Different fermentation processes produced variants of an anti-CD52 monoclonal antibody that have divergent in vitro and in vivo characteristics
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The anti-CD52 antibody has already been approved for the treatment of patients with resistant chronic lymphocytic leukemia, relapsing-remitting multiple sclerosis, and has demonstrable efficacy against stem cell transplantation rejection. A CHO cell line expressing a humanized anti-CD52 monoclonal antibody (mAb-TH) was cultivated in both fed-batch and perfusion modes, and then purified. The critical quality attributes of these mAb variants were characterized and the pharmacokinetics (PK) properties were investigated. Results showed that the perfusion culture achieved higher productivity, whereas the fed-batch culture produced more aggregates and acid components. Additionally, the perfusion culture produced similar fucose, more galactose and a higher proportion of sialic acid on the anti-CD52 mAb compared to the fed-batch culture. Furthermore, the perfusion process produced anti-CD52 mAb had higher complement-dependent cytotoxicity (CDC) efficacy than that produced by the fed-batch culture, a result probably linked to its higher galactose content. However, antibody produced by fed-batch and perfusion cultures showed similar PK profiles in vivo. In conclusion, perfusion is a more efficient method than fed-batch process in the production of functional anti-CD52 monoclonal antibody. Product quality variants of anti-CD52 mAb were found in different cell culture processes, which demonstrated different physiochemical and biological activities, but comparable PK properties. Whether these observations apply to all mAbs await further investigation.
KeywordsAntibody Process Quality Heterogeneity Pharmacokinetics
Compliance with ethical standards
A nonclinical PK comparability study was carried out in 16 naïve cynomolgus monkeys (aged 3–5 years, weight 2–5 kg, 50% male and 50% female) provided by Hainan Jingang Laboratory Animal Co. Ltd. (Hainan, China). Cynomolgus monkeys were chosen because anti-CD52 antibody only cross-reacts with baboons and macaques, wherein only some cynomolgus monkeys have CD52 negative erythrocytes. The in vivo procedures were conducted at the National Shanghai Center for New Drug Safety Evaluation Research Center (Shanghai, China) and were approved by the Institutional Animal Care and Use Committee (IACUC) of the National Shanghai Center for New Drug Safety Evaluation Research Center (Permit Number: IACUC-2015-M-012).
The housing and experimental procedures were carried out strictly according to the United States National Research Council’s Guide for the Care and Use of Laboratory Animals. Animals were maintained in separate cages of 1000 × 750 × 930 mm3 size during daytime and paired housing was applied overnight. The temperature was maintained at 16–24 °C, the relative humidity was 40∼70%, air changes occurred at least eight times per hour, and the illumination time was 7 am to 7 pm. The animals were fed monkey chow (Ke Ao Xie Li, Beijing, China) and fresh fruits and vegetables. The cages were composed of stainless steel with a lookout, perches, and a stainless steel mirror. To alleviate animal suffering, procedures were performed under sedation with anesthesia by intramuscular (I.M.) injection of ketamine HCl (Hengrui Medicine, Jiangsu, China), animals were trained prior to the venous sampling with positive reinforcement, and fruits were offered after sampling. No animal that participated in this study was sacrificed.
Conflict of interest statement
The authors declare that they have no conflict of interest.
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