Search for oral cancer specific genes and molecular cloning using in-house cDNA microaray

以口腔鳞状细胞癌(SCC)和涎腺肿瘤(SGT)手术标本为材料，采用寡核苷酸捕获的方法，构建了口腔鳞状细胞癌(SCC)和涎腺肿瘤(SGT)的cDNA文库。测定了克隆的5‘端核苷酸序列。排除重叠克隆，选择2201个独立克隆用于微阵列生产。与公开的核苷酸序列数据库相比，61%的克隆是全长的。利用该微阵列系统分析了鳞状细胞癌(SCC)和腺样囊性癌(ACC)中的mRNAs。微阵列检测到许多基因的表达上调，这些基因在人肿瘤组织中的表达是正常组织的两倍或两倍以上。通过基因芯片分析，我们可以获得全长的全长基因，并能够直接和即时地利用全长克隆来检测基因的功能。在鳞癌中表达差异上调/下调的代表全长克隆有白细胞介素1β、Centaurinγ2、腺苷…而在SGT中差异表达的基因有补体衰变加速因子(CD55)、可诱导信号通路蛋白2、胰岛素样生长因子结合蛋白3、Fos样抗原1、人波形蛋白基因、免疫球蛋白重链蛋白、T细胞受体β链、脱氢酶/还原酶成员2、18S rRNA基因和ET等。埃迪松-启动子系统功能分析和临床标本免疫组织化学染色，揭示了几个重要的生物标志物。Clorf 10基因是一种肿瘤抑制基因，它可以阻断细胞周期，Rab1a基因是预测肿瘤发生的候选基因，maspin基因是ACC特异的分子标志物，stathmin基因是恶性肿瘤的特异性分子标志物。较少

cDNA library was constructed from surgical specimens of oral squamous cell carcinoma(SCC) and salivary gland tumors(SGT) using an oligo-eapping method. The 5 '-end nucleotide sequences of the cDNA clones were determined. Overlapping clones were excluded, and 2,201 independent clones were selected and used for microarray production. Compared to the public nucleotide sequence database, 61% of our cDNA clones were full length. mRNAs obtained from squamous cell carcinomas(SCC) and adenoid cystic carcinomas(ACC) were analyzed using this microarray system. Microarray detected up-regulated expression of many genes, which expressed twice or more in tumors man in normal tissue. We could pick up the full length whole gene, which was focused on by microarray analysis, and were directly and immediately able to examine the gene function using the full length clone.Representative full length clones with differential up-/down-regulated expression in SCC were Interleukin 1 β, Centaurin gamma 2, Adenyl … More yl cyclase-associated protein, Rh type C glycoprotein, Spermine syntase, Copine 1, Cytokeratin 13, Paired-like homeodomain transcription factor 1, Hypothetical protein FLJ20150, Small proline-rich protein, Cartilage link protein, G-protein alpha 15, Interleukin receptor antagonist, Transglutamin and so on, whereas the genes differentially expressed in SGTs were decay accelerating factor for complement (CD55), Inducible signaling pathway protein 2, Insulin-like growth factor binding protein 3, Fos-like antigen 1,Human vimentin gene, Immunogloblin heavy constant mu, T cell receptor beta chain, Dehydrogenase/reductase member 2,18S rRNA gene and et. The fgene function -nalysis by edycsone-promoter system and immunohistochemical staining of clinical samples, revealed several important bio-markers. Clorf 10 gene is a tumor suppressor, which stops cell cycle, Rab la is a candidate for a predictor of carcinogenesis, maspin is a molecular marker specific to ACC, and stathmin is a molecular marker specific to malignancy. Less