Our understanding of the involvement of ASCT2 in the physiological processes involving glutamine is hampered by a lack of understanding of its pharmacology and the absence of high-affinity inhibitors.
In this study, we combined an in silico docking approach with experimental investigation of binding parameters to develop new ASCT2 inhibitors and substrates, a series of serine esters, and to determine structural parameters that govern their functional effects.
The series of compounds was synthesized using standard methods and exhibited a range of properties, from inhibitors to partial substrates and full substrates.
Our results suggest that amino acid derivatives with small side-chain volume and low side-chain hydrophobicity interact strongly with the closed-loop form of the binding site, in which re-entrant loop 2, the presumed extracellular gate for the substrate binding site, is closed off. However, these derivatives bind weakly to the open-loop form (external gate open to the extracellular side), acting as transported substrates.
In contrast, inhibitors bind preferentially to the open-loop form.
An aromatic residue in the side chain is required for high-affinity interaction.
One of the compounds, the l-serine ester serine biphenyl-4-carboxylate reversibly inhibits ASCT2 function with an apparent affinity of 30 μM.