https://orcid.org/0009-0001-0275-141X
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ABSTRACT
A major driver for the recent investment surge in bispecific antibody (bsAb) platforms and products is the multitude of distinct mechanisms of action that bsAbs offer compared to a combination of two monoclonal antibodies. Four bsAb products were granted first regulatory approvals in the US or EU during 2023 and the biopharmaceutical industry pipeline is brimming with bsAb candidates across a broad range of therapeutic applications. In previously reported bsAb discovery campaigns, following a hypothesis-based choice of two specific target proteins, selections and screening activities have often been performed in mono-specific formats. The conversion to bispecific modalities has usually been positioned toward the end of the discovery process and has involved small numbers of lead molecules, largely due to challenges in expressing, purifying, and analyzing large numbers of bsAbs. In this review, we discuss emerging strategies to facilitate the production of expanded bsAb panels, focusing particularly upon combinatorial methods to generate bsAb matrices. Such technologies will enable screening in. bispecific formats at earlier stages of discovery campaigns, not only widening the accessible protein space to maximize chances of success, but also advancing empirical bi-target validation activities to assess initial target selection hypotheses.
Acknowledgments
The authors would like to thank Joanne McGregor, Tomasz Klaus, Emma Harding, Paul Taylor, Craig Jamieson and Karina Chan for proofreading the manuscript and providing helpful comments and suggestions.
Disclosure statement
J.R.T. and C.H.C. are employees and stock-holders of GSK plc, Stevenage, UK.
Abbreviations
| ADC | = | antibody-drug conjugate |
| ADCC | = | antibody-dependent cellular cytotoxicity |
| bsAb | = | bispecific antibody |
| cAMP | = | cyclic adenosine monophosphate |
| CD20 | = | cluster of differentiation 20 cell surface protein |
| CD3 | = | cluster of differentiation 3 cell surface protein complex |
| CH1(2,3) | = | heavy chain constant region 1(2,3) |
| CL | = | light chain constant region |
| cLC | = | common light chain |
| CMC | = | Chemistry, Manufacturing and Controls |
| CODV-Ig | = | cross-over dual variable domain antibody |
| Da | = | Dalton |
| dAb | = | domain antibody |
| DNA | = | deoxyribonucleic acid |
| DR5 | = | death receptor 5 |
| E. coli | = | Escherichia coli |
| EGFR | = | epidermal growth factor receptor |
| ESI | = | electrospray ionization |
| Fab | = | fragment antigen-binding |
| (c)FAE | = | (controlled) Fab-arm exchange |
| Fc | = | fragment crystallizable |
| FcRn | = | neonatal Fc receptor |
| FDA | = | U.S. Food and Drug Administration |
| FORCE | = | format chain exchange |
| HC | = | antibody heavy chain |
| HEK | = | human embryonic kidney |
| HER1(2,3) | = | human epidermal growth factor receptor 1(2,3) |
| HIC | = | hydrophobic interaction chromatography; |
| HIV | = | human immunodeficiency virus |
| HTP | = | high throughput |
| IEX | = | ion exchange chromatography |
| IgG | = | immunoglobulin G |
| KiH | = | knob-in-hole |
| LC | = | antibody light chain |
| LCMS | = | liquid chromatography mass spectrometry |
| mAb | = | monoclonal antibody |
| MALDI | = | matrix-assisted laser desorption/ionization |
| MET | = | hepatocyte growth factor receptor |
| mmSEC | = | mixed mode size exclusion chromatography |
| MS | = | mass spectrometry |
| NGF | = | nerve growth factor |
| ppm | = | parts per million |
| PTM | = | post-translational modification |
| SARS-CoV-2 | = | Severe acute respiratory syndrome coronavirus 2 |
| scFv | = | single-chain variable fragment |
| SEC | = | size exclusion chromatography |
| SILAC | = | stable isotope labeling using amino acids in cell culture |
| TAA | = | tumor-associated antigen |
| TCE | = | T-cell engager |
| UV | = | ultraviolet |
| VH | = | heavy chain variable region |
| VL | = | light chain variable region |
| VHH | = | camelid heavy chain variable domain antibody |