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.

胃肠道的粘膜表面在免疫稳态和防御中起重要作用，并且可能因肠胃疾病或感染而受损。使用单克隆抗体（mAb）进行的治疗干预具有潜在的最小脱靶效应以及口服时全身暴露受限的可能性。至关重要的是，为了在腔表面获得功效，mAb必须在胃肠道环境中保持稳定并具有功能活性。为了更好地了解同种型，类别和分子结构对重组抗体肠稳定性的影响，我们使用了体外模拟肠液（SIF）分析，以评估针对基于肠mAb的治疗药物的一组候选抗体。重组IgG1在SIF孵育后最不稳定，而IgA的稳定性通常在聚合后增加，亚类之间存在细微差异。值得注意的是，mAb内部和之间的变异性模式表明可变区有助于mAb的稳定性，并可能介导mAb对蛋白酶的敏感性。尽管SIF降解相对较快，但靶向SIF处理后针对肠毒素的大肠杆菌（ETEC）的单克隆抗体仍显示出功能活性，与SIgA2相比，SIgA1的功能有所改善。这项研究的结果对肠内治疗药物的设计和随后基于以下药物的潜在候选药物的选择具有重要意义。体外肠道稳定性评估。