Dive C, Workman P, Watson J V
MRC Clinical Oncology Unit, The Medical School, Cambridge, U.K.
Biochem Pharmacol. 1987 Nov 1;36(21):3731-8. doi: 10.1016/0006-2952(87)90027-x.
We present a novel dynamic flow cytoenzymological demonstration of the potent inhibition by the antitumour chloroethylnitrosourea BCNU of the intracellular hydrolysis of fluorescein diacetate by esterases of viable, intact murine and human tumour cells in vitro. The BCNU metabonate chloroethyl isocyanate and the related compound n-butyl isocyanate were also potent inhibitors. I50 values were in the range 4.2 X 10(-5)-2.0 X 10(-4) M. Generally similar quantitative results were obtained for intact cells and sonicates by conventional spectrofluorimetry, and inhibition of purified porcine liver carboxyl esterase (EC 3.1.1.1) was demonstrated. Little or no inhibitory activity was seen with the alkylating agents methyl methane-sulphonate, melphalan and nitrogen mustard. The results are consistent with carbamoylation of the esterase molecules by isocyanates, and this may provide a basis for the flow cytometric determination of intracellular carbamoylation in discrete subpopulations of heterogeneous samples.
我们展示了一种新型的动态流动细胞酶学方法，用于证明体外培养的存活、完整的小鼠和人类肿瘤细胞的酯酶对荧光素二乙酸酯进行细胞内水解时，抗肿瘤药物氯乙基亚硝基脲（BCNU）具有强大的抑制作用。BCNU的代谢产物氯乙基异氰酸酯以及相关化合物正丁基异氰酸酯也是有效的抑制剂。半数抑制浓度（I50）值在4.2×10⁻⁵ - 2.0×10⁻⁴ M范围内。通过传统荧光分光光度法对完整细胞和超声破碎物进行检测，通常可获得类似的定量结果，并且证明了其对纯化的猪肝羧酸酯酶（EC 3.1.1.1）有抑制作用。甲磺酸甲酯、美法仑和氮芥等烷化剂几乎没有或没有抑制活性。这些结果与异氰酸酯使酯酶分子发生氨甲酰化作用相一致，这可能为流式细胞术测定异质样品中离散亚群细胞内的氨甲酰化作用提供基础。
