Wang Huan, Li Bin, Yan Kang, Wu Yonghong, Wen Yanhua, Liu Yunyan, Fan Pei, Ma Qiong
Orthopedic Oncology Institute, Department of Orthopedic Surgery, Tangdu Hospital, Fourth Military Medical University, Xi'an, China.
Department of Orthopedics, The Second Affiliated Hospital of Wenzhou Medical University, Yuying Children's Hospital, Wenzhou, China.
Front Oncol. 2021 Mar 9;11:602712. doi: 10.3389/fonc.2021.602712. eCollection 2021.
Lobaplatin is a third-generation platinum-based antineoplastic agent and is widely used for osteosarcoma treatment before and after tumor removal. However, treatment failure often results from lobaplatin drug resistance. In our study, we found that SaOS-2 and SOSP-9607 osteosarcoma cells became less sensitive to lobaplatin after treatment with exogenous interleukin (IL)-6. Quantitative proteomic analysis was performed to elucidate the underlying mechanism in SaOS-2 osteosarcoma cells. Cells were divided into a control group (CG), a lobaplatin treatment group (LG), a recombinant human IL-6 (rhIL-6), and a lobaplatin treatment group (rhILG). We performed three biological replicates in each group to compare the differential protein expression between groups using a tandem mass tag (TMT) labeling technology based on liquid chromatography-tandem mass spectrometry (LC-MS/MS). A total of 1,313 proteins with significant differential expression was identified and quantified. The general characteristics of the significantly enriched proteins were identified by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, and protein-protein interaction (PPI) analysis was conducted using IntAct and STRING. In total, 31 proteins were further verified by parallel reaction monitoring (PRM), among which ras GTPase-activating protein-binding protein 1 (G3BP1), fragile X mental retardation syndrome-related protein 1 (hFXR1p), and far upstream element-binding protein 1 (FUBP1) were significantly differentially expressed. Immunohistochemistry results showed that these three proteins are highly expressed in specimens from platinum-resistant osteosarcoma patients, while the proteins are negatively or weakly expressed in specimens from platinum-sensitive osteosarcoma patients. The immunofluorescence staining results were in accord with the immunohistochemistry staining results. siRNA knockdown of FUBP1 showed a strikingly decreased IC50 value for lobaplatin in FUBP1-silenced cells, which verified the role of FUBP1 in the drug susceptibility of osteosarcoma and the potential therapeutic value for increasing the sensitivity to lobaplatin. This is the first proteomic study on a rhIL-6 intervention before lobaplatin treatment in osteosarcoma cells.
洛铂是第三代铂类抗肿瘤药物，广泛用于骨肉瘤切除前后的治疗。然而，治疗失败往往是由于对洛铂产生耐药性。在我们的研究中，我们发现用外源性白细胞介素（IL）-6处理后，SaOS-2和SOSP-9607骨肉瘤细胞对洛铂的敏感性降低。进行了定量蛋白质组学分析以阐明SaOS-2骨肉瘤细胞中的潜在机制。细胞分为对照组（CG）、洛铂治疗组（LG）、重组人IL-6组（rhIL-6）和洛铂治疗组（rhILG）。我们在每组中进行了三次生物学重复，以基于液相色谱-串联质谱（LC-MS/MS）的串联质量标签（TMT）标记技术比较各组之间的差异蛋白表达。共鉴定和定量了1313种具有显著差异表达的蛋白质。通过基因本体论（GO）和京都基因与基因组百科全书（KEGG）富集分析鉴定了显著富集蛋白质的一般特征，并使用IntAct和STRING进行了蛋白质-蛋白质相互作用（PPI）分析。总共通过平行反应监测（PRM）进一步验证了31种蛋白质，其中ras GTP酶激活蛋白结合蛋白1（G3BP1）、脆性X智力低下综合征相关蛋白1（hFXR1p）和远上游元件结合蛋白1（FUBP1）有显著差异表达。免疫组织化学结果表明，这三种蛋白在铂耐药骨肉瘤患者的标本中高表达，而在铂敏感骨肉瘤患者的标本中阴性或弱表达。免疫荧光染色结果与免疫组织化学染色结果一致。FUBP1的siRNA敲低显示FUBP1沉默细胞中洛铂的IC50值显著降低，这证实了FUBP1在骨肉瘤药物敏感性中的作用以及提高对洛铂敏感性的潜在治疗价值。这是第一项关于在骨肉瘤细胞中洛铂治疗前进行rhIL-6干预的蛋白质组学研究。
