Yeast Cell-Based Transport Assay for the Functional Characterization of Human 4F2hc-LAT1 and ‐LAT2, and LAT1 and LAT2 Substrates and Inhibitors

In mammalian cells, the L-type amino acid transporters (LATs) LAT1 (SLC7A5) and LAT2 (SLC7A8) form heterodimeric (HATs) with ancillary protein 4F2hc are involved in cellular uptake of specific acids. The HAT 4F2hc-LAT1 is found upregulated various cancer cell types, while 4F2hc-LAT2 a transporter for non-cancer cells. Preclinical studies have highlighted that plays an important role tumor progression representing valid anticancer target. Consequently, current research focusing on development potent human inhibitors. On other hand, emerging as target diseases, thus also gaining clinical interest. To determine affinity specificity substrates inhibitors or 4F2hc-LAT2, robust transport assays indispensable. We optimized validated assay using cells methylotrophic yeast Pichia pastoris stably overexpressing HATs -LAT2, LATs alone. radioligand [ 3 H]L-leucine was used reporter L-leucine, triiodothyronine (T3) thyroxine (T4) well BCH JPH203 (KYT-0353) validation. Obtained half-maximal inhibitory concentrations provided new insights, e.g., into LAT inhibitor potency thyroid hormones T3 T4 to inhibit through 4F2hc-LAT2. particular interest possible implications influences ligand binding transport. summary, presented valuable characterization ligands, towards specificity, can be applied compound screening. Finally, our established approach would applicable