Two newly synthesized 5-methyltetrahydrofolate-like compounds inhibit methionine synthase activity accompanied by cell cycle arrest in G1/S phase and apoptosis in vitro
Two newly synthesized 5-methyltetrahydrofolate-like compounds inhibit methionine synthase activity accompanied by cell cycle arrest in G1/S phase and apoptosis in vitro
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DOI：
10.1097/cad.0b013e32830317f2
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发表时间：
2008-08-01
影响因子：
2.3
通讯作者：
Cui, Jingrong
中科院分区：
文献类型：
--
作者：
Tang, Cai;Zhang, Zhili;Cui, Jingrong
钴胺素依赖性甲硫氨酸合酶（Cobalamin-dependent methionine synthase，简称GS）是一种以维生素B12为辅助因子的酶，催化5-甲基四氢叶酸（5-methyltetrahydrofolate，简称MTH）与同型半胱氨酸（homocysteine，简称Hcy）反应生成甲硫氨酸和四氢叶酸（methionine and tetrahydrofolate），在叶酸循环、一碳单元转移和含硫氨基酸途径中占据核心地位。“甲基叶酸陷阱”假说表明，蛋氨酸合成酶是一个潜在的抗癌药物开发的目标。ZL 031和ZL 033是新合成的5-甲基四氢叶酸样化合物，作为酶的潜在抑制剂。为了确定这两种化合物对甲硫氨酸合成酶活性的影响，采用分光光度法测定，结果证明ZL 031和ZL 033在HL-60细胞中灭活甲硫氨酸合成酶，IC 50分别为10.0和1.4 μ mol/l。此外，观察到明显的抑制HL-60细胞增殖的作用，导致我们进一步探讨其潜在的抗癌机制。在甲硫氨酸合成酶缺乏和随后叶酸耗竭的情况下，细胞周期被阻滞在G1/S期，并观察到凋亡。细胞周期调控蛋白分析表明，细胞周期蛋白E和细胞周期蛋白依赖性激酶2都增加。此外，还观察到caspase-3、聚（ADP-核糖）聚合酶、caspase-8和caspase-9蛋白水平降低。在我们进行的所有生物学实验中，ZL 033显示出比ZL 031更好的功效。上述结果表明，ZL 031和ZL 033作为一种新型的蛋氨酸合成酶抑制剂，在体外可引起HL-60细胞G1/S期延迟和凋亡，并最终抑制HL-60细胞的增殖。羧酸取代的ZL 033可能比酯取代的ZL 031具有更好的药物开发潜力。
Cobalamin-dependent methionine synthase, with a cofactor of vitamin B12, catalyzes the reaction of 5-methyltetrahydrofolate and homocysteine to form methionine and tetrahydrofolate, which takes a core position in folate cycle, one-carbon-unit transfer, and sulfur amino acid pathways. The 'methyl folate trap' hypothesis suggests that methionine synthase is a potential target for anticancer drug development. ZL031 and ZL033 are 5-methyltetrahydrofolate-like compounds that have been newly synthesized as potential inhibitors of the enzyme. To identify the effect of these two compounds on methionine synthase activity, a spectrophotometric assay was used and the results proved that ZL031 and ZL033 inactivated methionine synthase in HL-60 cells with an IC50 dose of 10.0 and 1.4 mu mol/l, respectively. Moreover, obvious inhibitory effect on proliferation of HL-60 cells was observed, leading to our further investigation of the underlying anticancer mechanism. Under the circumstances of methionine synthase deficiency and subsequent folate depletion, cell cycle was arrested in G1/S phase and apoptosis was also observed. Analysis of cell cycle regulatory proteins demonstrated that cyclin E and cyclin-dependent kinase 2 were both increased. Furthermore, reduction of caspase-3, poly (ADP-ribose) polymerase, caspase-8, and caspase-9 protein levels were observed. In all the biological experiments we have performed, ZL033 has shown a better efficacy compared with ZL031. These results suggest that ZL031 and ZL033, as novel methionine synthase inhibitors, caused G1/S phase delay and apoptosis and eventually inhibit the proliferation of HL-60 cells in vitro. ZL033, with a carboxylic acid substituent, might have a better potential for drug development than ZL031 with an ester substituent.