Potent and Selective Inhibition of Plasma Membrane Monoamine Transporter by HIV Protease Inhibitors.
Plasma membrane monoamine transporter (PMAT) is a major uptake-2 monoamine transporter that shares extensive substrate and inhibitor overlap with organic cation transporters 1-3 (OCT1-3). Currently, there are no PMAT-specific inhibitors available that can be used in in vitro and in vivo studies to differentiate between PMAT and OCT activities. In this study, we showed that IDT307 (4-(4-(dimethylamino)phenyl)-1-methylpyridinium iodide), a fluorescent analog of 1-methyl-4-phenylpyridinium (MPP+), is a transportable substrate for PMAT and that IDT307-based fluorescence assay can be used to rapidly identify and characterize PMAT inhibitors. Using the fluorescent substrate-based assays, we analyzed the interactions of eight human immunodeficiency virus (HIV) protease inhibitors (PIs) with human PMAT and OCT1-3 in human embryonic kidney 293 (HEK293) cells stably transfected with individual transporters. Our data revealed that PMAT and OCTs exhibit distinct sensitivity and inhibition patterns toward HIV PIs. PMAT is most sensitive to PI inhibition whereas OCT2 and OCT3 are resistant. OCT1 showed an intermediate sensitivity and a distinct inhibition profile from PMAT. Importantly, lopinavir is a potent PMAT inhibitor and exhibited >120 fold selectivity toward PMAT (IC₅₀ = 1.4 ± 0.2 µM) over OCT1 (IC₅₀ = 174 ± 40 µM). Lopinavir has no inhibitory effect on OCT2 or OCT3 at maximal tested concentrations. Lopinavir also exhibited no or much weaker interactions with uptake-1 monoamine transporters. Together, our results reveal that PMAT and OCTs have distinct specificity exemplified by their differential interaction with HIV PIs. Further, we demonstrate that lopinavir can be used as a selective PMAT inhibitor to differentiate PMAT-mediated monoamine and organic cation transport from those mediated by OCT1-3.
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.).
Haichuan
H
Duan
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.).
Tao
T
Hu
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.).
Robert S
RS
Foti
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.).
Yongmei
Y
Pan
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.).
Peter W
PW
Swaan
Department of Pharmaceutics, University of Washington, Seattle, Washington (H.D., T.H., J.W.); Department of Pharmacokinetics and Drug Metabolism, Amgen Inc., Seattle, Washington (R.S.F.); and Department of Pharmaceutical Sciences, University of Maryland, Baltimore, Maryland (Y.P., P.W.S.) jowang@u.washington.edu.
Joanne
J
Wang
D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D033705:Equilibrative Nucleoside Transport Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D057809:HEK293 Cells; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D006801:Humans; D027701:Organic Cation Transport Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D027702:Organic Cation Transporter 1 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D000075102:Organic Cation Transporter 2; D000099254:Organic Cation Transporter 3
D033705:Equilibrative Nucleoside Transport Proteins; D017320:HIV Protease Inhibitors; D027701:Organic Cation Transport Proteins; D027702:Organic Cation Transporter 1; D000075102:Organic Cation Transporter 2; D000099254:Organic Cation Transporter 3; C491013:SLC22A2 protein, human; C495988:SLC29A4 protein, human