基于STAT1磷酸化/去磷酸化平衡研究益气扶正法协同IFN-α抑制肝癌术后复发转移的作用机制

IFN-α is a clinically conventional drug to prevent postoperative recurrence and metastasis of hepatocellular carcinoma(HCC) , but long-term use of high-doses of IFN-α can induce drug resistance, which limits its clinical application. We found that the representative drug of the “strengthening and supporting vital qi” method-Shenqi Fuzheng Injection(SFI) can effectively enhance the efficacy of IFN-α, and can affect the expression of regulatory genes which control the STAT1 phosphorylation / dephosphorylation balance in HCC. Because the phosphorylation of STAT1 is the key to reduce drug resistance of IFN-α, we hypothesize that by regulating STAT1 phosphorylation / dephosphorylation balance, SFI can enhance phosphorylation of STAT1 and activate the signaling pathway of JAK-STAT, which reduce the drug resistance of IFN-α, so can enhance the therapeutic effect of IFN-α on HCC. In this research, MTT, Transwell, RT-PCR, Western Blot, Co-Immunoprecipitation, Confocal laser scanning microscope, Protein chip and RNA Interference technology etc are taken into application. From two aspects of both in vivo and in vitro we reveal the funtion of SFI on regulating STAT1 phosphorylation / dephosphorylation balance and the mechanism of reducing the resistance of IFN-α. The findings could provide a theoretical basis for the prevention and treatment of HCC recurrence and metastasis by the use of SFI combined with IFN-α.

IFN-a是临床上预防肝癌术后复发、转移的常规药物，但长期大剂量应用所诱发的耐药性限制了其临床应用。我们研究发现益气扶正法的代表药物参芪扶正注射液可显著增强IFN-a的疗效，并能够影响肝癌中STAT1磷酸化/去磷酸化平衡的调控基因表达，鉴于STAT1的磷酸化激活是降低IFN-a耐药性的关键，我们推测参芪扶正注射液可通过调控肝癌中STAT1磷酸化/去磷酸化平衡，增强STAT1蛋白磷酸化修饰，激活JAK-STAT通路，减少IFN-a耐药性，进而提高IFN-α疗效。本研究运用MTT、Transwell、RT-PCR、Western Blot、免疫共沉淀、激光共聚焦、蛋白质芯片及RNA干扰等技术，从体内和体外两个层面系统揭示参芪扶正注射液调控STAT1磷酸化/去磷酸化平衡的作用，阐明参芪扶正注射液减少IFN-α耐药性的机制，为临床联合使用参芪扶正注射液和IFN-a防治肝癌术后复发转移提供理论依据。

IFN-a是临床上预防肝癌术后复发、转移的常规药物，但长期大剂量应用所诱发的耐药性限制了其临床应用。我们研究发现益气扶正法的代表药物参芪扶正注射液可显著增强IFN-a的疗效，并能够影响肝癌中STAT1磷酸化/去磷酸化平衡的调控基因表达，我们发现参芪扶正注射液可增强荷瘤裸鼠肝癌组织内STAT1、SHP1、p21、Bcl-2及BAX的磷酸化同时抑制JAK的磷酸化，通过调控肝癌中STAT1磷酸化/去磷酸化平衡，增强STAT1蛋白磷酸化修饰，激活JAK-STAT通路，减少IFN-a耐药性，进而提高IFN-α疗效。本研究运用MTT、Transwell、RT-PCR、Western Blot、二代基因测序及RNA干扰等技术，从体内和体外两个层面系统揭示参芪扶正注射液调控STAT1磷酸化/去磷酸化平衡的作用，阐明参芪扶正注射液减少IFN-α耐药性的机制，为临床联合使用参芪扶正注射液和IFN-a防治肝癌术后复发转移提供理论依据。