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Review
. 2016 Mar;270(1):113-31.
doi: 10.1111/imr.12385.

Engineered IgG1-Fc--one Fragment to Bind Them All

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Free PMC article
Review

Engineered IgG1-Fc--one Fragment to Bind Them All

Elisabeth Lobner et al. Immunol Rev. .
Free PMC article

Abstract

The crystallizable fragment (Fc) of the immunoglobulin class G (IgG) is a very attractive scaffold for the design of novel therapeutics due to its quality of uniting all essential antibody functions. This article reviews the functionalization of this homodimeric glycoprotein by diversification of structural loops of CH3 domains for the design of Fcabs, i.e. antigen-binding Fc proteins. It reports the design of libraries for the selection of nanomolar binders with wildtype-like in vivo half-life and correlation of Fc receptor binding and ADCC. The in vitro and preclinical biological activity of selected Fcabs is compared with that of clinically approved antibodies. Recently, the great potential of the scaffold for the development of therapeutics for clinical use has been shown when the HER2-binding Fcab FS102 entered clinical phase I. Furthermore, methods for the engineering of biophysical properties of Fcabs applicable to proteins in general are presented as well as the different approaches in the design of heterodimeric Fc-based scaffolds used in the generation of bispecific monoclonal antibodies. Finally, this work critically analyzes and compares the various efforts in the design of highly diverse and functional libraries that have been made in the engineering of IgG1-Fc and structurally similar scaffolds.

Keywords: Antibody engineering; Fcab; IgG1-Fc; Library design.

Figures

Figure 1
Figure 1
Schematic representation of homodimeric human IgG1‐Fc (PDBID 1OQO), generated using PyMOL, with the corresponding amino acid sequence. IgG1 is composed of two heavy and two light chains, with the Fab region carrying the antigen‐binding sites and the Fc part mediating various effector functions. The homodimeric Fc part comprises the CH2 and CH3 domains, where β strands are depicted in green, and α helices and random coils are shown in gray. The C‐terminal loops of the CH3 domain are colored in red (AB loop), orange (CD loop), and purple (EF loop). The asparagine at position 297 carries the glycan (NaNaFbi), graphically represented according to Anthony et al. 44. The amino acid sequence of the CH2 and CH3 domain of wildtype IgG1‐Fc is shown and numbered according to Eu numbering system 20. On top of the sequence, the secondary structure elements of CH2 and CH3 according to the crystal structure (PDBID 1OQO) are shown.
Figure 2
Figure 2
Representation of secondary structural elements with transparent molecular surface of IgG1‐Fc and its binding ligands. IgG1‐Fc (PDBID 1OQO) is colored in gray. Glycan structures are depicted as lines. Dashed arrows mark the binding site of the neonatal Fc receptor [PDBID 1I1A 50]; Fc gamma receptor I [PDBID 4W4O 119]; the mini‐Z domain of protein A (PDBID 1OQO); a globular head of the complement system protein C1q [PDBID 1PK6 47]; the C‐terminal PRYSPRY domain of TRIM21 [PDBID 2IWG 120]; and the anti‐CH2 antibody (clone MK 1 A6, AbD Serotec). The epitope of C1q was obtained by docking studies combined with MD simulations by Schneider et al. 121. Traxlmayr et al. 86 defined the binding site of the anti‐CH2 antibody to be located at the C‐terminal part of the CH2 domain. The proportions regarding size of all shown crystal structures are true to scale.
Figure 3
Figure 3
Schematic representation of a hypothetical mAb2 [based on PDB‐ID 1HZH ( 122 )] and one isolated CH3 domain thereof. Secondary structure elements of the antibody are colored in gray with the molecular surface of the CDR loops shown in green. Loop areas in the CH3 domains are displayed in red (AB loop), orange (CD loop), and purple (EF loop). The surface approximations of the three loops indicate the putative region for the generation of binding sites.

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