CDR grafting, or antibody reshaping, is the most clinically validated route to a successful therapeutic humanised monoclonal antibody, and is the process described in this chapter. Accurate determination of the rodent antibody variable region DNA sequences, amplified by RT-PCR, and homology modelling of their protein translation are key starting points. Human framework selection and the design of the DNA and protein sequences of the CDR-grafted humanised antibody, based on a thorough analysis of the rodent variable regions and comparison with existing human antibody sequences, are discussed. The central concept of the design and generation of a set of framework mutations, expressed as full-length human IgG-kappa in transiently transfected mammalian cells, for determining relative binding potency is described. This naturally leads to the objective of identifying the minimal set of framework mutations necessary to recapitulate the binding potency of the original rodent.
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Acknowledgements
The authors wish to acknowledge the valuable contributions of all previous workers in the Therapeutic Antibody Group (and its earlier incarnations, including AERES Biomedical Ltd.) at MRC Technology in developing and testing the methods outlined in this chapter, especially MRC Technology scientists (past and present) Mary Bendig PhD, Alex Brown MSc, Jon Chappel PhD, Margaret Cronin BSc, Dolores Crowley BSc, Eilish Cullen BSc, Vicky Heath BSc, Simon Keen BSc, Katy Kettleborough PhD, Olivier Léger PhD Alison Levy BSc, Charlotte Morrison BSc, Grant Munroe PhD, Siobhan O’Brien PhD, Sue Potts BSc, José Saldanha PhD, Alicia Sedo, BSc and Linda Smith BSc, as well as visiting scientists Dr. Hiroshi Maeda of Kaketsuken, Dr. Frank Kolbinger of Novartis, and Dr. Masa Tsuchiya and Dr. Koh Sato of Chugai Pharmaceuticals.4 to 5
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Williams, D.G., Matthews, D.J., Jones, T. (2010). Humanising Antibodies by CDR Grafting. In: Kontermann, R., Dübel, S. (eds) Antibody Engineering. Springer Protocols Handbooks. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-01144-3_21
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