Henderson G B, Strauss B P
Department of Molecular and Experimental Medicine, Scripps Clinic and Research Foundation, La Jolla, CA 92037.
Biochem Pharmacol. 1990 Jun 15;39(12):2019-25. doi: 10.1016/0006-2952(90)90624-t.
A subline (JT-1) of L1210 mouse leukemia cells that contains elevated levels of a high-affinity folate binding protein is sensitive to growth inhibition by homofolate. Inhibition was observed at nanomolar concentrations of folate or 5-formyltetrahydrofolate where the high-affinity binding protein is the predominant uptake route for folate compounds. At 1.0 nM folate, inhibition of growth by 50% occurred at 0.7 nM homofolate, and maximal inhibition exceeded 90% at homofolate concentrations above 10 nM. Homofolate also inhibited the uptake of 1.0 nM [3H]folate by L1210/JT-1 cells in 72-hr cultures, and the extent of uptake inhibition by 1.0 and 20 nM homofolate was comparable to the inhibition of cell growth by the same concentrations of homofolate. At a growth-limiting concentration of 5-formyltetrahydrofolate (0.5 nM), half-maximal inhibition of L1210/JT-1 cell growth occurred at 1.0 nM homofolate. When excess concentrations of folate (5 microM) or 5-formyltetrahydrofolate (0.5 microM) were added to the medium, no growth inhibition was observed for homofolate at concentrations up to 100 microM. Parental cells lacking the folate binding protein did not respond to homofolate either at growth-limiting (0.5 microM) or excess (5.0 microM) levels of folate. Binding measurements showed that homofolate has a high affinity for the folate-binding protein (Ki = 0.03 nM) but interacts poorly with the reduced-folate transport system (Ki = 203 microM). These results indicate that homofolate inhibits the growth of L1210 cells when intracellular folates are acquired via the high-affinity folate binding protein. The basis for this inhibition appears to be competition by homofolate for substrate binding and internalization.
L1210小鼠白血病细胞的一个亚系（JT - 1），其高亲和力叶酸结合蛋白水平升高，对高同型叶酸的生长抑制敏感。在纳摩尔浓度的叶酸或5 - 甲酰四氢叶酸下观察到抑制作用，此时高亲和力结合蛋白是叶酸化合物的主要摄取途径。在1.0 nM叶酸存在下，0.7 nM高同型叶酸导致50%的生长抑制，高同型叶酸浓度高于10 nM时，最大抑制超过90%。高同型叶酸还抑制72小时培养的L1210/JT - 1细胞对1.0 nM [³H]叶酸的摄取，1.0和20 nM高同型叶酸对摄取的抑制程度与相同浓度高同型叶酸对细胞生长的抑制程度相当。在5 - 甲酰四氢叶酸的生长限制浓度（0.5 nM）下，1.0 nM高同型叶酸使L1210/JT - 1细胞生长抑制达到半数最大抑制。当向培养基中添加过量浓度的叶酸（5 μM）或5 - 甲酰四氢叶酸（0.5 μM）时，高达100 μM的高同型叶酸浓度未观察到生长抑制。缺乏叶酸结合蛋白的亲本细胞在叶酸的生长限制水平（0.5 μM）或过量水平（5.0 μM）下对高同型叶酸均无反应。结合测量表明，高同型叶酸对叶酸结合蛋白具有高亲和力（Ki = 0.03 nM），但与还原型叶酸转运系统相互作用较弱（Ki = 203 μM）。这些结果表明，当细胞内叶酸通过高亲和力叶酸结合蛋白获取时，高同型叶酸会抑制L1210细胞的生长。这种抑制的基础似乎是高同型叶酸对底物结合和内化的竞争。
