人脐静脉内皮细胞增殖抑制法检测贝伐珠单抗活性的优化和应用

Optimization and application of HUVEC proliferation inhibitory assay

目的：优化人脐静脉内皮细胞（human umbilical vein endothelial cells,HUVEC）增殖抑制法,并用优化后方法比较6种贝伐珠单抗（bevacizumab）生物类似药（similar bio-therapeutic products,SBPs）候选药物与原研药的生物学活性。方法：对细胞接种量、VEGF165浓度和样品稀释浓度进行优化,提高方法精密性和准确性。使用优化后的方法,以贝伐珠单抗为参比品,测定6种贝伐珠SBPs的生物学活性,并与原研药进行比较。结果：优化后的HUVEC增殖抑制法,信噪比为1.5~1.8,板间变异度〈20%,在70%~130%活性范围内有良好的准确性。使用优化后方法检测,质控样品的相对活性均值为102.00%,6种贝伐珠SBPs的相对活性分别为95.12%,88.81%,96.96%,103.00%,105.70%和102.40%。结论：通过优化,HUVEC增殖抑制法的精密性和准确性较经典方法有显著提高,适用于贝伐珠单抗及其SBPs的可比性评价。

Objective： To optimize human umbilical vein endothelial cell（ HUVEC） proliferation inhibitory assay,and compare the biological potency of 6 bevacizumab similar bio-therapeutic products（ SBPs） with bevacizumab. Methods： Cell inoculum density,VEGF165 concentration and sample concentration were optimized to improve repeatability and accuracy of the method. Using optimized method,relative potencies of 6 bevacizumab SBPs were determined and compared with original drug. Results： After optimization,the signal / noise ratio was 1. 5 -1. 8,and variation 20% within plates. The optimized assay had a good repeatability and accuracy within the potency range of 70% - 130%. Using the optimized assay,the relative potency of product control was 102. 00%.Relative potencies of 6 bevacizumab SBPs were determined as 95. 12%,88. 81%,96. 96%,103. 00%,105. 70%and 102. 40%. Conclusion： By optimization,the precision and accuracy of HUVEC proliferation inhibition assay are significantly elevated,which meets the requirements of potency comparison studies on bevacizumab and its SBPs.