Abstract
Mucosal surfaces of the gastrointestinal tract play an important role in immune homeostasis and defense and may be compromised by enteric disorders or infection. Therapeutic intervention using monoclonal antibody (mAb) offers the potential for treatment with minimal off-target effects as well as the possibility of limited systemic exposure when administered orally. Critically, to achieve efficacy at luminal surfaces, mAb must remain stable and functionally active in the gastrointestinal environment. To better understand the impact of isotype, class, and molecular structure on the intestinal stability of recombinant antibodies, we used an in vitro simulated intestinal fluid (SIF) assay to evaluate a panel of antibody candidates for enteric mAb-based therapeutics. Recombinant IgG1 was the least stable following SIF incubation, while the stability of IgA generally increased upon polymerization, with subtle differences between subclasses. Notably, patterns of variability within and between mAbs suggest that variable regions contribute to mAb stability and potentially mediate mAb susceptibility to proteases. Despite relatively rapid degradation in SIF, mAbs targeting Enterotoxigenic Escherichia coli (ETEC) displayed functional activity following SIF treatment, with SIgA1 showing improved function compared to SIgA2. The results of this study have implications for the design of enteric therapeutics and subsequent selection of lead candidates based upon in vitro intestinal stability assessments.
Highlights
Immune responses at the mucosal surface of the gastrointestinal (GI) tract play a crucial role in defense against pathogens as well as regulating inflammatory responses against commensal gut microbes and food antigens
While all antibody therapeutics licensed to date have been of the immunoglobulin G (IgG) isotype, secretory IgA (sIgA)-based enteric therapeutics offer the potential for higher avidity, enhanced gastrointestinal stability and half-life, decreased systemic exposure, and reduced inflammatory responses
We assessed four monoclonal antibody (mAb) targeting bacterial toxoid, fimbrial, and viral antigens expressed as IgG and/or SIgA for gastrointestinal stability using an in vitro simulated intestinal fluid (SIF) assay (Fig. S1)
Summary
Immune responses at the mucosal surface of the gastrointestinal (GI) tract play a crucial role in defense against pathogens as well as regulating inflammatory responses against commensal gut microbes and food antigens. Over the last several decades the number of licensed monoclonal antibody (mAb)-based therapeutics has increased substantially, with 79 mAbs approved by the US Food and Drug Administration as of December 2019, including several targeting IBD [3]. While such successes support the further development of enteric mAb-based therapeutics, the gastrointestinal environment presents significant challenges for mAb delivery and potency. Primary design considerations for enteric mAb therapeutics must include formulation and optimization for protease resistance, retention in the GI tract, and sufficient functional half-life to achieve efficacy. Studies using IgG have shown limited success in this regard [5,6,7,8], and other antibody classes have been investigated with the goal of enhanced stability at luminal surfaces [9,10]
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