Establishment,Optimization And Validation Of Ipilimumab Assay In Vitro

Ipilimumab is the first-in-class and sole-approved drug as anti-CTLA-4 monoclonal antibody in the world.As a immune checkpoint inhibitor,Ipilimumab shows promising results in clinical trials and has been approved as a single drug or combined drug for the treatment of multiple solid tumors.In this study,a variety of methods wereestablished to detect the activity of Ipilimumab in vitro:the method to detect the binding activity of Ipilimumab and rhCTLA-4 by ELISA;to detect the inhibitory activity of Ipilimumab on B7-1 binding to CTLA-4 by FACS;as well as to measure the concentration of IL-2 in PBMC,which could indicate the activity of Ipilimumab.We verified the ELISA methods for detecting the binding activity of Ipilimumab to rhCTLA-4 and the inhibitory activity of Ipilimumab on B7-1/B7-2 binding to CTLA-4.The results showed that the specificity and durability of the two methods were good,and the recovery rates of multiple activity tests were in the range of 90-104%when the relative binding activity of Ipilimumab was 50%-200%,the repeatability and intermediate precision of the precision were less than 15.0%,and the linear correlation coefficients were more than 0.98.The results indicated that the two analytical methods can reflect the activity of Ipilimumab in a reliable,accurate and stable way.This study mainly established the activity methods of Ipilimumab in vitro,which provided the basis and reference for the development of Ipilimumab biosimilar drugs at home and abroad,as well as the research on CTLA-4 as the target of innovative antibody or bis-specific antibody and other products.