Substrate-Dependent Trans-Stimulation of Organic Cation Transporter 2 Activity.

Charles R Lefèvre,Caroline Moreau,Olivier Fardel,Arnaud Bruyere,Elodie Jouan,Sophie Gaubert,Marc Le Vée

doi:10.3390/ijms222312926

Abstract

The search of substrates for solute carriers (SLCs) constitutes a major issue, owing notably to the role played by some SLCs, such as the renal electrogenic organic cation transporter (OCT) 2 (SLC22A2), in pharmacokinetics, drug–drug interactions and drug toxicity. For this purpose, substrates have been proposed to be identified by their cis-inhibition and trans-stimulation properties towards transporter activity. To get insights on the sensitivity of this approach for identifying SLC substrates, 15 various exogenous and endogenous OCT2 substrates were analysed in the present study, using 4-(4-(dimethylamino)styryl)-N-methylpyridinium iodide (DiASP) as a fluorescent OCT2 tracer substrate. All OCT2 substrates cis-inhibited DiASP uptake in OCT2-overexpressing HEK293 cells, with IC50 values ranging from 0.24 µM (for ipratropium) to 2.39 mM (for dopamine). By contrast, only 4/15 substrates, i.e., acetylcholine, agmatine, choline and metformin, trans-stimulated DiASP uptake, with a full suppression of the trans-stimulating effect of metformin by the reference OCT2 inhibitor amitriptyline. An analysis of molecular descriptors next indicated that trans-stimulating OCT2 substrates exhibit lower molecular weight, volume, polarizability and lipophilicity than non-trans-stimulating counterparts. Overall, these data indicated a rather low sensitivity (26.7%) of the trans-stimulation assay for identifying OCT2 substrates, and caution with respect to the use of such assay may therefore be considered.

Highlights

Solute carriers (SLCs) constitute a superfamily of more than 450 membrane proteins, acting as transporters of a large spectrum of molecules, including nutrients, metabolites, xenobiotics, small molecule drugs and metal ions [1]
Fifteen known OCT2 substrates, corresponding to laboratory/chemical reagents (n = 2), drugs (n = 6) and endogenous compounds/metabolites (n = 7) and whose affinities/Michaelis constant (KM) values for OCT2 are listed in Table 1, were investigated for their potential cis-inhibitory effects towards OCT2-mediated transport of DiASP
Endogenous OCT2 substrates cis-inhibited OCT2 activity, with IC50 ranging from 0.18 mM to 2.39 mM (Figure 2)

Summary

Introduction

Solute carriers (SLCs) constitute a superfamily of more than 450 membrane proteins, acting as transporters of a large spectrum of molecules, including nutrients, metabolites, xenobiotics (such as phytochemicals), small molecule drugs and metal ions [1]. Some of them have to be regulatorily studied during the pharmaceutical development of new molecular entities [5] This is notably the case for organic cation transporter (OCT) 2 (SLC22A2), a uniporter expressed at the basolateral pole of proximal tubular cells [4]. OCT2 mediates uptake of cationic drugs such as metformin and cimetidine from the blood into renal cells [10]; it functions in conjunction with multidrug and toxin extrusion protein (MATE) 1 (SLC47A1) and MATE2-K (SLC47A2), which expel OCT2 substrates into the urine at the apical pole of proximal cells [11] In this way, OCT2 plays a major role in the renal secretion of cationic drugs [12]; it is involved in the nephrotoxicity of cisplatin [13]. Organic anion transporter (OAT) 1 (SLC22A6) and OAT3 (SLC22A8), expressed at the basolateral pole of renal proximal cells, such as OCT2, are implicated in their renal secretion [14]

Methods

Results

Conclusion