Substrate-Dependent Ligand Inhibition of the Human Organic Cation Transporter OCT2

Abstract

Organic cation transporter 2 (OCT2) mediates the initial step in renal secretion of organic cations: uptake from the blood, across the basolateral membrane, and into the renal proximal tubule cells. Because of its potential as a target for unwanted drug-drug interactions (DDIs), considerable attention has been directed toward understanding the basis of OCT2 selectivity. These studies typically assess selectivity based on ligand inhibition profiles for OCT2-mediated transport of a probe substrate. However, little attention has been given to the potential influence of the substrate on the profile of ligand inhibition. Here we compared the IC 50 values obtained for a set of structurally distinct inhibitors against OCT2-mediated transport of three structurally distinct substrates: 1-methyl-4-phenylpyridinium (MPP); metformin; and a novel fluorescent substrate, N,N,Ntrimethyl-2-[methyl (7-nitrobenzo[c][l,2,5]oxadiazol-4-yl)amino] ethanaminium iodide (NBD-MTMA). The median IC 50 value for inhibition of MPP transport was 9-fold higher than that for inhibition of metformin transport. Similarly, the median IC 50 value for inhibition of MPP transport was 5-fold higher than that for NBD-MTMA transport. However, this was not a systematic difference in inhibitory efficacy; the ratio of IC 50 values, MPP versus NBD-MTMA, ranged from 88-fold (ipratropium) to 0.3-fold (metformin). These data show that 1) the choice of OCT2 substrate significantly influences both quantitative and qualitative inhibitory interactions with cationic drugs; and 2) ligand interactions with OCT2 are not restricted to competition for a common ligand binding site, consistent with a binding surface characterized by multiple, possibly overlapping interaction sites. Development of predictive models of DDIs with OCT2 must take into account the substrate dependence of ligand interaction with this protein.

Lack of Influence of Substrate on Ligand Interaction with the Human Multidrug and Toxin Extruder, MATE1

“…With the information gained from these screens, plus IC 50 values determined for a structurally diverse subset of these compounds, we generated machine-learning and pharmacophore models, respectively. In contrast to the behavior observed with some other multidrug transporters (Ekins et al, 2002b;Garrigues et al, 2002;Westholm et al, 2009;Roth et al, 2011;Belzer et al, 2013;Hacker et al, 2015), the results suggest that substrate identity exerts comparatively little influence on ligand interaction with MATE1. (Aavula et al, 2006).…”

“…However, little attention has been given to a critical issue relevant to understanding the influence of MATE1 on unwanted DDI: the potential impact of substrate identity on the profile of drug interaction with MATE1. Increasing evidence suggests that the effectiveness of cationic drugs as inhibitors of multidrug transporters can be significantly influenced by the substrate used to monitor transport activity (Belzer et al, 2013;Thévenod et al, 2013;Hacker et al, 2015), which may complicate the interpretation of decision tree-based assays for assessing potential DDIs (Giacomini et al, 2010;Hillgren et al, 2013). However, the extent to which MATE transporters display such behavior is not clear.…”

Organic Cation Transporter 2 Overexpression May Confer an Increased Risk of Gentamicin-Induced Nephrotoxicity

“…The structural complexity of the OCT binding pocket is known, and for several substrates, including choline, tetraethylammonium (TEA), and 1-methyl-4-phenylpyridinium (MPP), two binding sites have been described, but no cooperativity was reported (48)(49)(50). Substrate-dependent inhibitory ligand interaction is a common characteristic for OCT2 (50). The search for inhibitors that can disrupt such allosteric binding may potentially reduce the accumulation of gentamicin in the cortex avoiding nephrotoxicity in the clinical setting.…”

Substrate-Dependent Inhibition of Human MATE1 by Cationic Ionic Liquids

“…In addition, we show that their quantitative inhibitory effect on MATE1 activity depends on the transported substrate, which is not consistent with simple competition for a common binding site and, instead, suggests that ligand interaction with MATE1 can involve binding to distinct sites within a larger binding surface. (Belzer et al, 2013); analysis by paper chromatography showed that the compound was .90% pure. The chloride salts of NBuPy, Bmim, and BmPy were obtained from Merck KGaA (Darmstadt, Germany).…”