Production, characterization, andin vivohalf-life extension of polymeric IgA molecules in mice

Abstract

IgA antibodies have broad potential as a novel therapeutic platform based on their superior receptor-mediated cytotoxic activity, potent neutralization of pathogens, and ability to transcytose across mucosal barriers via polymeric immunoglobulin receptor (pIgR)-mediated transport, compared to traditional IgG-based drugs. However, the transition of IgA into clinical development has been challenged by complex expression and characterization, as well as rapid serum clearance that is thought to be mediated by glycan receptor scavenging of recombinantly produced IgA monomer bearing incompletely sialylated N-linked glycans. Here, we present a comprehensive biochemical, biophysical, and structural characterization of recombinantly produced monomeric, dimeric and polymeric human IgA. We further explore two strategies to overcome the rapid serum clearance of polymeric IgA: removal of all N-linked glycosylation sites creating an aglycosylated polymeric IgA and engineering in FcRn binding with the generation of a polymeric IgG-IgA Fc fusion. While previous reports and the results presented in this study indicate that glycan-mediated clearance plays a major role for monomeric IgA, systemic clearance of polymeric IgA in mice is predominantly controlled by mechanisms other than glycan receptor clearance, such as pIgR-mediated transcytosis. The developed IgA platform now provides the potential to specifically target pIgR expressing tissues, while maintaining low systemic exposure.

IgA: Structure, Function, and Developability

“…The therapeutic antibody field is currently dominated by IgG-based mAbs. The advantages of opening up this arena to include IgA-based mAbs are becoming increasingly apparent, piquing interest in both academia and industry [79,[131][132][133]. One advantage is the new prospects it offers in terms of intellectual property, in what is already a complex landscape [134].…”

“…This shorter half-life is in part due to clearance mediated by the ASGPR, which recognises terminal galactose residues on the glycans of IgA. Efforts have been made to extend half-life by removing N-linked glycosylation sites [139], generating IgA with higher terminal sialylation of N-glycans [140], by attaching an albumin-binding domain to either the LC or HC in order to facilitate binding to the neonatal Fc receptor FcRn [141], or by engineering in FcRn binding by generating an IgG-IgA Fc fusion [133].…”

“…The production of polymeric forms of IgA or SIgA is particularly complex, given the requirement to co-express LC, HC, and J chain, and ensure attachment of SC. However, systems to achieve this have been explored and continue to be refined [133,155,156].…”

Human IgA Monoclonal Antibodies That Neutralize Poliovirus, Produced by Hybridomas and Recombinant Expression

“…Human IgA mAbs have been isolated from memory B cell populations using recombinant methods [13,[21][22][23], and IgAs have been produced in plants [26,29,47,48] and mammalian cells [25,28,30,49]. In this study, we produced PV-neutralizing IgA mAbs using two strategies: (1) de novo IgA cloning from primary human B cells using an updated hybridoma method; and (2) isotype switching of existing IgG mAbs, with recombinant expression in transiently transfected HEK293F cells.…”

Discovery and characterization of single-domain antibodies for polymeric Ig receptor-mediated mucosal delivery of biologics

“…The portion of hpIgR ECD (residues 1-547) that remains to be bound with dIgA is known as secretory component and complex between dIgA and secretory component is known as sIgA. 11,12 In the mucosal lumen, the secretory component has numerous functions, including conferring stability and mucophilic properties to dIgA, nonspecific microbial scavenging, neutralization of toxins and epithelial homeostasis. 13 The schematic of pIgR-mediated transport of dIgA is shown in Figure 1.…”