DNAメチル化の抑制と各種成長因子による細胞形質転換の抑制、抗癌剤耐性機構の克服

抑制多种生长因子诱导的DNA甲基化和细胞转化，克服抗癌耐药机制

• 批准号: 13770672
• 负责人: 前田 真悟
• 金额: $ 1.34万
• 依托单位: Keio University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Young Scientists (B)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13770672/
• 关键词: DNA methylation; 抗癌剤感受性; antisense oligonulcleotide; 胃癌細胞株; 5-FU; CDDP; methylation; 抗癌剤耐性

近年、DNA methylationと遺伝子の発現制御との関連が注目され、HDRなどの抗癌剤感受性因子との関連も報告されてきた。われわれは、DNA methyltransferaseに対するantisense oligonucleotideを作成し、DNA methylationが及ぼす5-FU及びCDDPへの感受性変化を検討した。胃癌細胞株TMH-1、MKN-45、MKN-74に対し、DNA methyltransferaseのantisense oligonucleotideまたはnonsense oligonucleotideを2.5uMの濃度で前処置した後、抗癌剤5-FU(1.100ug/ml)またはCDDP(0.5.50ug/ml)を接触させ、抗癌剤感受性をMTT assayまたはBrdU uptake incorporation assay(BrdU assay)で判定した。DNA methyltransferaseに対するantisense oligonucleotideの配列は、5-TCTATTTGAGTCTGCCATTT-3とした。nonsene oligonucleotideの配列は、5-TGTGATTCTCCTTATTCGA-3とし、antisense oligonucleotideの塩基配分と同様に作成した。結果として、antisense群はnonsense群と比較して、5-FUに関しては、TMK-1、MKN-45で約20%の感受性の増強がみられ、感受性因子として、TS mRNA、DPD mRNAの発現を定量的PCR法を用いて測定したが、TS mRNAは両群で差がなく、DPD mRNAはantisense群で有意に低下していた。CDDPでもTMK-1、MKN-45で約15%の感受性の増強が認められた。

In recent years, the relationship between DNA methylation and gene expression has been noticed, and the relationship between HDR and anti-cancer agent susceptibility factors has been reported. DNA methylation and sensitivity to 5-FU and CDDP are discussed. Gastric cancer cell lines TMH-1, MKN-45 and MKN-74 were exposed to 5-FU(1.100ug/ml) and CDDP(0.5.50ug/ml) at concentrations of 2.5uM before and after treatment, and the sensitivity of the anticancer agent was determined by MTT assay(BrdU uptake assay). The alignment of antisense oligonucleotides for DNA methyltransferase is 5-TCTATTTGAGTCTGCCATTT-3. 5-TGTGATTCTTCTATCGA-3 and 5-TGATTCTTCTATCGA-3 are the same as 5-TGATTCTTCTATCGA-3 and 5-TGATTCTTCTATCGA-3. The results showed that the sensitivity of 5-FU, TMK-1 and MKN-45 was increased by about 20% compared with that of nonsense group. The sensitivity factors, TS mRNA and DPD mRNA were quantitatively detected by PCR method. The TS mRNA was decreased compared with that of nonsense group. The DPD mRNA was decreased compared with that of antisense group. CDDP was used for TMK-1 and MKN-45. The sensitivity of CDDP was increased by about 15%.