Objective: To study and establish quality control methods for recombinant anti-cluster of differentiation 19 (CD19) monoclonal antibody (mAb). Methods: Peptide mapping was used for identification of the anti-CD19 mAb. The purity of the mab was controlled by the reduced/non-reduced capillary electrophoresis-sodium dodecyl sulfonate (CE-SDS) and size exclusion-high performance liquidchromatography (SEC-HPLC)methods. Imaging capillary isoelectric focusing electrophoresis (iCIEF) was used to determine the charge heterogeneity. The glycation of anti-CD19 mAb was analyzed by high performance liquid chromatography (HPLC) and the biological activity was measured by antibody-dependent cell-mediated cytotoxicity (ADCC) based on reporter assay. Other indicators should comply with corresponding requirements of Chinese Pharmacopeia 2020 edition and related guidelines. Results: Peptide mapping detection of anti-CD19 mAb had the corresponding characteristic map, which played a good role in the identification. The sum percentage of peak area of light chain and heavy chain of reduced CE-SDS was (98.77 ± 0.05) %, the main peak area percentage of non-reduced CESDS was (96.19 ± 0.05) %, and the fragment percentage was (3.81 ± 0.05) %. The peak area percentage of SECHPLC monomer was (99.42 ± 0.001) % and the percentage of polymer peak area was (0.54 ± 0.001) %. The peak area percentage of the main subtypes analyzed by iCIEF was (66.41 ± 0.38) %, the peak area percentage of acid subtype was (13. 69 ± 0.16)%, and that of alkaline subtype was (19.90 ± 0.46) %. In the analysis of sugar type, the three sugar types with the highest proportion were G0, G0-Gn and M5, respectively. The content of G0 was (64.64 ± 0.61) % , G0-Gn was (9.75 ± 0.42) %, M5 was (6.22 ± 0.35) %. The EC50 value of ADCC activity was (10.09 ± 1.40) ng·mL-1. Conclusions: According to the key quality attributes of anti-CD19 monoclonal antibody, the corresponding quality control methods are studied and established to ensure its safety, effectiveness and quality control , which provides references for the quality control methods and strategies of anti-CD19 mAb products.
Key words
anti-CD19 mAb； identification； purity determination； biological activity
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