六神丸干预“β-catenin与COX-2环路”的镇癌痛机制研究

Pain is a major health care problem for patients with cancer. Unfortunately, the existing analgesics for the treatment of pain in cancer often cause severe side effects. Hence, we have intensely focused on finding and providing the cancer pain relief by using Liu-Shen-Wan, a well-known traditional Chinese medicine (TCM) compound with powerful effects of heat-clearing and detoxifying,dissipating hard mass and removing blood stasis to ease pain. In this project, we will examine the expressions of β-catenin and COX-2 in liver tissues, and determine the amount of substance P in blood of mice with or without primary liver cancer for over time. Simultaneously, we will investigate the mRNA and protein expression levels of pain-related regulatory factors including neurokinin-1 receptor, etc in T7-10 or L4-6 dorsal root ganglions (DRGs) and spinal cords of both a murine model of primary liver cancer and a murine bone cancer pain model by using some techniques including radioimmounassay, image analysis and RT-PCR,etc. Furthermore, we will investigate the mRNA and protein expression levels of those factors described above and analyze the relationship between β-catenin signaling and those factors by mimicking of the β-catenin signaling pathway in mixed culture of HepG2 cells and DRG cells by Wnt-3a (the Wnt/β-catenin signaling pathway) or TWS119 (a distinct GSK-3β inhibitor). Taken together, this project may propose a “β-catenin and COX-2” regulatory loop-regulated molecular mechanism for the sensitization of peripheral and central spinal cord neurons according to the in vivo experimental data of both primary liver cancer and cancer pain models and the in vitro experimental data of cell model. The results described above will clarify pharmacological mechanisms of the inhibition of a “β-catenin and COX-2” regulatory loop in the treatment of cancer pain by Liu-Shen-Wan`s powerful effects of dissipating hard mass and removing blood stasis to ease pain. The results of this project contributes to a better understanding of the molecular mechanism of cancer pain, provide new targets for the development of drugs in the treatment of cancer pain.

癌症患者苦于疼痛，但临床上缺乏既抗癌又镇痛的有效治疗药物。中医药治疗癌痛有其特色，在控制肿瘤发展的同时可减轻癌痛。为此，本项目针对具有“清热解毒、软坚散结、祛瘀止痛”功效的中成药六神丸，研究其控制肿瘤基础上的镇痛机制。拟依次建立小鼠原发性肝癌模型、骨癌痛模型及HepG2细胞与背根神经节细胞的共培养系等多重研究平台，利用放射免疫法、免疫组化、RT-PCR等技术，监测成模前后小鼠肝组织中β-catenin、COX-2等表达以及血液中P物质等含量，研究癌细胞凋亡以及T7-10或L4-6节段背根神经节、脊髓背角等组织中与神经元敏化相关的P物质受体等表达变化，揭示六神丸“标本兼治”控瘤、镇痛之临床特色，是通过干预“β-catenin与COX-2环路”，促进肿瘤细胞凋亡，抑制神经元敏化而实现的。本研究结果能够为阐明中医临床控制肿瘤同时减轻癌痛的效应机制提供基础性数据，最终促进肿瘤患者生活质量的提高。

癌症患者苦于疼痛，但临床上缺乏既抗癌又镇痛的有效治疗药物。中医药治疗癌痛有其特色，在控制肿瘤发展的同时可减轻癌痛。为此，本项目针对具有“清热解毒、软坚散结、祛瘀止痛”功效的中成药六神丸，研究其控制肿瘤基础上的镇痛机制。.依据所定研究内容与计划，本项目团成员实施了相应研究工作，并发现：（1）六神丸乙醇提取物能有效改善模型动物的炎性以及神经病理性疼痛反应，且效果优于对照药物双氯芬酸钠；（2）相比亚硝胺，其纳米化诱癌剂（纳米亚硝胺）能基于“β-catenin/COX-2的炎性环路”的调控，促进炎症、脂肪化、纤维化等恶变，加快小鼠肝脏癌变的进程；（3）在肝癌模型动物和肝癌患者的肝组织样本中均存在雄性激素受体AR的激活可促进“β-catenin/COX-2的炎性环路”，引发病变细胞的迁移和浸润；（4）六神丸醇提取物及其纳米制剂可有效阻断毒物纳米亚硝胺引发的“β-catenin/COX-2的炎性环路”，抑制炎症、脂肪化、纤维化等恶变，消除小鼠肝脏的癌变进程，尤其是以低浓度的六神丸纳米制剂的效果最好，毒性最小；（5）发现了六神丸在大鼠血浆中的入血成分（胆酸类成分）9个，代谢产物（蟾蜍内酯类代谢产物和胆酸类代谢产物）19个和应激内源性成分（氨基酸类成分）6个，为今后六神丸的体内药效物质基础及作用机理研究提供了科学依据，也可为进一步研究六神丸的药效物质基础或减毒增效机制提供参考。.总而言之，本项目揭示六神丸“标本兼治”控瘤、镇痛之临床特色，是通过干预“β-catenin与COX-2环路”，既有效阻断肝脏细胞的癌变进程，又可抑制炎性及神经病理性疼痛而实现的。本研究结果能够为阐明中医临床控制肿瘤同时减轻癌痛的 效应机制提供基础性数据，最终促进肿瘤患者生活质量的提高。