Nek2 通过调节肝癌细胞中β-catenin 的泛素化和定位来增强索拉非尼耐药性。
Nek2 augments sorafenib resistance by regulating the ubiquitination and localization of β-catenin in hepatocellular carcinoma.
机构信息
Department of Radiation Oncology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, China.
Hepatology Unit and Department of Infectious Diseases, Nanfang Hospital, Southern Medical University, 1838 Guangzhoudadaobei Road, Guangzhou, Guangzhou, 510515, China.
出版信息
J Exp Clin Cancer Res. 2019 Jul 18;38(1):316. doi: 10.1186/s13046-019-1311-z.
BACKGROUND
Sorafenib is the first-line treatment for advanced-stage hepatocellular carcinoma (HCC). Several studies have shown that the up-regulation of β-catenin plays a role in sorafenib resistance in HCC; however, the mechanism associated with this phenomenon remains elusive.
METHODS
Western blotting, flow cytometry, and an evaluation of IC values were used to confirm the role of β-catenin in HCC sorafenib resistance. Immunoprecipitation and western blotting were then performed to identify regulatory interactions between β-catenin and Nek2. Further, western blotting, flow cytometry, and an in vivo xenograft model were used to evaluate the function of Nek2 in HCC sorafenib resistance, whereas rescue experiments were performed to confirm that Nek2 induces sorafenib resistance via β-catenin. Finally, western blotting and immunohistochemistry were used to evaluate the expression level of Nek2 in paired HCC and non-tumor tissues.
RESULTS
We showed that β-catenin could suppress sorafenib-induced apoptosis and cell growth inhibition in HCC cell lines. By screening β-catenin-interacting proteins, we found that Nek2 could bind β-catenin in sorafenib-treated HCC cell lines. Our results also showed that Nek2 stabilizes β-catenin and promotes its translocation to the nucleus, consequently activating the transcription of downstream target genes. We further confirmed that Nek2 could induce sorafenib resistance in HCC cell lines, and that β-catenin was the key element involved in this process. Further, a xenograft tumor model showed that Nek2 knockdown could improve the anti-tumor effect of sorafenib, whereas an analysis of tumor proteins showed that Nek2 regulates β-catenin protein levels and its nuclear translocation in vivo. In addition, Nek2 was found to be up-regulated in HCC tissue, and especially in advanced-stage disease.
CONCLUSIONS
Our study proves that Nek2 induces HCC sorafenib resistance via β-catenin and suggests a novel therapeutic strategy to improve the anti-tumor effects of sorafenib in HCC.
背景
索拉非尼是晚期肝细胞癌（HCC）的一线治疗药物。多项研究表明，β-连环蛋白的上调在 HCC 索拉非尼耐药中起作用；然而，与这种现象相关的机制仍不清楚。
方法
使用 Western blot、流式细胞术和 IC 值评估来确认 β-连环蛋白在 HCC 索拉非尼耐药中的作用。然后进行免疫沉淀和 Western blot 以鉴定β-连环蛋白和 Nek2 之间的调节相互作用。进一步使用 Western blot、流式细胞术和体内异种移植模型来评估 Nek2 在 HCC 索拉非尼耐药中的作用，而进行挽救实验以确认 Nek2 通过β-连环蛋白诱导索拉非尼耐药。最后，使用 Western blot 和免疫组织化学评估配对的 HCC 和非肿瘤组织中 Nek2 的表达水平。
结果
我们表明β-连环蛋白可以抑制 HCC 细胞系中索拉非尼诱导的细胞凋亡和生长抑制。通过筛选β-连环蛋白相互作用蛋白，我们发现 Nek2 可以在索拉非尼处理的 HCC 细胞系中与β-连环蛋白结合。我们的结果还表明，Nek2 稳定β-连环蛋白并促进其向核内易位，从而激活下游靶基因的转录。我们进一步证实，Nek2 可诱导 HCC 细胞系中的索拉非尼耐药，而β-连环蛋白是参与该过程的关键因素。此外，异种移植肿瘤模型表明，Nek2 敲低可改善索拉非尼的抗肿瘤作用，而对肿瘤蛋白的分析表明，Nek2 在体内调节β-连环蛋白蛋白水平及其核易位。此外，发现 Nek2 在 HCC 组织中上调，尤其是在晚期疾病中。
结论
我们的研究证明，Nek2 通过β-连环蛋白诱导 HCC 索拉非尼耐药，并提出了一种新的治疗策略，以提高索拉非尼在 HCC 中的抗肿瘤作用。
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