两种重组抗CD20人源化单克隆抗体定量分析方法的比较及其在药代动力学研究中的应用

摘要

An enzyme linked immunosorbent assay ( ELISA ) and a flow cytometry assay ( FCA ) based on Wil2-S cells were developed and systematically compared for quantification of recombinant anti-CD20 humanized monoclonal antibody ( rh-anti-CD20zumab) in biological matrix. The specificity, precision and accuracy of each method at correspondingly different linear range showed good results. For ELISA, the precisions of intra-day and inter-day were both <19 . 5%, the relative error was from-18 . 2% to 17 . 6%;For FCA, the precisions of intra-day and inter-day were both <19. 0%, the relative error was from -18. 9% to 18. 4%. The sensitivity of ELISA was significantly higher than that of FCA. The quantitative ranges of ELISA and FCA methods were 0. 04-5. 0 mg/L and 3. 1-200 mg/L, respectively. The concentrations in serum samples and pharmacokinetics analysis were determined by both of two methods after vein drip administration of rh-anti-CD20zumab in rhesus monkeys. Pharmacokinetics data showed that there was excellent consistency between results obtained by two methods at the given dose. We believe that the novel FCA with high speed and high sensitivity can be used to perform PK and PD study of cell surface antigen-targeted antibody derivatives.%采用酶联免疫吸附分析( ELISA)与基于Wil2-S细胞的流式细胞术( FCA)两种方法对生物基质中的重组抗CD20人源化单克隆抗体(rh-anti-CD20zumab)进行定量分析,并对两种方法进行系统比较。方法学验证结果表明,两种方法均具有良好的特异性、精密度和准确度,但定量范围存在明显差异。 ELISA法批内和批间精密度均小于19.5%,准确度为-18.2%~17.6%；FCA法批内和批间精密度均小于19.0%,准确度为-18.9%~18.4%。二者定量范围分别为0.04~5.0 mg/L和3.1~200 mg/L,ELISA法的灵敏度显著高于FCA法。利用两种方法测定猕猴静脉滴注 rh-anti-CD20zumab后的血药浓度-时间变化并进行药代动力学( PK)分析。结果表明,在给定剂量下,通过两种方法获得的PK结果具有良好的一致性。 FCA法是一种细胞表面抗原靶向性抗体类药物PK研究及药效动力学( PD)研究的快捷、理想的方法。