癌-睾丸抗原OY-TES-1 对脑肿瘤恶性生物学行为的影响及抗体血清学分析

Previous studies found that OY-TES-1 was highly expressed in brain tumor and not expressed in normal tissues. Therefore it has been recognized as a potential target for clinical application. Until now, it is still not known the effects and molecular mechanism of OY-TES-1 on the malignant biological behavior of brain tumor cells. OY-TES-1 antibody was observed in sera of some patients with brain tumors. The changes of OY-TES-1 antibody in sera of brain tumor patients before and after operation, the clinical significance of antibody production have not been reported. In order to investigate the biological function and immunological characteristics of OY-TES-1, this study focused on: ① OY-TES-1 gene transfection and RNAi experiment. To select the cell lines of brain tumor as subjects, analysis of cell cycle, apoptosis, growth, differentiation, proliferation, migration and invasion should be done. ② Dynamic monitoring of OY-TES-1 antibody in patients'sera. The serum antibody titer of OY-TES-1 should be investigated in different periods of before and after operation.The positive rate,specificity and clinical significance of serum OY-TES-1 should be explored. Furthermore, the relationship between OY-TES-1 antibody and prognosis was also analysed. The aim of this study was to assess the value of OY-TES-1 which can be used to aid in diagnose and therapy of brain tumor, or as a prognostic indicator through monitoring.

前期研究发现癌-睾丸抗原OY-TES-1高表达于脑肿瘤，在除睾丸以外的正常组织几乎不表达，具有潜在的临床应用前景。迄今为止，OY-TES-1对脑肿瘤恶性生物学行为的影响及相关的分子机制仍不清楚。部分脑肿瘤患者血清中可检测到OY-TES-1抗体，但是患者手术前后OY-TES-1血清抗体的滴度有否变化及抗体产生的临床意义也未见报道。 本课题从研究OY-TES-1 的生物学功能和免疫学特性入手，拟做以下研究：① OY-TES-1 基因转入和沉默实验。观察比较OY-TES-1在脑肿瘤细胞的生长、分化、增殖、迁移、侵袭、细胞周期及凋亡等方面的作用；② OY-TES-1血清抗体的动态监测。探讨脑肿瘤患者术前、术后不同时段的血清中OY-TES-1抗体出现的阳性率、特异性及与临床指标的关系，比较分析其预后相关因素。该课题的完成有助于初步论证OY-TES-1应用于脑肿瘤辅助诊疗和预后评估的可行性。

1.在生物学功能方面：RNAi下调OY-TES-1表达，使胶质瘤细胞的生长、增殖、迁移、粘附和侵袭能力下降，早期凋亡细胞增加，S期细胞减少，G2/M期细胞增加，提示OY-TES-1表达下调可减弱胶质瘤细胞的恶性生物学行为。Cyclin A、caspase 3及MMP2、MMP9可能参与OY-TES-1对胶质瘤细胞恶性生物学行为的调控。. 2.在免疫学方面：⑴体液免疫反应：OY-TES-1血清抗体在胶质瘤和脑膜瘤的阳性率分别为17.14%（12/70）和16.3%（5/31），尚未发现正常人血清中有OY-TES-1抗体产生。⑵细胞免疫反应：我们采用细胞因子联合诱导HLA-A2+健康人外周血单核细胞为DC，通过OY-TES-1重组蛋白致敏DC，诱导同体T淋巴细胞产生CTL，可产生特异性抗胶质瘤免疫应答，杀伤HLA-A2+OY-TES-1+胶质瘤细胞。. 3.在表达及其临床意义分析方面：RT-PCR检测发现，OY-TES-1mRNA在胶质瘤和脑膜瘤表达的阳性率分别为80.21%（18/91）和70.73%（58/82）。RT-qPCR检测发现，胶质瘤OY-TES-1mRNA的表达明显高于正常脑组织（P=0.0015），且WHOⅢ-Ⅳ级胶质瘤OY-TES-1的表达明显高于WHOⅠ-Ⅱ级（P=0.0002）。免疫组织化学法检测发现，OY-TES-1蛋白在正常脑组织中均未见表达，在胶质瘤中的表达率高达69.35%（86/124）。统计分析显示OY-TES-1蛋白的表达与胶质瘤WHO分级、P53和Ki67有关。我们对进行蛋白检测的124例胶质瘤患者成功随访84例，随访时间为1个月-69个月（24.8±20.9月），预后分析显示OY-TES-1蛋白的表达可作为胶质瘤患者的独立预后因素。. OY-TES-1能在脑肿瘤中特异性的表达并引起免疫反应，OY-TES-1表达下调可减弱胶质瘤的恶性生物学行为，提示将其用于脑肿瘤免疫治疗和预后评估具有潜在的应用开发前景。