A comparative study of the developability of full-length antibodies, fragments, and bispecific formats reveals higher stability risks for engineered constructs
Itzel Condado-Morales,
Fabian Dingfelder,
Isabel Waibel,
Oliver M. Turnbull,
Bhargav Patel,
Zheng Cao,
Jais Rose Bjelke,
Susanne Nedergaard Grell,
Anja Bennet,
Alissa M. Hummer,
Matthew I. J. Raybould,
Charlotte M. Deane,
Thomas Egebjerg,
Nikolai Lorenzen,
Paolo Arosio
Affiliations
Itzel Condado-Morales
Department of Biophysics and Injectable Formulation, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Fabian Dingfelder
Department of Biophysics and Injectable Formulation, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Isabel Waibel
Department of Chemistry and Applied Biosciences, Institute for Chemical and Bioengineering, Swiss Federal Institute of Technology, Zurich, Switzerland
Oliver M. Turnbull
Department of Statistics, University of Oxford, Oxford, UK
Bhargav Patel
Department of Chemistry and Applied Biosciences, Institute for Chemical and Bioengineering, Swiss Federal Institute of Technology, Zurich, Switzerland
Zheng Cao
Department of Bioanalysis, Beijing Novo Nordisk Pharmaceutical Science & Technology Co. Ltd (Novo Nordisk R&D China), Beijing, China
Jais Rose Bjelke
Department of Purification Technologies, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Susanne Nedergaard Grell
Department of Kidney Biology, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Anja Bennet
Department of Kidney Biology, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Alissa M. Hummer
Department of Statistics, University of Oxford, Oxford, UK
Matthew I. J. Raybould
Department of Statistics, University of Oxford, Oxford, UK
Charlotte M. Deane
Department of Statistics, University of Oxford, Oxford, UK
Thomas Egebjerg
Department of Mammalian Expression, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Nikolai Lorenzen
Department of Biophysics and Injectable Formulation, Global Research Technologies, Novo Nordisk A/S, Måløv, Denmark
Paolo Arosio
Department of Chemistry and Applied Biosciences, Institute for Chemical and Bioengineering, Swiss Federal Institute of Technology, Zurich, Switzerland
Engineered antibody formats, such as antibody fragments and bispecifics, have the potential to offer improved therapeutic efficacy compared to traditional full-length monoclonal antibodies (mAbs). However, the translation of these non-natural molecules into successful therapeutics can be hampered by developability challenges. Here, we systematically analyzed 64 different antibody constructs targeting Tumor Necrosis Factor (TNF) which cover 8 distinct molecular format families, encompassing full-length antibodies, various types of single chain variable fragments, and bispecifics. We measured 15 biophysical properties related to activity, manufacturing, and stability, scoring variants with a flag-based risk approach and a recent in silico developability profiler. Our comparative assessment revealed that overall developability is higher for the natural full-length antibody format. Bispecific antibodies, antibodies with scFv fragments at the C-terminus of the light chain, and single-chain Fv antibody fragments (scFvs) have intermediate developability properties, while more complicated formats, such as scFv- scFv, bispecific mAbs with one Fab exchanged with a scFv, and diabody formats are collectively more challenging. In particular, our study highlights the propensity for fragmentation and aggregation, both in bulk and at interfaces, for many current engineered formats.
