Ligand Discovery for the Alanine-Serine-Cysteine Transporter (ASCT2, SLC1A5) from Homology Modeling and Virtual Screening

Abstract

The Alanine-Serine-Cysteine transporter ASCT2 (SLC1A5) is a membrane protein that moves neutral amino acids into cells in exchange for outward movement of intracellular amino acid. ASCT2 is highly expressed in peripheral tissues such as the lung, kidney, intestines, and testis where it contributes to the homeostasis of intracellular concentrations of neutral amino acids. ASCT2 also plays an important role in the development of a variety of cancers such as melanoma, by transporting amino acid nutrients such as glutamine into the proliferating tumors. Therefore, ASCT2 is a key drug target with potentially great pharmacological importance. Here, we identify seven ASCT2 ligands by computational modeling and experimental testing. In particular, we construct homology models based on crystallographic structures of the aspartate transporter GltPh in two different conformations, and optimize the models’ binding sites for protein-ligand complementarity. Virtual screening of 594,166 compounds including drugs, metabolites, and fragment-like molecules from the ZINC database, followed by experimental testing of 11 top hits with functional measurements using electrophysiological methods reveals seven ligands including five activators and two inhibitors. For example, aminooxetane-3-carboxylate is a more efficient activator than any other known ASCT2 natural or unnatural substrate. Furthermore, two of the hits inhibited ASCT2 mediated glutamine uptake and proliferation of a melanoma cancer cell line. Our results improve our understanding on how substrate specificity is determined in amino acid transporters as well as provide novel scaffolds for developing drugs targeting ASCT2, an emerging therapeutic target for cancer and neurological disorders.