新型融合蛋白Ec-LDM-TF通过促进巨噬细胞吞噬发挥抗肿瘤效应的机制研究

Tumor-associated macropahages (TAMs) promoting angiogenesis and tumor cell growth, as well as suppressing the adaptive immune response, are therefore potential targets for adjuvant anticancer therapies. In the previous study, the fusion protein Ec-LDM-TF was generated, composed of an enedyne-energized lidamycin (LDM), which displays extremely potent cytotoxicity to cancer cells, linked to an EGFR-targeting oligopeptide Ec, and a tetrepeptide tuftsin (TF). Ec-LDM-TF effectively bound to tumor cells and MФ (binding to the receptor Nrp1), and augmented the therapeutic efficacy compared to Ec-LDM, Tuftsin or their combination. More importantly, the Ec-LDM-TF caused marked down-regulation of CD47 in cancer cells. CD47 has caused much attention for its role in the suppression of phagocytosis. Because the growth of tumor requires avoidance of being phagocytosed by the MФ, the expression of CD47 in cell surface is a common mechanism by which cells protect themselves from phagocytosis. Ec-LDM-TF showed even higher therapeutic efficacy. The efficacy of Ec-LDM-TF may related to the activation of MФ and the down-regulation of CD47. Therefore, this project aims to explore Ec-LDM-TF that caused MФ (TAM) activation, differentiation and regulation of secretion of cytokines, and to investigate the underlying mechanisims of Ec-LDM-TF caused marked down-regulation of CD47 in tumor cells, providing theoretical basis for Ec-LDM-TF tumor therapy.

肿瘤相关巨噬细胞（TAMs）可以促使肿瘤的生长和血管的生成，抑制抗肿瘤免疫应答，是新的肿瘤治疗靶点。我们前期构建的融合蛋白Ec-LDM-TF，是将有强细胞毒活性的力达霉素与EGF分子EGFR结合域（Ec）及促吞噬肽（TF）联结，具有结合EGFR和MФ（表面受体Nrp-1）的多靶向性，比Ec-LDM、Tuftsin或二者联合抗肿瘤作用更显著。进一步研究发现，Ec-LDM-TF能显著降低肿瘤细胞表面CD47表达。CD47抑制MФ的吞噬，是肿瘤细胞逃避免疫监视的机制之一。因此我们提出Ec-LDM-TF可能通过抑制CD47表达活化MФ吞噬增强自身的抗肿瘤作用。本项目拟在此基础上，进一步探索Ec-LDM-TF对MФ（特别是TAMs）的激活、分化和细胞因子分泌的调控及机制，研究Ec-LDM-TF降低CD47表达进而解除肿瘤细胞逃避MФ吞噬的机制，为临床应用Ec-LDM-TF治疗肿瘤提供理论依据。

肿瘤相关巨噬细胞（TAMs）可以促使肿瘤的生长和血管的生成，抑制抗肿瘤免疫应答，是新的肿瘤治疗靶点。我们前期构建的融合蛋白Ec-LDM-TF，是将有强细胞毒活性的力达霉素（LDM）与EGF分子的EGFR结合域（Ec）及促吞噬肽（TF）联结，具有结合EGFR和MФ（表面受体Nrp-1）的多靶向性，比Ec-LDM、Tuftsin或二者联合抗肿瘤作用更显著。进一步研究发现，Ec-LDM-TF能显著降低肿瘤细胞表面CD47表达。.我们获得融合蛋白（Ec-LDP-hBD1）及其烯二炔整合的强化融合蛋白（Ec-LDP(AE)-DF）并观察其对肺癌细胞以及人肺癌异种移植瘤小鼠模型的抗肿瘤作用。结果显示多功能融合蛋白Ec-LDP-DF可以有效结合表达EGFR的肿瘤细胞。此外，Ec-LDP(AE)-DF在NSCLC细胞系中表现出极强的细胞毒性，IC50小于10-10mol/L。Ec-LDP(AE)-DF显着抑制人肺癌A549和H460异种移植瘤裸鼠肿瘤的生长。肿瘤细胞通过Ec-LDP(AE)-DF治疗以剂量依赖性方式导致细胞周期停滞和细胞凋亡。Ec-LDP(AE)-DF也阻断了EGF刺激的EGFR磷酸化信号通路。总之，防御素在癌症发展和进展中的作用不断扩大，Ec-LDP(AE)-DF是一种有前景的NSCLC癌细胞靶向药物。