CD73 sustained cancer-stem-cell traits by promoting SOX9 expression and stability in hepatocellular carcinoma
CD73 sustained cancer-stem-cell traits by promoting SOX9 expression and stability in hepatocellular carcinoma
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CD73 通过促进肝细胞癌中 SOX9 的表达和稳定性来维持癌症干细胞特征
DOI：
10.1186/s13045-020-0845-z
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发表时间：
2020-02-05
影响因子：
28.5
通讯作者：
Lu, Ren-Quan
中科院分区：
文献类型：
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作者：
Ma, Xiao-Lu;Hu, Bo;Lu, Ren-Quan
背景AKT异常激活与肝细胞癌的肿瘤干细胞(CSC)特征有关。我们先前报道CD73通过Rap1/p110β级联激活AKT信号。在此，我们进一步探讨了CD73在调节肝细胞癌CSC特性中的作用。方法慢病毒转染法进行CD73的表达调控。采用磁选法分离纯化CD73+组分，并用荧光激活细胞分选法评价其分化潜能。用球体形成法评价CSC的体外特性，建立NOD/SCID小鼠皮下模型评价体内CSC的特性，用集落形成试验评价耐药能力。还测定了与茎相关的基因表达，并通过评估免疫沉淀和泛素化来研究其潜在的机制。结果CD73的表达与肝细胞癌球体形成能力呈正相关，且在肝细胞癌球体中表达增强。CD73基因敲除阻碍了球体的形成、Lenvatinib抗性和茎相关基因的表达，而CD73过表达则达到了相反的效果。此外，CD73基因敲除显著抑制了肿瘤的体内增殖能力。值得注意的是，我们发现CD73+细胞比CD73-细胞表现出更强的CSC特征。机制上，CD73通过AKT依赖的两种机制发挥其原茎活性：通过c-Myc激活SOX9转录，通过抑制糖原合成酶K3β抑制SOX9降解。临床上，联合检测CD73和SOX9可更准确地预测预后。结论CD73通过上调SOX9的表达和增强其蛋白质的稳定性，在维持CSCs的特性方面发挥了关键作用。靶向CD73可能是根除CSCs和逆转Lenvatinib耐药的一种有前景的策略。
Background Aberrant AKT activation contributes to cancer stem cell (CSC) traits in hepatocellular carcinoma (HCC). We previously reported that CD73 activated AKT signaling via the Rap1/P110 beta cascade. Here, we further explored the roles of CD73 in regulating CSC characteristics of HCC. Methods CD73 expression modulations were conducted by lentiviral transfections. CD73+ fractions were purified by magnetic-based sorting, and fluorescent-activated cell sorting was used to assess differentiation potentials. A sphere-forming assay was performed to evaluate CSC traits in vitro, subcutaneous NOD/SCID mice models were generated to assess in vivo CSC features, and colony formation assays assessed drug resistance capacities. Stemness-associated gene expression was also determined, and underlying mechanisms were investigated by evaluating immunoprecipitation and ubiquitylation. Results We found CD73 expression was positively associated with sphere-forming capacity and elevated in HCC spheroids. CD73 knockdown hindered sphere formation, Lenvatinib resistance, and stemness-associated gene expression, while CD73 overexpression achieved the opposite effects. Moreover, CD73 knockdown significantly inhibited the in vivo tumor propagation capacity. Notably, we found that CD73+ cells exhibited substantially stronger CSC traits than their CD73- counterparts. Mechanistically, CD73 exerted its pro-stemness activity through dual AKT-dependent mechanisms: activating SOX9 transcription via c-Myc, and preventing SOX9 degradation by inhibiting glycogen synthase kinase 3 beta. Clinically, the combined analysis of CD73 and SOX9 achieved a more accurate prediction of prognosis. Conclusions Collectively, CD73 plays a critical role in sustaining CSCs traits by upregulating SOX9 expression and enhancing its protein stability. Targeting CD73 might be a promising strategy to eradicate CSCs and reverse Lenvatinib resistance in HCC.