Molecular analysis of flg and its products for targeted therapy on brain tumors

已有研究表明，碱性成纤维细胞生长因子在脑胶质瘤细胞的生长中起重要作用。本研究对碱性成纤维细胞生长因子受体基因即FMS样基因(FMS-like gene，Flg)进行了分析，旨在阐明其在胶质瘤细胞生长中的生物学活性，并探讨其作为治疗恶性胶质瘤的分子靶点的可能性。首先，我们利用RT-PCR技术分析了手术获得的脑肿瘤组织中Flg基因的表达。对65例脑胶质瘤组织的研究表明，脑胶质瘤组织中Flg基因的表达明显高于正常脑组织。此外，在恶性转化或复发的病例中，其表达进一步增强。在组织病理学上，胶质瘤组织中增强的Flg表达与高细胞性相关。基于这些事实，我们利用Southern和Northern印迹技术对培养的胶质母细胞瘤细胞系进行了更详细的分子分析。Southern分析未观察到Flg基因的扩增，而Northern分析发现其表达增强，提示其表达可能受上游未知转录因子的调控。此外，免疫组织化学分析表明，在胶质母细胞瘤细胞系中检测到的Flg-产物比正常成纤维细胞更多。反义寡核苷酸在20µM时可有效抑制胶质母细胞瘤细胞的生长。这些结果表明，在胶质瘤细胞的增殖过程中，成纤维细胞生长因子-Flg系统的自分泌或旁分泌环是必不可少的，这也表明Flg可能是治疗胶质瘤的一个良好的分子靶点。

It has been reported that basic fibroblast growth factor (bFGF) plays an important role in the growth of glioma cells. The gene encoding receptor of bFGF,namely fms-like gene (flg), was analyzed in this study not only to clarify its biological activity in the growth of glioma cells but also to examine its potential to be a molecular target in treatment of malignant gliomas. First we analyzed expression of flg mRNA in brain tumor tissues obtained at surgery using RT-PCR technique. Sixty-five samples were studied, and it was indicated that expression of flg mRNA in glioma tissues were more enhanced than normal brain tissues. Moreover, it's expression was further enhanced in cases with malignant transformation or recurrence. Histopathologically, enchanced flg expression was in correlation with high cellularity in glioma tissues. Based on these facts, we made more detailed molecular analyzes by Southern and Northern blot techniques on cultured glioblastoma cell lines. flg gene amplification was not observed in Southern analysis, while its expression was increased in Northern, indicating its expression should be controlled by upstream unknown transcription factors. In addition, immunohistochemical analyzes indicated that the flg-product was detected more in glioblastoma cell lines than normal fibroblasts. Anti-sense oligo nucleotide effectively suppressed the growth of glioblastoma cells at concentration of 20 muM.All these results indicate that autocrine or paracrine loops of FGF-flg system is essential in glioma cell proliferation, which also indicates that flg can be a good candidate as a molecular target in treatment of gliomas.