Intestinal permeability of N-acetylcysteine is driven by gut microbiota-dependent cysteine palmitoylation

Trillions of intestinal microbiota are essential to the permeability of orally administered drugs. However, identifying microbial-drug interactions remains challenging due to the highly variable composition of intestinal flora among individuals. Using single-pass intestinal perfusion (SPIP) platform, we established the microbiota-based permeability screening framework involving germ-free (GF) and specific-pathogen-free (SPF) rats to compare in-situ Peff-values and metabolomic profiles of 32 orally administered drugs with disputable classifications of permeability. In contrast with SPF controls, N-Acetylcysteine (NAC) exhibited significantly increased permeability in GF rats, which was inversely related to reduced cysteine-3-ketosphinganine by Bacteroides. To further validate these microbiome features, we integrated clinical descriptors from a prospective cohort of 319 participants to optimize a 15-feature eXtreme Gradient Boosting (XGB) model. This machine learning (ML) model of clinical prediction revealed that cysteine palmitoylation by intestinal microbiota has significantly affected NAC permeability, thus leading to discordant biopharmaceutics classification.