Molecular Determinants of Substrate/Inhibitor Binding to the Human and Rabbit Renal Organic Cation Transporters hOCT2 and rbOCT2

Suhre, Wendy M.; Ekins, Sean; Chang, Cheng; Swaan, Peter W.; Wright, Stephen H.

doi:10.1124/mol.104.004713

Cited by 89 publications

(78 citation statements)

References 30 publications

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“…The results presented in this article demonstrate that, in addition to serving as substrates, NBuPy-Cl, BmPy-Cl, and Bmim-Cl are also potent inhibitors of rOCT1/2 and hOCT2. The IC 50 values for inhibition of hOCT2-mediated TEA transport by these three ILs ranged from 0.5 to 2.3 M. These inhibitory values are similar to those observed for MPP (2 M) and tetrapentylammonium (10 M), compounds considered to be potent inhibitors of hOCT2 (Suhre et al, 2005). When metformin, the widely prescribed type 2 antidiabetic drug, was used as the probe substrate for hOCT2, these ILs demonstrated even stronger inhibitory effects.…”

Section: Inhibitory Effects Of Ionic Liquids On Octs and Matesmentioning

confidence: 49%

Characterization of the Inhibitory Effects ofN-Butylpyridinium Chloride and Structurally Related Ionic Liquids on Organic Cation Transporters 1/2 and Human Toxic Extrusion Transporters 1/2-K In Vitro and In Vivo

Cheng¹,

Martinez-Guerrero²,

Wright³

et al. 2011

Drug Metab Dispos

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ABSTRACT:Ionic liquids (ILs) are a class of salts that are expected to be used as a new source of solvents and for many other applications. Our previous studies revealed that selected ILs, structurally related organic cations, are eliminated exclusively in urine as the parent compound, partially mediated by renal transporters. This study investigated the inhibitory effects of N-butylpyridinium chloride (NBuPy-Cl) and structurally related ILs on organic cation transporters (OCTs) and multidrug and toxic extrusion transporters (MATEs) in vitro and in vivo. After Chinese hamster ovary cells expressing rat (r) OCT1, rOCT2, human (h) OCT2, hMATE1, or hMATE2-K were constructed, the ability of NBuPy-Cl, 1-methyl-3-butylimidazolium chloride (Bmim-Cl), N-butyl-N-methylpyrrolidinium chloride (BmPyCl), and alkyl substituted pyridinium ILs to inhibit these transporters was determined in vitro. NBuPy-Cl (0, 0.5, or 2 mg/kg per hour) was also infused into rats to assess its effect on the pharmacokinetics of metformin, a substrate of OCTs and MATEs. NBuPy-Cl, Bmim-Cl, and BmPy-Cl displayed strong inhibitory effects on these transporters (IC 50 ‫؍‬ 0.2-8.5 M). In addition, the inhibitory effects of alkyl-substituted pyridinium ILs on OCTs increased dramatically as the length of the alkyl chain increased. The IC 50 values were 0.1, 3.8, 14, and 671 M (hexyl-, butyl-, and ethyl-pyridinium and pyridinium chloride) for rOCT2-mediated metformin transport. Similar structurally related inhibitory kinetics were also observed for rOCT1 and hOCT2. The in vivo coadministration study revealed that NBuPy-Cl reduced the renal clearance of metformin in rats. These results demonstrate that ILs compete with other substrates of OCTs and MATEs and could alter the in vivo pharmacokinetics of such substrates.

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Section: Inhibitory Effects Of Ionic Liquids On Octs and Matesmentioning

confidence: 49%

Characterization of the Inhibitory Effects ofN-Butylpyridinium Chloride and Structurally Related Ionic Liquids on Organic Cation Transporters 1/2 and Human Toxic Extrusion Transporters 1/2-K In Vitro and In Vivo

Cheng¹,

Martinez-Guerrero²,

Wright³

et al. 2011

Drug Metab Dispos

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“…Since the membrane potential assay we used is likely driven by CHT-dependent sodium flux, we reasoned that the failure to produce enhanced membrane depolarization despite elevated choline uptake could derive from endogenous electroneutral choline transport pathways. Organic cation transport systems are expressed in HEK293 cells, 64 and these transporters that could be responsible for the elevation in choline flux rates observed when endogenous choline levels are depleted. Supporting these ideas, incubation of ChAT/CHT LV cotransfected cells with the OCT2 inhibitor clonidine 65 ( Figure 6F) reduced the gain achieved in choline uptake relative to CHT LV-AA cells.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

Nonoisotopic Assay for the Presynaptic Choline Transporter Reveals Capacity for Allosteric Modulation of Choline Uptake

Ruggiero

Wright

Ferguson

et al. 2012

ACS Chem. Neurosci.

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Current therapies to enhance CNS cholinergic function rely primarily on extracellular acetylcholinesterase (AChE) inhibition, a pharmacotherapeutic strategy that produces dose-limiting side effects. The Na + -dependent, high-affinity choline transporter (CHT) is an unexplored target for cholinergic medication development. Although functional at the plasma membrane, CHT at steady-state is localized to synaptic vesicles such that vesicular fusion can support a biosynthetic response to neuronal excitation. To identify allosteric potentiators of CHT activity, we mapped endocytic sequences in the C-terminus of human CHT, identifying transporter mutants that exhibit significantly increased transport function. A stable HEK-293 cell line was generated from one of these mutants (CHT LV-AA) and used to establish a high-throughput screen (HTS) compatible assay based on the electrogenic nature of the transporter. We established that the addition of choline to these cells, at concentrations appropriate for high-affinity choline transport at presynaptic terminals, generates a hemicholinium-3 (HC-3)-sensitive, membrane depolarization that can be used for the screening of CHT inhibitors and activators. Using this assay, we discovered that staurosporine increased CHT LV-AA choline uptake activity, an effect mediated by a decrease in choline K M with no change in V max . As staurosporine did not change surface levels of CHT, nor inhibit HC-3 binding, we propose that its action is directly or indirectly allosteric in nature. Surprisingly, staurosporine reduced choline-induced membrane depolarization, suggesting that increased substrate coupling to ion gradients, arising at the expense of nonstoichiometric ion flow, accompanies a shift of CHT to a higher-affinity state. Our findings provide a new approach for the identification of CHT modulators that is compatible with high-throughput screening approaches and presents a novel model by which small molecules can enhance substrate flux through enhanced gradient coupling.

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“…Before we conducted these experiments, we fortuitously noticed that one of these compounds, palbociclib, exists as a cation protonated on the nitrogen atom of the pyridine ring (52) and has a hydrophobicaromatic site separated by a short distance from the positive charge, as well as multiple double-bonded oxygen moieties. These three structural motifs previously were found to be overrepresented in potent inhibitors of the organic cation transporter OCT2 (53)(54)(55). The presence of these motifs is of potential interest because OCT2 is responsible for accumulation of cisplatin in renal cells (35,37,56).…”

mentioning

confidence: 99%

Mitigation of acute kidney injury by cell-cycle inhibitors that suppress both CDK4/6 and OCT2 functions

Pabla

Gibson

Buege

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Acute kidney injury (AKI) is a potentially fatal syndrome characterized by a rapid decline in kidney function caused by ischemic or toxic injury to renal tubular cells. The widely used chemotherapy drug cisplatin accumulates preferentially in the renal tubular cells and is a frequent cause of drug-induced AKI. During the development of AKI the quiescent tubular cells reenter the cell cycle. Strategies that block cell-cycle progression ameliorate kidney injury, possibly by averting cell division in the presence of extensive DNA damage. However, the early signaling events that lead to cell-cycle activation during AKI are not known. In the current study, using mouse models of cisplatin nephrotoxicity, we show that the G1/S-regulating cyclin-dependent kinase 4/6 (CDK4/6) pathway is activated in parallel with renal cell-cycle entry but before the development of AKI. Targeted inhibition of CDK4/6 pathway by small-molecule inhibitors palbociclib (PD-0332991) and ribociclib (LEE011) resulted in inhibition of cell-cycle progression, amelioration of kidney injury, and improved overall survival. Of additional significance, these compounds were found to be potent inhibitors of organic cation transporter 2 (OCT2), which contributes to the cellular accumulation of cisplatin and subsequent kidney injury. The unique cell-cycle and OCT2-targeting activities of palbociclib and LEE011, combined with their potential for clinical translation, support their further exploration as therapeutic candidates for prevention of AKI.

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Molecular Determinants of Substrate/Inhibitor Binding to the Human and Rabbit Renal Organic Cation Transporters hOCT2 and rbOCT2

Cited by 89 publications

References 30 publications

Characterization of the Inhibitory Effects ofN-Butylpyridinium Chloride and Structurally Related Ionic Liquids on Organic Cation Transporters 1/2 and Human Toxic Extrusion Transporters 1/2-K In Vitro and In Vivo

Characterization of the Inhibitory Effects ofN-Butylpyridinium Chloride and Structurally Related Ionic Liquids on Organic Cation Transporters 1/2 and Human Toxic Extrusion Transporters 1/2-K In Vitro and In Vivo

Nonoisotopic Assay for the Presynaptic Choline Transporter Reveals Capacity for Allosteric Modulation of Choline Uptake

Mitigation of acute kidney injury by cell-cycle inhibitors that suppress both CDK4/6 and OCT2 functions

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