Yeh Chao-Chi, Hsu Chih-Hung, Shao Yu-Yun, Ho Wen-Ching, Tsai Mong-Hsun, Feng Wen-Chi, Chow Lu-Ping
From the ‡Graduate Institute of Biochemistry and Molecular Biology.
§Graduate Institute of Oncology, College of Medicine, ‖Department of Oncology, National Taiwan University Hospital, Taipei, Taiwan.
Mol Cell Proteomics. 2015 Jun;14(6):1527-45. doi: 10.1074/mcp.M114.046417. Epub 2015 Apr 7.
Sorafenib has become the standard therapy for patients with advanced hepatocellular carcinoma (HCC). Unfortunately, most patients eventually develop acquired resistance. Therefore, it is important to identify potential biomarkers that could predict the efficacy of sorafenib. To identify target proteins associated with the development of sorafenib resistance, we applied stable isotope labelling with amino acids in cell culture (SILAC)-based quantitative proteomic approach to analyze differences in protein expression levels between parental HuH-7 and sorafenib-acquired resistance HuH-7 (HuH-7(R)) cells in vitro, combined with an isobaric tags for relative and absolute quantitation (iTRAQ) quantitative analysis of HuH-7 and HuH-7(R) tumors in vivo. In total, 2,450 quantified proteins were identified in common in SILAC and iTRAQ experiments, with 81 showing increased expression (>2.0-fold) with sorafenib resistance and 75 showing decreased expression (<0.5-fold). In silico analyses of these differentially expressed proteins predicted that 10 proteins were related to cancer with involvements in cell adhesion, migration, and invasion. Knockdown of one of these candidate proteins, galectin-1, decreased cell proliferation and metastasis in HuH-7(R) cells and restored sensitivity to sorafenib. We verified galectin-1 as a predictive marker of sorafenib resistance and a downstream target of the AKT/mTOR/HIF-1α signaling pathway. In addition, increased galectin-1 expression in HCC patients' serum was associated with poor tumor control and low response rate. We also found that a high serum galectin-1 level was an independent factor associated with poor progression-free survival and overall survival. In conclusion, these results suggest that galectin-1 is a possible biomarker for predicting the response of HCC patients to treatment with sorafenib. As such, it may assist in the stratification of HCC and help direct personalized therapy.
索拉非尼已成为晚期肝细胞癌（HCC）患者的标准治疗药物。不幸的是，大多数患者最终会产生获得性耐药。因此，识别能够预测索拉非尼疗效的潜在生物标志物非常重要。为了识别与索拉非尼耐药发展相关的靶蛋白，我们应用基于细胞培养中氨基酸稳定同位素标记（SILAC）的定量蛋白质组学方法，在体外分析亲本HuH-7细胞和获得索拉非尼耐药的HuH-7（HuH-7(R)）细胞之间的蛋白质表达水平差异，并结合体内HuH-7和HuH-7(R)肿瘤的相对和绝对定量等压标签（iTRAQ）定量分析。在SILAC和iTRAQ实验中总共鉴定出2450种定量蛋白质，其中81种在索拉非尼耐药时表达增加（>2.0倍），75种表达减少（<0.5倍）。对这些差异表达蛋白质的计算机分析预测，有10种蛋白质与癌症相关，涉及细胞粘附、迁移和侵袭。敲低这些候选蛋白之一的半乳糖凝集素-1，可降低HuH-7(R)细胞的增殖和转移，并恢复对索拉非尼的敏感性。我们验证了半乳糖凝集素-1是索拉非尼耐药的预测标志物以及AKT/mTOR/HIF-1α信号通路的下游靶点。此外，HCC患者血清中半乳糖凝集素-1表达增加与肿瘤控制不佳和低反应率相关。我们还发现高血清半乳糖凝集素-1水平是与无进展生存期和总生存期差相关的独立因素。总之，这些结果表明半乳糖凝集素-1可能是预测HCC患者对索拉非尼治疗反应的生物标志物。因此，它可能有助于HCC的分层并指导个性化治疗。
