Genetic diagnosis of gliomas and its clinical application

胶质瘤的基因诊断及其临床应用

• 批准号: 05454397
• 负责人: HAYAKAWA Toru
• 金额: $ 4.22万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (B)
• 财政年份: 1993
• 资助国家: 日本
• 起止时间: 1993 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05454397/
• 关键词: Glioma; Genetic Diagnosis; Glioma-derived Motility Factor (GMF); Oncogenes; Monoclonal Antibodies; Huminized Antibodies; 癌抑制遺伝子; 染色体; ベクター

We had already made mouse monoclonal antibodies(ONS-M21) for the antigens which were shared in both glioma and medulloblastoma(Br J Cancer 68 : 831-837,1993). Then we succeed in humanization of their antibodies (Mol Immunol, in press) and their single-chained antibodies in cooperation with Chugai Pharmaceutical Co.Ltd. Using these humanized antibodies and their single-chained antibodies, we might develop the correct diagnose of tumor sites, and will try to apply to immunotherapy of malignant glioma patients. And now it is in the last steps to extract the malignant glioma-associated antigens which were recognized with these mouse monoclonal antibodies. After extraction of their antigens, we will identify their gene arrangement and mechanisms of their expression.We publishied two papers about selective expression of foreign genes in glioma cells by glial-specific promoters (Jpn J Cancer Res 83 : 1244-1247,1992 ; J Neurosci Res 38 : 415-423,1994). At present, we repeats the in vivo gene t … More herapy in mouse glioma models under these in vitro data with the mouse myelin basic protein (MBP) gene promoters to direct toxic gene expression (J Neurosci Res 36 : 472-479,1993). Then, if we could elucidate the glioma-associated antigens accordig to our plans, we will analyze the promoters controlling their manifestation and will apply them in gene therapy with tissuespecificity in the future.On the other hand, to understand the cellular mechanism of brain invasion by glioma cells, two molecular species of glioma-derived motility factor (GMF), GMF-I and GMF-II,have been purified to homogeneity from the serum-free conditioned medium of a highly invasive human glioma cell line, T98G,by gelatin affinity chromatography and heparin affinity-, DEAE-, hydroxyapatite-, gel permeation- and sulfopropyl high performance liquid chromatography. GMF-I and GMF-II both stimulated the migration of T98G cells in a concentration-dependent manner, and the activity of GMF-I was about 5 times as strong as that of GMF-II.C_6 glioma cells, both of which showed high invasiveness in an in vitro invasion assay with reconstituted basement membrane, Matrigel, migrated to the GMFs with great intensity, while A172 and 9L glioma cells and normal glial cells, all of which weakly infiltrated the Matrigel barrier, migrated to the GMFs with much less intensity. These results indicate that migratory response of glioma cells to the GMFs correlates well with invasiveness, suggesting an important roles of the GMFs in the process of glioma cell invasion (Biochem Biophys Res Commun 193 : 518-525,1993). Less

我们已经制备了针对神经胶质瘤和成神经管细胞瘤共有抗原的小鼠单克隆抗体（ONS-M21） [J] .中华肿瘤杂志，68:831-837,1993)。随后，我们与初盖制药有限公司合作，成功地将其抗体（Mol immuno1，即将出版）和单链抗体人源化。利用这些人源化抗体及其单链抗体，可能对肿瘤部位进行正确的诊断，并将尝试应用于恶性胶质瘤患者的免疫治疗。现在是提取恶性胶质瘤相关抗原的最后一步这些抗原被这些小鼠单克隆抗体识别。提取它们的抗原后，我们将确定它们的基因排列和表达机制。我们发表了两篇关于外源基因在胶质瘤细胞中通过胶质特异性启动子选择性表达的论文（Jpn J Cancer Res 83: 1244-1247,1992; J Neurosci Res 38: 415-423,1994）。目前，我们在小鼠神经胶质瘤模型中重复了体内基因t…在这些体外数据下，使用小鼠髓鞘碱性蛋白（MBP）基因启动子来指导毒性基因表达（神经科学杂志36:472-479,1993）。然后，如果我们能够按照我们的计划阐明胶质瘤相关抗原，我们将分析控制其表现的启动子，并在未来将其应用于具有组织特异性的基因治疗。另一方面，为了了解胶质瘤细胞侵袭大脑的细胞机制，从高度侵袭性胶质瘤细胞系T98G的无血清条件培养基中，采用明胶亲和层析和肝素亲和-、DEAE-、羟基磷灰石-、凝胶渗透-和磺胺丙基高效液相层析纯化了两种胶质瘤源性运动因子（GMF）： GMF- i和GMF- ii。GMF-I和GMF-II均以浓度依赖性的方式刺激T98G细胞的迁移，GMF-I的活性是GMF-II. c_6胶质瘤细胞的5倍左右，在体外重建基底膜Matrigel的侵袭实验中，GMF-I和GMF-II胶质瘤细胞都表现出很强的向gmf的迁移，而A172和9L胶质瘤细胞和正常胶质细胞都有弱的向Matrigel屏障的浸润。迁移到转基因食品的强度要小得多这些结果表明，胶质瘤细胞对GMFs的迁移反应与侵袭性密切相关，表明GMFs在胶质瘤细胞侵袭过程中发挥了重要作用（Biochem Biophys Res comm193: 518-525,1993）。少

项目成果

Takai T: "Anti-proliferative effects of TNP-470 on human malignant glioma in vivo:potent inhibition of tumor angiogenesis." J Neuro-Oncol. 19. 251-258 (1994)

Takai T：“TNP-470 对人体内恶性神经胶质瘤的抗增殖作用：有效抑制肿瘤血管生成。”

Yamada M,et al: "Migration of genetically labeled glioma cells after inplantation into murine brain." J Neurosci Res. 38. 415-423 (1994)

Yamada M 等人：“基因标记的神经胶质瘤细胞植入小鼠大脑后的迁移。”

Ohnishi T: "Purification of motility factor(GMF)from human malignant glioma cells and its biological significance in tumor invasion." Biochem Biophys Res Commun. 193. 518-525 (1993)

Ohnishi T：“从人恶性胶质瘤细胞中纯化运动因子（GMF）及其在肿瘤侵袭中的生物学意义。”

Moriuchi S: "Characterization of a new mouse monoclonal antibody(ONS-M21)reactive with both medulloblastomas and gliomas." Br J Cancer. 68. 831-837 (1993)

Moriuchi S：“一种与髓母细胞瘤和神经胶质瘤均具有反应性的新型小鼠单克隆抗体 (ONS-M21) 的表征。”

Miyao Y: "Selective expression of foregin in glioma cells:Use of the mouse myelin basic protein gene promoter to direct toxic gene expression." J Neurosci Res. 36. 472-479 (1993)

Miyao Y：“胶质瘤细胞中foregin的选择性表达：利用小鼠髓磷脂碱性蛋白基因启动子来指导毒性基因表达。”