CD59异构体的鉴定及其促进肿瘤侵袭转移的作用机制

CD59的经典功能为在补体级联反应末期抑制膜攻击复合物的形成，它在肿瘤细胞中高表达并与肿瘤抗体治疗耐药密切相关。大量研究表明CD59尚具有其它难以解释的功能，包括参与肿瘤侵袭转移、血管生成、抑制凋亡以及参与生殖发育等。前体mRNA的可变剪接是产生蛋白质多样性的重要原因，我们前期发现人类hCD59和小鼠mCd59各存在四种异构体，且很可能异构体hCD59-IV和mCd59b-III与上述非经典功能有关，但机制尚有待于揭示。本申请将制备特异性单抗，与定量PCR方法结合，观察这两种异构体在正常组织与肿瘤组织中的表达水平，进一步在细胞与动物水平改变这两个CD59异构体的表达水平，通过多种手段在体内外观察对细胞增殖与迁移能力的影响并揭示其分子机制，在分子、细胞及整体水平重点阐述hCD59-IV促进肿瘤侵袭转移的功能与机制，探讨其作为诊断标记物和治疗靶点的潜在性，为肿瘤侵袭转移的诊治提供新思路新方法。

CD59 tipically functions for restricting the assembly of membrane attack complex at the late stage of complement activation. It has been demonstrated to be up-regulated in cancer and highly correlates with resistance to antibody-based cancer therapy. Further, numerous evidence demonstrates the alternative roles of CD59 including involvement in tumor metastasis, angiogenesis and anti-apoptosis, as well as participation in reproduction. The alternative splicing of pre-mRNA is an important determinant for protein diversity, and we previously found each human CD59 or mouse Cd59 express four isoforms, in which hCD59-IV and mCd59b-III most likely contribute to the alternative roles of CD59. However, the underlying mechanisms for their functions and molecular mechanisms require future investigation. In this proposal, we will prepare the specific monoclonal antibodies, in combined with quantitative RT-PCR, to detect the expression levels of the two CD59 isoforms in normal and tumor tissues. In addition, we will alter their expression levels by transducing shRNA and exogenous coding DNA of hCD59-IV in cancer cells, or by knocking-out and exogenously expressing mCd59b-III in mice, by which we will reveal the isoforms’ functions and mechanisms for regulating cell proliferation and mobility, particularly for hCD59-IV in tumor metastasis. We expect that our findings may help to investigate the potential of hCD59-IV as diagnostic biomarker and/or therapeutic target for cancer metastasis.