Human monoclonal IgG isotypes differ in complement activating function at the level of C4 as well as C1q.

Abstract

Humanized antibodies are likely to have a major role in therapy and it is important to define their interaction with physiological effectors. By comparing a matched series of chimeric human mAbs we found that igG1 was most efficient in complement lysis, although IgG3 bound more C1q. To resolve this paradox we compared the ability of human IgG1, IgG2, IgG3, IgG4, and IgE and rat IgG2b to cause C1q binding, C1 binding and activation, C4 activation, C4b binding, and C3b binding. Rat IgG2b was included because this isotype has already successfully been used for therapy. Human IgG1 was less efficient than IgG3 and fixing C1q and C1 on the cell surface, but the number of C4 molecules bound per C1 was 10-fold greater for IgG1 than for IgG3. This difference, amplified through later stages of the complement cascade, can account for the superiority of IgG1 for cell lysis. The efficiency of IgG1 in fixing C4 was not due to a favored binding site on the antibody molecule, since virtually all of the bound C4b was attached to the cells. Rather, it appeared that the activation of C4 by C1s was greatly favored by IgG1 compared with IgG3. It should be possible to combine the optimal properties of IgG1 and IgG3 antibodies to produce an improved therapeutic reagent.

Glomerular Autoimmune Multicomponents of Human Lupus Nephritis In Vivo (2)

“…IgG2 were unexpected, because these antibodies are weak complement activators compared with IgG1 and IgG3. 31 Immunofluorescence on the same biopsies used for the proteomic approach confirmed IgG2 prevalence but also showed IgG1 and IgG3 (Supplemental Figure 3). We cannot exclude that IgG1 and IgG3 are directed versus other glomerular autoantigens, and research should now focus on this point, because there are at least 11 podocyte antigens that are good candidates for being investigated.…”

Engineering an Improved IgG4 Molecule with Reduced Disulfide Bond Heterogeneity and Increased Fab Domain Thermal Stability

“…IgG1 and IgG3 are generally described as active isotypes because they perform antibody-dependent cellmediated cytotoxicity and complement-dependent cell-mediated cytotoxicity. This is because these isotypes have the greatest affinity to Fc␥ receptors (4 -6), whereas complement 1q binds strongly to C H 2 of these two isotypes (7,8). Conversely, IgG2 and IgG4 bind poorly to these effector molecules, resulting in relatively low effector function induction.…”

IgG2m4, an engineered antibody isotype with reduced Fc function

“…Here, we report a novel engineered IgG isotype, IgG2m4, with reduced Fc functionality. IgG2m4 is based on the IgG2 isotype with four key amino acid residue changes derived from IgG4 (H268Q, V309L, A330S and P331S 10 Interactions between Fc motifs or residues and cellular components have been extensively studied using both human and mouse IgGs. The key amino acid residues in the Fc region that interact with Fcγ receptors (CD64 for FcγRI, CD32 FcγRII and CD16 FcγRIII), complement C1q and neonatal FcRn receptors have been determined.…”