Proteomic Analysis of Chemonaive Pediatric Osteosarcomas and Corresponding Normal Bone Reveals Multiple Altered Molecular Targets
Proteomic Analysis of Chemonaive Pediatric Osteosarcomas and Corresponding Normal Bone Reveals Multiple Altered Molecular Targets
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DOI：
10.1021/pr900113w
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发表时间：
2009-08-01
影响因子：
4.4
通讯作者：
Patino-Garcia, Ana
中科院分区：
文献类型：
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作者：
Folio, Cecilia;Mora, Maria I.;Patino-Garcia, Ana
为了确定与骨肉瘤的转化、转移或化疗耐药相关的蛋白质，我们开展了一种新的实验方法，该方法基于对同一患者的骨肉瘤和正常骨组织的蛋白质组图谱的比较。用双向差示凝胶电泳法获得5对细胞系(正常和肿瘤)的蛋白质组图谱。我们检测到56个差异蛋白质点(t检验，p<；0.05)。随后的纳米LC-ESI-MS/MS蛋白质表征使我们能够鉴定其中的一些蛋白质，其中16个蛋白质是根据骨肉瘤和配对的正常骨之间它们的相对丰度的变化而选择的，也因为它们的参与得到了基因组分析的支持。16个蛋白质中的两个，α-晶体蛋白B链(CryAB)和Ezrin(EZR1)被选作进一步的研究：对14对骨肉瘤/正常骨进行TMA(组织芯片)的免疫组织化学分析和实时PCR。这项第二级研究的结果证实，CryAB和Ezrin的数量显著增加，特别是在疾病的晚期。我们的总体结论是，对同一患者的骨肉瘤和正常骨组织的配对样本进行蛋白质组学分析是启动和继续寻找与儿童骨肉瘤相关的蛋白质和基因的一种可行且潜在的有效方法。
With a view to identify the proteins involved in transformation, metastasis or chemoresistance in pediatric osteosarcoma, we carried out a new experimental approach based on comparison of the proteomic profile of paired samples of osteosarcoma and normal bone tissues from the same patient. The proteomic profiles of five pairs of cell lines (normal vs tumoral) were obtained by two-dimensional difference gel electrophoresis. We detected 56 differential protein spots (t test, p < 0.05). Subsequent protein characterization by nano-LC-ESI-MS/MS enabled us to identify some of these proteins, 16 of which were chosen on the basis of the change of their relative abundance between osteosarcomas and paired normal bones and also because their involvement was supported by the genomic analysis. Two of the 16 proteins, Alpha-crystallin B chain (CRYAB) and ezrin (EZR1), were selected for further studies: an immunohistochemical analysis of a TMA (tissue microarray) and real-time PCR for a set of 14 osteosarcoma/normal-bone pairs. The results of this second tier of studies confirmed that there were significant increases in the amounts of CRYAB and ezrin, especially in advanced stages of the disease. Our overall conclusion is that proteomic profiling of paired samples of osteosarcoma and normal bone tissues from the same patient is a practicable and potentially powerful way of initiating and proceeding with a search for proteins and genes involved in pediatric osteosarcoma.