Functional analysis of gastric cancer-specific genes identified by SAGE data and its clinical implication

本研究的目的是通过SAGE数据分析筛选新的胃癌特异性基因，建立以分子靶向治疗为导向的肿瘤诊断新体系。结果表明：（1）将胃癌（GSE 545）SAGE文库与14个正常组织SAGE文库进行比较，结合RT-PCR，共扩增出9个胃癌特异性基因（APIN、MIA、MMP-10、DKK 4、REGIV等）。2）免疫组化结果显示，MIA和MMP-10的表达与乳腺癌的恶性程度和预后有关。REGIV不仅在胃癌中表达，而且在结直肠癌、胰腺癌和胃肠道类癌中也有表达; 3）ELISA检测结果显示，胃癌患者血清中REGIV和MMP-10水平分别为36%和95%; 4）功能分析显示，EGFR的磷酸化和caspase-9的抑制参与了5-FU对细胞凋亡的抑制。SCP 18的表达与胃癌的分期和转移有关，通过在胃癌细胞系中强制表达促进细胞侵袭。5）关于肿瘤相关基因的遗传多态性与胃癌的关系，发现新的胃癌候选抑癌基因CLDN 18的5 'UTR中的SNP（G/A）影响胃癌的风险（OR-5.17，95%CI：2.01-13.3）。6）我们开发了定制的寡聚DNA芯片，其中包含207个cDNA，包括通过SAGE分析鉴定的特异性基因、与癌症发生和进展相关的已知基因以及参与DNA损伤反应和修复的基因。通过检测20例胃癌的基因表达谱，我们发现了23个与侵袭、淋巴结转移和肿瘤分期相关的基因。

The purpose of the study is to identify novel gastric cancer-specific genes through SAGE data analysis and to establish new system of cancer diagnosis oriented towards molecular target therapy. Following results were obtained:1) By comparison between SAGE libraries of gastric cancer (GSE545) and 14 normal organs in combination with RT-PCR, 9 gastric cancer-specific genes (APIN, MIA, MMP-10, DKK4, REGIV, etc.) have been identified.2) By immunostaining, expression of MIA and MMP-10 were correlated with grade of malignancy and poor prognosis. REGIV expression was detected not only in gastric cancer but in colorectal cancer, pancreas cancer and GI carcinoid.3) By ELISA, high levels of REGIV and MMP-10 in sera were detected in 36% and 95% of gastric cancer patients, respectively.4) Through functional analyses, phosphorylation of EGFR and inhibition of caspase-9 were found to participate in apoptosis inhibition by 5-FU. SCP18, whose expression is associated with tumor stage and metastasis, promoted cell invasion by forced expression in gastric cancer cell line.5) Regarding the relation between genetic polymorphism of tumor-related genes and gastric cancer, SNP (G/A) in 5'UTR of the CLDN18, new candidate tumor suppressor of gastric cancer, was found to affect gastric cancer risk (OR-5.17, 95%CI: 2.01-13.3).6) We have developed custom-made oligo-DNA microarray with 207 cDNAs including specific genes identified by SAGE analysis, known genes related to development and progression of cancer, and genes participating in DNA damage response and repair. By examining gene expression profiles of 20 gastric cancers, we identified 23 genes related invasion, nodal metastasis and tumor stage.