Sun Jiaqi, Wang Xiyang, Shen Qingsong, Wang Min, Chen Shuxian, Zhang Xuechun, Huang Yongping, Zhang Zhonglin, Li Wenhua, Yuan Yufeng, Huang Zan
College of Life Sciences, Hubei Key Laboratory of Cell Homeostasis, Wuhan University, Wuhan, China.
Department of Hepatobiliary and Pancreatic Surgery, Zhongnan Hospital of Wuhan University, Wuhan, China.
Cell Oncol (Dordr). 2022 Dec;45(6):1187-1202. doi: 10.1007/s13402-022-00709-1. Epub 2022 Nov 3.
Dysregulated cell cycle targeting is a well-established therapeutic strategy against hepatocellular carcinoma (HCC). Dissecting the underlying mechanism may improve the efficacy of HCC therapy.
HCC data from TCGA and new clinical samples were used for DNASE1L3 expression analysis and for assessing its correlation with HCC development. The in vitro function of DNASE1L3 in HCC cell proliferation, colony formation, migration and invasion was assessed using RTCA, CCK-8 and transwell assays and the in vivo function in subcutaneous tumor formation in a xenograft nude mouse model. The role of DNASE1L3 in HCC tumorigenesis was further verified in AKT/NRASV12-induced and DEN/CCl-induced primary liver cancers in wildtype and Dnase1l3 mice. Finally, RNA-Seq analysis followed by biochemical methods including cell cycle, immunofluorescence, co-immunoprecipitation and Western blotting assays were employed to reveal the underlying mechanism.
We found that DNASE1L3 was significantly downregulated and served as a favorable prognostic factor in HCC. DNASE1L3 dramatically attenuated HCC cell proliferation, colony formation, migration and invasion in vitro and reduced subcutaneous tumor formation in nude mice in vivo. Furthermore, DNASE1L3 overexpression dampened AKT/NRASV12-induced mouse liver cancer in wildtype mice and DNASE1L3 deficiency worsened DEN/CCl-induced liver cancer in Dnase1l3 mice. Systemic analysis revealed that DNASE1L3 impaired HCC cell cycle progression by interacting with CDK2 and inhibiting CDK2-stimulated E2F1 activity. C-terminal deletion (DNASE1L3) diminished the interaction with CDK2 and abrogated the inhibitory function against HCC.
Our study unveils DNASE1L3 as a novel HCC cell cycle regulator and tumor suppressor. DNASE1L3 impairs HCC tumorigenesis by delaying cell cycle progression possibly through disrupting the positive E2F1-CDK2 regulatory loop. DNASE1L3 may serve as a target for the development of novel therapeutic strategies against HCC.
失调的细胞周期靶向是一种成熟的抗肝细胞癌（HCC）治疗策略。剖析其潜在机制可能会提高HCC治疗的疗效。
来自TCGA的HCC数据和新的临床样本用于DNASE1L3表达分析，并评估其与HCC发生的相关性。使用RTCA、CCK-8和transwell试验评估DNASE1L3在HCC细胞增殖、集落形成、迁移和侵袭中的体外功能，以及在异种移植裸鼠模型中皮下肿瘤形成的体内功能。在野生型和Dnase1l3小鼠的AKT/NRASV12诱导和DEN/CCl诱导的原发性肝癌中进一步验证DNASE1L3在HCC肿瘤发生中的作用。最后，采用RNA-Seq分析，随后通过包括细胞周期、免疫荧光、免疫共沉淀和蛋白质印迹分析在内的生化方法来揭示潜在机制。
我们发现DNASE1L3在HCC中显著下调，并作为一个有利的预后因素。DNASE1L3在体外显著减弱HCC细胞的增殖、集落形成、迁移和侵袭，并在体内减少裸鼠皮下肿瘤的形成。此外，DNASE1L3过表达抑制野生型小鼠中AKT/NRASV12诱导的小鼠肝癌，而DNASE1L3缺陷则使Dnase1l3小鼠中DEN/CCl诱导的肝癌恶化。系统分析表明，DNASE1L3通过与CDK2相互作用并抑制CDK2刺激的E2F1活性来损害HCC细胞周期进程。C末端缺失（DNASE1L3）减少了与CDK2的相互作用，并消除了对HCC 的抑制功能。
我们的研究揭示DNASE1L3是一种新型的HCC细胞周期调节因子和肿瘤抑制因子。DNASE1L3可能通过破坏正向E2F1-CDK2调节环来延迟细胞周期进程，从而损害HCC肿瘤发生。DNASE1L3可能作为开发新型抗HCC治疗策略的靶点。
