Chen Yu-Chia, Chen I-Shu, Huang Guan-Jin, Kang Chi-Hsiang, Wang Kuo-Chiang, Tsao Min-Jen, Pan Hung-Wei
Department of Surgery, Division of General Surgery, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan.
Department of Pathology, National Chung Kung University Hospital, Tainan, Taiwan.
Onco Targets Ther. 2018 Mar 21;11:1601-1616. doi: 10.2147/OTT.S147453. eCollection 2018.
Hepatocellular carcinoma (HCC) has an increasing incidence and high mortality. Surgical operation is not a comprehensive strategy for liver cancer. Moreover, tolerating systemic chemotherapy is difficult for patients with HCC because hepatic function is often impaired due to underlying cirrhosis. Therefore, a comprehensive strategy for cancer treatment should be developed. DTL (Cdc10-dependent transcript 2) is a critical regulator of cell cycle progression and genomic stability. In our previous study, the upregulation of DTL expression in aggressive HCC correlated positively with tumor grade and poor patient survival. We hypothesize that targeting DTL may provide a novel therapeutic strategy for liver cancer. DTL small interference RNAs were used to knock down DTL protein expression.
A clonogenic assay, immunostaining, double thymidine block, imaging flow cytometry analysis, and a tumor spheroid formation assay were used to analyze the role of DTL in tumor cell growth, cell cycle progression, micronucleation, ploidy, and tumorigenicity.
Our results demonstrated that targeting DTL reduced cell cycle regulators and chromosome segregation genes, resulting in increased cell micronucleation. DTL depletion inhibited liver cancer cell growth, increased senescence, and reduced tumorigenesis. DTL depletion resulted in the disruption of the mitotic proteins cyclin B, CDK1, securin, seprase, Aurora A, and Aurora B as well as the upregulation of the cell cycle arrest gene . A rescue assay indicated that DTL should be targeted through TPX2 downregulation for cancer cell growth inhibition. Moreover, DTL silencing inhibited the growth of patient-derived primary cultured HCC cells.
Our study results indicate that DTL is a potential novel target gene for treating liver cancer through liver cancer cell senescence induction. Furthermore, our results provide insights into molecular mechanisms for targeting DTL in liver cancer cells. The results also indicate several other starting points for future preclinical and clinical studies on liver cancer treatment.
肝细胞癌（HCC）的发病率日益上升且死亡率很高。手术并非肝癌的综合治疗策略。此外，HCC患者难以耐受全身化疗，因为潜在的肝硬化常导致肝功能受损。因此，应制定癌症治疗的综合策略。DTL（细胞分裂周期蛋白10依赖性转录物2）是细胞周期进程和基因组稳定性的关键调节因子。在我们之前的研究中，侵袭性HCC中DTL表达上调与肿瘤分级及患者不良生存呈正相关。我们推测靶向DTL可能为肝癌提供一种新的治疗策略。使用DTL小干扰RNA来敲低DTL蛋白表达。
采用克隆形成试验、免疫染色、双胸腺嘧啶核苷阻断、成像流式细胞术分析和肿瘤球形成试验来分析DTL在肿瘤细胞生长、细胞周期进程、微核形成、倍性和致瘤性中的作用。
我们的结果表明，靶向DTL可减少细胞周期调节因子和染色体分离基因，导致细胞微核形成增加。DTL缺失抑制肝癌细胞生长，增加衰老，并降低肿瘤发生。DTL缺失导致有丝分裂蛋白细胞周期蛋白B、细胞周期蛋白依赖性激酶1、分离酶抑制蛋白、分离酶、极光激酶A和极光激酶B的破坏以及细胞周期阻滞基因的上调。一项拯救试验表明，应通过下调TPX2来靶向DTL以抑制癌细胞生长。此外，DTL沉默抑制了患者来源的原代培养HCC细胞的生长。
我们的研究结果表明，DTL是通过诱导肝癌细胞衰老来治疗肝癌的潜在新靶基因。此外，我们的结果为肝癌细胞中靶向DTL的分子机制提供了见解。这些结果还指出了未来肝癌治疗临床前和临床研究的其他几个起点。
