Khanna C, Khan J, Nguyen P, Prehn J, Caylor J, Yeung C, Trepel J, Meltzer P, Helman L
Pediatric Oncology, National Cancer Institute, and Cancer Genetics Branch, Human Genome Research Institute, NIH , Bethesda, Maryland 20892, USA.
Cancer Res. 2001 May 1;61(9):3750-9.
Despite advances in the management of osteosarcoma (OSA) and other solid tumors, the development of metastasis continues to be the most significant problem and cause of death for cancer patients. To define genetic determinants of pulmonary metastasis, we have applied cDNA microarrays to a recently described murine model of OSA that is characterized by orthotopic tumor growth, a period of minimal residual disease, spontaneous pulmonary metastasis, and cell line variants that differ in metastatic potential. Microarray analysis defined 53 genes (of 3166 unique cDNAs) that were differentially expressed between the primary tumors of the more aggressive (K7M2) and less aggressive (K12) OSA models. By review of the literature, these differentially expressed genes were assigned to six nonmutually exclusive metastasis-associated categories (proliferation and apoptosis, motility and cytoskeleton, invasion, immune surveillance, adherence, and angiogenesis). Functional studies to evaluate K7M2 and K12 for differences in each of these metastasis-associated processes revealed enhanced motility, adherence, and angiogenesis in the more aggressive K7M2 model. For this reason, 10 of the 53 differentially expressed genes that were assigned to the motility and cytoskeleton, adherence, and angiogenesis categories were considered as most likely to define differences in the metastatic behavior of the two models. Ezrin, a gene not described previously in OSA, with functions in motility, invasion, and adherence, was 3-fold overexpressed in K7M2 compared with K12 by microarray. Differential expression for RNA was confirmed by Northern analysis and for protein by immunostaining. Alterations in ezrin protein levels and concomitant cytoskeletal changes in our model confirmed predictions from the arrays. The potential relevance of ezrin in OSA was suggested by its expression in five of five human OSA cell lines. This work represents a rationale approach to the evaluation of microarray data and will be useful to identify genes that may be causally associated with metastasis.
尽管骨肉瘤（OSA）和其他实体瘤的治疗取得了进展，但转移的发生仍然是癌症患者最严重的问题和死亡原因。为了确定肺转移的遗传决定因素，我们将cDNA微阵列应用于最近描述的一种OSA小鼠模型，该模型的特点是原位肿瘤生长、一段微小残留病期、自发肺转移以及具有不同转移潜能的细胞系变体。微阵列分析确定了3166个独特cDNA中的53个基因，这些基因在侵袭性较强（K7M2）和侵袭性较弱（K12）的OSA模型的原发性肿瘤之间存在差异表达。通过文献综述，这些差异表达基因被归为六个非相互排斥的转移相关类别（增殖与凋亡、运动与细胞骨架、侵袭、免疫监视、黏附以及血管生成）。评估K7M2和K12在这些转移相关过程中差异的功能研究显示，侵袭性较强的K7M2模型具有增强的运动性、黏附性和血管生成能力。因此，被归为运动与细胞骨架、黏附以及血管生成类别的53个差异表达基因中的10个被认为最有可能界定这两种模型转移行为的差异。埃兹蛋白（ezrin）是一种此前未在OSA中描述过的基因，具有运动、侵袭和黏附功能，通过微阵列分析发现其在K7M2中的表达比K12高3倍。通过Northern分析证实了RNA的差异表达，通过免疫染色证实了蛋白质的差异表达。我们模型中埃兹蛋白水平的改变以及伴随的细胞骨架变化证实了微阵列的预测。埃兹蛋白在五种人类OSA细胞系中的表达表明了其在OSA中的潜在相关性。这项工作代表了一种评估微阵列数据的合理方法，将有助于识别可能与转移有因果关系的基因。
