的应用前景。

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.