Shen Mingfang, Zhang Yunfeng, Tang Lun, Fu Qinyan, Zhang Jiawei, Xu Yang, Zeng Hui, Li Yuan
Department of Hematology, the First Hospital of Jiaxing, 314001, Zhejiang, China.
Department of Hematology, the Second Affiliated Hospital, Zhejiang University School of Medicine, 310009, Zhejiang, China.
BMC Cancer. 2023 Oct 13;23(1):978. doi: 10.1186/s12885-023-11186-6.
Cell division cycle 6 (CDC6) is a key licensing factor in the assembly of pre-replicative complexes at origins of replication. The role of CDC6 in the pathogenesis of in diffuse larger B-cell lymphoma (DLBCL) remains unknown. We aim to investigate the effects of CDC6 on the proliferation, apoptosis and cell cycle regulation in DLBCL cells, delineate its underlying mechanism, and to correlate CDC6 expression with clinical characteristics and prognosis of patients with DLBCL.
Initial bioinformatic analysis was performed to screen the potential role of CDC6 in DLBCL. Lentiviral constructs harboring CDC6 or shCDC6 was transfected to overexpress or knockdown CDC6 in SUDHL4 and OCI-LY7 cells. The cell proliferation was evaluated by CCK-8 assay, cell apoptosis was detected by Annexin-V APC/7-AAD double staining, and cell cycle was measured by flow cytometry. Real time quantitative PCR and western blot was used to characterize CDC6 expression and its downstream signaling pathways. The clinical data of DLBCL patients were retrospectively reviewed, the CDC6 expression in DLBCL or lymph node reactive hyperplasia tissues was evaluated by immunohistochemistry.
In silico data suggest that CDC6 overexpression is associated with inferior prognosis of DLBCL. We found that CDC6 overexpression increased SUDHL4 or OCI-LY7 cell proliferation, while knockdown of CDC6 inhibited cell proliferation in a time-dependent manner. Upon overexpression, CDC6 reduced cells in G1 phase and did not affect cell apoptosis; CDC6 knockdown led to significant cell cycle arrest in G1 phase and increase in cell apoptosis. Western blot showed that CDC6 inhibited the expression of INK4, E-Cadherin and ATR, accompanied by increased Bcl-2 and deceased Bax expression. The CDC6 protein was overexpressed DLBCL compared with lymph node reactive hyperplasia, and CDC6 overexpression was associated with non-GCB subtype, and conferred poor PFS and OS in patients with DLBCL.
CDC6 promotes cell proliferation and survival of DLBCL cells through regulation of G1/S cell cycle checkpoint and apoptosis. CDC6 is overexpressed and serves as a novel prognostic marker in DLBCL.
细胞分裂周期6（CDC6）是复制起始点预复制复合物组装中的关键许可因子。CDC6在弥漫性大B细胞淋巴瘤（DLBCL）发病机制中的作用尚不清楚。我们旨在研究CDC6对DLBCL细胞增殖、凋亡和细胞周期调控的影响，阐明其潜在机制，并将CDC6表达与DLBCL患者的临床特征和预后相关联。
进行初步生物信息学分析以筛选CDC6在DLBCL中的潜在作用。将携带CDC6或shCDC6的慢病毒构建体转染至SUDHL4和OCI-LY7细胞中，以过表达或敲低CDC6。通过CCK-8法评估细胞增殖，通过Annexin-V APC/7-AAD双染检测细胞凋亡，并通过流式细胞术测量细胞周期。使用实时定量PCR和蛋白质印迹来表征CDC6表达及其下游信号通路。回顾性分析DLBCL患者的临床数据，通过免疫组织化学评估DLBCL或淋巴结反应性增生组织中CDC6的表达。
计算机数据表明，CDC6过表达与DLBCL的不良预后相关。我们发现，CDC6过表达增加了SUDHL4或OCI-LY7细胞的增殖，而敲低CDC6则以时间依赖性方式抑制细胞增殖。过表达时，CDC6减少了G1期细胞，且不影响细胞凋亡；CDC6敲低导致显著的G1期细胞周期阻滞并增加细胞凋亡。蛋白质印迹显示，CDC6抑制INK4、E-钙黏蛋白和ATR的表达，同时Bcl-2表达增加，Bax表达降低。与淋巴结反应性增生相比，DLBCL中CDC6蛋白过表达，且CDC6过表达与非生发中心B细胞（GCB）亚型相关，并导致DLBCL患者的无进展生存期（PFS）和总生存期（OS）较差。
CDC6通过调节G1/S细胞周期检查点和凋亡促进DLBCL细胞增殖和存活。CDC6在DLBCL中过表达，是一种新的预后标志物。
