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CC BY 4.0 · Pharmaceutical Fronts 2020; 02(04): e160-e167
DOI: 10.1055/s-0041-1728817
Original Article

Extending the in vivo Half-Life of Adalimumab Fab via Sortase A-Mediated Conjugation of Adalimumab Fab with Modified Fatty Acids

Qing-Bin Zhang
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
,
Si-Da Ruan
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
,
Yong Wu
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
,
Jin-Hua Zhang
3   School of Pharmacy, Fudan University, Shanghai, People's Republic of China
,
Jian-Guang Lu
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
,
Jun Feng
1   State Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of China
2   Shanghai Duomirui Bio-Technology Co., Ltd., Shanghai, People's Republic of China
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Abstract

Adalimumab, a full-length monoclonal antibody, is widely used as an anti-tumor necrosis factor-α (anti-TNF-α) agent. In this article, we aimed to prolong the in vivo half-life of adalimumab antigen-binding fragment (Fab) through Sortase A (SrtA)-mediated conjugation of its Fab with fatty acid (FA). In our study, adalimumab Fab analog was prepared by adding an SrtA recognition sequence (LPETGG) and His6 tag to the heavy chain C-terminal of the Fab via (G4S)3 linker. Four FA motifs with different linkers were designed and synthesized by solid-phase methodology, then conjugated with the Fab analog using SrtA to produce Fab bioconjugates. The successful generation of four Fab bioconjugates (Fab–FA1, Fab–FA2, Fab–FA3, and Fab–FA4) was confirmed by SDS-PAGE (sodium dodecyl sulfate polyacrylamide gel electrophoresis) and mass spectrometry. Then, the bioactivities and half-life of these Fab bioconjugates were examined using TNF-α-/human serum albumin (HSA)-binding enzyme-linked immunosorbent assay, cytotoxicity assay, and pharmacokinetic study, respectively. All Fab bioconjugates exhibited similar TNF-α-neutralizing activities when compared with the Fab analog, even in the presence of albumin, indicating that there were no apparent influences on the functional site of Fab after FA modification. However, different degrees of affinity for HSA were observed among these Fab–FA bioconjugates, with Fab–FA3 exhibiting the maximal affinity. An in vivo study in mice further revealed remarkably improved pharmacokinetics of Fab– FA3 with a 15.2-fold longer plasma half-life (19.86 hours) compared with that of the Fab analog (1.31 hours). In summary, we have developed a novel long-acting adalimumab Fab bioconjugate, Fab–FA3, with more sustained in vivo activity, which can be used for drug development targeting TNF-α-mediated inflammatory diseases.

Keywords

TNF-α - adalimumab Fab - half-life - FA motifs - sortase A


Publikationsverlauf

Eingereicht: 02. Februar 2021

Angenommen: 16. Februar 2021

Artikel online veröffentlicht:
10. Mai 2021

© 2021. The Author(s). This is an open access article published by Thieme under the terms of the Creative Commons Attribution License, permitting unrestricted use, distribution, and reproduction so long as the original work is properly cited. (https://creativecommons.org/licenses/by/4.0/)

Georg Thieme Verlag KG
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  • References

  • 1 Baud V, Karin M. Signal transduction by tumor necrosis factor and its relatives. Trends Cell Biol 2001; 11 (09) 372-377
    CrossrefPubMedGoogle Scholar
  • 2 Mitoma H, Horiuchi T, Tsukamoto H, Ueda N. Molecular mechanisms of action of anti-TNF-α agents - comparison among therapeutic TNF-α antagonists. Cytokine 2018; 101: 56-63
    CrossrefPubMedGoogle Scholar
  • 3 Steeland S, Libert C, Vandenbroucke RE. A new venue of TNF targeting. Int J Mol Sci 2018; 19 (05) 1442
    CrossrefPubMedGoogle Scholar
  • 4 Hansel TT, Kropshofer H, Singer T, Mitchell JA, George AJ. The safety and side effects of monoclonal antibodies. Nat Rev Drug Discov 2010; 9 (04) 325-338
    CrossrefPubMedGoogle Scholar
  • 5 Chames P, Van Regenmortel M, Weiss E, Baty D. Therapeutic antibodies: successes, limitations and hopes for the future. Br J Pharmacol 2009; 157 (02) 220-233
    CrossrefPubMedGoogle Scholar
  • 6 Rader C. Overview on concepts and applications of Fab antibody fragments. Curr Protoc Protein Sci 2009; Chapter 6: Unit 6.9
    Google Scholar
  • 7 Flanagan RJ, Jones AL. Fab antibody fragments: some applications in clinical toxicology. Drug Saf 2004; 27 (14) 1115-1133
    CrossrefPubMedGoogle Scholar
  • 8 Rosa J, Sabelli M, Soriano ER. Prefilled certolizumab pegol (Cimzia(®)) syringes for self-use in the treatment of rheumatoid arthritis. Med Devices (Auckl) 2010; 3: 25-31
    PubMedGoogle Scholar
  • 9 Schlapschy M, Binder U, Börger C. et al. PASylation: a biological alternative to PEGylation for extending the plasma half-life of pharmaceutically active proteins. Protein Eng Des Sel 2013; 26 (08) 489-501
    CrossrefPubMedGoogle Scholar
  • 10 Nguyen A, Reyes II AE, Zhang M. et al. The pharmacokinetics of an albumin-binding Fab (AB.Fab) can be modulated as a function of affinity for albumin. Protein Eng Des Sel 2006; 19 (07) 291-297
    CrossrefPubMedGoogle Scholar
  • 11 Schlapschy M, Theobald I, Mack H, Schottelius M, Wester HJ, Skerra A. Fusion of a recombinant antibody fragment with a homo-amino-acid polymer: effects on biophysical properties and prolonged plasma half-life. Protein Eng Des Sel 2007; 20 (06) 273-284
    CrossrefPubMedGoogle Scholar
  • 12 van Witteloostuijn SB, Pedersen SL, Jensen KJ. Half-life extension of biopharmaceuticals using chemical methods: alternatives to PEGylation. ChemMedChem 2016; 11 (22) 2474-2495
    CrossrefPubMedGoogle Scholar
  • 13 Zorzi A, Linciano S, Angelini A. Non-covalent albumin-binding ligands for extending the circulating half-life of small biotherapeutics. MedChemComm 2019; 10 (07) 1068-1081
    CrossrefPubMedGoogle Scholar
  • 14 Bech EM, Pedersen SL, Jensen KJ. Chemical strategies for half-life extension of biopharmaceuticals: lipidation and its alternatives. ACS Med Chem Lett 2018; 9 (07) 577-580
    CrossrefPubMedGoogle Scholar
  • 15 Ilangovan U, Ton-That H, Iwahara J, Schneewind O, Clubb RT. Structure of sortase, the transpeptidase that anchors proteins to the cell wall of Staphylococcus aureus. Proc Natl Acad Sci U S A 2001; 98 (11) 6056-6061
    CrossrefPubMedGoogle Scholar
  • 16 Lieser RM, Yur D, Sullivan MO, Chen W. Site-specific bioconjugation approaches for enhanced delivery of protein therapeutics and protein drug carriers. Bioconjug Chem 2020; 31 (10) 2272-2282
    CrossrefPubMedGoogle Scholar
  • 17 Levary DA, Parthasarathy R, Boder ET, Ackerman ME. Protein-protein fusion catalyzed by sortase A. PLoS One 2011; 6 (04) e18342
    CrossrefPubMedGoogle Scholar
  • 18 Popplewell AG, Sehdev M, Spitali M, Weir AN. Expression of antibody fragments by periplasmic secretion in Escherichia coli. Methods Mol Biol 2005; 308: 17-30
    PubMedGoogle Scholar
  • 19 Newton JM, Vlahopoulou J, Zhou Y. Investigating and modelling the effects of cell lysis on the rheological properties of fermentation broths. Biochem Eng J 2017; 121: 38-48
    CrossrefGoogle Scholar
  • 20 Li Q, Aucamp JP, Tang A, Chatel A, Hoare M. Use of focused acoustics for cell disruption to provide ultra scale-down insights of microbial homogenization and its bioprocess impact--recovery of antibody fragments from rec E. coli. Biotechnol Bioeng 2012; 109 (08) 2059-2069
    CrossrefPubMedGoogle Scholar
  • 21 Lau J, Bloch P, Schäffer L. et al. Discovery of the once-weekly glucagon-like peptide-1 (GLP-1) analogue semaglutide. J Med Chem 2015; 58 (18) 7370-7380
    CrossrefPubMedGoogle Scholar
  • 22 Chen X, Zaro JL, Shen WC. Fusion protein linkers: property, design and functionality. Adv Drug Deliv Rev 2013; 65 (10) 1357-1369
    CrossrefPubMedGoogle Scholar
  • 23 Gosselin ML, Fabrizio D, Swain JF. et al. Serum albumin binding molecules. US Patent 8969289. March, 2015
    Google Scholar
  • 24 Zhang QB, Huang ZQ, Lu JG, Zhao WJ, Feng J. Prokaryotic expression, purification and activity determination of Sortase A enzyme mutant [in Chinese]. Carol J Pharm 2020; 51 (01) 37-47
    Google Scholar
  • 25 Pishesha N, Ingram JR, Ploegh HL. Sortase A: a model for transpeptidation and its biological applications. Annu Rev Cell Dev Biol 2018; 34: 163-188
    CrossrefPubMedGoogle Scholar
  • 26 Lau J, Hansen TK, Madsen K. et al. Novel GLP-1 derivatives. WO Patent 2005027978. March, 2005
    Google Scholar