Quantitative proteomics identification of phosphoglycerate mutase 1 as a novel therapeutic target in hepatocellular carcinoma
Quantitative proteomics identification of phosphoglycerate mutase 1 as a novel therapeutic target in hepatocellular carcinoma
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磷酸甘油酸变位酶 1 作为肝细胞癌新治疗靶点的定量蛋白质组学鉴定
DOI：
10.1186/1476-4598-9-81
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发表时间：
2010-04-19
期刊：
影响因子：
37.3
通讯作者：
Huang, Canhua
中科院分区：
文献类型：
--
作者：
Ren, Fenglian;Wu, Hong;Huang, Canhua
背景资料：肝细胞癌（Hepatocellular carcinoma，HCC）是世界上最常见的恶性肿瘤之一，由于对常规化疗耐药和放疗疗效有限，预后差。目前迫切需要开发新的生物标志物用于早期诊断，以及确定新的药物靶点用于治疗干预。患者和方法：54例配对肝癌标本和21例正常肝组织均来自四川大学华西医院。在分析前获得所有患者或其亲属的知情同意书，并获得四川大学机构伦理委员会的批准。采用基于细胞培养中氨基酸稳定同位素标记（SILAC）的蛋白质组学方法，对人肝癌细胞系HepG 2和永生化肝细胞系L02之间的差异表达蛋白质进行分析。使用临床样本通过半定量RT-PCR、免疫印迹和免疫组织化学进行PGAM 1表达的验证。由GenePharma Corporation（Shanghai，China）设计并构建特异性靶向PGAM 1的shRNA表达质粒，并用于在体外和体内沉默PGAM 1的表达。通过集落形成测定和Ki 67染色的组合测量细胞增殖。结果：共检测到63个蛋白质表达异常，其中51个蛋白质表达上调，12个蛋白质表达下调（2倍以上，p < 0.01）。发现磷酸甘油酸酯β 1（PGAM 1）显著上调。临床病理分析表明，66.7%的HCC与PGAM 1过表达有关，且PGAM 1过表达与低分化和低生存率密切相关（p < 0.01）。结论：PGAM 1在肝癌的发生发展过程中起重要作用，有望成为肝癌诊断的生物标志物和治疗的靶点。
Background: Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with poor prognosis due to resistance to conventional chemotherapy and limited efficacy of radiotherapy. There is an urgent need to develop novel biomarkers for early diagnosis, as well as to identify new drug targets for therapeutic interventions.Patients and methods: 54 paired HCC samples and 21 normal liver tissues were obtained from West China Hospital of Sichuan University. Informed consent was obtained from all the patients or their relatives prior to analysis, and the project was approved by the Institutional Ethics Committee of Sichuan University. Stable Isotope Labeling with Amino Acids in Cell Culture (SILAC)-based proteomics was employed to profile the differentially expressed proteins between a HepG2 human hepatoma cell line and an immortal hepatic cell line L02. Validation of PGAM1 expression was performed by semi-quantitative RT-PCR, immunoblot and immunohistochemistry using clinical samples. shRNA expressing plasmids specifically targeting PGAM1 were designed and constructed by GenePharma Corporation (Shanghai, China), and were utilized to silence expression of PGAM1 in vitro and in vivo. Cell proliferation was measured by a combination of colony formation assay and Ki67 staining. Apoptosis was examined by flow cytometry and TUNEL assay.Results: A total of 63 dysregulated proteins were identified, including 51 up-regulated proteins, and 12 down-regulated proteins (over 2-fold, p < 0.01). Phosphoglycerate mutase 1 (PGAM1) was found markedly upregulated. Clinico-pathological analysis indicated that overexpression of PGAM1 was associated with 66.7% HCC, and strongly correlated with poor differentiation and decreased survival rates (p < 0.01). shRNAs-mediated repression of PGAM1 expression resulted in significant inhibition in liver cancer cell growth both in vitro and in vivo.Conclusion: Our studies suggested that PGAM1 plays an important role in hepatocarcinogenesis, and should be a potential diagnostic biomarker, as well as an attractive therapeutic target for hepatocellular carcinoma.