An efficient process of generating bispecific antibodies via controlled Fab-arm exchange using culture supernatants

Bispecific antibody generation is actively pursued for therapeutic and research antibody development. Although there are multiple strategies for generating bispecific antibodies (bsAbs); the common challenge is to develop a scalable method to prepare bsAbs with high purity and yield. The controlled...

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Veröffentlicht in:Protein expression and purification 2016-05, Vol.121, p.133-140
Hauptverfasser: Paul, Suparna, Connor, Judy, Nesspor, Tom, Haytko, Peter, Boakye, Ken, Chiu, Mark L., Jiang, Haiyan
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Sprache:eng
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Zusammenfassung:Bispecific antibody generation is actively pursued for therapeutic and research antibody development. Although there are multiple strategies for generating bispecific antibodies (bsAbs); the common challenge is to develop a scalable method to prepare bsAbs with high purity and yield. The controlled Fab-arm exchange (cFAE) method combines two parental monoclonal antibodies (mAbs), each with a matched point mutation, F405L and K409R in the respective CH3 domains. The conventional process employs two steps: the purification of two parental mAbs from culture supernatants followed by cFAE. Following a reduction/oxidation reaction, the bispecific mAb is formed with greater than 95% heterodimerization efficiency. In this study, cFAE was initiated in culture supernatants expressing the two parental mAbs, thereby eliminating the need to first purify the parental mAbs. The bsAbs formed in culture supernatant was then purified using a Protein A affinity chromatography. The BsAbs generated in this manner had efficiency comparable to the conventional method using purified parental mAbs. BsAbs prepared by two different routes showed indistinguishable characteristics by SDS capillary electrophoresis, analytical size exclusion, and cation exchange chromatography. This alternative method significantly shortened timelines and reduced resources required for bsAb generation, providing an improved process with potential benefits in large-scale bsAb preparation, as well as for HTP small-scale bsAb matrix selection. •Bispecific antibodies made by using recombinant monoclonal antibody culture supernatants.•Conventional Fab-arm exchange process time can be shortened more than 2 fold.•Procedure maintained high heterodimerization efficiency and protein yield.•No loss in the bispecific antibody characteristics when using this method.
ISSN:1046-5928
1096-0279
DOI:10.1016/j.pep.2016.01.014