Multidrug And Toxin Extrusion Protein 1 Research Articles

Inhibitory effect of flavonoids on multidrug and toxin extrusion protein 1 function: Implications for food/herb-drug interaction and drug-induced kidney injury.

Multidrug and toxin extrusion protein 1 (MATE1), an efflux transporter mainly expressed in renal proximal tubules, mediates the renal secretion of organic cationic drugs. The inhibition of MATE1 will impair the excretion of drugs into the tubular lumen, leading to the accumulation of nephrotoxic drugs in the kidney and consequently potentiating nephrotoxicity. Screening and identifying potent MATE1 inhibitors can predict or minimize the risk of drug-induced kidney injury. Flavonoids, a group of polyphenols commonly found in foodstuffs and herbal products, have been reported to cause transporter-mediated food/herb-drug interactions. Our objective was to investigate the inhibitory effects of flavonoids on MATE1 in vitro and in vivo and to assess the effects of flavonoids on cisplatin-induced kidney injury. Thirteen flavonoids exhibited significant transport activity inhibition (>50%) on MATE1 in MATE1-MDCK cells. Among them, the six strongest flavonoid inhibitors, including irisflorentin, silymarin, isosilybin, sinensetin, tangeretin, and nobiletin, markedly increased cisplatin cytotoxicity in these cells. In cisplatin-induced in vivo renal injury models, irisflorentin, isosilybin, and sinensetin also increased serum creatinine and blood urea nitrogen levels to different degrees, especially irisflorentin, which exhibited the most potent nephrotoxicity with cisplatin. The pharmacophore model indicated that the hydrogen bond acceptors at the 3, 5, and 7 positions may play a critical role in the inhibitory effect of flavonoids on MATE1. Our findings provide helpful information for predicting the potential risks of flavonoid-containing food/herb-drug interactions and avoiding the exacerbation of drug-induced kidney injury via MATE1 mediation.

Inhibition of human drug transporter activities by succinate dehydrogenase inhibitors

Succinate dehydrogenase inhibitors (SDHIs) are widely-used fungicides, to which humans are exposed and for which putative health risks are of concern. In order to identify human molecular targets for these environmental chemicals, the interactions of 15 SDHIs with activities of main human drug transporters implicated in pharmacokinetics were investigated in vitro. 5/15 SDHIs, i.e., benzovindiflupyr, bixafen, fluxapyroxad, pydiflumetofen and sedaxane, were found to strongly reduce activity of the renal organic anion transporter (OAT) 3, in a concentration-dependent manner (with IC50 values in the 1.0–3.9 μM range), without however being substrates for OAT3. Moreover, these 5/15 SDHIs decreased the membrane transport of estrone-3 sulfate, an endogenous substrate for OAT3, and sedaxane was predicted to inhibit in vivo OAT3 activity in response to exposure to the acceptable daily intake (ADI) dose. In addition, pydiflumetofen strongly inhibited the renal organic cation transporter (OCT) 2 (IC50 = 2.0 μM) and benzovindiflupyr the efflux pump breast cancer resistance protein (BCRP) (IC50 = 3.9 μM). Other human transporters, including organic anion transporting polypeptide (OATP) 1B1 and OATP1B3 as well as multidrug and toxin extrusion protein (MATE) 1 and MATE2-K were moderately or weakly inhibited by SDHIs, whereas P-glycoprotein, multidrug resistance-associated protein (MRP), OCT1 and OAT1 activities were not or only marginally impacted. Then, some human drug transporters, especially OAT3, constitute molecular targets for SDHIs. This could have toxic consequences, notably with respect to levels of endogenous compounds and metabolites substrates for the considered transporters or to potential SDHI-drug interactions. This could therefore contribute to putative health risk of these fungicides.

Identification and characterization of an endogenous biomarker of the renal vectorial transport (OCT2-MATE1).

The renal tubular organic cation transporter 2 (OCT2) and multidrug and toxin extrusion protein 1 (MATE1) mediate the vectorial elimination of many drugs and toxins from the kidney, and endogenous biomarkers for vectorial transport (OCT2-MATE1) would allow more accurate drug dosing and help to characterize drug-drug interactions and toxicity. Human serum uptake in OCT2-overexpressing cells and metabolomics analysis were carried out. Potential biomarkers were verified in vitro and in vivo. The specificity of biomarkers was validated in renal transporter overexpressing cells and the sensitivity was investigated by Km . The results showed that the uptake of thiamine, histamine, and 5-hydroxytryptamine was significantly increased in OCT2-overexpressing cells. In vitro assays confirmed that thiamine, histamine, and 5-hydroxytryptamine were substrates of both OCT2 and MATE1. In vivo measurements indicated that the serum thiamine level was increased significantly in the presence of the rOCT2 inhibitor cimetidine, and the level in renal tissue was increased significantly by the rMATE1 inhibitor pyrimethamine. There were no significant changes in the uptake or efflux of thiamine in cell lines overexpressed OAT1, OAT2, OAT3, MRP4, organic anion transporting polypeptide 4C1, P-gp, peptide transporter 2, urate transporter 1, and OAT4. The Km for thiamine with OCT2 and MATE1 were 71.2 and 10.8μM, respectively. In addition, the cumulative excretion of thiamine at 2 and 4h was strongly correlated with metformin excretion (R2 >0.6). Thus, thiamine is preferentially secreted by the OCT2 and MATE1 in renal tubules and can provide a reference value for evaluating the function of the renal tubular OCT2-MATE1.

Regulation of Transporters for Organic Cations by High Glucose.

Endogenous positively charged organic substances, including neurotransmitters and cationic uremic toxins, as well as exogenous organic cations such as the anti-diabetic medication metformin, serve as substrates for organic cation transporters (OCTs) and multidrug and toxin extrusion proteins (MATEs). These proteins facilitate their transport across cell membranes. Vectorial transport through the OCT/MATE axis mediates the hepatic and renal excretion of organic cations, regulating their systemic and local concentrations. Organic cation transporters are part of the remote sensing and signaling system, whose activity can be regulated to cope with changes in the composition of extra- and intracellular fluids. Glucose, as a source of energy, can also function as a crucial signaling molecule, regulating gene expression in various organs and tissues. Its concentration in the blood may fluctuate in specific physiological and pathophysiological conditions. In this work, the regulation of the activity of organic cation transporters was measured by incubating human embryonic kidney cells stably expressing human OCT1 (hOCT1), hOCT2, or hMATE1 with high glucose concentrations (16.7 mM). Incubation with this high glucose concentration for 48 h significantly stimulated the activity of hOCT1, hOCT2, and hMATE1 by increasing their maximal velocity (Vmax), but without significantly changing their affinity for the substrates. These effects were independent of changes in osmolarity, as the addition of equimolar concentrations of mannitol did not alter transporter activity. The stimulation of transporter activity was associated with a significant increase in transporter mRNA expression. Inhibition of the mechanistic target of rapamycin (mTOR) kinase with Torin-1 suppressed the transporter stimulation induced by incubation with 16.7 mM glucose. Focusing on hOCT2, it was shown that incubation with 16.7 mM glucose increased hOCT2 protein expression in the plasma membrane. Interestingly, an apparent trend towards higher hOCT2 mRNA expression was observed in kidneys from diabetic patients, a pathology characterized by high serum glucose levels. Due to the small number of samples from diabetic patients (three), this observation must be interpreted with caution. In conclusion, incubation for 48 h with a high glucose concentration of 16.7 mM stimulated the activity and expression of organic cation transporters compared to those measured in the presence of 5.6 mM glucose. This stimulation by a diabetic environment could increase cellular uptake of the anti-diabetic drug metformin and increase renal tubular secretion of organic cations in an early stage of diabetes.

#3475 NOVEL OPTIMISED PROTOCOL FOR ISOLATION OF PTEC AND SYSTEMATIC CHARACTERISATION FROM HUMAN KIDNEY BIOPSY

Abstract Background and Aims Human kidneys have a role in water homeostasis, acid-base control, reabsorption of compounds, and secretion of xenobiotics and endogenous metabolites, exposing them to substances that could cause harm. This results in an alarming number of acute kidney injuries (AKI) worldwide, estimated at 13%. Furthermore, one-quarter of hospitalised cases are due to drug-induced AKI [1]. Current methods for nephrotoxicity assays are based on animal testing and/or the use of simple human cell lines. Meta-analyses show that we can correctly predict human drug responses in only 10–50% [2]. Our work aimed to develop a novel and optimised protocol for isolating proximal tubular epithelial cells (PTEC) from human kidney biopsy to aid future research regarding AKI and nephrotoxicity studies. Method Isolation and cultivation of primary human adult PTEC obtained with biopsy during the regular diagnostic procedure was performed. We used a protocol consisting of micro-dissection of a tissue sample to get ∼1 mm3 fragments, enzymatic dissociation with 0.2% collagenase type 1, and use of selective culture media (Advanced DMEM/F12 with added insulin, transferrin, and selenite (all three together termed ITS), epidermal growth factor (EGF), and hydrocortisone). Light microscopy was used for morphologic characterisation. Some cells were cultured on Transwell inserts, and the transepithelial electric resistance (TEER) was measured in mature cells that formed a confluent culture. For phenotypic characterisation, several markers characteristic of PTEC were chosen [3], and immunocytochemical staining was performed using a fluorescent microscope to evaluate the PTEC phenotype. Results Following the described protocol resulted in isolating cells that formed first colonies after 24 h. Using light microscopy, the cells exhibited a cobblestone appearance, reached confluence after eight days, and showed dome (hemicysts) formation after 13 days. TEER reached 169 Ω/cm2 after 14 days. The isolated cells were marked positive using immunocytochemistry for sodium-glucose cotransporter 2 (SGLT2), multidrug-resistant protein 4 (MRP4), organic anionic transporter 1 and 3 (OAT1 and OAT3), organic cationic transporter 2 (OCT2), p-glycoprotein (p-gp), multidrug and toxin extrusion protein 1 (MATE1), and N-cadherin. Conclusion In this study, we developed a protocol for isolating and cultivating primary human PTEC from biopsy samples. To the best of our knowledge, we have performed the most extensive systematic characterisation following the isolation of PTEC from kidney biopsy reported to date.

MATE1 expression in the cochlea and its potential involvement in cisplatin cellular uptake and ototoxicity

Background Cisplatin appears to enter the cochlear cells through the organic cation transporter 2 (OCT2). There is recent evidence that multidrug and toxin extrusion protein 1 (MATE1) is involved in cisplatin-induced nephrotoxicity. Its presence and role in the ear are unknown. Aims/Objectives Evaluate the presence and localization of MATE1, and determine the localization of OCT2, in the cochlea. Evaluate cisplatin uptake with regard to MATE1 and OCT2 expression. Material and Methods Murine cochlear explants and paraffin-embedded cochleae were evaluated with immunohistochemistry for OCT2 and MATE1. Explant cultures were also treated with Texas Red cisplatin to determine their cellular uptake. Results MATE1 is present in the cochlea. Most intense labeling of MATE1 and OCT2 was seen in the outer hair cells (OHCs) and pillar cells, respectively. Both transporters were observed in the spiral ganglion neurons and stria vascularis. Expression levels of OCT2 and MATE1 decreased following cisplatin exposure. Texas Red cisplatin staining was strong in OHCs and pillar cells. Conclusions and Significance To the best of our knowledge, this is the first study demonstrating the presence and localization of MATE1 in the cochlea. OCT2 labeling was seen in pillar cells. Consistently, OHCs and pillar cells uptake Texas Red cisplatin.

1136. No Change of Pharmacokinetics of Metformin by Concomitant Use of Ensitrelvir

Abstract Background Ensitrelvir is a novel oral SARS-CoV-2 3C-like protease inhibitor, and under late clinical development stage for COVID-19 diseases. Ensitrelvir exhibited an inhibition potency for organic cation transporter 1 (OCT1) and multidrug and toxin extrusion protein 1 (MATE1) in in vitro study and clinical drug-drug interaction (DDI) study is required judging from DDI guidance. Metformin is widely used for treatment of diabetes, and is a sensitive substrate for OCT1 and MATE1. We evaluated the effect of ensitrelvir on the pharmacokinetics (PK) of metformin with physiologically-based pharmacokinetic (PBPK) modeling and simulation and clinical DDI study. Methods The PBPK model of ensitrelvir was developed based on the physicochemical parameters, in vitro transporter inhibition parameters, and estimated PK parameters for human. DDI simulations between ensitrelvir and metformin were performed. Simcyp PBPK Simulator (Version 20, Certara UK Limited, UK) was used to develop PBPK model and simulate the DDIs. The in vitro 50% inhibitory concentration (IC50) values of each transporter were used as inhibition constant (Ki) for DDI simulations. Based on the PBPK analysis, the clinical DDI study planed. Results PBPK analysis: As the result of DDI simulation, ensitrelvir increased the area under the curve (AUC) of metformin by 12%. The result suggests that in vivo DDI potency of ensitrelvir via inhibition of OCT1 or MATE1 would be low at a single dose of ensitrelvir 1000 mg. Clinical DDI study: The plasma concentration-time profile of metformin and ensitrelvir were monitored after 96 hours from a single dose of metformin with or without ensitrelvir. Ensitrelvir does not have effect on the PK of metformin (a geometric mean of AUC ratio was 1.02, Japanese healthy subjects, N=14), suggesting no MATE1 and OCT1 inhibition by ensitrelvir at a clinical dose. The PBPK analysis could well predict the clinical DDI study result. Conclusion The results of PBPK analysis and the clinical DDI study suggest that no OCT1 and MATE1 inhibition by ensitrelvir is in the clinical dose. Therefore, ensitrelvir does not have a clinically meaningful effect on the pharmacokinetic profile of OCT1 and/or MATE1 substrates including metformin. Disclosures Kana Horiuchi, Shionogi & Co., Ltd.: employee Hiroki Koshimichi, n/a, Shionogi & Co., Ltd.: employee Takanobu Matsuzaki, Ph.D., Shionogi & Co., Ltd.: Employee Ryosuke Shimizu, Shionogi & Co., Ltd.: employee Shinpei Yoshida, Ph.D., Shionogi & Co., Ltd.: employee Ryuji Kubota, Ph.D., Shionogi & Co., Ltd.: employee|Shionogi & Co., Ltd.: employee Shingo Sakamoto, n/a, Shionogi & Co., Ltd.: employee.

Association of SLC22A1, SLC22A2, SLC47A1, and SLC47A2 Polymorphisms with Metformin Efficacy in Type 2 Diabetic Patients.

Response to metformin, first-line therapy for type 2 diabetes mellitus (T2DM), exists interindividual variation. Considering that transporters belonging to the solute carrier (SLC) superfamily are determinants of metformin pharmacokinetics, we evaluated the effects of promoter variants in organic cation transporter 1 (OCT1) (SLC22A1 rs628031), OCT2 (SLC22A2 rs316019), multidrug and toxin extrusion protein 1 (MATE1) (SLC47A1 rs2289669), and MATE2 (SLC47A2 rs12943590) on the variation in metformin response. The glucose-lowering effects and improvement of insulin resistance of metformin were assessed in newly diagnosed, treatment-naive type 2 diabetic patients of Han nationality in Chaoshan China (n = 93) receiving metformin. Fasting plasma glucose (FPG), fasting insulin (FINS), glycated hemoglobin A1 (HbA1C), homeostasis model assessment-insulin sensitivity (HOMA-IS), and homeostasis model assessment-insulin resistance (HOMA-IR) were the main metformin efficacy measurements. There were significant correlations between both SLC47A1 rs2289669 and SLC47A2 rs12943590 and the efficacy of metformin in individuals with T2DM. In normal weight T2DM patients, significant associations between the AA and GG genotypes of the rs2289669 variant of SLC47A1 and a greater reduction in FINS and HOMA-IR were detected. A significant correlation was observed between the AG genotype of the rs12943590 polymorphism of SLC47A2 and a greater reduction in HOMA-IR. Gene–environment interaction analysis showed that in the FINS interaction model, the second-order of dose30_g-SLC47A2 rs12943590 was statistically significant. The variants of SLC47A1 rs2289669 and SLC47A2 rs12943590 could be predictors of insulin resistance in type 2 diabetic patients treated with metformin. The second-order interaction of dose30_g-SLC47A2 rs12943590 may have a significant effect on FINS in patients with T2DM on metformin treatment. These findings suggest that promoter variants of SLC47A1 and SLC47A2 are important determinants of metformin transport and response in type 2 diabetes mellitus.

Renal Transporter-Mediated Drug-Biomarker Interactions of the Endogenous Substrates Creatinine and N1 -Methylnicotinamide: A PBPK Modeling Approach.

Endogenous biomarkers for transporter-mediated drug-drug interaction (DDI) predictions represent a promising approach to facilitate and improve conventional DDI investigations in clinical studies. This approach requires high sensitivity and specificity of biomarkers for the targets of interest (e.g., transport proteins), as well as rigorous characterization of their kinetics, which can be accomplished utilizing physiologically-based pharmacokinetic (PBPK) modeling. Therefore, the objective of this study was to develop PBPK models of the endogenous organic cation transporter (OCT)2 and multidrug and toxin extrusion protein (MATE)1 substrates creatinine and N1 -methylnicotinamide (NMN). Additionally, this study aimed to predict kinetic changes of the biomarkers during administration of the OCT2 and MATE1 perpetrator drugs trimethoprim, pyrimethamine, and cimetidine. Whole-body PBPK models of creatinine and NMN were developed utilizing studies investigating creatinine or NMN exogenous administration and endogenous synthesis. The newly developed models accurately describe and predict observed plasma concentration-time profiles and urinary excretion of both biomarkers. Subsequently, models were coupled to the previously built and evaluated perpetrator models of trimethoprim, pyrimethamine, and cimetidine for interaction predictions. Increased creatinine plasma concentrations and decreased urinary excretion during the drug-biomarker interactions with trimethoprim, pyrimethamine, and cimetidine were well-described. An additional inhibition of NMN synthesis by trimethoprim and pyrimethamine was hypothesized, improving NMN plasma and urine interaction predictions. To summarize, whole-body PBPK models of creatinine and NMN were built and evaluated to better assess creatinine and NMN kinetics while uncovering knowledge gaps for future research. The models can support investigations of renal transporter-mediated DDIs during drug development.

Transcriptional Regulation of Solute Carrier (SLC) Drug Transporters.

Facilitated transport is necessitated for large size, charged, and/or hydrophilic drugs to move across the membrane. The drug transporters in the solute carrier (SLC) superfamily, mainly including organic anion-transporting polypeptides (OATPs), organic anion transporters (OATs), organic cation transporters (OCTs), organic cation/carnitine transporters (OCTNs), peptide transporters (PEPTs), and multidrug and toxin extrusion proteins (MATEs), are critical facilitators of drug transport and distribution in human body. The expression of these SLC drug transporters is found in tissues throughout the body, with high abundance in the epithelial cells of major organs for drug disposition, such as intestine, liver, and kidney. These SLC drug transporters are clinically important in drug absorption, metabolism, distribution, and excretion. The mechanisms underlying their regulation have been revealing in recent years. Epigenetic and nuclear receptor-mediated transcriptional regulation of SLC drug transporters have particularly attracted much attention. This review focuses on the transcriptional regulation of major SLC drug transporter genes. Revealing the mechanisms underlying the transcription of those critical drug transporters will help us understand pharmacokinetics and pharmacodynamics, ultimately improving drug therapeutic effectiveness while minimizing drug toxicity. Significance Statement It has become increasingly recognized that solute carrier (SLC) drug transporters play a crucial, and sometimes determinative, role in drug disposition and response, which is reflected in decision-making during not only clinical drug therapy but also drug development. Understanding the mechanisms accounting for the transcription of these transporters is critical to interpret their abundance in various tissues under different conditions, which is necessary to clarify the pharmacological response, adverse effects, and drug-drug interactions for clinically used drugs.

Evaluation of drug-drug interaction potential for pemigatinib using physiologically based pharmacokinetic modeling.

Pemigatinib is a potent inhibitor of fibroblast growth factor receptor being developed for oncology indications. It is primarily metabolized by cytochrome P450 (CYP) 3A4, and the ratio of estimated concentration over concentration required for 50% inhibition ratio for pemigatinib as an inhibitor of P‐glycoprotein (P‐gp), organic cation transporter‐2 (OCT2), and multidrug and toxin extrusion protein‐1 (MATE1) exceeds the cutoff values established in regulatory guidance. A Simcyp minimal physiologically based pharmacokinetic (PBPK) with advanced dissolution, absorption, and metabolism absorption model for pemigatinib was developed and validated using observed clinical pharmacokinetic (PK) data and itraconazole/rifampin drug–drug interaction (DDI) data. The model accurately predicted itraconazole DDI (approximate 90% area under the plasma drug concentration–time curve [AUC] and approximate 20% maximum plasma drug concentration [Cmax] increase). The model underpredicted rifampin induction by 100% (approximate 6.7‐fold decrease in AUC and approximate 2.6‐fold decrease in Cmax in the DDI study), presumably reflecting non‐CYP3A4 mechanisms being impacted. The verified PBPK model was then used to predict the effect of other CYP3A4 inhibitors/inducers on pemigatinib PK and pemigatinib as an inhibitor of P‐gp or OCT2/MATE1 substrates. The worst‐case scenario DDI simulation for pemigatinib as an inhibitor of P‐gp or OCT2/MATE1 substrates showed only a modest DDI effect. The recommendation based on this simulation and clinical data is to reduce pemigatinib dose for coadministration with strong and moderate CYP3A4 inhibitors. No dose adjustment is required for weak CYP3A4 inhibitors. The coadministration of strong and moderate CYP3A4 inducers with pemigatinib should be avoided. PBPK modeling suggested no dose adjustment with P‐gp or OCT2/MATE1 substrates.

Interactions Between Meropenem and Renal Drug Transporters.

Meropenem is a carbapenem antibiotic and is commonly used with other antibiotics for the treatment of bacterial infections. It is primarily eliminated renally by glomerular filtration and renal tubular secretion. This study aimed to evaluate the roles of renal uptake and efflux transporters in the excretion of meropenem and potential drug interactions mediated by renal drug transporters. Uptake and inhibition studies were conducted in human embryonic kidney 293 cells stably transfected with Organic Anion Transporter (OAT) 1, OAT3, Multidrug and Toxin Extrusion Protein (MATE) 1, and MATE2K, as well as membrane vesicles containing breast cancer resistance-related protein (BCRP), multidrug resistance protein 1 (MDR1), and Multidrug Resistance-associated Protein 2 (MRP2). Probenecid and piperacillin were used to assess potential drug interactions with meropenem in rats. We observed that meropenem was a low-affinity substrate of OAT1/3 and had a weak inhibitory effect on OAT1/3 and MATE2K. BCRP, MDR1, MRP2, MATE1, and MATE2K could not mediate renal excretion of meropenem. Moreover, meropenem was not an inhibitor of BCRP, MDR1, MRP2, or MATE1. Among five tested antibiotics, moderate inhibition on OAT3-mediated meropenem uptake was observed for linezolid (IC50 value was 69.2 μM), weak inhibition was observed for piperacillin, benzylpenicillin, and tazobactam (IC50 values were 282.2, 308.0 and 668.1 μM, respectively), and no inhibition was observed for sulbactam. Although piperacillin had a relatively high drug-drug interaction index (ratio of maximal unbound plasma concentration to IC50 was 1.42) in vitro, no meaningful impact was reported on the pharmacokinetics of meropenem in rats. Our results indicated that clinically significant interactions between meropenem and these five antibiotics are low.

Localization of Xenobiotic Transporters Expressed at the Human Blood-Testis Barrier.

The blood-testis barrier (BTB) is formed by basal tight junctions between adjacent Sertoli cells (SCs) of the seminiferous tubules and acts as a physical barrier to protect developing germ cells in the adluminal compartment from reproductive toxicants. Xenobiotics, including antivirals, male contraceptives, and cancer chemotherapeutics, are known to cross the BTB, although the mechanisms that permit barrier circumvention are generally unknown. This study used immunohistological staining of human testicular tissue to determine the site of expression for xenobiotic transporters that facilitate transport across the BTB. Organic anion transporter (OAT) 1, OAT2, and organic cation transporter, novel (OCTN) 1 primarily localized to the basal membrane of SCs, whereas OCTN2, multidrug resistance protein (MRP) 3, MRP6, and MRP7 localized to SC basal membranes and peritubular myoid cells (PMCs) surrounding the seminiferous tubules. Concentrative nucleoside transporter (CNT) 2 localized to Leydig cells (LCs), PMCs, and SC apicolateral membranes. Organic cation transporter (OCT) 1, OCT2, and OCT3 mostly localized to PMCs and LCs, although there was minor staining in developing germ cells for OCT3. Organic anion transporting polypeptide (OATP) 1A2, OATP1B1, OATP1B3, OATP2A1, OATP2B1, and OATP3A1-v2 localized to SC basal membranes with diffuse staining for some transporters. Notably, OATP1C1 and OATP4A1 primarily localized to LCs. Positive staining for multidrug and toxin extrusion protein (MATE) 1 was only observed throughout the adluminal compartment. Definitive staining for CNT1, OAT3, MATE2, and OATP6A1 was not observed. The location of these transporters is consistent with their involvement in the movement of xenobiotics across the BTB. Altogether, the localization of these transporters provides insight into the mechanisms of drug disposition across the BTB and will be useful in developing tools to overcome the pharmacokinetic and pharmacodynamic difficulties presented by the BTB. SIGNIFICANCE STATEMENT: Although the total mRNA and protein expression of drug transporters in the testes has been explored, the localization of many transporters at the blood-testis barrier (BTB) has not been determined. This study applied immunohistological staining in human testicular tissues to identify the cellular localization of drug transporters in the testes. The observations made in this study have implications for the development of drugs that can effectively use transporters expressed at the basal membranes of Sertoli cells to bypass the BTB.

Prediction of drug\u2013drug interaction potential mediated by transporters between dasatinib and metformin, pravastatin, and rosuvastatin using physiologically based pharmacokinetic modeling

PurposeRecent in vitro studies demonstrated that dasatinib inhibits organic cation transporter 2 (OCT2), multidrug and toxin extrusion proteins (MATEs), and organic anion transporting polypeptide 1B1/1B3 (OATP1B1/1B3). We developed a physiologically based pharmacokinetic (PBPK) model to assess drug–drug interaction (DDI) potential between dasatinib and known substrates for these transporters in a virtual population.MethodsThe dasatinib PBPK model was constructed using Simcyp® Simulator by combining its physicochemical properties, in vitro data, in silico predictions, and pharmacokinetic (PK) results from clinical studies. Model validation against three independent clinical trials not used for model development included dasatinib DDI studies with ketoconazole, rifampin, and simvastatin. The validated model was used to simulate DDIs of dasatinib and known substrates for OCT2 and MATEs (metformin) and OATP1B1/1B3 (pravastatin and rosuvastatin).ResultsSimulations of metformin PK in the presence and absence of dasatinib, using inhibitor constant (Ki) values measured in vitro, produced estimated geometric mean ratios (GMRs) of the maximum observed concentration (Cmax) and area under the concentration–time curve (AUC) of 1.05 and 1.06, respectively. Sensitivity analysis showed metformin exposure increased < 30% in both AUC and Cmax when dasatinib Ki was reduced by tenfold for OCT2 and MATEs simultaneously, and < 40% with a 20-fold Ki reduction. The estimated GMRs of Cmax and AUC for pravastatin and rosuvastatin with co-administration of dasatinib were unity (1.00).ConclusionsThis PBPK model accurately described the observed PK profiles of dasatinib. The validated PBPK model predicts low risk of clinically significant DDIs between dasatinib and metformin, pravastatin, or rosuvastatin.

The L125F MATE1 variant enriched in populations of Amerindian origin is associated with increased plasma levels of metformin and lactate

Genetic factors that affect variability in metformin response have been poorly studied in the Latin American population, despite its being the initial drug therapy for type 2 diabetes, one of the most prevalent diseases in that region. Metformin pharmacokinetics is carried out by members of the membrane transporters superfamily (SLCs), being the multidrug and toxin extrusion protein 1 (MATE1), one of the most studied. Some genetic variants in MATE1 have been associated with reduced in vitro metformin transport. They include rs77474263 p.[L125F], a variant present at a frequency of 13.8% in Latin Americans, but rare worldwide (less than 1%). Using exome sequence data and TaqMan genotyping, we revealed that the Mexican population has the highest frequency of this variant: 16% in Mestizos and 27% in Amerindians, suggesting a possible Amerindian origin. To elucidate the metformin pharmacogenetics, a children cohort was genotyped, allowing us to describe, for the first time, a MATE1 rs77474263 TT homozygous individual. An additive effect of the L125F variant was observed on blood metformin accumulation, revealing the highest metformin and lactate serum levels in the TT homozygote, and intermediate metformin values in the heterozygotes. Moreover, a molecular dynamics analysis suggested that the genetic variant effect on metformin efflux could be due to a decreased protein permeability. We conclude that pharmacogenetics could be useful in enhancing metformin pharmacovigilance in populations having a high frequency of the risk genotype, especially considering that these populations also have a higher susceptibility to the diseases for which metformin is the first-choice drug.

Islatravir Is Not Expected to Be a Victim or Perpetrator of Drug-Drug Interactions via Major Drug-Metabolizing Enzymes or Transporters.

Islatravir (MK-8591) is a nucleoside reverse transcriptase translocation inhibitor in development for the treatment and prevention of HIV-1. The potential for islatravir to interact with commonly co-prescribed medications was studied in vitro. Elimination of islatravir is expected to be balanced between adenosine deaminase–mediated metabolism and renal excretion. Islatravir did not inhibit uridine diphosphate glucuronosyltransferase 1A1 or cytochrome p450 (CYP) enzymes CYP1A2, 2B6, 2C8, 2C9, 2C19, 2D6, or 3A4, nor did it induce CYP1A2, 2B6, or 3A4. Islatravir did not inhibit hepatic transporters organic anion transporting polypeptide (OATP) 1B1, OATP1B3, organic cation transporter (OCT) 1, bile salt export pump (BSEP), multidrug resistance-associated protein (MRP) 2, MRP3, or MRP4. Islatravir was neither a substrate nor a significant inhibitor of renal transporters organic anion transporter (OAT) 1, OAT3, OCT2, multidrug and toxin extrusion protein (MATE) 1, or MATE2K. Islatravir did not significantly inhibit P-glycoprotein and breast cancer resistance protein (BCRP); however, it was a substrate of BCRP, which is not expected to be of clinical significance. These findings suggest islatravir is unlikely to be the victim or perpetrator of drug-drug interactions with commonly co-prescribed medications, including statins, diuretics, anti-diabetic drugs, proton pump inhibitors, anticoagulants, benzodiazepines, and selective serotonin reuptake inhibitors.

Assessment of the Effect of Pyrimethamine, a Potent Inhibitor of Multidrug and Toxin Extrusion Protein 1/2K, on the Pharmacokinetics of Gefapixant (MK-7264), a P2X3 Receptor Antagonist.

Gefapixant (MK-7264, AF-219) is a first-in-class P2X3 antagonist in development for refractory or unexplained chronic cough. Gefapixant is primarily cleared by renal excretion. To assess the importance of the multidrug and toxin extrusion protein 1 (MATE1) and MATE2K transporters in the elimination of gefapixant, a drug-drug interaction study was conducted evaluating the effect of coadministration of a single dose of pyrimethamine, a competitive inhibitor of MATE1 and MATE2K, on the single-dose pharmacokinetics of gefapixant in healthy participants. Safety and tolerability were also assessed. In this open-label, 2-period, fixed-sequence study, a 45-mg dose of gefapixant was administered to 12 participants in period 1. After a 7-day washout, a 50-mg dose of pyrimethamine was administered 3hours before a 45-mg dose of gefapixant in period 2. Compared with the administration of gefapixant alone, concomitant dosing of gefapixant with pyrimethamine increased the total gefapixant plasma exposure (area under the plasma concentration-time curve from time 0 to infinity) by 24%, reduced gefapixant renal clearance by 30%, and increased gefapixant mean terminal half-life from 7.7 to 10.3hours. The most frequently reported adverse events were dysgeusia, hypogeusia, and dry mouth; all adverse events were considered of mild intensity and resolved by the end of the study. These results support that MATE1 and/or MATE2K contribute to the renal clearance of gefapixant, but the effect of inhibition of these transporters on gefapixant pharmacokinetics is not considered clinically meaningful.

In Vitro Inhibition of Renal OCT2 and MATE1 Secretion by Antiemetic Drugs.

The organic cation transporter 2 (OCT2) and multidrug and toxin extrusion protein 1 (MATE1) mediate the renal secretion of drugs. Recent studies suggest that ondansetron, a 5-HT3 antagonist drug used to prevent nausea and vomiting, can inhibit OCT2- and MATE1-mediated transport. The purpose of this study was to test the ability of five 5-HT3 antagonist drugs to inhibit the OCT2 and MATE1 transporters. The transport of the OCT2/MATE1 probe substrate ASP+ was assessed using two models: (1) HEK293 kidney cells overexpressing human OCT2 or MATE1, and (2) MDCK cells transfected with human OCT2 and MATE1. In HEK293 cells, the inhibition of ASP+ uptake by OCT2 listed in order of potency was palonosetron (IC50: 2.6 μM) > ondansetron > granisetron > tropisetron > dolasetron (IC50: 85.4 μM) and the inhibition of ASP+ uptake by MATE1 in order of potency was ondansetron (IC50: 0.1 μM) > palonosetron = tropisetron > granisetron > dolasetron (IC50: 27.4 μM). Ondansetron (0.5–20 μM) inhibited the basolateral-to-apical transcellular transport of ASP+ up to 64%. Higher concentrations (10 and 20 μM) of palonosetron, tropisetron, and dolasetron similarly reduced the transcellular transport of ASP+. In double-transfected OCT2-MATE1 MDCK cells, ondansetron at concentrations of 0.5 and 2.5 μM caused significant intracellular accumulation of ASP+. Taken together, these data suggest that 5-HT3 antagonist drugs may inhibit the renal secretion of cationic drugs by interfering with OCT2 and/or MATE1 function.

Drug–drug interaction potential and clinical pharmacokinetics of enerisant, a novel potent and selective histamine H3 receptor antagonist

We evaluated the in vitro drug–drug interaction (DDI) potential of enerisant (TS-091), a histamine H3 receptor antagonist/inverse agonist, mediated by cytochrome P450 (CYP) and transporters, as well as the pharmacokinetics of enerisant in healthy male subjects. Enerisant did not inhibit CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1, or CYP3A4 and did not induce CYP1A2, CYP2B6, or CYP3A4. Enerisant inhibited organic cation transporter 2, multidrug and toxin extrusion protein (MATE) 1, and MATE2-K, but not P-glycoprotein (P-gp), breast cancer resistance protein, organic anion transporting polypeptide (OATP) 1B1, OATP1B3, organic anion transporter (OAT) 1, or OAT3. Enerisant was a substrate for P-gp, but not for eight other transporters. In healthy male subjects, enerisant was rapidly absorbed after oral administration, and the plasma concentration increased dose-dependently. The urinary excretion of enerisant within 48 h after administration was 64.5% to 89.9% of the dose, indicating that most of the absorbed enerisant was excreted in the urine without being metabolized. Based on the plasma concentrations at the estimated clinical dose, enerisant is unlikely to cause CYP-mediated, clinically relevant DDI. Although the possibility of transporter-mediated, clinically relevant DDI cannot be ruled out, there is little or no risk of side effects.

Divergent Regulation of OCT and MATE Drug Transporters by Cadmium Exposure

Coordinated transcellular transport by the uptake via organic cation transporters (OCTs) in concert with the efflux via multidrug and toxin extrusion proteins (MATEs) is an essential system for hepatic and renal drug disposition. Despite their clinical importance, the regulation of OCTs and MATEs remains poorly characterized. It has been reported that cadmium (Cd2+) increase the activities of OCTs while being a substrate of MATEs. Here, we found that human (h) OCT2 protein, as compared with hMATE1, was more active in trafficking between the plasma membrane and cytoplasmic storage pool. Cd2+ exposure could significantly enhance the translocation of hOCT2 and hOCT1, but not hMATE1, to the plasma membrane. We further identified that candesartan, a widely prescribed angiotensin II receptor blocker, behaved similarly toward OCT2 and MATE1 as Cd2+ did. Importantly, Cd2+ and candesartan treatments could lead to an enhanced accumulation of metformin, which is a well-characterized substrate of OCTs/MATEs, in mouse kidney and liver, respectively. Altogether, our studies have uncovered possible divergent regulation of OCTs and MATEs by certain xenobiotics, such as Cd2+ and candesartan due to the different cellular trafficking of these two families of transporter proteins, which might significantly affect drug disposition in the liver and kidney.