Affinity of human IgG subclasses to mouse Fc gamma receptors

Abstract

Human IgG is the main antibody class used in antibody therapies because of its efficacy and longer half-life, which are completely or partly due to FcγR-mediated functions of the molecules. Preclinical testing in mouse models are frequently performed using human IgG, but no detailed information on binding of human IgG to mouse FcγRs is available. The orthologous mouse and human FcγRs share roughly 60-70% identity, suggesting some incompatibility. Here, we report binding affinities of all mouse and human IgG subclasses to mouse FcγR. Human IgGs bound to mouse FcγR with remarkably similar binding strengths as we know from binding to human ortholog receptors, with relative affinities IgG3>IgG1>IgG4>IgG2 and FcγRI>>FcγRIV>FcγRIII>FcγRIIb. This suggests human IgG subclasses to have similar relative FcγR-mediated biological activities in mice.

Keywords: Fc-receptors; FcγR; IgG subclasses; mouse models; surface plasmon resonance.

Figure 1.

Calculation of K_D by interpolation of equilibrium fit. Sensorgrams (a) and derived affinity plots (b) of human IgG1 anti-RhD binding to ligands FcγRI, FcγRIIb, FcγRIII and FcγRIV for 4 different ligand (FcγR) concentrations; 30, 10, 3 and 1nM, c) The affinity plot derived K_D and R_max of each ligand concentration are plotted for interpolation of K_D to a constant R_max of 500.

Figure 2.

Binding of mouse IgG subclasses to mouse FcγR. The mouse IgG1, IgG2a, IgG2b and IgG3 with anti-Kell specificity were assessed for their binding affinity in K_D (M) to the mouse FcγRI, FcγRIIb, FcγRIII and FcγRIV by SPR. −/− denotes no binding detected. n = 3. The horizontal dashed line represents the maximum concentration of IgG which is used, and the threshold required to accurately calculate the K_D.

Figure 3.

Binding of human IgG subclasses to mouse FcγR. The human IgG1, IgG2, IgG3 and IgG4 with anti-RhD specificity (clone 19A10) were assessed for their binding affinity in K_D (M) to the mouse FcγRI, FcγRIIb, FcγRIII and FcγRIV by SPR. −/− means no binding detected; *denotes binding > 2 μM or > 3 μM for hIgG2 or hIgG4, respectively, represented by the horizontal dashed lines, which represent the maximum concentration of IgG that is used, and the threshold required to accurately calculate the K_D, 2 μM for IgG2 and 3 μM for IgG1, IgG3 and IgG4; **K_Ds for FcγRIII binding of hIgG1, hIgG2 and hIgG4 represent the average of 2 alternative estimates (see Material & Methods section for details). n = 3.