Ohnami S, Matsumoto N, Nakano M, Aoki K, Nagasaki K, Sugimura T, Terada M, Yoshida T
Genetics Division, National Cancer Center Research Institute, Tokyo, Japan.
Cancer Res. 1999 Nov 1;59(21):5565-71.
K-ras point mutation occurs in >80% of pancreatic cancer. We reported previously that the transduction of an antisense K-ras RNA expression vector suppressed the growth of pancreatic cancer cells with K-ras point mutations in vitro and in vivo. The RNA differential display method (DD) was used to compare the mRNA expression profile of the pancreatic cancer cell line AsPC-1 and that of the antisense K-ras-transduced, growth-retarded AsPC-1 cells. cDNA fragments were isolated from 20 bands on the DD gel, and their differential expression between the two cell lines was confirmed. A sequence analysis revealed that all of the 11 clones up-regulated in the antisense-transduced cells were mitochondrial genes. The other nine cDNA clones that were down-regulated in the antisense-transduced AsPC-1 cells included an oncogene PTI-1 (prostate tumor inducing gene-1), matrix metalloproteinase (MMP)-7, the beta3 chain of laminin-5, lysosome-associated membrane protein-2, the H chain of apoferritin, ribosomal protein S6, proteasome subunit XAPC7, and two cDNA fragments with no homology to the GenBank database. In addition to the AsPC-1 cells, reverse transcription-PCR analysis on surgical specimens of pancreatic cancer revealed that the PTI-1 and MMP-7 genes were overexpressed in three and four cases, respectively, of five cases examined. This method offers a unique opportunity to identify a set of genes that may be modulated by K-ras activation, at least in a subset of the pancreatic cancer. The information on such genes may facilitate our understanding of the spectrum of the functional genetic changes in pancreatic cancer.
K-ras点突变发生在超过80%的胰腺癌中。我们之前报道过，反义K-ras RNA表达载体的转导在体外和体内均抑制了具有K-ras点突变的胰腺癌细胞的生长。采用RNA差异显示法（DD）比较胰腺癌细胞系AsPC-1与反义K-ras转导、生长受抑制的AsPC-1细胞的mRNA表达谱。从DD凝胶上的20条带中分离出cDNA片段，并证实了它们在两种细胞系之间的差异表达。序列分析显示，在反义转导细胞中上调的11个克隆均为线粒体基因。在反义转导的AsPC-1细胞中下调的其他9个cDNA克隆包括一个癌基因PTI-1（前列腺肿瘤诱导基因-1）、基质金属蛋白酶（MMP）-7、层粘连蛋白-5的β3链、溶酶体相关膜蛋白-2、脱铁铁蛋白的H链、核糖体蛋白S6、蛋白酶体亚基XAPC7以及两个与GenBank数据库无同源性的cDNA片段。除AsPC-1细胞外，对胰腺癌手术标本进行的逆转录PCR分析显示，在检测的5例病例中，PTI-1和MMP-7基因分别在3例和4例中过度表达。该方法为鉴定一组可能受K-ras激活调节的基因提供了独特的机会，至少在一部分胰腺癌中是这样。关于这些基因的信息可能有助于我们了解胰腺癌中功能性基因变化的范围。
