Aspartate β-hydroxylase modulates cellular senescence through glycogen synthase kinase 3β in hepatocellular carcinoma.
Aspartate β-hydroxylase modulates cellular senescence through glycogen synthase kinase 3β in hepatocellular carcinoma.
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DOI：
10.1002/hep.28411
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发表时间：
2016-04
期刊：
影响因子：
--
通讯作者：
Wands JR
中科院分区：
文献类型：
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作者：
Iwagami Y;Huang CK;Olsen MJ;Thomas JM;Jang G;Kim M;Lin Q;Carlson RI;Wagner CE;Dong X;Wands JR
天冬氨酸β-羟化酶（ASPH）是一种在人肝细胞癌（HCC）肿瘤中过表达的酶，并参与恶性转化过程。我们确定ASPH是否是一个通过影响细胞衰老来延缓HCC进展的治疗靶点。ASPH敲低或敲除分别通过shRNA或CRISPR/Cas9系统实现，而酶抑制由ASPH的有效的第二代小分子抑制剂（SMI）提供。在人HCC细胞系中评价细胞增殖、集落形成和细胞衰老的改变。使用小鼠皮下和原位异种移植模型探索了激活细胞衰老的潜在机制。抑制ASPH表达和酶活性显著降低细胞增殖和集落形成，但诱导肿瘤细胞衰老。ASPH活性抑制后，GSK 3 β磷酸化水平和p16表达增加，促进细胞衰老，cyclin D1和PCNA表达减少，抑制细胞增殖。所涉及的机制表明，ASPH与GSK 3 β结合，并抑制其随后与AKT和p38上游激酶的相互作用，如免疫共沉淀所示。体内实验表明，SMI治疗肝癌荷瘤小鼠导致显著的剂量依赖性肿瘤生长减少，诱导GSK 3 β磷酸化，增强肿瘤细胞中p16表达并促进细胞衰老。我们已经确定了一种新的机制，通过调节肿瘤细胞的衰老来促进HCC的生长和进展。这些发现表明ASPH酶活性是HCC的新治疗靶点。
Aspartate β-hydroxylase (ASPH) is an enzyme overexpressed in human hepatocellular carcinoma (HCC) tumors and participates in the malignant transformation process. We determined if ASPH was a therapeutic target by exerting effects on cellular senescence to retard HCC progression. ASPH knockdown or knockout was achieved by shRNAs or CRISPR/Cas9 system, respectively, whereas enzymatic inhibition was rendered by a potent 2nd generation small molecule inhibitor (SMI) of ASPH. Alterations of cell proliferation, colony formation and cellular senescence were evaluated in human HCC cell lines. The potential mechanisms for activating cellular senescence were explored using murine subcutaneous and orthotopic xenograft models. Inhibition of ASPH expression and enzymatic activity significantly reduced cell proliferation and colony formation, but induced tumor cell senescence. Following inhibition of ASPH activity, phosphorylation of GSK3β and p16 expression were increased to promote senescence whereas cyclin D1 and PCNA were decreased to reduce cell proliferation. The mechanisms involved demonstrate that ASPH binds to GSK3β and inhibits its subsequent interactions with AKT and p38 upstream kinases as shown by co-immunoprecipitation. In vivo experiments demonstrated that the SMI treatment of HCC bearing mice resulted in significant dose-dependent reduced tumor growth, induced phosphorylation of GSK3β, enhanced p16 expression in tumor cells and promoted cellular senescence. We have identified a new mechanism that promotes HCC growth and progression by modulating senescence of tumor cells. These findings suggest that ASPH enzymatic activity is a novel therapeutic target for HCC.