Proton pump inhibitors inhibit metformin uptake by organic cation transporters (OCTs)

Abstract

Metformin, an oral insulin-sensitizing drug, is actively transported into cells by organic cation transporters (OCT) 1, 2, and 3 (encoded by SLC22A1, SLC22A2, or SLC22A3), which are tissue specifically expressed at significant levels in various organs such as liver, muscle, and kidney. Because metformin does not undergo hepatic metabolism, drug-drug interaction by inhibition of OCT transporters may be important. So far, comprehensive data on the interaction of proton pump inhibitors (PPIs) with OCTs are missing although PPIs are frequently used in metformin-treated patients. Using in silico modeling and computational analyses, we derived pharmacophore models indicating that PPIs (i.e. omeprazole, pantoprazole, lansoprazole, rabeprazole, and tenatoprazole) are potent OCT inhibitors. We then established stably transfected cell lines expressing the human uptake transporters OCT1, OCT2, or OCT3 and tested whether these PPIs inhibit OCT-mediated metformin uptake in vitro. All tested PPIs significantly inhibited metformin uptake by OCT1, OCT2, and OCT3 in a concentration-dependent manner. Half-maximal inhibitory concentration values (IC(50)) were in the low micromolar range (3-36 µM) and thereby in the range of IC(50) values of other potent OCT drug inhibitors. Finally, we tested whether the PPIs are also transported by OCTs, but did not identify PPIs as OCT substrates. In conclusion, PPIs are potent inhibitors of the OCT-mediated metformin transport in vitro. Further studies are needed to elucidate the clinical relevance of this drug-drug interaction with potential consequences on metformin disposition and/or efficacy.

Figure 1. Pharmacophore models of OCT1, OCT2, and OCT3.

Ligand-based pharmacophore models were generated with igandScout using the 15 most potent OCT1, OCT2, or OCT3 inhibitors as training set and 5 PPIs (omeprazole, pantoprazole, lansoprazole, rabeprazole, tenatoprazole) as test set. The 5 PPIs are mapped to each pharmacophore model. Yellow sphere: hydrophobic interaction site (Hy), red sphere: H-bond acceptor site (HBA), green sphere: H-bond donor site (HBD); 2 stacked blue rings: aromatic ring. On the right, the centers of the pharmacophore features are redrawn and the interfeature distances are shown in Ångstroms.

Figure 2. Characterization of HEK cells stably transfected with cDNAs encoding human OCT1, OCT2, or OCT3.

A. Time-dependent uptake of 100 µM TEA, a prototypic OCT substrate, by HEK-OCT1, HEK-OCT2, or HEK-OCT3. Data are means ± SD of 3 determinations. B. Time-dependent uptake of 5 µM metformin by HEK-OCT1, HEK-OCT2, or HEK-OCT3 and assessment of non-specific uptake by performing uptake studies in the presence of 2 mM unlabeled MPP, a high-affinity OCT substrate. Data are means ± SD of 3 determinations. C. Confocal laser scanning micrographs of HEK-OCT1, HEK-OCT2, and HEK-OCT3 cells after incubation with the OCT isoform-specific antisera KEN, KEK, and CGR, respectively , . Bars, 20 µm. D. Immunoblot analyses of membrane fractions from HEK-OCT1, HEK-OCT2, and HEK-OCT3 cells in comparison with those from vector-transfected control cells (HEK-Co) using the OCT isoform-specific antisera KEN, KEK, and CGR, respectively.

Figure 3. Inhibition of OCT-mediated metformin uptake by proton pump inhibitors.

Uptake of 5 µM [¹⁴C]metformin was measured after 5 min (OCT1, OCT3) or 5 sec (OCT2) to remain within the linear uptake phase of each transporter. OCT-specific metformin uptake was obtained by subtracting metformin uptake in the presence of 2 mM unlabeled MPP from that in its absence. OCT-specific metformin uptake in the presence of different concentrations of omeprazole, pantoprazole, lansoprazole, rabeprazole, or tenatoprazole was then calculated as percentage of that in the absence of the respective PPI (100% uninhibited). Data points are the means ± SE of 3 individual experiments, with 3 parallel measurements performed in each individual experiment. Lines result from direct fits of the Hill equation to the data points.

Figure 4. Evaluation of proton pump inhibitors as transport substrates of OCTs.

Intracellular accumulation of 10 µM [¹⁴C]omeprazole, 10 µM [¹⁴C]pantoprazole, 5 µM lansoprazole, 5 µM rabeprazole, or 5 µM tenatoprazole by vector-transfected control HEK cells (white bars) or OCT-expressing HEK cells (black bars) was measured after 1 or 5 min of incubation. Data are means ± SD of 3 determinations.