Circulating tumour cells escape from EpCAM-based detection due to epithelial-to-mesenchymal transition.
Circulating tumour cells escape from EpCAM-based detection due to epithelial-to-mesenchymal transition.
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DOI：
10.1186/1471-2407-12-178
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发表时间：
2012-05-16
期刊：
影响因子：
3.8
通讯作者：
von Ahsen O
中科院分区：
文献类型：
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作者：
Gorges TM;Tinhofer I;Drosch M;Röse L;Zollner TM;Krahn T;von Ahsen O
循环肿瘤细胞（CTC）已显示出与转移性乳腺癌、前列腺癌、结肠癌和胰腺癌的预后相关性。为了进一步开发CTC作为生物标志物，我们比较了在鼠乳腺癌异种移植模型（MDA-MB-231、MDA-MB-468和KPL-4）中CTC检测的不同方案的性能。从携带肿瘤的动物（20至200 mm 2）采集血液样品，并使用1.基于上皮标志物的富集方法（AdnaTest），2.抗体非依赖性技术，靶向人基因转录物（定性PCR），和3.一种不依赖抗体的方法，靶向人DNA序列（定量PCR）。此外，与上皮-间充质转化（EMT）相关的基因表达变化用EMT特异性PCR测定。我们使用市售的Adna测试、对人类管家基因的RT-PCR和对HCTJ序列的PCR来检测异种移植物模型中的CTC。CTC的表型变化用市售的“人上皮至间充质转化RT-Profiler PCR阵列”测试。尽管AdnaTest在1 ml小鼠血液加标实验中检测到少至1个肿瘤细胞，但尽管有可见的转移形成，但用该方法在体内检测不到CTC。然而，CTC的存在可以通过靶向人转录物或DNA序列的PCR来证明，而无需上皮预富集。AdnaTest检测CTC的失败是由于EpCAM的下调，而间充质标志物如Twist和EGFR在CTC上上调。已经报道了在肿瘤细胞转移扩散期间表达谱的这种变化，并且与称为上皮-间充质转化（EMT）的生物学程序有关。使用基于EpCAM的富集技术导致无法检测已经经历EMT的CTC群体。我们的研究结果可以解释即使在晚期转移性癌症患者中也报告了低CTC数量的临床结果。这些结果是鉴定用于检测或捕获CTC（包括间充质样亚群）的新标志物的起点。
Circulating tumour cells (CTCs) have shown prognostic relevance in metastatic breast, prostate, colon and pancreatic cancer. For further development of CTCs as a biomarker, we compared the performance of different protocols for CTC detection in murine breast cancer xenograft models (MDA-MB-231, MDA-MB-468 and KPL-4). Blood samples were taken from tumour bearing animals (20 to 200 mm2) and analysed for CTCs using 1. an epithelial marker based enrichment method (AdnaTest), 2. an antibody independent technique, targeting human gene transcripts (qualitative PCR), and 3. an antibody-independent approach, targeting human DNA-sequences (quantitative PCR). Further, gene expression changes associated with epithelial-to-mesenchymal transition (EMT) were determined with an EMT-specific PCR assay. We used the commercially available Adna Test, RT-PCR on human housekeeping genes and a PCR on AluJ sequences to detect CTCs in xenografts models. Phenotypic changes in CTCs were tested with the commercially available “Human Epithelial to Mesenchymal Transition RT-Profiler PCR Array”. Although the AdnaTest detects as few as 1 tumour cell in 1 ml of mouse blood spiking experiments, no CTCs were detectable with this approach in vivo despite visible metastasis formation. The presence of CTCs could, however, be demonstrated by PCR targeting human transcripts or DNA-sequences - without epithelial pre-enrichment. The failure of CTC detection by the AdnaTest resulted from downregulation of EpCAM, whereas mesenchymal markers like Twist and EGFR were upregulated on CTCs. Such a change in the expression profile during metastatic spread of tumour cells has already been reported and was linked to a biological program termed epithelial-mesenchymal transition (EMT). The use of EpCAM-based enrichment techniques leads to the failure to detect CTC populations that have undergone EMT. Our findings may explain clinical results where low CTC numbers have been reported even in patients with late metastatic cancers. These results are a starting point for the identification of new markers for detection or capture of CTCs, including the mesenchymal-like subpopulations.