从肿瘤炎性微环境角度探讨清热解毒法在原发性肝癌治疗中的价值

“Qingre Jiedu” method is a commonly used method in the treatment of primary liver cancer (PLC), not only for early stage disease, but also for all the stages of PLC. However, many patients who do not show “heat symptom” according to Traditional Chinese Medicine are also treated with “Qingre Jiedu” herbs. We have developed a "Jiedu Decoction" that has been used in the whole process of the treatment of PLC and has showed good effects. Although the "Cancer toxin theory" provides a theoretical basis for the above phenomenon from TCM theory, the experimental evidences related to above phenomenon are still deficient. Recent studies have revealed that tumor inflammatory microenvironment is tightly related to the occurrence and progression of PLC. Our previous studies also suggest "Jiedu Decoction" is able to regulate the expression of chemokines in the PLC inflammation microenvironment, suggesting inflammatory microenvironment may be a new target for the prevention and treatment of liver cancer by “Qingre Jiedu” method. However, whether there are any differences in regulating the inflammatory microenvironment in different stages of liver cancer by “Qingre Jiedu” method? What are the differences between “Qingre Jiedu” method and “Huoxue”, “Huare” and “Jianpi” methods? Is there any crosstalk between these methods in regulating the inflammatory microenvironment? Therefore, we intend to investigate the effects of “Qingre Jiedu” method on the inflammatory microenvironment of different stages of PLC, and then explore the scientific basis of the compatibility of different treatment method in TCM through regulating inflammatory microenvironment. The results may provide scientific experimental basis for the application of “Qingre Jiedu” method in the treatment of PLC.

清热解毒法是肝癌常用治法之一，不仅被用于早期“邪盛正不亏”阶段，亦常被用于肝癌治疗的各个阶段。然而，很多患者按中医辨证并无“热毒蕴结”之象，亦常配伍清热解毒中药。我们研制的“解毒方”已被用于肝癌治疗的全过程并表现出良好的疗效。“癌毒学说”虽从中医基础的角度为以上现象提供了理论依据，但相关实验依据迄今欠缺。炎性微环境与肝癌的发生发展密切相关。我们前期研究也表明，“解毒方”能调控肝癌炎性微环境中趋化因子的表达，提示炎性微环境可能是清热解毒法治疗肝癌的新靶点。但清热解毒法对肝癌不同阶段炎性微环境的影响有否不同？肝癌证候与炎性微环境有否联系？清热解毒药与活血、化湿、健脾等治则方药对炎性微环境的影响有何异同？配伍后有何相互影响？因此，我们拟观察清热解毒法对肝癌不同阶段炎性微环境的影响，从炎性微环境的角度探讨其他治则方药与清热解毒药配伍的现代理论基础，为清热解毒法在肝癌治疗中的应用提供科学的实验依据。

我国的肝癌多发生在肝炎、肝硬化的基础上，炎性微环境是促进肝癌发展的重要因素。手术仍是肝癌治疗的首选方法，然而，术后复发是临床治疗中的一大难题。我们前期临床研究显示解毒方（JDF）能够有效抑制小肝癌术后复发。本研究中，我们首先探索了JDF预防肝癌术后复发的作用及对肿瘤炎性微环境的影响。在大鼠肝纤维化基础上构建肝癌原位移植瘤模型，切除术后予JDF干预，结果提示JDF能显著降低大鼠肝癌肺转移率。PCR芯片分析表明JDF能够抑制CCL9等在癌旁组织中的高表达，并显著抑制巨噬细胞向肿瘤组织的浸润，提示其机制可能与抑制这些炎性因子分泌有关。体外实验提示JDF既抑制肝癌细胞诱导的巨噬细胞迁移，及M2型巨噬细胞诱导的肝癌细胞迁移。经动脉化疗栓塞（TACE）是不能切除的肝癌常用的一种治疗方法，但TACE术造成肿瘤周围缺血缺氧，能够促进血管新生及残癌转移，JDF能够显著抑制缺氧诱导的多种炎性因子表达和内皮细胞小管生成，并抑制缺氧诱导的肝癌细胞迁移和EMT，其分子机制可能与抑制IL-8对AKT和ERK信号通路的激活有关。肿瘤转移是肝癌治疗的主要难题，在大鼠肝纤维化-肝癌移植瘤模型中，RNA芯片分析显示癌症相关的差异基因在疾病与功能方面被显著富集，JDF能抑制NF-κB、IL-6、IL-8、JAK/Stat等与炎症相关信号通路，在其上游因子调控中，JDF显著抑制了TGF-β、LPS、STAT1、IL1B、肿瘤坏死因子、P38MAPK等，其中以TGF-β调控基因数最多。JDF能够显著抑制TGF-β诱导的肝癌细胞迁移侵袭，其机制Smad依赖和非依赖的机制有关。另一方面，JDF能够显著抑制M2型巨噬细胞分泌IGFBP2蛋白，对IGFBP2诱导的肝癌细胞增殖、迁移和侵袭，JDF也有显著的抑制作用。此外，JDF抑制肝癌迁移侵袭的效应还与抑制肝星状细胞的增殖和活化有关。索拉菲尼是晚期肝癌的常用治疗药物，耐药是临床治疗的难题，而JDF能增强索拉菲尼对耐药细胞的抑制作用，其机制与抑制内质网应激及Caspase8的抑制蛋白C-FLIP的表达有关。本研究初步阐明了JDF调节炎性微环境抑制肝癌的部分靶点，为清热解毒法在肝癌治疗中的应用提供科学的实验依据。