Regorafenib联合TAS102新策略治疗肝细胞癌的研究

A novel strategy of regorafenib in combination with TAS102 against hepatocellular carcinoma

背景与目的:瑞格菲尼(regorafenib,REG)和TAS102是治疗消化道肿瘤的新型药物。嘧啶类药物5-氟尿嘧啶(5-fluorouracil,5-FU)联合REG可抑制多药耐药转移性结肠癌的进展。肿瘤干细胞(cancer stem cell,CSC)决定肿瘤的自我更新,异质性及治疗耐受等,靶向CSC为治愈肿瘤提供了可能途径。探讨REG联合TAS102治疗肝细胞癌(hepatocellular carcinoma,HCC)的联合效应及潜在机制。方法:以人HepG2、Huh7和SK-Hep1肝癌细胞系为实验对象。REG和TAS102单药或联合处理肝癌细胞及荷瘤动物。采用细胞计数试剂盒(cell counting kit-8,CCK-8)检测肝癌细胞活力;流式细胞术分析肝癌细胞中CD133阳性细胞亚群比例;采用蛋白质印迹法(Western blot)分析不同药物处理后肝癌细胞中蛋白表达差异;含不同药物的干细胞培养基培养肝癌干细胞球(HCC sphere);荷瘤动物实验评估药物联合治疗效果。结果:REG及TAS102单药显著抑制肝癌细胞活力。TAS102联合使用减低REG单药时诱导增加的CD133阳性细胞亚群比例及干细胞标志分子SRY相关的高迁移率族盒蛋白-2(SRY-related high mobility group box protein-2,SOX2)和乙醛脱氢酶1A(aldehyde dehydrogenase 1A,ALDH1A)的蛋白水平。同时REG的联合使用下调TAS102单药处理时诱导活化的抗凋亡蛋白髓细胞白血病-1蛋白(myeloid cell leukemia-1,MCL1)信号。REG联合TAS102显著抑制了HCC sphere的形成。动物实验证实REG联合TAS102显著抑制肝癌瘤体的生长。结论:REG联合TAS102通过调控肝癌细胞的干细胞性及抗凋亡信号,发挥联合用药增强抗HCC的活性,为临床治疗难治性HCC提供了一种新的治疗策略。

Background and purpose:Both regorafenib(REG)and TAS102 are novel anti-tumor drugs in the treatment of the digestive cancers.REG in combination with 5-fluorouracil(5-FU),a pyrimidine analogue,could inhibit the progression of multidrug resistant metastatic colorectal cancer.Cancer stem cells(CSCs)determine tumor self-renewal,heterogeneity and therapeutic resistance,and targeting CSCs provides a curable approach to multiple malignancies.This study aimed to investigate the therapeutic effect and potential mechanism of REG in combination with TAS102 against hepatocellular carcinoma(HCC).Methods:A panel of human liver cancer cell lines HepG2,Huh7 and SK-Hep1 were used in this study.Liver cancer cells and xenograft tumor animals were treated with REG and TAS102 monotherapy or combining regimen.Cell counting kit-8(CCK-8)was used to assess HCC cell viability.The subpopulations of CD133 positive cells were detected and analyzed using flow cytometry.Western blot was used to evaluate these protein contents in HCC cells with diverse drug treatments.HCC spheres were cultured in growing sphere medium with diverse drugs.Xenograft tumor animals were used to assess the therapeutic effect in vivo.Results:REG or TAS102 monotherapy significantly inhibited HCC cell viability.TAS102 in combination therapy reduced the increase of CD133+cell subpopulation,SRY-related high mobility group box protein-2(SOX2)and aldehyde dehydrogenase 1A(ALDH1A)expressions induced by REG alone.Moreover,REG attenuated TAS102-induced myeloid cell leukemia-1(MCL-1)expression.REG in combination with TAS102 ameliorated the formation of HCC sphere.Animal experiment demonstrated that REG in combination with TAS102 significantly eliminated HCC tumor growth in xenograft animals.Conclusion:REG in combination with TAS102 regulates HCC stemness and MCL-1 signaling and improves the therapeutic effect against HCC.The combination therapy may provide a novel strategy for refractory HCC patients.