A novel matrine derivate inhibits differentiated human hepatoma cells and hepatic cancer stem-like cells by suppressing PI3K/AKT signaling pathways
A novel matrine derivate inhibits differentiated human hepatoma cells and hepatic cancer stem-like cells by suppressing PI3K/AKT signaling pathways
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DOI：
10.1038/aps.2016.104
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发表时间：
2017-01-01
影响因子：
8.2
通讯作者：
Zhang, Jun-ping
中科院分区：
文献类型：
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作者：
Liu, Ying;Qi, Yang;Zhang, Jun-ping
苦参碱是一种从中草药苦参中提取的生物碱，对多种癌症具有化学预防作用。本研究对苦参碱的一种新型衍生物(6aS, 10S, 11aR, 11bR, 11cS)-10-甲氨基-十二氢-3a，7 -二氮杂苯并（de）蒽-8-硫酮（MASM）在体外和体内对人肝细胞癌（HCC）细胞及其相应的球细胞的抗癌作用进行了研究。用MASM处理人肝癌细胞株Hep3B和Huh7。采用CCK8和菌落测定法评估细胞增殖；流式细胞术检测细胞凋亡和细胞周期分布。Western blot和qRT-PCR检测细胞标志物和信号分子的表达。采用球培养技术在Hep3B和Huh7细胞中富集肿瘤干细胞（CSC）。在BALB/c裸鼠Huh7细胞异种移植模型中，以10 mg.kg（-1）的剂量给药MASM，评价MASM的体内抗肿瘤效果。D (-1), ig) 3周。治疗结束后，切除肿瘤并称重。一部分肿瘤组织被酶解得到单细胞悬浮液用于球体形成试验。MASM（2、10、20 mu mol/L）呈剂量依赖性地抑制HCC细胞增殖，诱导细胞凋亡，与Bcl-2表达降低和细胞凋亡相关。PARP裂解增加。MASM还诱导细胞周期阻滞在G(0)/G(1)期，并伴有p27升高和Cyclin D1表达降低。有趣的是，MASM（2、10和20 mu mol/L）显著降低EpCAM(+)/CD133（+）细胞数量，抑制球体形成，抑制干细胞标记基因的表达，促进Hep3B和Huh7球体中成熟肝细胞标记的表达。此外，MASM剂量依赖性地抑制Hep3B和Huh7细胞中的PI3K/AKT/mTOR和AKT/GSK3 β / β -连环蛋白信号通路。在携带Huh7异种移植瘤的裸鼠中，给药MASM显著抑制Huh7异种移植瘤的生长，并显著减少肿瘤中存活的癌干细胞样细胞的数量。MASM还降低了干细胞标记物的表达，同时增加了肿瘤组织中成熟肝细胞标记物的表达。苦参碱的新衍生物MASM通过多种机制显著抑制HCC肿瘤生长，可能是治疗肝细胞癌的有希望的候选药物。
Matrine is an alkaloid extracted from a Chinese herb Sophora flavescens Ait, which has shown chemopreventive potential against various cancers. In this study, we evaluated the anticancer efficacy of a novel derivative of matrine, (6aS, 10S, 11aR, 11bR, 11cS)-10-methylamino-dodecahydro-3a,7a-diazabenzo (de) anthracene-8-thione (MASM), against human hepatocellular carcinoma (HCC) cells and their corresponding sphere cells in vitro and in vivo. Human HCC cell lines (Hep3B and Huh7) were treated with MASM. Cell proliferation was assessed using CCK8 and colony assays; cell apoptosis and cell cycle distributions were examined with flow cytometry. The expression of cell markers and signaling molecules was detected using Western blot and qRT-PCR analyses. A sphere culture technique was used to enrich cancer stem cells (CSC) in Hep3B and Huh7 cells. The in vivo antitumor efficacy of MASM was evaluated in Huh7 cell xenograft model in BALB/c nude mice, which were administered MASM (10 mg.kg(-1).d(-1), ig) for 3 weeks. After the treatment was completed, tumor were excised and weighed. A portion of tumor tissue was enzymatically dissociated to obtain a single cell suspension for the spheroid formation assays. MASM (2, 10, 20 mu mol/L) dose-dependently inhibited the proliferation of HCC cells, and induced apoptosis, which correlated with a reduction in Bcl-2 expression and. an increase in PARP cleavage. MASM also induced cell cycle arrest in G(0)/G(1) phase, which was accompanied by increased p27 and decreased Cyclin D1 expression. Interestingly, MASM (2, 10, and 20 mu mol/L) drastically reduced the EpCAM(+)/CD133(+) cell numbers, suppressed the sphere formation, inhibited the expression of stem cell marker genes and promoted the expression of mature hepatocyte markers in the Hep3B and Huh7 spheroids. Additionally, MASM dose-dependently suppressed the PI3K/AKT/mTOR and AKT/GSK3 beta/beta-catenin signaling pathways in Hep3B and Huh7 cells. In Huh7 xenograft bearing nude mice, MASM administration significantly inhibited Huh7 xenograft tumor growth and markedly reduced the number of surviving cancer stem-like cells in the tumors. MASM administration also reduced the expression of stem cell markers while increasing the expression of mature hepatocyte markers in the tumor tissues. The novel derivative of matrine, MASM, markedly suppresses HCC tumor growth through multiple mechanisms, and it may be a promising candidate drug for the treatment of hepatocellular carcinoma.