mRNA sequencing of human colorectal cancer organoids. mRNA sequencing of human colorectal cancer organoids

Given that colorectal cancer stem cells (CCSCs) play key roles in the tumor dormancy, metastasis and relapse, targeting CCSCs is a promising strategy in cancer therapy. Here, we aimed to identify new regulators of CCSCs and found that Cullin 4B (CUL4B), which possesses oncogenic properties in multiple solid tumors, drives the development and metastasis of colon cancer by sustaining cancer stem-like features. Elevated expression of CUL4B was confirmed in colon tumors and was associated with poor overall survival. Inhibition of CUL4B in cancer cell lines and patient-derived tumor organoids led to reduced sphere formation, proliferation and metastasis capacity. Mechanistically, CUL4B coordinates with PRC2 complex to repress miR34a expression, thus up-regulates oncogenes including MYCN and NOTCH1, which are targeted by miR34a. Mutation of miR34a binding sites in the 3'UTR of the oncogenes rescues the phenotype caused by CUL4B depletion. Significant correlations were found between expression of CUL4B and miR34a (negatively), miR34a target genes (positively) in clinical samples from colon cancer patients. Collectively, our work demonstrates that CUL4B functions to repress miR34a in maintaining cancer stemness in CRC and provides a potential therapeutic target. Overall design: We generated transciptome data from human colorectal cancer organoids with knocked-down CUL4B or over-expressed CUL4B

结直肠癌干细胞（colorectal cancer stem cells, CCSCs）在肿瘤休眠、转移与复发过程中发挥关键作用，靶向CCSCs是癌症治疗中极具前景的策略。本研究旨在鉴定CCSCs的新型调控因子，结果发现，在多种实体瘤中具有致癌特性的Cullin 4B（CUL4B）可通过维持肿瘤干细胞样特征，推动结肠癌的发生与转移。研究证实，CUL4B在结肠肿瘤组织中呈高表达，且与患者不良总生存期显著相关。在癌细胞系及患者来源的肿瘤类器官中抑制CUL4B表达，可降低细胞成球能力、增殖活性与转移潜能。机制研究表明，CUL4B可与多梳抑制复合体2（PRC2 complex）协同，抑制miR34a的表达，进而上调miR34a的靶基因——包括MYCN与NOTCH1在内的致癌基因。将上述致癌基因3'非翻译区（3'UTR）中的miR34a结合位点突变，可挽救CUL4B敲低所引发的表型。在结肠癌患者的临床样本中，CUL4B的表达与miR34a呈负相关，与miR34a靶基因呈正相关，二者均存在显著关联。综上，本研究证实CUL4B通过抑制miR34a的表达，维持结直肠癌的肿瘤干细胞干性，为结直肠癌治疗提供了潜在靶点。实验整体设计：本研究对CUL4B敲低或过表达的人类结直肠癌类器官进行转录组（transcriptome）测序，获取了其转录组数据。