Non-Steroidal Anti-Inflammatory Drugs Are Inhibitors of the Intestinal Proton-Coupled Amino Acid Transporter (PAT1): Ibuprofen and Diclofenac Are Non-Translocated Inhibitors.

: The proton-coupled amino acid transporter (PAT1) is an intestinal absorptive solute carrier responsible for the oral bioavailability of some GABA-mimetic drug substances such as vigabatrin and gaboxadol. In the present work, we investigate if non-steroidal anti-inflammatory drug substances (NSAIDs) interact with substrate transport via human (h)PAT1. : The transport of substrates via hPAT1 was investigated in Caco-2 cells using radiolabeled substrate uptake and in oocytes injected with , measuring induced currents using the two-electrode voltage clamp technique. The molecular interaction between NSAIDs and hPAT1 was investigated using an AlphaFold2 model and molecular docking. NSAIDs such as ibuprofen, diclofenac, and flurbiprofen inhibited proline uptake via hPAT1, with IC values of 954 (logIC 2.98 ± 0.1) µM, 272 (logIC 2.43 ± 0.1) µM, and 280 (logIC 2.45 ± 0.1) µM, respectively. Ibuprofen acted as a non-competitive inhibitor of hPAT1-mediated proline transport. In hPAT1-expressing oocytes, ibuprofen and diclofenac did not induce inward currents, and inhibited inward currents caused by proline. Molecular modeling pointed to a binding mode involving an allosteric site. NSAIDs interact with hPAT1 as non-translocated non-competitive inhibitors, and molecular modeling points to a binding mode involving an allosteric site distinct from the substrate binding site. The present findings could be used as a starting point for developing specific hPAT1 inhibitors.