A MONOCLONAL ANTIBODY-BETA-GLUCURONIDASE CONJUGATE AS ACTIVATOR OF THE PRODRUG EPIRUBICIN-GLUCURONIDE FOR SPECIFIC TREATMENT OF CANCER
A MONOCLONAL ANTIBODY-BETA-GLUCURONIDASE CONJUGATE AS ACTIVATOR OF THE PRODRUG EPIRUBICIN-GLUCURONIDE FOR SPECIFIC TREATMENT OF CANCER
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DOI：
10.1038/bjc.1992.298
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发表时间：
1992-09-01
影响因子：
8.8
通讯作者：
PINEDO, HM
中科院分区：
文献类型：
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作者：
HAISMA, HJ;BOVEN, E;PINEDO, HM
用抗PAN癌单抗(MAb)323/A3与大肠杆菌衍生的β-葡萄糖醛酸酶(GUS)偶联，研究了前药表阿霉素-葡萄糖醛酸苷(Epi-Glu)对肿瘤部位的选择性激活作用。用硅胶C18柱反相色谱从表阿霉素(Epi)治疗患者的尿液中分离EPI-Glu。EPI-Glu在人血中是稳定的，不能被A2780、MCF-7或OVCAR-3癌细胞转化为EPI，尽管细胞内存在GUS。该前药在BALB/c小鼠体内的稳定性得到证实。单抗323/A3与GUS通过稳定的硫醚键相连。结合物(1：1)经离子交换和凝胶过滤层析纯化。与靶细胞的结合显示至少60%的免疫反应性和良好的酶活性保留。蛋白染料(磺胺蓝B)法检测细胞毒作用。当癌细胞暴露于药物作用4或24小时时，EPI(IC50值为0.003-0.2mU-M)的毒性是EPI-Glu(IC50值为20-mU-M)的100-1,000倍。Epi-Glu的低细胞毒性很可能是由于前体药物的细胞摄取率(2.7pmol10(-6)细胞min-1)低于母体化合物(25pmol10(-6)细胞min-1)所致。在前药暴露前用323/A3-GUS结合物对抗原阳性细胞进行预处理，完全恢复了由于Epi-Glu水解为Epi而产生的细胞毒性。我们的结果表明，323/A3-GUS结合物可以在肿瘤细胞水平上特异性地激活稳定的无毒前药Epi-Glu，
The anti-pan carcinoma monoclonal antibody (MAb) 323/A3, linked to E. coli-derived beta-glucuronidase (GUS) was used to study the tumour-site-selective activation of the prodrug Epirubicin-glucuronide (Epi-glu). Epi-glu was isolated from the urine of patients treated with Epirubicin (Epi) by reversed phase chromatography on a silica-C18 column. Epi-glu was stable in human blood and was not converted into Epi by A2780, MCF-7, or OVCAR-3 cancer cells, despite the presence of intracellular GUS. The stability of the prodrug was confirmed in BALB/c mice. MAb 323/A3 and GUS were linked through a stable thioether bond. The conjugate (1:1) was purified by ion exchange and gel filtration chromatography. Binding to target cells revealed an immunoreactivity of at least 60% and good retention of enzyme activity. A protein dye (sulforhodamine B) assay was used to analyse cytotoxicity. Epi (IC50 of 0.003-0.2-mu-M) was 100-1,000 times more toxic than Epi-glu (IC50 of > 20-mu-M), when cancer cells were exposed for 4 or 24 h to the drugs. The low cytotoxicity of Epi-glu was most likely due to the reduced cellular uptake rate of the prodrug (2.7 pmol 10(-6) cells min-1) as compared to that of the parent compound (25 pmol 10(-6) cells min-1). Pretreatment of antigen-positive cells with the 323/A3-GUS conjugate prior to prodrug exposure completely restored cytotoxicity as a result from hydrolysis of Epi-glu into Epi. Our results demonstrate that the 323/A3-GUS conjugate can specifically activate the stable non-toxic prodrug Epi-glu at the tumour cell level,