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Aristolochic acid (AA) ingestion causes Balkan nephropathy, characterized by tubular injury and progression to chronic kidney disease (CKD). AA is taken up by proximal tubule cells via organic anion transport and induces p21-mediated DNA damage response, but little is known about dietary modulating factors. Western diet (WD) is rich in saturated fats and sugars and can promote metabolic disorders and CKD progression. Here, we determined the impact of WD on AA-induced kidney injury. Five-week-old male C57BL/6J mice were fed WD or normal chow (NC) for 8 wk, followed by administration of AA every 3 days for 3 wk. Measurements were performed after the last injection and following a 3-wk recovery. Independent of dosing AA by body weight (3 mg/kg/day) or same dose/mouse (0.1125 mg/day), the AA-induced increase in plasma creatinine and reduction of hematocrit were greater in WD versus NC. This was associated with increased kidney gene expression in WD vs. NC of markers of DNA damage (p21), injury (Kim1 and Ngal), and inflammation (Tnfa) and kidney fibrosis staining. WD alone increased fractional excretion of indoxyl sulfate by 7.5-fold, indicating enhanced kidney organic anion transport. Kidney proteomics identified further WD-induced changes that could increase kidney sensitivity to AA and contribute to the altered response to AA including weakening of energy metabolism, potentiation of immune and infection pathways, and disruption in RNA regulation. In conclusion, WD can increase the susceptibility of mice to Balkan nephropathy, possibly in part through facilitating kidney uptake of the organic anion AA. NEW & NOTEWORTHY: This study shows that a Western diet (WD) aggravates a murine model of Balkan nephropathy induced by the application of the organic anion and nephrotoxin aristolochic acid (AA). Mechanistically, this may involve WD-induced kidney organic anion secretion, which can facilitate the AA uptake into proximal tubular cells and thereby contribute to the injury. Kidney proteomics identified further changes induced by feeding a WD that could have increased the sensitivity of the kidney to stress and injury. [ABSTRACT FROM AUTHOR]

The solute carrier 17 (SLC17) family contains anion transporters that accumulate neurotransmitters in secretory vesicles, remove carboxylated monosaccharides from lysosomes, or extrude organic anions from the kidneys and liver. We combined classical molecular dynamics simulations, Markov state modeling and hybrid first principles quantum mechanical/classical mechanical (QM/MM) simulations with experimental approaches to describe the transport mechanisms of a model bacterial protein, the d-galactonate transporter DgoT, at atomic resolution. We found that protonation of D46 and E133 precedes galactonate binding and that substrate binding induces closure of the extracellular gate, with the conserved R47 coupling substrate binding to transmembrane helix movement. After isomerization to an inward-facing conformation, deprotonation of E133 and subsequent proton transfer from D46 to E133 opens the intracellular gate and permits galactonate dissociation either in its unprotonated form or after proton transfer from E133. After release of the second proton, apo DgoT returns to the outward-facing conformation. Our results provide a framework to understand how various SLC17 transport functions with distinct transport stoichiometries can be attained through subtle variations in proton and substrate binding/unbinding. Synopsis: The bacterial D-galactonate transporter (DgoT) and the eukaryotic solute carrier 17 (SLC17) family share key functional residues, but differ in both substrate selectivity and transport stoichiometry. Here, molecular dynamics simulations combined with experimental approaches describe the transport mechanisms of DgoT and the molecular mechanisms that underlie its specificity. DgoT transports negatively charged galactonate in symport with two protons. Protonation of D46 and E133 permits galactonate binding, followed by closure of the extracellular gate. The conserved R47 couples substrate binding to the conformational changes in the protein. Deprotonation of D46 opens the intracellular gate and permits galactonate dissociation, either protonated or deprotonated. Key amino acid residues in the coupled transport cycle of a major facilitator-superfamily protein are revealed. [ABSTRACT FROM AUTHOR]

The organic anion transporter (OAT) family of over 10 members within the solute carrier (SLC) superfamily of membrane proteins plays critical roles in facilitating the flux of negatively charged molecules in and out of cell membranes. These anionic molecules include various endogenous and exogenous compounds such as signaling molecules, nutrients, metabolites, toxins, and drugs. Therefore, OATs actively contribute to the systemic homeostasis and efficacy of therapeutics. This article provides a brief overview on recent advances in the understanding of the regulatory mechanisms that control the expression and activity of OATs in both health and diseases. [ABSTRACT FROM AUTHOR]

• Metabolome analysis revealed that plasma concentration of 3-indoxyl sulfate was increased in Slc17a3−/− mice compared to that in wild-type mice. • Urinary excretion of 3-indoxyl sulfate was reduced in Slc17a3−/− mice compared to that in wild-type mice. • An uptake study using NPT4-expressing oocytes revealed that 3-indoxyl sulfate is a physiological substrate of NPT4. • Our findings suggest that NPT4 regulates systemic exposure to 3-indoxyl sulfate by regulating its urinary excretion. Sodium-phosphate transporter NPT4 (SLC17A3) is a membrane transporter for organic anionic compounds localized on the apical membranes of kidney proximal tubular epithelial cells and plays a role in the urinary excretion of organic anionic compounds. However, its physiological role has not been sufficiently elucidated because its substrate specificity is yet to be determined. The present study aimed to comprehensively explore the physiological substrates of NPT4 in newly developed Slc17a3−/− mice using a metabolomic approach. Metabolomic analysis showed that the plasma concentrations of 11 biological substances, including 3-indoxyl sulfate, were more than two-fold higher in Slc17a3−/− mice than in wild-type mice. Moreover, urinary excretion of 3-indoxyl sulfate was reduced in Slc17a3−/− mice compared to that in wild-type mice. The uptake of 3-indoxyl sulfate by NPT4-expressing Xenopus oocytes was significantly higher than that by water-injected oocytes. The calculated K m and V max values for NPT4-mediated 3-indoxyl sulfate uptake were 4.52 ± 1.18 mM and 1.45 ± 0.14 nmol/oocyte/90 min, respectively. In conclusion, the present study revealed that 3-indoxyl sulfate is a novel substrate of NPT4 based on the metabolomic analysis of Slc17a3−/− mice, suggesting that NPT4 regulates systemic exposure to 3-indoxyl sulfate by regulating its urinary excretion. [ABSTRACT FROM AUTHOR]

Objective To investigate the changes of OAT expression in bone cells in chronic renal failure (CRF) and involved mechanism, and to explore the effect of OAT expression on bone metabolism. Methods Randomly divide the rats into a control group (n=6) and a model group (n=6) .The model group established a rat chronic renal failure model using "single nephrectomy+adenine gavage" method, and the red blood cell ( RBC) and hemoglobin ( HGB) of the rat body were measured using a blood routine analyzer; Measure indicators such as creatinine (Cr), urea nitrogen (BUN), uric acid (UA), blood calcium (Ca2+), and blood phosphorus (P3+) using a fully automated biochemical analyzer; Pathological examination of rat kidneys; X-ray examination of rat tibia; Immunohistochemical examination of bone tissue OAT1 level. Results The bone density of the model group rats is lower than that of the control group; The calcium and phosphorus metabolism of rats in the model group was in metabolic disorder, and the OAT1 value of bone tissue binding was much lower than that of the control group (P= 0.001 8), which was statistically significant. (P=0.001 8) Conclusion Chronic renal failure affected the binding ability of OAT1 in bone tissue, leading to the metabolic disorder for calcium absorption and phosphorus metabolism, thus aggravating renal osteodystrophy (P<0.05). [ABSTRACT FROM AUTHOR]

Objective: Thyroid hormone (TH) transport represents a critical first step in governing intracellular TH regulation. It is still unknown whether the full repertoire of TH transporters has been identified. Members of the solute carrier (SLC) 22 family have substrates in common with the known TH transporters of the organic anion-transporting peptide family. Therefore, we screened the SLC22 family for TH transporters Methods: Uptake of 1 nM of iodothyronines or sulfated iodothyronines in COS1 cells expressing SLC22 proteins was performed. Results: We first tested 25 mouse (m) SLC22 proteins for TH uptake and fo und that the majority of the organic anion transporter (OAT) clade were capable of 3,3’,5-triiodothyronine and/or thyroxine (T4) transport. Based on phylogenetic tree analysis of the mouse and human (h) SLC22 family, we selected eight hSLC22s that grouped with the newly identified mouse TH transporters. Of thes e, four tested positive for uptake of one or more substrates, particularly hSLC22A11 showed robust (3-fold over control) uptake of T4. Uptake of sulfated iodothyronines was strongly (up to 17-fold) induced by some SLC22s, most notably SLC22A8, hSLC22A9, mSLC22A27 and mSLC22A29. Finally, the zebrafish orthologues of SLC22A6/8 drOat x and drSlc22a6l also transported almost all (sulfated) iodothyronines tested. The OAT inhibitors lesinurad and probenecid inhibited most SLC22 proteins. Conclusions: Our results demonstrated that members of the OAT clade of the SLC22 family constitute a novel, evolutionary conserved group of transporters for (sulfated) iodothyronines. Future studies should reveal the relevance of these transporters in TH homeostasis and physiology.

Organic anion transporter 1 (OAT1/SLC22A6) is a drug transporter with numerous xenobiotic and endogenous substrates. The Remote Sensing and Signaling Theory suggests that drug transporters with compatible ligand preferences can play a role in "organ crosstalk," mediating overall organismal communication. Other drug transporters are well known to transport lipids, but surprisingly little is known about the role of OAT1 in lipid metabolism. To explore this subject, we constructed a genome-scale metabolic model using omics data from the Oat1 knockout mouse. The model implicated OAT1 in the regulation of many classes of lipids, including fatty acids, bile acids, and prostaglandins. Accordingly, serum metabolomics of Oat1 knockout mice revealed increased polyunsaturated fatty acids, diacylglycerols, and long-chain fatty acids and decreased ceramides and bile acids when compared with wildtype controls. Some aged knockout mice also displayed increased lipid droplets in the liver when compared with wildtype mice. Chemoinformatics and machine learning analyses of these altered lipids defined molecular properties that form the structural basis for lipid-transporter interactions, including the number of rings, positive charge/volume, and complexity of the lipids. Finally, we obtained targeted serum metabolomics data after short-term treatment of rodents with the OAT-inhibiting drug probenecid to identify potential drug-metabolite interactions. The treatment resulted in alterations in eicosanoids and fatty acids, further supporting our metabolic reconstruction predictions. Consistent with the Remote Sensing and Signaling Theory, the data support a role of OAT1 in systemic lipid metabolism.

In patients with liver or kidney disease, it is especially important to consider the routes of metabolism and elimination of small-molecule pharmaceuticals. Once in the blood, numerous drugs are taken up by the liver for metabolism and/or biliary elimination, or by the kidney for renal elimination. Many common drugs are organic anions. The major liver uptake transporters for organic anion drugs are organic anion transporter polypeptides (OATP1B1 or SLCO1B1; OATP1B3 or SLCO1B3), whereas in the kidney they are organic anion transporters (OAT1 or SLC22A6; OAT3 or SLC22A8). Since these particular OATPs are overwhelmingly found in the liver but not the kidney, and these OATs are overwhelmingly found in the kidney but not liver, it is possible to use chemoinformatics, machine learning (ML) and deep learning to analyze liver OATP-transported drugs versus kidney OAT-transported drugs. Our analysis of >30 quantitative physicochemical properties of OATP- and OAT-interacting drugs revealed eight properties that in combination, indicate a high propensity for interaction with "liver" transporters versus "kidney" ones based on machine learning (e.g., random forest, k-nearest neighbors) and deep-learning classification algorithms. Liver OATPs preferred drugs with greater hydrophobicity, higher complexity, and more ringed structures whereas kidney OATs preferred more polar drugs with more carboxyl groups. The results provide a strong molecular basis for tissue-specific targeting strategies, understanding drug-drug interactions as well as drug-metabolite interactions, and suggest a strategy for how drugs with comparable efficacy might be chosen in chronic liver or kidney disease (CKD) to minimize toxicity.

The multispecific organic anion transporters, OAT1 (SLC22A6) and OAT3 (SLC22A8), the main kidney elimination pathways for many common drugs, are often considered to have largely-redundant roles. However, whereas examination of metabolomics data from Oat-knockout mice (Oat1 and Oat3KO) revealed considerable overlap, over a hundred metabolites were increased in the plasma of one or the other of these knockout mice. Many of these relatively unique metabolites are components of distinct biochemical and signaling pathways, including those involving amino acids, lipids, bile acids, and uremic toxins. Cheminformatics, together with a "logical" statistical and machine learning-based approach, identified a number of molecular features distinguishing these unique endogenous substrates. Compared with OAT1, OAT3 tends to interact with more complex substrates possessing more rings and chiral centers. An independent "brute force" approach, analyzing all possible combinations of molecular features, supported the logical approach. Together, the results suggest the potential molecular basis by which OAT1 and OAT3 modulate distinct metabolic and signaling pathways in vivo As suggested by the Remote Sensing and Signaling Theory, the analysis provides a potential mechanism by which "multispecific" kidney proximal tubule transporters exert distinct physiological effects. Furthermore, a strong metabolite-based machine-learning classifier was able to successfully predict unique OAT1 versus OAT3 drugs; this suggests the feasibility of drug design based on knockout metabolomics of drug transporters. The approach can be applied to other SLC and ATP-binding cassette drug transporters to define their nonredundant physiological roles and for analyzing the potential impact of drug-metabolite interactions.

The SLC22 family of transporters is widely expressed, evolutionarily conserved, and plays a major role in regulating homeostasis by transporting small organic molecules such as metabolites, signaling molecules, and antioxidants. Analysis of transporters in fruit flies provides a simple yet orthologous platform to study the endogenous function of drug transporters in vivo. Evolutionary analysis of Drosophila melanogaster putative SLC22 orthologs reveals that, while many of the 25 SLC22 fruit fly orthologs do not fall within previously established SLC22 subclades, at least four members appear orthologous to mammalian SLC22 members (SLC22A16:CG6356, SLC22A15:CG7458, CG7442 and SLC22A18:CG3168). We functionally evaluated the role of SLC22 transporters in Drosophila melanogaster by knocking down 14 of these genes. Three putative SLC22 ortholog knockdowns-CG3168, CG6356, and CG7442/SLC22A-did not undergo eclosion and were lethal at the pupa stage, indicating the developmental importance of these genes. Additionally, knocking down four SLC22 members increased resistance to oxidative stress via paraquat testing (CG4630: p < 0.05, CG6006: p < 0.05, CG6126: p < 0.01 and CG16727: p < 0.05). Consistent with recent evidence that SLC22 is central to a Remote Sensing and Signaling Network (RSSN) involved in signaling and metabolism, these phenotypes support a key role for SLC22 in handling reactive oxygen species.

7-Hydroxycoumarin (7-HC) is a natural coumarin compound rich in Chinese herbal medicines and has various pharmacological activities. After oral administration of 7-HC in rodents, its conjugated metabolites 7-hydroxycoumarin-β-D-glucuronide (7-HCG) and 7-hydroxycoumarin sulfate (7-HCS), exhibit high systemic exposure and urinary excretion. Organic anion transporters 1 and 3 (OAT1 and OAT3), mainly expressed in the proximal renal tubules, play an important role in drug-drug interactions and drug-induced kidney injury. We aimed to explore the mechanisms of OAT-mediated drug interactions and renal protective mechanisms of 7-HC and its conjugates. OAT-overexpressing cell models revealed that 7-HC was not a substrate for OAT1 and OAT3, while 7-HCG was specifically transported by OAT3. In contrast, 7-HCS can be transported by both OATs. Besides, 7-HC significantly inhibited the activity of OAT1 and OAT3, while 7-HCS had a strong inhibitory effect on OAT1 (IC 50 < 10 μM). After co-administration of 100 mg/kg of 7-HC to mice, systemic exposure and clearance of furosemide (a clinical substrate of OATs) were significantly increased and decreased, respectively. In addition, 7-HC decreased OAT-mediated cytotoxicity and reduced the renal distribution of adefovir in mice. Together, these findings will provide support for OAT-mediated drug interactions and the renal protection of 7-HC. [Display omitted] • 7-Hydroxycoumarin conjugates were transported by renal OAT1 and/or OAT3. • 7-Hydroxycoumarin strongly inhibited OAT1 and OAT3 in vitro with IC 50 values comparable to probenecid. • 7-Hydroxycoumarin increased systemic exposure to furosemide in mice. • 7-Hydroxycoumarin reduced OAT-mediated cytotoxicity and decreased renal concentration of adefovir in mice. [ABSTRACT FROM AUTHOR]

Chiglitazar, a pan agonist of non‐thiazolidinedione peroxisome proliferator‐activated receptor, has the potential to regulate blood sugar, improve lipid metabolism, and reduce cardiovascular complications. This study aimed to examine the effect of cytochrome P450 (CYP) 3A4 inhibitors/inducers on the in vivo metabolism of chiglitazar and provide a reference for the clinical combination use of chiglitazar. A single‐center, open‐label, sequential crossover, and self‐control study was carried out in 24 healthy subjects to determine the pharmacokinetics of chiglitazar dosed with and without CYP3A4 inhibitors and inducers. The findings showed that the CYP3A4 inhibitor itraconazole had no apparent pharmacokinetic drug interaction with chiglitazar, whereas rifampicin did. When combined with rifampicin after continuous dosing, chiglitazar exposure was not theoretically reduced but increased compared to a single dose of chiglitazar. The possible explanation may be the transporters of bile salt export pump, but this needs to be confirmed. The safety of chiglitazar in single or combination doses was well tolerated. The findings of this study provide a basis for clinical combinations of chiglitazar with CYP3A4 inhibitors or inducers. [ABSTRACT FROM AUTHOR]

Organic anion transporter 1 (OAT1) expressed in the kidney plays an important role in the elimination of numerous anionic drugs used in the clinic. We report here that insulin, a pancreas-secreted hormone, regulated the expression and activity of kidney-specific OAT1 both in cultured cells and in rats. We showed that treatment of OAT1-expressing cells with insulin led to an increase in OAT1 expression, transport activity, and SUMOylation. Such insulin-induced increase was blocked by afuresertib, a specific inhibitor for protein kinase B (PKB), suggesting insulin regulates OAT1 through PKB signaling pathway. Furthermore, insulin stimulated transport activity and SUMOylation of endogenously expressed OAT1 in rat kidneys. In conclusion, our data support a remote sensing and signaling model, in which OAT1 plays an essential role in intercellular and inter-organ communication and in maintaining local and whole-body homeostasis. Such complex and dedicated communication is carried out by insulin, and PKB signaling and membrane sorting. [ABSTRACT FROM AUTHOR]

Background: Polydatin has significant uricosuric effects. This study was conducted to examine the xanthine oxidase inhibition and to detect protein levels of urate transporter 1 (URAT1), organic anion transporters 1 (OAT1), and organic anion transporters 3 (OAT3) in the hyperuricemic mice to understand the role of antihyperuricemic mechanism of polydatin. Materials and Methods: Hyperuricemia mice model was established with uricase inhibitor (potassium oxonate), and uric acids in serum were observed. Kidney tissues were used to detect gene contents of URAT1, OAT1, and OAT3 by real-time-PCR (polymerase chain reaction) and to detect pathological features. The activity of xanthine oxidase (XOD) in the liver tissues of mice was detected. Results: The polydatin experimental groups and the positive control group (benzbromarone) showed significantly inhibited levels of serum uric acid when compared with the model group. Polydatin significantly inhibited the increasing tendency of the mRNA and the protein levels of OAT1 and OAT3, and decreased the tendency of the mRNA and the protein level of URAT1. Polydatin significantly inhibited the level of XOD in the liver tissues of mice in a concentration-dependent manner. Polydatin showed a protective effect on the pathological injury of the kidney. Conclusion: Polydatin has an antihyperuricemic effect in oxonate-induced hyperuricemic mice. The effect was related to the downregulation of renal URAT1, upregulation of renal OAT1 and OAT3, and inhibition of XOD in the hyperuricemic mice. [ABSTRACT FROM AUTHOR]

Mirogabalin is a α2δ ligand as well as pregabalin. The aim of this study was to clarify whether mirogabalin is a substrate of human LAT1, which involved in absorption and disposition of pregabalin, and to investigate transporters involved in renal secretion and absorption of mirogabalin using transporter-expressing cells and fresh human kidney slices. We employed uptake assay of [3H]mirogabalin by HEK293T or HEK293 cells transiently overexpress human OAT1, OAT3, OCT2, LAT1/4F2hc, LAT2/4F2hc, PEPT1, and PEPT2 proteins. Transport assay of MDCKII cells transiently overexpress OCT2/MATE1, and OCT2/MATE2-K proteins was conducted. Contribution of transporters to renal secretion was investigated by uptake assay using human kidney slices. Uptake clearances of [3H]mirogabalin by OAT1-, OAT3-, OCT2-, PEPT1-, and PEPT2-expressing cells were higher than that by vector cells, but by LAT1/4F2hc and LAT2/4F2hc-expressing cells were not. In transport assay using OCT2/MATE1 and OCT2/MATE2-K cells, [3H]mirogabalin showed directional transport from basolateral to apical side. Contribution of OAT1, OAT3, and OCT2 was observed by uptake of [3H]mirogabalin into the kidney slices. These results indicate that mirogabalin is not a substrate of LAT1, but of PEPT1 and PEPT2 involved in absorption and of OAT1, OAT3, OCT2, MATE1 and/or MATE2-K involved in its urinary secretion. [ABSTRACT FROM AUTHOR]

1. We investigated the changes in the expression of drug-metabolising enzymes and drug transporters in the liver, small intestine and kidney of mice with collagen antibody-induced arthritis (CAIA) to determine whether changes in these expressions affect pharmacokinetics of drugs in patients with rheumatoid arthritis. 2. mRNA expression levels of cytochrome P450 (Cyp) 2b10, Cyp2c29 and Cyp3a11 were observed to be lower in the liver and small intestine of CAIA mice than in control mice. Compared with control mice, mRNA expression levels of multidrug resistance 1 b, peptide transporter 2 and organic anion transporter (Oat) 2 were high in the liver of CAIA mice. Changes in these expression levels were different among organs. However, elevated expression of Oat2 mRNA was not associated with an increase in protein expression and transport activity evaluated using [3H]cGMP as a substrate. 3. These results suggest that arthritis can change the expression of pharmacokinetics-related genes, but the changes may not necessarily be linked to the pharmacokinetics in patients with rheumatoid arthritis. On the other hand, we found Oat2 mRNA expression level was positively correlated with plasma interleukin-6 level, indicating that transcriptional activation of Oat2 may occur in inflammatory state. [ABSTRACT FROM AUTHOR]

One of the characteristics of chronic kidney disease (CKD) is the accumulation of uremic solutes in the plasma. Less is known about the effects of uremic solutes on transporters that may play critical roles in pharmacokinetics. We evaluated the effect of 72 uremic solutes on organic anion transporter 1 and 3 (OAT1 and OAT3) using a fluorescent probe substrate, 6-carboxyfluorescein. A total of 12 and 13 solutes were identified as inhibitors of OAT1 and OAT3, respectively. Several of them inhibited OAT1 or OAT3 at clinically relevant concentrations and reduced the transport of other OAT1/3 substrates in vitro. Review of clinical studies showed that the active secretion of most drugs that are known substrates of OAT1/3 deteriorated faster than the renal filtration in CKD. Collectively, these data suggest that through inhibition of OAT1 and OAT3, uremic solutes contribute to the decline in renal drug clearance in patients with CKD.

Context Relinqing granules (RLQ) are being used alone or in combination with antibacterial drugs to treat urological disorders. Objective This study investigates the pharmacokinetics of RLQ in humans and the potential for RLQ-perpetrated interactions on transporters. Materials and methods Twelve healthy subjects (six women and six men) participated to compare single- and multiple-dose pharmacokinetics of RLQ. In the single-dose study, all 12 subjects received 8 g of RLQ orally. After a 7-d washout period, the subjects received 8 g of RLQ for seven consecutive days (t.i.d.) and then a single dose. Gallic acid (GA) and protocatechuic acid (PCA) in plasma and urine samples were analysed using LC-MS/MS. The transfected cells were used to study the inhibitory effect of GA (50–5000 μg/L) and PCA (10–1000 μg/L) on transporters OAT1, OAT3, OCT2, OATP1B1, P-gp and BCRP. Results GA and PCA were absorbed into the blood within 1 h after administration and rapidly eliminated with a half-life of less than 2 h. The mean peak concentrations of GA (102 and 176 μg/L) and PCA (4.54 and 7.58 μg/L) were lower in males than females, respectively. The 24 h urine recovery rates of GA and PCA were about 10% and 5%, respectively. The steady-state was reached in 7 d without accumulation. GA was a potent inhibitor of OAT1 (IC50 = 3.73 μM) and OAT3 (IC50 = 29.41 μM), but not OCT2, OATP1B1, P-gp or BCRP. Discussion and conclusions GA and PCA are recommended as PK-markers in RLQ-related pharmacokinetic and drug interaction studies. We should pay more attention to the potential for RLQ-perpetrated interactions on transporters.

Over the course of more than 500 million years, the kidneys have undergone a remarkable evolution from primitive nephric tubes to intricate filtration-reabsorption systems that maintain homeostasis and remove metabolic end products from the body. The evolutionarily conserved solute carriers organic cation transporter 2 (OCT2) and organic anion transporters 1 and 3 (OAT1/3) coordinate the active secretion of a broad range of endogenous and exogenous substances, many of which accumulate in the blood of patients with kidney failure despite dialysis. Harnessing OCT2 and OAT1/3 through functional preservation or regeneration could alleviate the progression of kidney disease. Additionally, it would improve current in vitro test models that lose their expression in culture. With this review, we explore OCT2 and OAT1/3 regulation from different perspectives: phylogenetic, ontogenetic, and cell dynamic. Our aim is to identify possible molecular targets both to help prevent or compensate for the loss of transport activity in patients with kidney disease and to enable endogenous OCT2 and OAT1/3 induction in vitro in order to develop better models for drug development. [ABSTRACT FROM AUTHOR]

Indoxyl sulfate, closely related to indigo, a dye valued for it binding to cloth, has been recognized as a protein-bound solute bound to albumin, present in increased concentration in the serum of patients with impaired glomerular filtration (13). The early studies of Niwa identified indoxyl sulfate as a toxin capable of accelerating the rate of renal damage in subtotal nephrectomized rats (18). Over the past decade other protein-bound solutes have been identified in the plasma of patients with impaired glomerular filtration. Although the early studies, focused on the kidney, identified indoxyl sulfate as a toxic waste product dependent on the kidney for its removal, subsequent observations have identified organic anion transporters on many non-renal tissue, leading to the view that indoxyl sulfate is part of a systemic signaling system. [ABSTRACT FROM AUTHOR]

Aims: Tenofovir and para‐aminosalicylic acid (PAS) may be coprescribed to treat patients with concomitant infections of human immunodeficiency virus and Mycobacterium tuberculosis bacteria. Both drugs are known to have remarkable renal uptake transporter‐mediated clearance. Owing to the lack of clinical studies on drug–drug interaction between the 2 drugs, we conducted a translational clinical study to investigate the effect of PAS on tenofovir pharmacokinetics (PK). Methods: Initially, we studied in vitro renal uptake transporter‐mediated drug–drug interactions using stably transfected cells with human organic anion transporters (OAT1 and OAT3). Later, we estimated clinical drug interactions using static and physiologically based PK modelling. Finally, we investigated the effects of PAS‐calcium formulation (PAS‐Ca) on tenofovir disoproxil fumarate PK in healthy male Korean subjects. Results: PAS inhibited OAT1‐ and OAT3‐mediated tenofovir uptake in vitro. The physiologically based PK drug–drug interaction model suggested a 1.26‐fold increase in tenofovir peak plasma concentration when coadministered with PAS. By contrast, an open‐label, randomized, crossover clinical trial evaluating the effects of PAS‐Ca on tenofovir PK showed significantly altered geometric mean ratio (90% confidence intervals) of maximum plasma concentration (Cmax) and area under the curve (AUC0‐inf) by 0.33 (0.28–0.38) and 0.29 (0.26–0.33), respectively. Conclusion: Our study findings suggest that the PAS‐Ca formulation significantly reduced systemic exposure to tenofovir through an unexplained mechanism, which was contrary to the initial prediction. Caution should be exercised while predicting in vivo PK profiles from in vitro data, particularly when there are potential confounders such as pharmaceutical interactions. [ABSTRACT FROM AUTHOR]

Background/objectives: Cladophora glomerata extract (CGE), rich in polyphenols, was reported to exhibit antidiabetic and renoprotective effects by modulating the functions of protein kinases-mediated organic anion transporter 1 (Oat1) and 3 (Oat3) in rats with type 2 diabetes mellitus (T2DM). Nevertheless, the antioxidant effects of CGE on such renoprotection have not been investigated. This study examined the mechanisms involved in the antioxidant effects of CGE on renal organic anion transport function in an in vivo study., Materials/methods: Diabetes was induced in the rats through a high-fat diet combined with a single dose of 40 mg/kg body weight (BW) streptozotocin. Subsequently, normal-diet rats were supplemented with a vehicle or 1,000 mg/kg BW of CGE, while T2DM rats were supplemented with a vehicle, CGE, or 200 mg/kg BW of vitamin C for 12 weeks. The study evaluated the general characteristics of T2DM and renal oxidative stress markers. The renal organic transport function was assessed by measuring the para-aminohippurate (PAH) uptake using renal cortical slices and renal inflammatory cytokine expression in the normal diet (ND) and ND + CGE treated groups., Results: CGE supplementation significantly reduced hyperglycemia, hypertriglyceridemia, insulin resistance, and renal lipid peroxidation in T2DM rats. This was accompanied by the normalization of high expressions of renal glutathione peroxidase and nuclear factor kappa B by CGE and vitamin C. The renal anti-inflammation of CGE was evidenced by the reduction of tumor necrosis factor-1α and interleukin-1β. CGE directly blunted sodium nitroprusside-induced renal oxidative/nitrosative stresses and mediated the PAH uptake in the normally treated CGE in rats was particularly noteworthy. These data also correlated with reduced nitric oxide production, highlighting the potential of CGE as a therapeutic agent for managing T2DM-related renal complications., Conclusion: These findings suggest that CGE has antidiabetic effects and directly prevents diabetic nephropathy through oxidative/nitrosative stress pathways., Competing Interests: Conflict of Interest: The authors declare no potential conflicts of interests., (©2024 The Korean Nutrition Society and the Korean Society of Community Nutrition.)

Relinqing granules (RLQ) are being used alone or in combination with antibacterial drugs to treat urological disorders. This study investigates the pharmacokinetics of RLQ in humans and the potential for RLQ-perpetrated interactions on transporters. Twelve healthy subjects (six women and six men) participated to compare single- and multiple-dose pharmacokinetics of RLQ. In the single-dose study, all 12 subjects received 8 g of RLQ orally. After a 7-d washout period, the subjects received 8 g of RLQ for seven consecutive days (t.i.d.) and then a single dose. Gallic acid (GA) and protocatechuic acid (PCA) in plasma and urine samples were analysed using LC-MS/MS. The transfected cells were used to study the inhibitory effect of GA (50–5000 μg/L) and PCA (10–1000 μg/L) on transporters OAT1, OAT3, OCT2, OATP1B1, P-gp and BCRP. GA and PCA were absorbed into the blood within 1 h after administration and rapidly eliminated with a half-life of less than 2 h. The mean peak concentrations of GA (102 and 176 μg/L) and PCA (4.54 and 7.58 μg/L) were lower in males than females, respectively. The 24 h urine recovery rates of GA and PCA were about 10% and 5%, respectively. The steady-state was reached in 7 d without accumulation. GA was a potent inhibitor of OAT1 (IC50 = 3.73 μM) and OAT3 (IC50 = 29.41 μM), but not OCT2, OATP1B1, P-gp or BCRP. GA and PCA are recommended as PK-markers in RLQ-related pharmacokinetic and drug interaction studies. We should pay more attention to the potential for RLQ-perpetrated interactions on transporters. [ABSTRACT FROM AUTHOR]

URAT1 (slc22a12) was identified as the transporter responsible for renal reabsorption of the medically important compound, uric acid. However, subsequent studies have indicated that other transporters make contributions to this process, and that URAT1 transports other organic anions besides urate (including several in common with the closely related multi-specific renal organic anion transporters, OAT1 (slc22a6) and OAT3 (slc22a8)). These findings raise the possibility that urate transport is not the sole physiological function of URAT1. We previously characterized mice null for the murine ortholog of URAT1 (mURAT1; previously cloned as RST), finding a relatively modest decrement in urate reabsorptive capacity. Nevertheless, there were shifts in the plasma and urinary concentrations of multiple small molecules, suggesting significant metabolic changes in the knockouts. Although these molecules remain unidentified, here we have computationally delineated the biochemical networks consistent with transcriptomic data from the null mice. These analyses suggest alterations in the handling of not only urate but also other putative URAT1 substrates comprising intermediates in nucleotide, carbohydrate, and steroid metabolism. Moreover, the analyses indicate changes in multiple other pathways, including those relating to the metabolism of glycosaminoglycans, methionine, and coenzyme A, possibly reflecting downstream effects of URAT1 loss. Taken together with the available substrate and metabolomic data for the other OATs, our findings suggest that the transport and biochemical functions of URAT1 overlap those of OAT1 and OAT3, and could contribute to our understanding of the relationship between uric acid and the various metabolic disorders to which it has been linked.

Effects of cholecalciferol (VitD3) and calcitriol (1,25-VitD3), on the expression and function of major vitamin D metabolizing enzymes (cytochrome P450 [CYP]2R1, CYP24A1) and select drug transport pathways (ABCB1/P-gp, SLCO4C1/OATP4C1) were evaluated in human kidney proximal tubule epithelial cells (hPTECs) under normal and uraemic serum conditions. hPTECs were incubated with 10% normal or uraemic serum for 24 h followed by treatment with 2% ethanol vehicle, or 100 and 240 nM doses of VitD3, or 1,25-VitD3 for 6 days. The effects of treatment on mRNA and protein expression and functional activity of select CYP enzymes and transporters were assessed Under uraemic serum, treatment with 1,25-VitD3 resulted in increased mRNA but decreased protein expression of CYP2R1. Activity of CYP2R1 was not influenced by serum or VitD analogues. CYP24A1 expression was increased with 1,25-VitD3 under normal as well as uraemic serum, although to a lesser extent. ABCB1/P-gp mRNA expression increased under normal and uraemic serum, with exposure to 1,25-VitD3. SLCO4C1/OATP4C1 exhibited increased mRNA but decreased protein expression, under uraemic serum + 1,25-VitD3. Functional assessments of transport showed no changes regardless of exposure to serum or 1,25-VitD3. Key findings indicate that uraemic serum and VitD treatment led to differential effects on the functional expression of CYPs and transporters in hPTECs. [ABSTRACT FROM AUTHOR]

Organic anion transporter 3 (OAT3, SLC22A8), a transporter expressed on the basolateral membrane of the proximal tubule, plays a critical role in the renal excretion of organic anions including many therapeutic drugs. The goal of this study was to evaluate the in vivo effects of the OAT3-Ile305Phe variant (rs11568482), present at 3.5% allele frequency in Asians, on drug disposition with a focus on cefotaxime, a cephalosporin antibiotic. In HEK293-Flp-In cells, the OAT3-Ile305Phe variant had a lower maximum cefotaxime transport activity, Vmax , [159 ± 3 nmol*(mg protein)(-1) /min (mean ± SD)] compared with the reference OAT3 [305 ± 28 nmol*(mg protein)(-1) /min, (mean ± SD), p < 0.01], whereas the Michaelis-Menten constant values (Km ) did not differ. In healthy volunteers, we found volunteers that were heterozygous for the Ile305Phe variant and had a significantly lower cefotaxime renal clearance (CLR ; mean ± SD: 84.8 ± 32.1 mL/min, n = 5) compared with volunteers that were homozygous for the reference allele (158 ± 44.1 mL/min, n = 10; p = 0.006). Furthermore, the net secretory component of cefotaxime renal clearance (CLsec ) was reduced in volunteers heterozygous for the variant allele [33.3 ± 31.8 mL/min (mean ± SD)] compared with volunteers homozygous for the OAT3 reference allele [97.0 ± 42.2 mL/min (mean ± SD), p = 0.01]. In summary, our study suggests that a low-frequency reduced-function polymorphism of OAT3 associates with reduced cefotaxime CLR and CL(sec) .

The mixture of Salvia miltiorrhiza and Carthamus tinctorius (Danhong injection, DHI) is widely prescribed in China for the treatment of cardiovascular and cerebrovascular diseases. In most cases, DHI is used in combination with acetylsalicylic acid (aspirin, ASA). However, the interaction between DHI and ASA remains largely undefined. The purpose of this study is to explore the interaction profile and mechanism between DHI and ASA. The frequency of drug combination of DHI and ASA was analyzed based on 5,183 clinical cases. The interaction characteristics were evaluated by analyzing the pharmacokinetics and disposition profile of salicylic acid (SA, the primary metabolite of ASA) in rats. The interaction mechanisms were explored through evaluating the hydrolysis of ASA regulated by ASA esterase, the tubular secretion of SA mediated by influx and efflux transporters, and the tubular reabsorption of SA regulated by urinary acidity-alkalinity. The inhibitory potential of DHI on organic anion transporters (OATs) was further verified in aristolochic acid I (AAI) induced nephropathy. Clinical cases analysis showed that DHI and ASA were used in combination with high frequency of 70.73%. In drug combination of DHI and ASA, the maximum plasma concentration of SA was significantly increased by 1.37 times, while the renal excretion of SA was significantly decreased by 32.54%. The mechanism study showed that DHI significantly inhibited the transport function, gene transcription and protein expression of OATs. In OATs mediated AAI nephropathy, DHI significantly reduced the renal accumulation of AAI by 55.27%, and alleviated renal damage such as glomerulus swelling, tubular blockage and lymphocyte filtration. In drug combination of DHI and ASA, DHI increased the plasma concentration of SA not through enhancing the hydrolysis of ASA, and the tubular reabsorption of SA was not significantly affected. Inhibition of tubular secretion of SA mediated by OATs might be the reason that contributes to the decrease of SA renal excretion.

The third trimester of pregnancy is related to physiological changes that can modify the process of absorption, distribution, metabolism, and excretion and, consequently, the efficacy and toxicity of drugs. However, little is known about furosemide pharmacokinetics and placental transfer in pregnancy. This study evaluated the maternal‐fetal pharmacokinetics and distribution to amniotic fluid of furosemide in hypertensive parturient women under cesarean section. Twelve hypertensive parturient women under methyldopa (250 mg/8 h) and/or pindolol (10 mg/12 h) treatment received a 40‐mg single oral dose of furosemide 1 to 10 hours before delivery by cesarean section. Blood and urine samples were collected for 12 hours after furosemide administration. At delivery, samples were obtained from maternal and umbilical cord blood (n = 8) to assess the transplacental transfer. Amniotic fluid (n = 4) was collected at the time of delivery. The following furosemide pharmacokinetic parameters were obtained as median (interquartile range): Cmax, 403 ng/mL (229 to 715 ng/mL); Tmax, 2.00 hours (1.50 to 4.83 hours); elimination half‐life (t1/2), 2.50 hours (1.77 to 2.97 hours); AUC0‐12 h, 1366 ng⋅h/mL (927 to 2531 ng⋅h/mL); AUC0‐∞, 1580 ng⋅h/mL (1270 to 2881 ng⋅h/mL); CL/F 25.3 L/h (13.8 to 31.4 L/h); CLR, 2.50 L/h (1.77 to 2.97 L/h); CLNR, 22.7 L/h (12.1 to 25.6 L/h); and Vd/F 82.8 L (34.4 to 173 L). The transplacental transfer of furosemide was 0.43 (0.10 to 0.73), and the amniotic fluid concentration was 11.0 ng/mL (5.51 to 14.6 ng/mL). From a clinical point of view, these results suggest that substrates of uridine diphosphate‐glucuronosyltransferase isoenzymes such as furosemide may have increased clearance during pregnancy and could require dose adjustment in this population. [ABSTRACT FROM AUTHOR]

The mixture of Salvia miltiorrhiza and Carthamus tinctorius (Danhong injection, DHI) is widely prescribed in China for the treatment of cardiovascular and cerebrovascular diseases. In most cases, DHI is used in combination with acetylsalicylic acid (aspirin, ASA). However, the interaction between DHI and ASA remains largely undefined. The purpose of this study is to explore the interaction profile and mechanism between DHI and ASA. The frequency of drug combination of DHI and ASA was analyzed based on 5,183 clinical cases. The interaction characteristics were evaluated by analyzing the pharmacokinetics and disposition profile of salicylic acid (SA, the primary metabolite of ASA) in rats. The interaction mechanisms were explored through evaluating the hydrolysis of ASA regulated by ASA esterase, the tubular secretion of SA mediated by influx and efflux transporters, and the tubular reabsorption of SA regulated by urinary acidity-alkalinity. The inhibitory potential of DHI on organic anion transporters (OATs) was further verified in aristolochic acid I (AAI) induced nephropathy. Clinical cases analysis showed that DHI and ASA were used in combination with high frequency of 70.73%. In drug combination of DHI and ASA, the maximum plasma concentration of SA was significantly increased by 1.37 times, while the renal excretion of SA was significantly decreased by 32.54%. The mechanism study showed that DHI significantly inhibited the transport function, gene transcription and protein expression of OATs. In OATs mediated AAI nephropathy, DHI significantly reduced the renal accumulation of AAI by 55.27%, and alleviated renal damage such as glomerulus swelling, tubular blockage and lymphocyte filtration. In drug combination of DHI and ASA, DHI increased the plasma concentration of SA not through enhancing the hydrolysis of ASA, and the tubular reabsorption of SA was not significantly affected. Inhibition of tubular secretion of SA mediated by OATs might be the reason that contributes to the decrease of SA renal excretion. [ABSTRACT FROM AUTHOR]

Organic anion transporters 1 and 3 (OAT1 and OAT3) play a critical role in renal drug-drug interactions and are involved in the nephrotoxicity of many anionic xenobiotics. To date, relatively little is known about the interaction of natural compounds with OAT1 and OAT3. Of the 270 natural compounds screened in the present study, 21 compounds inhibited OAT1 and 45 compounds inhibited OAT3. Further concentration-dependent studies identified 7 OAT1 inhibitors and 10 OAT3 inhibitors with IC50 values of <10 μM, and most of them were flavonoids, the most commonly ingested polyphenolic compounds in the diet and herbal products. Computational modeling of OAT1 and OAT3 revealed the important residues for the recognition of inhibitors. The two strong OAT inhibitors, namely wedelolactone and wogonin, were evaluated for their in vivo interactions with the OAT substrate aristolochic acid I (AAI), a natural compound causing aristolochic acid-induced nephropathy (AAN) in many species. The cytotoxicity of AAI increased in two OAT-overexpressing cell lines, with more cytotoxicity in OAT1-overexpressing cells, suggesting a more important role of OAT1 than OAT3 in AAN. Both wedelolactone and wogonin markedly increased serum AAI concentrations in AAI-treated rats and ameliorated kidney injuries in AAI-treated mice. To conclude, the present findings are of significant value in understanding natural compound-drug interactions and provide a natural source for developing treatments for AAN. [ABSTRACT FROM AUTHOR]

Indoxyl sulfate (IS), a highly protein-bound nephro-cardiovascular toxin, was poorly removed by hemodialysis. IS exists as anions in the body and the renal excretion is mediated by organic anion transporter 1 (OAT1) and OAT3. Acidic antibiotics such as cephalosporins and fluoroquinolones were putative substrates/inhibitors of OATs. We hypothesized that cephalosporins and fluoroquinolones might compete with IS for OAT1- and/or OAT3-mediated renal excretions. This study investigated the effects of ciprofloxacin, cefuroxime, cefotaxime, cefazolin and ofloxacin on the intravenous pharmacokinetics of IS in rats. IS was intravenously injected with and without each individual antibiotics, and the concentrations of IS in serum and lysate were determined by HPLC. The results showed that ciprofloxacin significantly increased AUC0-t and T1/2 of IS by 272% and 491%, respectively, and decreased the clearance by 71%. However, ofloxacin, cefuroxime, cefotaxime and cefazolin did not alter the pharmacokinetics of IS. Furthermore, cell line study showed that ciprofloxacin inhibited the OAT3-mediated transport of IS. This study indicates 30 mg/kg of ciprofloxacin decreased the clearance of IS through inhibition on the OAT3-mediated transport, whereas 50 mg/kg of ofloxacin, cefuroxime, cefotaxime and cefazolin did not show significant influence. [ABSTRACT FROM AUTHOR]

With the internationally growing popularity of traditional Chinese medicine (TCM), TCM-induced nephropathy has attracted public attention. Minimizing this toxicity is an important issue for future research. Typical nephrotoxic TCM drugs such as Aristolochic acid, Tripterygium wilfordii Hook. f, Rheum officinale Baill, and cinnabar mainly damage renal proximal tubules or cause interstitial nephritis. Transporters in renal proximal tubule are believed to be critical in the disposition of xenobiotics. In this review, we provide information on the alteration of renal transporters by nephrotoxic TCMs, which may be helpful for understanding the nephrotoxic mechanism of TCMs and reducing adverse effects. Studies have proven that when administering nephrotoxic TCMs, the expression or function of renal transporters is altered, especially organic anion transporter 1 and 3. The alteration of these transporters may enhance the accumulation of toxic drugs or the dysfunction of endogenous toxins and subsequently sensitize the kidney to injury. Transporters-related drug combination and clinical biomarkers supervision to avoid the risk of future toxicity are proposed. [ABSTRACT FROM AUTHOR]

The renal elimination of uremic toxins (UTs) can be potentially altered by drugs that inhibit organic anion transporters 1/3 (OAT1/OAT3). The objective of the present study was to determine whether the prescription of at least one OAT1/OAT3 inhibitor was associated with the plasma accumulation of certain UTs in kidney transplant recipients. We included 403 kidney transplant recipients. For each patient, we recorded all prescription drugs known to inhibit OAT1/OAT3. Plasma levels of four UTs (trimethylamine N-oxide (TMAO), indole acetic acid (IAA), para-cresylsulfate (pCS), and indoxylsulfate (IxS) were assayed using liquid chromatography-tandem mass spectrometry. Plasma UT levels were significantly higher among patients prescribed at least one OAT inhibitor (n = 311) than among patients not prescribed any OAT inhibitors (n = 92). Multivariate analysis revealed that after adjustment for age, estimated glomerular filtration rate (eGFR), plasma level of albumin and time since transplantation, prescription of an OAT1/OAT3 inhibitor was independently associated with the plasma accumulation of pCS (adjusted odds ratio (95% confidence interval): 2.11 (1.26; 3.61]). Our results emphasize the importance of understanding the interactions between drugs and UTs and those involving UT transporters in particular.

In patients with liver or kidney disease, it is especially important to consider the routes of metabolism and elimination of small-molecule pharmaceuticals. Once in the blood, numerous drugs are taken up by the liver for metabolism and/or biliary elimination, or by the kidney for renal elimination. Many common drugs are organic anions. The major liver uptake transporters for organic anion drugs are organic anion transporter polypeptides (OATP1B1 or SLCO1B1; OATP1B3 or SLCO1B3), whereas in the kidney they are organic anion transporters (OAT1 or SLC22A6; OAT3 or SLC22A8). Since these particular OATPs are overwhelmingly found in the liver but not the kidney, and these OATs are overwhelmingly found in the kidney but not liver, it is possible to use chemoinformatics, machine learning (ML) and deep learning to analyze liver OATP-transported drugs versus kidney OAT-transported drugs. Our analysis of >30 quantitative physicochemical properties of OATP- and OAT-interacting drugs revealed eight properties that in combination, indicate a high propensity for interaction with “liver” transporters versus “kidney” ones based on machine learning (e.g., random forest, k-nearest neighbors) and deep-learning classification algorithms. Liver OATPs preferred drugs with greater hydrophobicity, higher complexity, and more ringed structures whereas kidney OATs preferred more polar drugs with more carboxyl groups. The results provide a strong molecular basis for tissue-specific targeting strategies, understanding drug–drug interactions as well as drug–metabolite interactions, and suggest a strategy for how drugs with comparable efficacy might be chosen in chronic liver or kidney disease (CKD) to minimize toxicity.

Abstract Organic anion transporter 1 (OAT1) is a polyspecific transport protein located in the basolateral membrane of renal proximal tubule cells. OAT1 plays a pivotal role in drug clearance. Adverse drug reactions (ADR) are observed more frequently in women than in men, especially ADR are higher in women for drugs which are known interactors of OAT1. Sex‐dependent expression of Oat1 has been observed in rodents with a tendency to male‐dominant expression. This study aims at elucidating the transcriptional regulation of human OAT1 and tests the effect of estrogen receptor α (ERα). Promoter activation of OAT1 was assessed by luciferase assays carried out by Opossum kidney (OK) cells, transiently transfected with promoter constructs of human OAT1 and expression vectors for ERα and exposed to 100 nmol/L 17β‐estradiol. Furthermore, a transcription factor array and proteomic analysis was performed to identify estrogen‐induced transcription factors. Human OAT1 was significantly activated by ligand activated ERα. However, activation occurred without a direct interaction of ERα with the OAT1 promoter. Our data rather show an activation of the transcription factors CCAAT‐box‐binding transcription factor (CBF) and heterogeneous nuclear ribonucleoprotein K (HNRNPK) by ERα, which in turn bind and initiate OAT1 promoter activity. Herewith, we provide novel evidence of estrogen‐dependent, transcriptional regulation of polyspecific drug transporters including the estrogen‐induced transcription factors CBF and HNRNPK.

Organic anion transporter 3 (OAT3) plays a vital role in removing a broad variety of anionic drugs from kidney, thus avoiding their possible toxicity in the body. In the current study, we investigated the role of insulin-like growth factor 1 (IGF-1) in the regulation of OAT3. We showed that IGF-1 induced a dose- and time-dependent increase in OAT3 transport activity, which correlated well with an increase in OAT3 expression. The IGF-1-induced increase in OAT3 expression was blocked by protein kinase A (PKA) inhibitor H89. Moreover, IGF-1 induced an increase in OAT3 phosphorylation, which was also blocked by H89. These data suggest that the IGF-1 modulation of OAT3 occurred through PKA signaling pathway. To further confirm the involvement of PKA, we treated OAT3-expressing cells with PKA activator Bt2-cAMP, followed by examining OAT activity and phosphorylation. We showed that OAT3 activity and phosphorylation were much enhanced in Bt2-cAMP-treated cells as compared to that in control cells. Finally, linsitinib, an anticancer drug that blocks the IGF-1 receptor, abrogated IGF-1-stimulated OAT3 transport activity. In conclusion, our study demonstrated that IGF-1 regulates OAT3 expression and transport activity through PKA signaling pathway, possibly by phosphorylating the transporter. IGF-1/PKA signaling pathway regulates OAT3 phosphorylation, expression and transport activity. Image 1 [ABSTRACT FROM AUTHOR]

To assess the mechanism of the pharmacokinetic interaction between piperacillin and tazobactam, renal excretion and pharmacokinetic studies of piperacillin/tazobactam were investigated in normal and bacteremia rats. A bacteremia model was established to investigate the pharmacokinetic properties of piperacillin and tazobactam under different conditions. Renal slices were taken to examine the uptake of piperacillin and tazobactam. Pharmacokinetic studies of β-lactamase in rats were performed to study the contribution of rOat1/3 to the inhibition of tazobactam on β-lactamase. The AUC (from 2.93 ±0.58 to 6.52 ±1.44 mg ·min/ml) and the plasma clearance (CLP) (from 2.41 ±1.20 to 0.961 ±0.212 ml/min/kg) of tazobactam were both altered after the intravenous coadministration of piperacillin and tazobactam in the bacteremia rats. The renal clearance (CLR) of tazobactam decreased from 1.30 ±0.50 to 0.361 ±0.043 ml/min/kg. In summary, there was a beneficial interaction between piperacillin and tazobactam mediated by rOat1 and rOat3. Piperacillin enhances the inhibitory effect of tazobactam on β-lactamase through the inhibition of rOat1 and rOat3 in rats. The contribution rate of rOat1/3 for the synergistic effect was 20% when the two drugs were coadministered. [ABSTRACT FROM AUTHOR]

Methotrexate is a widely used chemotherapy agent with a propensity for drug interactions placing the patient at risk for toxicities. There are several modes for altering methotrexate clearance including concomitant drugs leading to toxic effects on the kidneys, medications producing an acidic urine pH, and agents interfering with methotrexate transporters including the organic anion transporter and breast cancer resistance protein efflux pump. We report a case of a patient with central nervous system non-Hodgkin's lymphoma receiving high-dose methotrexate and being concomitantly treated for a urinary tract infection with nitrofurantoin. Subsequently, her initial methotrexate clearance was altered by the introduction of nitrofurantoin and returned to baseline when her nitrofurantoin was discontinued. This is the first case report describing the altered methotrexate clearance from concurrent administration of methotrexate and nitrofurantoin. [ABSTRACT FROM AUTHOR]

Organic anion transporter 3 (OAT3) plays a vital role in removing a broad array of anionic drugs from kidney, thereby avoiding their possibly toxic side effects in the body. We earlier demonstrated that OAT3 is subjected to a specific type of post-translational modification called SUMOylation. SUMOylation is a dynamic event, where de-SUMOylation is catalyzed by a class of SUMO-specific proteases. In the present investigation, we assessed the role of SUMO-specific protease Senp2 in OAT3 SUMOylation, expression and function. We report here that overexpression of Senp2 in COS-7 cells led to a reduced OAT3 SUMOylation, which correlated well with a decreased OAT3 expression and transport activity. Such phenomenon was not observed in cells overexpressing an inactive mutant of Senp2. Furthermore, transfection of cells with Senp2-specific siRNA to knockdown the endogenous Senp2 resulted in an increased OAT3 SUMOylation, which correlated well with an enhanced OAT3 expression and transport activity. Coimmunoprecipitation experiments showed that Senp2 directly interacted with OAT3 in the kidneys of rats. Together these results provided first demonstration that Senp2 is a significant regulator for OAT3-mediated organic anion/drug transport. Unlabelled Image • Overexpression of Senp2 in renal cells led to a decreased OAT3 SUMOylation, expression, and transport activity. • Knocking-down the endogenous Senp2 in renal cells increased OAT3 SUMOylation, expression, and transport activity. • Senp2 interacts directly with OAT3 both in cultured cells and in rat kidneys. [ABSTRACT FROM AUTHOR]

Niclosamide is an anthelmintic drug with a long history of use and is generally safe and well tolerated in humans. As the conventional dose of niclosamide results in a low but certain level in systemic circulation, drug interactions with concomitant drugs should be considered. We aimed to investigate the interaction between niclosamide and drug transporters, as such information is currently limited. Niclosamide inhibited the transport activity of OATP1B1, OATP1B3, OAT1, OAT3, and OCT2 in vitro. Among them, the inhibitory effects on OAT1, OAT3, and OCT2 were strong, with IC 50 values of less than 1 μM. When 3 mg/kg of niclosamide was co-administered to rats, systemic exposure to furosemide (a substrate of OAT1/3) and metformin (a substrate of OCT2) increased, and the renal clearance (CL r) of the drugs significantly decreased. These results suggest that niclosamide inhibits renal transporters, OAT1/3 and OCT2, not only in vitro but also in vivo , resulting in increased systemic exposure to the substrates of the transporters by strongly blocking the urinary elimination pathway in rats. The findings of this study will support a meticulous understanding of the transporter-mediated drug interactions of niclosamide and consequently aid in effective and safe use of niclosamide. [Display omitted] • Niclosamide strongly inhibited OAT 1 and 3 and OCT 2 in vitro with IC 50 values less than 1 μM. • Inhibition mechanisms of niclosamide on OAT1/3 and OCT2 were determined to be non-competitive mode. • Niclosamide increased systemic exposure to furosemide or metformin significantly increased in rats. • Renal clearance of furosemide and metformin significantly decreased in the presence of niclosamide in rats. • These findings will support a meticulous understanding of the transporter-mediated drug interactions of niclosamide. [ABSTRACT FROM AUTHOR]

Acamprosate is an anionic drug substance widely used in treating symptoms of alcohol withdrawal. It was recently shown that oral acamprosate absorption is likely due to paracellular transport. In contrast, little is known about the eliminating mechanism clearing acamprosate from the blood in the kidneys, despite the fact that studies have shown renal secretion of acamprosate. The hypothesis of the present study was therefore that renal organic anion transporters (OATs) facilitate the renal excretion of acamprosate in humans. The aim of the present study was to establish and apply OAT1 (gene product of SLC22A6) and OAT3 (gene product of SLC22A8) expressing cell lines to investigate whether acamprosate is a substrate or inhibitor of OAT1 and/or OAT3. The studies were performed in HEK293-Flp-In cells stably transfected with SLC22A6 or SLC22A8. Protein and functional data showed that the established cell lines are useful for studying OAT1- and OAT3-mediated transport in bi-laboratory studies. Acamprosate inhibited OAT1-mediated p-aminohippuric acid (PAH) uptake but did not inhibit substrate uptake via OAT3 expressing cells, neither when applied concomitantly nor after a 3 h preincubation with acamprosate. The uptake of PAH via OAT1 was inhibited in a competitive manner by acamprosate and cellular uptake studies showed that acamprosate is a substrate for OAT1 with a Km-value of approximately 700 µM. Probenecid inhibited OAT1-mediated acamprosate uptake with a Ki-value of approximately 13 µM, which may translate into an estimated clinically significant DDI index. In conclusion, acamprosate was identified as a substrate of OAT1 but not OAT3.

Abstract Organic anion transporters (OATs in humans, Oats in rodents) play an important role in the distribution and excretion of numerous endogenous metabolic products and exogenous organic anions, including a host of widely prescribed drugs. Their ligand recognition is also important for drug therapy and development. In this study, the n -butanol and dichloromethane soluble fractions of Juniperus oblonga were found to inhibit OAT3 in vitro and three biflavonoids were found to be responsible for this activity. One of these compounds, amentoflavone exhibited stronger inhibition than probenecid, a known strong inhibitor of OAT3. Biological characterization of amentoflavone in vivo also showed inhibition of Oat3. Preliminary observations of structure-activity relationships suggest that the biflavonoids are more potent inhibitors of this transporter than their corresponding monomer, and that methylation of even a single hydroxyl group results in a substantial decrease in activity. This greater potency of the biflavonoids may indicate the need for a more in-depth investigation of the distribution of biflavonoids in plants used as foodstuffs and herbal medicines, due to their potential for causing interactions with OAT3 substrate drugs. Highlights • Three biflavonoids were identified as potent inhibitors of OAT3. • Amentoflavone, which inhibits OAT3 possesses the potential for clinical interaction with drug substrates of OAT3. • Biflavones with different locations of their inter-flavone linkage showed different levels of inhibition. [ABSTRACT FROM AUTHOR]

Highlights • Cinnabar differs significantly from other mercury containing compounds in transport and accumulation in the cell. • Cinnabar is less cytotoxic than HgCl 2 and MeHg due to lower intracellular uptake. • OAT1 and OAT3 are partially involved in the regulation of the transport of HgCl 2 and MeHg, but not in cinnabar. • Knockdown of OAT1 by siRNA treatment induces resistance to the toxicity of HgCl 2 , but not to that of cinnabar, MeHg, and HgS. Abstract Background Cinnabar has a long history of uses in Chinese traditional medicines as an ingredient in various remedies. However, the detailed mechanism of cinnabar in medication remains unclear, and the toxicity of cinnabar has been a debate due to its containing mercury sulfide. This study was designed to investigate the differential transport mechanism of cinnabar and other Hg-containing compounds HgCl 2 , MeHg and HgS, and to determine if organic anion transporters OAT1 and OAT3 were involved in the differential transport mechanism. Materials and methods The 293T cells were employed to investigate and compare the differential transport mechanism of cinnabar and HgCl 2 , MeHg and HgS. Cells were incubated with a low dose (5 μM HgCl 2 and MeHg, 200 μM HgS and cinnabar), medium dose (10 μM HgCl 2 and MeHg, 400 μM HgS and cinnabar), and high dose (20 μM HgCl 2 and MeHg, 800 μM HgS and cinnabar) of HgCl 2 , MeHg, HgS and cinnabar for 24 h. Following treatment, the cells were collected and the cell viability was determined by MTT assay. The intracellular mercury content was measured at 1, 4, and 24 h after treatment with 10 μM of the tested agents by an atomic fluorescence spectrophotometer. The effect of these tested agents on mitochondrial respiration was determined in a high-resolution oxygraphyat 24 h following treatment. Furthermore, the effect of modulation of expression of transporters OAT1 and OAT3 on the transport and cytotoxicity of the tested agents was evaluated. The up and down regulation of OAT1 and OAT3 were achieved by overexpression and siRNA transfection, respectively. Results Compared with HgCl 2 and MeHg, the cytotoxicity of cinnabar and HgS was lower, with cell viability at the high dose cinnabar and HgS being about 65%, while MeHg and HgCl 2 were 40% and 20%, respectively. The intracellular mercury accumulation was time-dependent. At 24 h the intracellular concentrations of HgCl 2 and MeHg were about 7 and 5 times higher, respectively, than that of cinnabar. No significant difference was found in the intracellular mercury content in cells treated with cinnabar compared to HgS. The knockdown and overexpression of the transporter OAT1 resulted in significant reduction and increase, respectively, in mercury accumulation in HgCl 2 -treated cells in relative to control cells, while no significant changes were observed in cells treated with cinnabar, MeHg, and HgS. In addition, the knockdown and overexpression of the transporter OAT3 caused significant reduction and increase, respectively, in mercury accumulation in both HgCl 2 and MeHg-treated cells in relative to control cells, while no significant changes were observed in cells treated with cinnabar and HgS. Furthermore, it was found that cells transfected with siOAT1 caused significant resistance to the cytotoxicity induced by HgCl 2 , while no noticeable changes in cell viability were observed in cells treated with other tested agents. Additionally, cells transfected with OAT3 did not change cell sensitivity to cytotoxicity induced by all of the four tested agents. Conclusion This study demonstrates that differential transport and accumulation of mercury in 293T cells exists among cinnabar and the three mercury-containing compounds HgCl 2 , MeHg and HgS, leading to distinct sensitivity to mercury induced cytotoxicity. The kidney organic anion transporters OAT1 and OAT3 are partially involved in the regulation of the transport of HgCl 2 and MeHg, but not in the regulation of the transport of cinnabar. [ABSTRACT FROM AUTHOR]