Xian Jingrong, Wang Shiwen, Jiang Yanyu, Li Lihui, Cai Lili, Chen Ping, Liu Yue, Zeng Xiaofei, Chen Guoan, Ding Chen, Hoffman Robert M, Jia Lijun, Zhao Hu, Zhang Yanmei
Department of Laboratory Medicine, Huadong Hospital Affiliated to Fudan University, Shanghai 200040, China.
Cancer Institute, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China.
Cancer Biol Med. 2021 Mar 18;19(4):504-17. doi: 10.20892/j.issn.2095-3941.2020.0484.
The hyperactivated neddylation pathway plays an important role in tumorigenesis and is emerging as a promising anticancer target. We aimed to study whether NEDD8 (neural precursor cell expressed, developmentally down-regulated 8) might serve as a therapeutic target in esophageal squamous cell carcinoma (ESCC).
The clinical relevance of NEDD8 expression was evaluated by using The Cancer Genome Atlas (TCGA) database and tissue arrays. NEDD8-knockdown ESCC cells generated with the CRISPR/Cas9 system were used to explore the anticancer effects and mechanisms. Quantitative proteomic analysis was used to examine the variations in NEDD8 knockdown-induced biological pathways. The cell cycle and apoptosis were assessed with fluorescence activated cell sorting. A subcutaneous-transplantation mouse tumor model was established to investigate the anticancer potential of NEDD8 silencing .
NEDD8 was upregulated at both the mRNA and protein expression levels in ESCC, and NEDD8 overexpression was associated with poorer overall patient survival (mRNA level: = 0.028, protein level: = 0.026, log-rank test). Downregulation of NEDD8 significantly suppressed tumor growth both and . Quantitative proteomic analysis revealed that downregulation of NEDD8 induced cell cycle arrest, DNA damage, and apoptosis in ESCC cells. Mechanistic studies demonstrated that NEDD8 knockdown led to the accumulation of cullin-RING E3 ubiquitin ligases (CRLs) substrates through inactivation of CRLs, thus suppressing the malignant phenotype by inducing cell cycle arrest and apoptosis in ESCC. Rescue experiments demonstrated that the induction of apoptosis after NEDD8 silencing was attenuated by DR5 knockdown.
Our study elucidated the anti-ESCC effects and underlying mechanisms of NEDD8 knockdown, and validated NEDD8 as a potential target for ESCC therapy.
过度激活的NEDD8化途径在肿瘤发生过程中起重要作用，正成为一个有前景的抗癌靶点。我们旨在研究NEDD8（神经前体细胞表达，发育下调8）是否可作为食管鳞状细胞癌（ESCC）的治疗靶点。
利用癌症基因组图谱（TCGA）数据库和组织芯片评估NEDD8表达的临床相关性。使用CRISPR/Cas9系统构建的NEDD8敲低ESCC细胞来探究抗癌作用及机制。采用定量蛋白质组学分析来检测NEDD8敲低诱导的生物学途径变化。通过荧光激活细胞分选评估细胞周期和凋亡。建立皮下移植小鼠肿瘤模型以研究NEDD8沉默的抗癌潜力。
ESCC中NEDD8在mRNA和蛋白表达水平均上调，NEDD8过表达与患者总体生存率较差相关（mRNA水平：P = ０.０２８，蛋白水平：P = ０.０２６，对数秩检验）。NEDD8下调在体内和体外均显著抑制肿瘤生长。定量蛋白质组学分析显示，NEDD8下调诱导ESCC细胞的细胞周期停滞、DNA损伤和凋亡。机制研究表明，NEDD8敲低通过使Cullin-RING E3泛素连接酶（CRLs）失活导致CRLs底物积累，从而通过诱导ESCC细胞周期停滞和凋亡来抑制恶性表型。挽救实验表明，DR5敲低减弱了NEDD8沉默后诱导的凋亡。
我们的研究阐明了NEDD8敲低的抗ESCC作用及其潜在机制，并验证了NEDD8作为ESCC治疗潜在靶点的地位。
