PMID- 32869839
OWN - NLM
STAT- MEDLINE
DCOM- 20210223
LR  - 20210223
IS  - 1470-8728 (Electronic)
IS  - 0264-6021 (Print)
IS  - 0264-6021 (Linking)
VI  - 477
IP  - 18
DP  - 2020 Sep 30
TI  - Monoclonal antibody stability can be usefully monitored using the 
      excitation-energy-dependent fluorescence edge-shift.
PG  - 3599-3612
LID - 10.1042/BCJ20200580 [doi]
AB  - Among the major challenges in the development of biopharmaceuticals are 
      structural heterogeneity and aggregation. The development of a successful 
      therapeutic monoclonal antibody (mAb) requires both a highly active and also 
      stable molecule. Whilst a range of experimental (biophysical) approaches exist to 
      track changes in stability of proteins, routine prediction of stability remains 
      challenging. The fluorescence red edge excitation shift (REES) phenomenon is 
      sensitive to a range of changes in protein structure. Based on recent work, we 
      have found that quantifying the REES effect is extremely sensitive to changes in 
      protein conformational state and dynamics. Given the extreme sensitivity, 
      potentially this tool could provide a 'fingerprint' of the structure and 
      stability of a protein. Such a tool would be useful in the discovery and 
      development of biopharamceuticals and so we have explored our hypothesis with a 
      panel of therapeutic mAbs. We demonstrate that the quantified REES data show 
      remarkable sensitivity, being able to discern between structurally identical 
      antibodies and showing sensitivity to unfolding and aggregation. The approach 
      works across a broad concentration range (microg-mg/ml) and is highly consistent. We 
      show that the approach can be applied alongside traditional characterisation 
      testing within the context of a forced degradation study (FDS). Most importantly, 
      we demonstrate the approach is able to predict the stability of mAbs both in the 
      short (hours), medium (days) and long-term (months). The quantified REES data 
      will find immediate use in the biopharmaceutical industry in quality assurance, 
      formulation and development. The approach benefits from low technical complexity, 
      is rapid and uses instrumentation which exists in most biochemistry laboratories 
      without modification.
CI  - (c) 2020 The Author(s).
FAU - Knight, Michael J
AU  - Knight MJ
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Woolley, Rachel E
AU  - Woolley RE
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Kwok, Anthony
AU  - Kwok A
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Parsons, Stuart
AU  - Parsons S
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Jones, Hannah B L
AU  - Jones HBL
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Gulacsy, Christina E
AU  - Gulacsy CE
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Phaal, Polly
AU  - Phaal P
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Kassaar, Omar
AU  - Kassaar O
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
FAU - Dawkins, Kieran
AU  - Dawkins K
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Rodriguez, Elizabeth
AU  - Rodriguez E
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Marques, Andreia
AU  - Marques A
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Bowsher, Leo
AU  - Bowsher L
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Wells, Stephen A
AU  - Wells SA
AD  - Department of Physics, University of Bath, Bath, U.K.
FAU - Watts, Andrew
AU  - Watts A
AD  - Department of Pharmacy and Pharmacology, University of Bath, Bath, U.K.
AD  - Centre for Therapeutic Innovation, University of Bath, Bath, U.K.
FAU - van den Elsen, Jean M H
AU  - van den Elsen JMH
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
AD  - Centre for Therapeutic Innovation, University of Bath, Bath, U.K.
FAU - Turner, Alison
AU  - Turner A
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - O'Hara, John
AU  - O'Hara J
AD  - UCB, 216 Bath Road, Slough SL1 3WE, U.K.
FAU - Pudney, Christopher R
AU  - Pudney CR
AD  - Department of Biology and Biochemistry, University of Bath, Bath, U.K.
AD  - Centre for Therapeutic Innovation, University of Bath, Bath, U.K.
LA  - eng
GR  - BB_/Biotechnology and Biological Sciences Research Council/United Kingdom
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - England
TA  - Biochem J
JT  - The Biochemical journal
JID - 2984726R
RN  - 0 (Antibodies, Monoclonal)
SB  - IM
MH  - Antibodies, Monoclonal/*chemistry
MH  - Protein Conformation
MH  - Protein Stability
MH  - Spectrometry, Fluorescence
PMC - PMC7527260
OTO - NOTNLM
OT  - antibodies
OT  - edge-shift
OT  - fluorescence spectroscopy
OT  - protein dynamics
OT  - protein stability
COIS- We declare that CRP has commercial interests relating to the work presented in 
      the manuscript, pertaining to a directorship of Bloc Laboratories Limited.
EDAT- 2020/09/02 06:00
MHDA- 2021/02/24 06:00
PMCR- 2020/09/28
CRDT- 2020/09/02 06:00
PHST- 2020/07/27 00:00 [received]
PHST- 2020/08/27 00:00 [revised]
PHST- 2020/09/01 00:00 [accepted]
PHST- 2020/09/02 06:00 [pubmed]
PHST- 2021/02/24 06:00 [medline]
PHST- 2020/09/02 06:00 [entrez]
PHST- 2020/09/28 00:00 [pmc-release]
AID - 226283 [pii]
AID - BCJ-477-3599 [pii]
AID - 10.1042/BCJ20200580 [doi]
PST - ppublish
SO  - Biochem J. 2020 Sep 30;477(18):3599-3612. doi: 10.1042/BCJ20200580.