Mechanistic benchmarking of SNAC, DDM, and LLC reveals enhancer-dependent pathways for intestinal semaglutide delivery.

The oral delivery of peptide therapeutics remains constrained by poor intestinal permeability and the challenge of enhancing epithelial transport without compromising mucosal integrity. Although sodium N-(8-[2-hydroxybenzoyl]amino) caprylate (SNAC) enables gastric absorption of oral semaglutide and has also been explored for small-intestinal absorption of other peptide drugs, its suitability for enteric intestinal delivery of semaglutide remains unexplored. Here, we performed a comparative mechanistic evaluation of three permeation enhancers (SNAC, N-dodecyl-β-D-maltoside, and lauroyl-l-carnitine) for enteric delivery of semaglutide. Mechanisms were screened in epithelial monolayers and in situ intestinal absorption in rats, and the enhancers carried forward were formulated into enteric-coated capsules and evaluated in beagle dogs, with the leading candidate taken in to cynomolgus monkeys. In differentiated Caco-2 monolayers, all enhancers increased semaglutide transport but exhibited distinct transport mechanisms and barrier-response profiles. SNAC increased permeability without increasing mannitol transport or disrupting ZO-1, indicating a largely non-paracellular, cargo-dependent profile. DDM markedly increased paracellular marker transport and disrupted tight-junction organization, consistent with a primarily paracellular mode of action associated with reduced barrier recoverability. LLC showed a more favorable balance between transport enhancement and barrier recovery, while also increasing intracellular semaglutide-associated fluorescence and FD4 permeability. Rab5 immunostaining and dynasore inhibition indicated a dynamin-sensitive uptake component alongside partial tight-junction modulation. In rats, LLC produced the greatest increase in semaglutide absorption, and this effect was substantially attenuated by dynasore co-treatment. Transmission electron microscopy and tissue immunofluorescence further supported the presence of LLC-associated micelle-like structures, Rab5-related intracellular trafficking and junctional remodelling, with no overt tissue injury on local or systemic histology. For large-animal translation, citric acid and L-arginine improved semaglutide stability in intestinal fluids, and the resulting LLC-containing enteric formulation markedly increased systemic exposure in orally dosed beagle dogs and in cynomolgus monkeys following intraduodenal delivery. Collectively, these findings identify LLC as a promising permeation enhancer within the present SNAC/DDM/LLC comparison for enteric intestinal delivery of semaglutide and highlight how coordinated uptake-related and barrier-modulating mechanisms can support intestinal semaglutide absorption.