肺癌外泌体USP5对肺癌耐药性的作用及机理研究

Gain of resistance to chemo-, radiation, and targeted therapies in tumor cells appears to be the major stumbling block in cancer therapeutic. Our recent findings showed that, tumor cells could release exported vesicles (EVs)/exosome to re-form drug resistance for drug-sensitive cells. But the mechanism is unclear. To address this issue, we have built up drug-resistant lung cancer cells in vitro and validated them in vivo. Those cells actively produced a significant increased number of exosome. Analysis of the purified exosome in mass spectrometry revealled a specific component named USP5 in the exosome from drug tolerated lung cancer cells. Our preliminary results showed that USP5 might be delivered through exosome into drug-sensitive tumor cells, in which it could enhance TGF-β signaling thus maintain drug-resistance. Mechanistically, USP5 might interact with SMAD4. We thus hypothesized that USP5 might via deubiquitinating SMAD4 to activate TGF-β signaling. .We are determined to elucidate the function of exosomal USP5 and explore the mechanism by which USP5 regulates TGF-β signaling and drug resistance. Upon successful conclusion of this work, we should have proposed a novel regulatory model of drug resistance and provided a new strategy for cancer.

肿瘤治疗当中放化疗和靶向治疗药物的耐药性问题一直是亟待解决的问题。肿瘤释放的外泌体很可能介导了新的耐药性的形成，但机制尚不清楚。为了回答这一问题，本项目构建了肺癌细胞耐药株，发现耐药株细胞外泌体分泌异常活跃。外泌体纯化和微量质谱分析显示耐药株外泌体含有USP5蛋白。通过外泌体的传递，USP5能够激活药物敏感细胞中的TGF-β信号从而增强肿瘤耐药性。USP5很可能通过去泛素化底物SMAD4发挥以上作用；并因此导致形成的肺癌耐药细胞株中TGF-β信号高度激活。本项目旨在从体内、体外实验验证外泌体USP5激活TGF-β信号通路致使肺癌肿瘤获得耐药性的分子机制。此项目的顺利完成将提出新的肺癌耐药机制，为肿瘤治疗提供新的策略。

肿瘤治疗当中放化疗和靶向治疗药物的耐药性问题一直是亟待解决的问题。肿瘤释放的外泌体很可能介导了新的耐药性的形成，但其分子机制尚不清楚。本项目旨在筛选出肺癌耐药肿瘤细胞以及肺癌耐药患者分泌的外泌体所含有特异性蛋白，并以此为靶点筛选有效的小分子药物靶点，为逆转肺癌耐药提供理论基础。本项目提取了构建好的肺癌细胞耐药株培养液和肺癌耐药患者血清中的外泌体，发现耐药肿瘤中外泌体分泌异常活跃。微量质谱分析显示耐药株外泌体以及肺癌耐药患者外泌体中特异性含有USP5蛋白。我们的研究结果显示，肺癌耐药肿瘤中TGF-β信号显著激活，且外泌体USP5能够促进肿瘤产生耐药性，这个过程依赖于TGF-β信号的激活；体内和体外泛素化实验表明USP5能去泛素化底物SMAD4，稳定SMAD4蛋白，从而促进TGF-β信号；我们在本项目中还建立了以USP5去泛素化能力为读出的药物筛选体系，同时结合斑马鱼快速体内筛选系统，将USP5作为药物靶点筛选出了特异针对USP5活性的小分子抑制剂，结果显示阻断USP5的活性能够有效抑制肺癌的耐药性。我们的研究不仅进一步揭示了肺癌耐药的特征和肿瘤外泌体介导耐药性形成的分子机制，而且为临床逆转肺癌耐药药物的研发提供新的思路。