Single-step Protein A and Protein G avidity purification methods to support bispecific antibody discovery and development.
Antibody
BEAT
Protein A
Protein G
bispecific
engineering
heterodimer
purification
Journal
mAbs
ISSN: 1942-0870
Titre abrégé: MAbs
Pays: United States
ID NLM: 101479829
Informations de publication
Date de publication:
Historique:
pubmed:
30
8
2019
medline:
18
6
2020
entrez:
30
8
2019
Statut:
ppublish
Résumé
Heavy chain (Hc) heterodimers represent a majority of bispecific antibodies (bsAbs) under clinical development. Although recent technologies achieve high levels of Hc heterodimerization (HD), traces of homodimer contaminants are often present, and as a consequence robust purification techniques for generating highly pure heterodimers in a single step are needed. Here, we describe two different purification methods that exploit differences in Protein A (PA) or Protein G (PG) avidity between homo- and heterodimers. Differential elution between species was enabled by removing PA or PG binding in one of the Hcs of the bsAb. The PA method allowed the avidity purification of heterodimers based on the VH3 subclass, which naturally binds PA and interferes with separation, by using a combination of IgG3 Fc and a single amino acid change in VH3, N82aS. The PG method relied on a combination of three mutations that completely disrupts PG binding, M428G/N434A in IgG1 Fc and K213V in IgG1 CH1. Both methods achieved a high level of heterodimer purity as single-step techniques without Hc HD (93-98%). Since PA and PG have overlapping binding sites with the neonatal Fc receptor (FcRn), we investigated the effects of our engineering both
Identifiants
pubmed: 31462177
doi: 10.1080/19420862.2019.1660564
pmc: PMC6816383
doi:
Substances chimiques
Antibodies, Bispecific
0
Antibodies, Monoclonal
0
Bacterial Proteins
0
IgG Fc-binding protein, Streptococcus
0
Immunoglobulin Fc Fragments
0
Staphylococcal Protein A
0
Types de publication
Journal Article
Langues
eng
Sous-ensembles de citation
IM
Pagination
1464-1478Références
J Virol. 2001 Dec;75(24):12161-8
pubmed: 11711607
J Biotechnol. 2008 Apr 30;134(3-4):222-30
pubmed: 18304667
Nucleic Acids Res. 2015 Jan;43(Database issue):D413-22
pubmed: 25378316
Mol Cell. 2001 Apr;7(4):867-77
pubmed: 11336709
MAbs. 2010 Sep-Oct;2(5):480-99
pubmed: 20647768
Sci Rep. 2017 Nov 14;7(1):15521
pubmed: 29138497
J Mol Biol. 1997 Jul 4;270(1):26-35
pubmed: 9231898
Protein Eng. 2002 Oct;15(10):835-42
pubmed: 12468718
Br J Pharmacol. 2006 Aug;148(8):1116-23
pubmed: 16847437
Nat Rev Immunol. 2007 Sep;7(9):715-25
pubmed: 17703228
J Biol Chem. 2009 May 1;284(18):12373-83
pubmed: 19269963
J Biol Chem. 2006 Jan 13;281(2):951-61
pubmed: 16278208
Nat Biotechnol. 2016 Dec 7;34(12):1215-1217
pubmed: 27926715
Protein Expr Purif. 1992 Oct;3(5):368-73
pubmed: 1458050
Drugs. 2018 Feb;78(2):269-274
pubmed: 29357074
Nat Rev Drug Discov. 2014 Nov;13(11):799-801
pubmed: 25359367
FEMS Immunol Med Microbiol. 1998 Jan;20(1):69-78
pubmed: 9514577
Expert Opin Biol Ther. 2007 Mar;7(3):405-18
pubmed: 17309332
MAbs. 2016 May-Jun;8(4):828-38
pubmed: 26963837
MAbs. 2010 May-Jun;2(3):309-19
pubmed: 20418662
Cancer Cell. 2004 Apr;5(4):317-28
pubmed: 15093539
J Biol Chem. 2010 Jul 2;285(27):20850-9
pubmed: 20444694
Biochem Biophys Res Commun. 2007 Apr 13;355(3):751-7
pubmed: 17321501
Structure. 1995 Mar 15;3(3):265-78
pubmed: 7788293
Sci Rep. 2015 Dec 11;5:17943
pubmed: 26659273
Front Immunol. 2014 Feb 05;5:22
pubmed: 24600447
Adv Biosci Biotechnol. 2013 Apr;4(4a):73-84
pubmed: 25635232
J Biomol Tech. 2011 Jul;22(2):50-2
pubmed: 21738436
J Biol Chem. 2001 Mar 2;276(9):6591-604
pubmed: 11096108
J Chromatogr A. 2015 Aug 28;1409:60-9
pubmed: 26189670
Proc Natl Acad Sci U S A. 1992 May 15;89(10):4285-9
pubmed: 1350088
MAbs. 2012 Nov-Dec;4(6):653-63
pubmed: 22925968
PLoS One. 2013;8(2):e57479
pubmed: 23468998
Cancer Treat Rev. 2010 Oct;36(6):458-67
pubmed: 20347527
Proc Natl Acad Sci U S A. 2000 May 9;97(10):5399-404
pubmed: 10805799
Adv Immunol. 2004;82:155-215
pubmed: 14975257
Drug Discov Today. 2015 Jul;20(7):838-47
pubmed: 25728220
Basic Clin Pharmacol Toxicol. 2015 Oct;117(4):219-25
pubmed: 25683750
Biochemistry. 1981 Apr 28;20(9):2361-70
pubmed: 7236608
Nat Biotechnol. 2015 Mar;33(3):219-21
pubmed: 25748895
J Biol Chem. 2017 Jun 9;292(23):9745-9759
pubmed: 28450393
Protein Cell. 2018 Jan;9(1):86-120
pubmed: 28822103
FEBS Lett. 2014 Jan 21;588(2):269-77
pubmed: 24291820
J Immunol. 1995 Jul 1;155(1):219-25
pubmed: 7602098
J Biol Chem. 2014 Mar 14;289(11):7812-24
pubmed: 24469444
MAbs. 2017 Feb/Mar;9(2):182-212
pubmed: 28071970
MAbs. 2015;7(2):331-43
pubmed: 25658443
J Mol Biol. 1994 Nov 11;243(5):906-18
pubmed: 7966308
J Mol Biol. 2003 Jan 17;325(3):531-53
pubmed: 12498801
MAbs. 2011 Jan-Feb;3(1):61-6
pubmed: 20962582
J Immunol. 2002 Nov 1;169(9):5171-80
pubmed: 12391234