PMID- 37540995
OWN - NLM
STAT- MEDLINE
DCOM- 20230911
LR  - 20230911
IS  - 1873-264X (Electronic)
IS  - 0731-7085 (Linking)
VI  - 235
DP  - 2023 Oct 25
TI  - Comparison of middle- and bottom-up mass spectrometry in forced degradation 
      studies of bevacizumab and infliximab.
PG  - 115596
LID - S0731-7085(23)00365-5 [pii]
LID - 10.1016/j.jpba.2023.115596 [doi]
AB  - Monoclonal antibodies (mAbs) used as therapeutics need comprehensive 
      characterization for appropriate quality assurance. For analysis, cost-effective 
      methods are of high importance, especially when it comes to biosimilar 
      development which is based on extended physicochemical characterization. The use 
      of forced degradation to study the occurrence of modifications for analysis is 
      well established in drug development and may be used for the evaluation of 
      critical quality attributes (CQAs). For mAb analysis different procedures of 
      liquid chromatography hyphenated with mass spectrometry (LC-MS) analyses are 
      commonly applied. In this study the middle-up approach is compared to the more 
      expensive bottom-up analysis in a forced oxidation biosimilar comparability 
      study. Bevacizumab and infliximab as well as biosimilar candidates for the two 
      mAbs were forcefully oxidized by H(2)O(2) for 24, 48 and 72 h. For bottom-up, the 
      reduced and alkylated trypsin or Lys-C digested samples were analysed by LC-MS 
      with quadrupole time-of-flight mass analyser (LC-QTOF-MS) to detect susceptible 
      residues. By middle-up analysis several species of every subunit (Fc/2, light 
      chain and Fd') were detected which differed in the number of oxidations. For the 
      most abundant species, results from middle-up were in line with results from 
      bottom-up analysis, confirming the strength of middle-up analysis. However, for 
      less abundant species of some subunits, results differed between the two 
      approaches. In both mAbs, the Fc was extensively oxidized. In infliximab, 
      additional extensive oxidation was found in the Fab. Assignment to specific amino 
      acid residues was finally possible using the results from bottom-up analyses. 
      Interestingly, the C-terminal cysteine of the light chain was partially found 
      triply oxidized in both mAbs. The comparison of susceptibility to oxidation 
      showed high similarity between the reference products and their biosimilar 
      candidates. It is suggested that the findings of middle-up experiments should be 
      complemented by bottom-up analysis to confirm the assignments of the localization 
      of modifications. Once the consistency of results has been established, middle-up 
      analyses are sufficient in extended forced degradation biosimilar studies.
CI  - Copyright (c) 2023 Elsevier B.V. All rights reserved.
FAU - Dyck, Yan Felix Karl
AU  - Dyck YFK
AD  - Department of Pharmaceutical & Medicinal Chemistry, Institute of Pharmacy, Freie 
      Universitat Berlin, Konigin-Luise-Strasse 2+4, 14195 Berlin, Germany; Department 
      of Life Sciences & Technology, Berlin University of Applied Science, Seestrasse 
      64, 13347 Berlin, Germany.
FAU - Rehm, Daniel
AU  - Rehm D
AD  - Department of Pharmaceutical & Medicinal Chemistry, Institute of Pharmacy, Freie 
      Universitat Berlin, Konigin-Luise-Strasse 2+4, 14195 Berlin, Germany; ProBioGen 
      AG, Herbert-Bayer-Strasse 8, 13086 Berlin, Germany.
FAU - Winkler, Karsten
AU  - Winkler K
AD  - ProBioGen AG, Herbert-Bayer-Strasse 8, 13086 Berlin, Germany.
FAU - Sandig, Volker
AU  - Sandig V
AD  - ProBioGen AG, Herbert-Bayer-Strasse 8, 13086 Berlin, Germany.
FAU - Jabs, Wolfgang
AU  - Jabs W
AD  - Department of Life Sciences & Technology, Berlin University of Applied Science, 
      Seestrasse 64, 13347 Berlin, Germany.
FAU - Parr, Maria Kristina
AU  - Parr MK
AD  - Department of Pharmaceutical & Medicinal Chemistry, Institute of Pharmacy, Freie 
      Universitat Berlin, Konigin-Luise-Strasse 2+4, 14195 Berlin, Germany. Electronic 
      address: maria.parr@fu-berlin.de.
LA  - eng
PT  - Journal Article
PT  - Review
DEP - 20230722
PL  - England
TA  - J Pharm Biomed Anal
JT  - Journal of pharmaceutical and biomedical analysis
JID - 8309336
RN  - B72HH48FLU (Infliximab)
RN  - 2S9ZZM9Q9V (Bevacizumab)
RN  - 0 (Biosimilar Pharmaceuticals)
RN  - BBX060AN9V (Hydrogen Peroxide)
RN  - 0 (Antibodies, Monoclonal)
SB  - IM
MH  - Infliximab/chemistry
MH  - Bevacizumab
MH  - *Biosimilar Pharmaceuticals/chemistry
MH  - Hydrogen Peroxide
MH  - Antibodies, Monoclonal/chemistry
MH  - Mass Spectrometry/methods
OTO - NOTNLM
OT  - Accelerated stability testing
OT  - Avastin
OT  - Bottom-up approach
OT  - Liquid chromatography-high resolution mass spectrometry (LC-MS)
OT  - Middle-up approach
OT  - Remicade
COIS- Declaration of Competing Interest The authors declare the following financial 
      interests/personal relationships which may be considered as potential competing 
      interests, Karsten Winkler reports a relationship with ProBioGen AG that 
      includes: employment and equity or stocks. Volker Sandig reports a relationship 
      with ProBioGen AG that includes: employment and equity or stocks. Yan Dyck 
      reports equipment, drugs, or supplies were provided by ProBioGen AG. Daniel Rehm 
      reports financial support and equipment, drugs, or supplies were provided by 
      ProBioGen AG. Daniel Rehm reports a relationship with ProBioGen AG that includes: 
      employment.
EDAT- 2023/08/05 05:42
MHDA- 2023/09/11 06:42
CRDT- 2023/08/04 18:03
PHST- 2023/02/28 00:00 [received]
PHST- 2023/06/29 00:00 [revised]
PHST- 2023/07/19 00:00 [accepted]
PHST- 2023/09/11 06:42 [medline]
PHST- 2023/08/05 05:42 [pubmed]
PHST- 2023/08/04 18:03 [entrez]
AID - S0731-7085(23)00365-5 [pii]
AID - 10.1016/j.jpba.2023.115596 [doi]
PST - ppublish
SO  - J Pharm Biomed Anal. 2023 Oct 25;235:115596. doi: 10.1016/j.jpba.2023.115596. 
      Epub 2023 Jul 22.