The chemopreventive agent α-difluoromethylornithine blocks Ki-ras-dependent tumor formation and specific gene expression in Caco-2 cells
The chemopreventive agent α-difluoromethylornithine blocks Ki-ras-dependent tumor formation and specific gene expression in Caco-2 cells
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DOI：
10.1002/mc.20008
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发表时间：
2004-04-01
影响因子：
4.6
通讯作者：
Gerner, EW
中科院分区：
文献类型：
--
作者：
Ignaternko, NA;Zhang, H;Gerner, EW
Kirsten-ras (Ki-ras) 原癌基因突变在结直肠癌中频繁发生。 α-二氟甲基鸟氨酸 (DFMO) 是多胺生物合成酶鸟氨酸脱羧酶 (ODC) 的不可逆抑制剂，可抑制经致癌物处理的动物模型中的 Ki-ras 转化和结肠肿瘤发生，其机制尚未阐明。转染激活的 Ki-ras 的 Caco-2 细胞（而非亲代细胞）在严重联合免疫缺陷 (SCID) 小鼠中形成肿瘤。 DFMO 治疗（2% 的饮用水）可防止肿瘤生长。进行基因表达谱分析以鉴定依赖于 Ki-ras 和 DFMO 的基因表达模式。微阵列结果通过实时或半定量 RT-PCR 和/或蛋白质印迹分析进行验证。 Caco-2 细胞中表达激活的 Ki-ras 的基因上调，编码细胞骨架、转运、蛋白酶和间隙连接相关蛋白。这些基因对于结肠上皮组织的正常发育和维持很重要。转染激活的 Ki-ras 的 Caco-2 细胞显示整合素 α 1 (INGA1) 表达增加，层粘连蛋白上的细胞迁移增强。这些参数不受 DFMO 影响，但 Ki-ras 依赖性迁移受到 INGA1 抗体的抑制。其他依赖 Ki-ras 但不依赖 DFMO 的基因包括转谷氨酰胺酶 (TGase) 和激肽释放酶 6 (KLK6)。 Ki-ras 转染细胞的连接蛋白 43 (Cx43)（RNA 和蛋白质）、紧密连接蛋白和内皮素 1 的表达水平也有所增加。DFMO 逆转了这些增加。结果表明，Ki-ras 癌基因引起实验细胞迁移和细胞间通讯基因的变化，并且其中一些变化可以通过 DFMO 逆转。 (C) 2004 Wiley-Liss, Inc.
Mutation of the Kirsten-ras (Ki-ras) proto-oncogene occurs frequently in colorectal cancers. alpha-Difluoromethylornithine (DFMO), an irreversible inhibitor of the polyamine biosynthetic enzyme, ornithine decarboxylase (ODC), inhibits Ki-ras transformation and colon tumorigenesis in carcinogen-treated animal models by mechanisms yet to be elucidated. Caco-2 cells transfected with an activated Ki-ras, but not parental cells, formed tumors in severe combined immunodeficient (SCID) mice. DFMO treatment (2% in drinking water) prevented tumor growth. Gene expression profiling was performed to identify Ki-ras- and DFMO-depenclent patterns of gene expression. Microarray results were validated with real-time or semi-quantitative RT-PCR and/or Western blot analysis. Genes upregulated in Caco-2 cells expressing an activated Ki-ras encoded cytoskeletal-, transport-, protease-, and gap junction-associated proteins. These genes are important for normal development and maintenance of colonic epithelial tissue. Caco-2 cells transfected with an activated Ki-ras displayed increased expression of the integrin alpha 1 (INGA1) and enhanced cell migration on laminin. These parameters were unaffected by DFMO, but Ki-ras-dependent migration was inhibited by INGA1 antibodies. Other Ki-ras-dependent, but DFMO-independent, genes included transglutaminase (TGase) and kallikrein 6 (KLK6). Ki-ras-transfected cells also expressed increased levels of connexin43 (Cx43) (RNA and protein), tight junction protein, and endothelin 1. DFMO reversed these increases. The results indicated that the Ki-ras oncogene caused changes in experimental cell migration and cell-cell communication genes and that some of these changes could be reversed by DFMO. (C) 2004 Wiley-Liss, Inc.