Inaba M, Mitsuhashi J, Sawada H, Miike N, Naoe Y, Daimon A, Koizumi K, Tsujimoto H, Fukushima M
Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo, Japan.
Jpn J Cancer Res. 1996 Feb;87(2):212-20. doi: 10.1111/j.1349-7006.1996.tb03161.x.
The mechanism of resistance to 5-fluorourcil (5-FU) was studied with NUGC-3/5FU/L, a human stomach cancer cell line which had acquired resistance as a consequence of repeated 5-day exposures to stepwise-increasing concentrations of 5-FU in vitro. NUGC-3/5FU/L was 200-fold and over 16-fold resistant to 96-h and 1-h exposures to 5-FU, respectively. NUGC-3/5FU/L incorporated less 5-FU into RNA, indicating resistance to the RNA-directed action of 5-FU. On the other hand, NUGC-3/5/5FU/L also showed resistance to in situ thymidylate synthase (TS) inhibition by 5-FU. Polymerase chain reaction-single-strand conformation polymorphism analysis of TS cDNA and a FdUMP ligand binding assay showed that quantitative and qualitative alterations of TS are not responsible for this resistance. In contrast, the ability to metabolize 5-FU to its active metabolites, FUTP and FdUMP, was reduced in NUGC-3/5FU/L. We found that not only the activities of uridine phosphorylase/kinase and orotate phosphoribosyl-transferase (OPRT), but also the level of phosphoribosyl pyrophosphate, a cosubstrate for OPRT, were significantly lower in NUGC-3/5FU/L than in the parent NUGC-3. These results indicated that resistance to 5-FU in NUGC-3/5FU/L is due to reduced activities of 5-FU-anabolizing enzymes, but not to an alteration of TS. 2'-Deoxyinosine effectively enhanced TS inhibition by 5-FU in the resistant cells, thus markedly sensitizing them to 5-FU.
利用NUGC-3/5FU/L研究了对5-氟尿嘧啶（5-FU）的耐药机制。NUGC-3/5FU/L是一种人胃癌细胞系，通过在体外将其反复暴露于逐步增加浓度的5-FU中5天而获得耐药性。NUGC-3/5FU/L对96小时和1小时暴露于5-FU的耐药性分别为200倍和超过16倍。NUGC-3/5FU/L将较少的5-FU掺入RNA，表明对5-FU的RNA定向作用具有耐药性。另一方面，NUGC-3/5/5FU/L对5-FU原位抑制胸苷酸合成酶（TS）也表现出耐药性。TS cDNA的聚合酶链反应-单链构象多态性分析和FdUMP配体结合试验表明，TS的定量和定性改变与这种耐药性无关。相反，NUGC-3/5FU/L中5-FU代谢为其活性代谢产物FUTP和FdUMP的能力降低。我们发现，不仅尿苷磷酸化酶/激酶和乳清酸磷酸核糖基转移酶（OPRT）的活性，而且OPRT的共底物磷酸核糖焦磷酸的水平在NUGC-3/5FU/L中均显著低于亲本NUGC-3。这些结果表明，NUGC-3/5FU/L对5-FU的耐药性是由于5-FU合成酶活性降低，而不是由于TS的改变。2'-脱氧肌苷有效地增强了耐药细胞中5-FU对TS的抑制作用，从而显著提高了它们对5-FU的敏感性。
