患者来源的结直肠癌类器官模型的构建及其在有毒中药抗癌活性评价中的应用

Establishment of patient-derived colorectal cancer organoids and application in evaluating the anti-cancer activity of toxic monomers of traditional Chinese medicines

目的构建患者来源的结直肠癌类器官(CCOs)与正常组织类器官(CNOs),评价几种有毒中药对CCOs及CNOs活性的影响。方法选自南京中医药大学附属南京中医院肛肠中心收治的结直肠癌患者术中肿瘤组织及癌旁组织,运用3D培养法分别构建CCOs与CNOs,检测结直肠癌组织标本与类器官样本免疫组化指标Ki-67表达水平;采用CCK8法检测不同浓度的奥沙利铂、雷公藤甲素、斑蝥素以及蟾毒灵对4例CCOs与1例CNO活性的影响,绘制活性曲线并计算IC50值。结果奥沙利铂对4例CCOs和1例CNO的IC50值分别为72.21μM、46.78μM、54.88μM、128.20μM、122.10μM;雷公藤甲素的IC50值分别为1.54μM、0.19μM、0.24μM、0.19μM、1.46μM;斑蝥素的IC50值分别为34.64μM、1.60μM、162.61μM、13.29μM、146.47μM;蟾毒灵的IC50值分别为0.12μM、0.32μM、0.43μM、0.23μM、0.65μM,CCOs对蟾毒灵具有更高的敏感性。结论CCOs与CNOs成功构建并应用于有毒中药单体活性评价,为有毒中药在结直肠癌中应用的有效性及安全性提供理论依据及方法参考。

Objective To establish patient-derived colorectal cancer organoids(CCOs)and colorectal normal organoids(CNOs),and evaluate the anti-cancer activity of toxic monomers of Traditional Chinese Medicines(TCMs).Methods Tissue samples were collected from patients with colorectal cancer undergoing surgical treatment in Nanjing Hospital of Chinese Medicine.CCOs and CNOs were established by 3D cell culture.The expression level of Ki-67 in tissue samples and organoids was detected with immunohistochemistry.The sensitivity of CCOs and CNOs to L-OHP,triptolide,cantharidin and bufalin was detected by CCK8 assay kits,and IC50 value was calculated.Results The IC50 values of L-OHP to four CCOs and one CNO were 72.21μM、46.78μM、54.88μM、128.20μM、122.10μM,respectively.The IC50 values of triptolide were 1.54μM、0.19μM、0.24μM、0.19μM、1.46μM.The IC50 values of cantharidin were 34.64μM、1.60μM、162.61μM、13.29μM、146.47μM,and that of bufalin were 0.12μM、0.32μM、0.43μM、0.23μM、0.65μM,respectively.The four CCOs showed more sensitive to bufalin.Conclusion CCOs and CNOs were established successfully and applied in evaluation of activity of toxic components in TCMs,which provided a methodological reference.