KDR受体磷酸化及其调节机制在垂体瘤发生发展中的作用研究

Pituitary adenomas make up a significant number of intracranial tumors. The space-occupying lesions and hormone disorder could have heavy consequences for patients about growth, development, labor ability and fertility function. VEGF receptor 2 (KDR) is critical for the process of angiogenesis a tumor formation. Based on sequence analysis, we identified Ser-229 as the only consensus phosphorylation site for Cyclin dependent kinase 5 (Cdk5). In the Preliminary experiments, we detected abundant phosphorylated KDR protein in pituitary adenomas. pSer-229 KDR expression increased with differentiation from normal pituitary tissue, to noninvasive and to invasive prolactin pituitary adenomas. The expression of functional KDR Ser229A was significantly less than that of WT-DOR on cell surface in pituitary cell lines. This mutation is thought to prevent KDR from moving efficiently through the Golgi apparatus to the cell surface and mutation of KDR at Ser-229 impairs the invasiveness ability of pituitary cell lines. The present study indicates Cdk5 may regulate membrane trafficking of KDR through phosphorylation at Ser-229, but the mechanisms involved still need to be studied intensively. In the present study, through further investigation the effect of phosphorylation of KDR on Ser-229 on the function of receptor and its signal transduction pathway in pituitary adenomas, reveal the regulation mechanisms of phosphorylation of the KDR Ser229 in pituitary adenomas and provide new potential specific biomarkers of molecular diagnosis and molecular therapy of pituitary adenomas.

垂体瘤是临床最常见的神经系统肿瘤之一，对患者的生长、发育、劳动能力和生育功能等产生严重影响。VEGFR2（KDR）在血管生成和肿瘤形成中起主要作用。我们发现在KDR的229位丝氨酸为细胞周期素依赖激酶5（Cdk5）潜在的特异性磷酸化位点。前期实验表明垂体瘤组织样本中有丰富的磷酸化KDR表达，KDR的Ser229位磷酸化水平在侵袭性垂体瘤、非侵袭性垂体瘤、正常垂体瘤中依次递减，将KDR 229位丝氨酸突变后可能影响受体从高尔基体到细胞膜的膜转运，同时发现该位点的突变降低了垂体瘤细胞的迁移和侵袭能力。这些研究提示该位点磷酸化水平对于受体及受体后信号转导发挥重要作用。本研究主要探讨KDR 229位丝氨酸磷酸化对受体功能及后续信号转导的影响，并将在大规模垂体瘤样本中筛查KDR229位丝氨酸磷酸化水平与垂体瘤分子分类、侵袭、复发与预后的关系，并期望以此为线索探寻垂体瘤的个性化诊疗中有效的分子靶点。

垂体腺瘤是最常见的颅内肿瘤之一，约占颅内肿瘤的15-20%。虽然垂体瘤在病理类型上属于良性肿瘤，但约有15%的垂体瘤呈侵袭性生长，表现出恶性肿瘤的生长特性。。在前期工作中，本课题组首次发现在细胞周期依赖性蛋白激酶5 （cyclin-dependent kinase 5， CDK5）存在于人正常垂体和垂体瘤中。在本项目中，我们探讨了CDK5信号在垂体腺瘤中可能的作用机制。结果表明，侵袭性垂体腺瘤中CDK5最重要的激活子p35的表达和CDK5活性均显著高于非侵袭性垂体腺瘤。抑制CDK5活性可抑制大鼠垂体瘤细胞GH3的迁移和侵袭能力。我们发现在VEGFR2，即KDR （kinase insert domain receptor，KDR）的229位丝氨酸为CDK5潜在的特异性磷酸化位点，垂体瘤组织样本中有丰富的磷酸化KDR表达，KDR的Ser229位磷酸化水平在侵袭性垂体瘤、非侵袭性垂体瘤、正常垂体瘤中依次递减，我们发现CDK5可以通过磷酸化垂体瘤KDR的Ser229促进垂体瘤细胞的增殖，迁移和侵袭，且正确的KDR细胞膜表面定位需要Ser229的磷酸化。此外，我们的数据显示， KDR的Ser229高水平磷酸化与垂体瘤患者的预后较差相关。本项目首次从临床垂体瘤的肿瘤样本中筛查KDR受体特异性位点磷酸化的水平，而这种特异性位点磷酸化水平的变化，可能为垂体瘤的临床分子诊断和分子治疗提供潜在靶点，具有较为重要的理论意义和实际意义。