Validation and Assessment of The Stability Indicating capability of a CEX-HPLC For Charge Variant Analysis And a SEC-HPLC Method For size Heterogeneity Analysis of a Monoclonal Antibody; Sunmab

Abstract

Monoclonal antibodies (mAb) are prone to different kinds of Post Translational Modifications (PTMs) that may lead to different kinds of heterogeneities which result in the formation of mAb variants. These heterogeneities may arise at different stages of the life cycle of mAb. Two of the most common heterogeneities that are encountered are charge and size heterogeneity. On the hand, charge heterogeneity leads to the generation of charge variants in the form of acidic and basic species. The analysis of such variants can be analyzed with the help of Ion Exchange chromatography. On the other hand, size heterogeneity leads to the generation of size variants in the form of HMWs and LMWs species. The analysis of such variants can be easily done with the help of Size Exclusion Chromatography method. In reference to the current prospects, the CEX and SEC methods were developed for Sunmab which is a therapeutic monoclonal antibody developed and marketed by SUN Pharmaceuticals Industries Limited. The methods were validated for different validation parameters according to the ICH guidelines. The methods were also evaluated for their stability indicating capabilities with the help of a forced degradation study based on different stress parameters. Upon validation of the CEX method, the given analytical was found to be precise and accurate. The method was linear over a concentration range of 0.5-375% of the reference standard concentration with a correlation co-efficient of 0.9997. The method was able to quantify (LOQ) the main species up to a concentration of 0.4 mg/ml and detect (LOD) the same up to concentration of 0.08 mg/ml. Similarly, the SEC method was also precise and accurate. The range over which the method was linear was 0.5-125% of the reference standard concentration with a co-efficient of 0.9909. The LOD and LOQ of the method was found to be 0.5 ug/ml and 1.25 ug/ml respectively. Both the methods showed significant prowess in their stability indicating capabilities upon forced degradation of the sample via different stress parameters.