Identification of novel inhibitors of the amino acid transporter B0AT1 (SLC6A19), a potential target to induce protein restriction and to treat type 2 diabetes.
The neutral amino acid transporter B0AT1 (SLC6A19) has recently been identified as a possible target to treat type 2 diabetes and related disorders. B0AT1 mediates the Na+-dependent uptake of all neutral amino acids. For surface expression and catalytic activity, B0AT1 requires coexpression of collectrin (TMEM27). In this study, we established tools to identify and evaluate novel inhibitors of B0AT1.



A CHO-based cell line was generated, stably expressing collectrin and B0AT1. Using this cell line, a high-throughput screening assay was developed, which uses a fluorescent dye to detect depolarisation of the cell membrane during amino acid uptake via B0AT1. In parallel to these functional assays, we ran a computational compound screen using AutoDock4 and a homology model of B0AT1 based on the high-resolution structure of the highly homologous Drosophila dopamine transporter.



We characterized a series of novel inhibitors of the B0AT1 transporter. Benztropine was identified as a competitive inhibitor of the transporter showing an IC50of 44 ± 9 μM. The compound was selective with regard to related transporters and blocked neutral amino acid uptake in inverted sections of mouse intestine.



The tools established in this study can be widely used to identify new transport inhibitors. Using these tools, we were able to identify compounds that can be used to study epithelial transport, to induce protein restriction, or be developed further through medicinal chemistry.
Research School of Biology, The Australian National University, Canberra, Australia.
Qi
Q
Cheng
Research School of Biology, The Australian National University, Canberra, Australia.
Nishank
N
Shah
Research School of Biology, The Australian National University, Canberra, Australia.
Angelika
A
Bröer
Research School of Biology, The Australian National University, Canberra, Australia.
Stephen
S
Fairweather
Research School of Biology, The Australian National University, Canberra, Australia.
Yang
Y
Jiang
Industriepark Hoechst, Sanofi-Aventis Deutschland GmbH, Frankfurt am Main, Germany.
Dieter
D
Schmoll
Research School of Biology, The Australian National University, Canberra, Australia.
Ben
B
Corry
Research School of Biology, The Australian National University, Canberra, Australia.
Stefan
0000-0002-8040-1634
S
Bröer
D026905:Amino Acid Transport Systems / Q000737:chemistry / Q000378:metabolism; D026921:Amino Acid Transport Systems, Neutral / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000818:Animals; D001590:Benztropine / Q000737:chemistry / Q000494:pharmacology*; D016466:CHO Cells; D003412:Cricetulus; D003924:Diabetes Mellitus, Type 2 / Q000188:drug therapy* / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004330:Drosophila; D005260:Female; D057166:High-Throughput Screening Assays; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D062105:Molecular Docking Simulation; D013329:Structure-Activity Relationship
D026905:Amino Acid Transport Systems; D026921:Amino Acid Transport Systems, Neutral; C505021:SLC6A19 protein, mouse; D001590:Benztropine