Severe folate restriction results in depletion of and alteration in the composition of the intracellular folate pool, moderate sensitization to methotrexate and trimetrexate, upregulation of endogenous DHFR activity, and overexpression of metallothionein
Severe folate restriction results in depletion of and alteration in the composition of the intracellular folate pool, moderate sensitization to methotrexate and trimetrexate, upregulation of endogenous DHFR activity, and overexpression of metallothionein
复制标题
严重的叶酸限制会导致细胞内叶酸库的消耗和组成的改变、对甲氨蝶呤和三甲曲沙的中度过敏、内源性 DHFR 活性的上调以及金属硫蛋白的过度表达
DOI：
10.1046/j.1359-4117.2002.01049.x
复制
发表时间：
2002
影响因子：
--
通讯作者：
Melera,PeterW
中科院分区：
文献类型：
--
作者：
Zhu,Wei-Yong;Bunni,M;Priest,DG;DiCapua,JL;Dressler,JM;Chen,Z;Melera,PeterW
DC-3F/FA3 细胞 (FA3) 源自抗叶酸敏感 CHL 细胞，通过选择在含有 15 pM [6S]-5CHOFH4 的无叶酸培养基中生长。这些细胞的细胞内叶酸减少了 30 倍，过度表达叶酸受体 α 和金属硫蛋白 II，并且对二氢叶酸还原酶 (DHFR) 靶向抗叶酸药物甲氨蝶呤 (MTX) 和三甲曲沙 (TMTX) 的敏感性增加，这主要归因于叶酸池状态。在 15 nM [6S]-5CHOFH4 中生长补充叶酸后，它们对两种药物的耐药性分别增加了 5 倍和 10 倍，尽管叶酸库仅恢复了 43%。在纳摩尔叶酸中培养的转染子中强制过度表达 FRα 并不会赋予对 MTX 的抗性，但确实支持对 TMTX 的抗性适度增加 2 倍。 MTII 的强制过度表达具有类似的效果，但当两者一起过度表达时，没有发现抗性增加超过各自单独赋予的抗性，这表明两者通过共同的下游机制赋予 TMTX 抗性。对三个独立的低叶酸选择克隆 FA3、FA7 和 FA14 的分析表明，每个克隆的 DHFR 活性增加了 5 至 6 倍，同时 DHFR 蛋白水平也有类似的增加。然而，在 DHFR 基因拷贝数或 DHFR mRNA 的稳态量方面没有检测到差异，这表明转录后机制是导致 DHFR 表达增加的原因。
DC‐3F/FA3 cells (FA3) were derived from antifolate‐sensitive CHL cells by selection for growth in folate‐free media containing 15 pM [6S]‐5CHOFH4. These cells undergo a 30‐fold decrease in intracellular folates, overexpress folate receptor alpha and metallothionein II, and display increased sensitivity to the dihydrofolate reductase (DHFR) targeted antifolates methotrexate (MTX) and trimetrexate (TMTX), which can be attributed primarily to the folate pool status. Upon folate repletion by growth in 15 nM [6S]‐5CHOFH4, they display a 5‐ and 10‐fold increase in resistance to both drugs, respectively, even though folate pools are restored by only 43%. Enforced overexpression of FRα in transfectants cultured in nanomolar folate did not confer resistance to MTX but did support a modest 2‐fold increase in resistance to TMTX. Enforced overexpression of MTII had a similar effect, but when both were overexpressed together no increase in resistance beyond that conferred by each one separately was noted, suggesting that both confer resistance to TMTX through a common downstream mechanism. Analysis of three independent low folate selected clones, FA3, FA7, and FA14, showed that each had a 5‐ to 6‐fold increase in DHFR activity accompanied by a similar increase in DHFR protein level. However, no differences were detected in the DHFR gene copy number or in the steady‐state amount of DHFR mRNA, suggesting that a posttranscriptional mechanism was responsible for the increase in DHFR expression.