Presenting Author:

Jeffrey Schneider, Ph.D.

Principal Investigator:

Thomas Hope, Ph.D.

Department:

Cell and Molecular Biology

Keywords:

Antibodies, HIV, Immunology

Location:

Third Floor, Feinberg Pavilion, Northwestern Memorial Hospital

B19 - Basic Science Women's Health Research

Mucin binding HIV IgG, a novel antibody effector function

Background: MUC16 is a major component of the glycocalyx that protects the epithelial barrier of the upper female reproductive tract (FRT). We have recently published an IgG binding function in a fragment of MUC16 (MUC16RnD). Increased IgG binding was observed in the sera of HIV infected individuals. Surface Plasmon Resonance (SPR) analysis revealed the IgG interaction with the MUC16RnD is Fc-mediated. Removal of the Fc glycan leads to increased binding while enzymatically modifying the antibodies to the G0 glycoform gave tightest binding (6.2 ± 3.2 nM KD for VRC01). In contrast treating MUC16RnD with PNGaseF to remove N-linked glycans caused an ablation of IgG binding, revealing that mucin glycosylation is necessary for binding. Methods: We isolated IgG from the serum of SIV infected rhesus macaques (RM) and obtained IgG from serum of chronically HIV infected individuals. Human and RM IgG populations that associate with MUC16 were then interrogated for their antigen specificity through a Luminex assay and effector function through FcγR ELISA and a fluorescent ADCC assay. Finally we subcloned out smaller fragments of MUC16 to precisely identify the IgG binding region and mutated N-linked glycosylation residues that could contribute to binding. Results: Analysis of the MUC16 associated IgG from SIV infected macaques revealed that MUC16 associating antibodies were enriched approximately 8-fold for gp41 specificity. Reduced ADCC and FcγR engagement were observed in these MUC16 associated IgG populations relative to the input IgG, highlighting Fc specialization and a unique effector function profile.   Subcloning revealed a ~300 amino acid region of MUC16 containing the major IgG binding activity. Mutagenesis of the single N-glycosylation site therein greatly decreased IgG binding. Conclusions: These results reveal that MUC16 has at least one Fc receptor with nanomolar binding to the G0 IgG glycoform. The population of IgG binding to MUC16 is distinct from the FcγR binding and ADCC acting population. The increased amounts of gp41 specific IgG binding to MUC16 indicates that a specific antigen response is naturally targeted to MUC16. Through dissection of the precise mucin domain where IgG binds we can exploit the glycocalyx to create an antibody shield to prevent HIV from breaching mucosal barriers.