Development of novel mass-screening methods by three-dimension microarray for endometrial cancer

在本研究中，我们的最终目标是将三维微阵列技术应用于子宫内膜癌（EC）的筛查。首先，我们使用22K DNA芯片（Agilent）进行激光捕获显微解剖，分析了子宫内膜（正常）组织和EC（1级和3级）组织中切除的腺细胞中改变的基因表达。随后，我们利用实时荧光定量PCR技术对DNA芯片的有效性进行了评估，结果表明这27个基因可能成为EC的肿瘤标志物。我们从正常和EC（1级和3级）组织的10个样本中选择27个基因的mRNA水平进行聚类分析。聚类分析将30个样本分为正常组织和EC组织两组。因此，我们制作了定制芯片（EM3D）固定DNA探针，用于筛选筛选出的27个基因和已知的10个在EC中重要的基因。我们利用EM3D分析了EC组织和正常组织，8个样本和4个样本的基因表达。EM3D将样品明显分为EC组织和正常组织两组。此外，为了探索EM3D在筛选中的应用可能性，我们利用EM3D对3例正常细胞学标本和6例EC标本的总RNA进行了阵列和聚类分析。EC和正常标本分为两组。现在，我们已经使用EM3D分析了组织和细胞学标本的37个基因表达，以增加检查的数量。另一方面，我们发现在本分析中选择的蛋白质如midkine可能成为癌症的肿瘤标志物。未来通过分析EM3D数据与临床病理因素之间的关系，M3D可能成为显示EC恶性诊断、药物敏感性预测、复发转移预测等重要临床信息的方法。

In this study, our final goal is application of three-dimensional Microarray system which is new microarray technology to endometrial cancer (EC) screening. First, we analyzed the altered genes expression in the glandular cell cut out of endometrium (normal) tissue and EC (grade 1 and 3) tissue with laser-capture microdissection using 22K DNA microarray (Agilent). Following that, we evaluated the validity of DNA microarray using real-time PCR, and the results showed that the 27 genes may become the tumor marker in EC. We performed a cluster analysis using mRNA level of selected-27 genes from ten each samples in normal and EC (grade 1 and 3) tissue. The cluster analysis classified total 30 samples into two different groups as normal tissue or EC tissue. Therefore, we produced custom chip (EM3D) fixed DNA probe of selected-27 genes and well-known 10 genes which is important in EC for EC screening. We analyzed the gene expression of EC tissue and normal tissue, 8 samples and 4 samples, respectively, using EM3D. EM3D classified sample distinctly into two groups as EC tissue or normal tissue. Also, to explore application possibility of EM3D for screening, we performed array and cluster analysis using EM3D in total RNA from cytologic 3 normal specimens and 6 EC specimens. EC and normal specimen were classified into two different groups. Now, we have analyzed 37 gene expressions of tissue and cytologic specimen using EM3D to increase the number of the examination. On the other hand, we found that protein such as midkine selected in this analysis may become a tumor marker in cancer. In future by analyzing relationship between data of EM3D and the clinicopathologic factor, M3D may become the method to show important clinical information such as a malignancy diagnosis, a medicine sensitivity prediction, and recurrence and metastasis prediction in EC.