Low doses of antigen coupled to anti-CR2 mAbs induce rapid and enduring IgG immune responses in mice and in cynomolgus monkeys.

The complement system and B cell complement receptor 2 (CR2), specific for C component C3dg, play important roles in both the innate and adaptive immune response. We used hapten and protein conjugates of anti-CR2 mAbs as models for C3dg-opsonized antigens and immune complexes to examine the handling of and immune response to these reagents in mice and in non-human primates (NHP). Mice immunized and boosted i.v. with only 100 ng of Alexa 488 rat anti-mouse CR1/2 mAb 7G6 had strong IgG immune responses to the Alexa 488 hapten and to rat IgG, compared to very weak immune responses in mice treated with a comparable isotype control; larger doses of Alexa 488 mAb 7G6 did not increase the immune response. A vaccine constructed by cross-linking anthrax protective antigen to mAb 7G6 proved to be effective at low doses in generating sufficiently high titer serum IgG antibodies to neutralize anthrax lethal toxin in vitro and to protect mice from i.v. challenge with anthrax lethal toxin. When biotinylated HB135, a mouse mAb specific for human CR2, was injected i.v. into NHP, the probe manifested the same initial marginal zone B cell binding and subsequent localization to follicular dendritic cells as we have previously reported for comparable experiments in mice. Moreover, i.v. immunization of NHP with 1 microg/kg of Alexa 488 mAb HB135 promoted an IgG immune response to the Alexa 488 hapten and to mouse IgG. Taken together, these results demonstrate the efficacy of using anti-CR2 mAbs as antigen carriers for i.v. immunization with small amounts of antigens without adjuvant.

Analyses of the in vivo trafficking of stoichiometric doses of an anti-complement receptor 1/2 monoclonal antibody infused intravenously in mice.

Complement plays a critical role in the immune response by opsonizing immune complexes (IC) and thymus-independent type 2 Ags with C3 breakdown product C3dg, a CR2-specific ligand. We used a C3dg-opsonized IC model, anti-CR1/2 mAb 7G6, to investigate how such substrates are processed. We used RIA, whole body imaging, flow cytometry, and fluorescence immunohistochemistry to examine the disposition of 0.1- to 2-microg quantities of mAb 7G6 infused i.v. into BALB/c mice. The mAb is rapidly taken up by the spleen and binds preferentially to marginal zone (MZ) B cells; within 24 h, the MZ B cells relocate and transfer mAb 7G6 to follicular dendritic cells (FDC). Transfer occurs coincident with loss of the extracellular portion of MZ B cell CR2, suggesting that the process may be mediated by proteolysis of CR2. Intravenous infusion of an FDC-specific mAb does not induce comparable splenic localization or cellular reorganization, emphasizing the importance of MZ B cells in intrasplenic trafficking of bound substrates. We propose the following mechanism: binding of C3dg-opsonized IC to noncognate MZ B cells promotes migration of these cells to the white pulp, followed by CR2 proteolysis, which allows transfer of the opsonized IC to FDC, thus facilitating presentation of intact Ags to cognate B cells.