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OBJECTIVE To study the correlation of novel organic cation transporter 2 （OCTN2） with the chemosensitivity of prostate cancer cells to oxaliplatin. METHODS Tumor samples of patients receiving radical prostatectomy were collected， and OCTN2 protein was detected with immunohistochemistry； the primary cells of the specimen were cultivated to obtain prostate cancer cell line. Inductively coupled plasma mass spectrometry was used to detect the uptake of low concentration （0.1 μmol/L） of oxaliplatin by cancer cells. Real-time PCR and Western blot were used to detect the mRNA and protein expressions of OCTN2 in cancer cells； the prostate cancer cells with the highest and lowest expression of OCTN2 protein were selected， and IC50 of oxaliplatin to prostate cancer cells was analyzed by ATP-TCA method. The inhibitory rate of plasma peak concentration of oxaliplatin （50 μmol/L） to prostate cancer cells was detected by MTT assay. Spearman method was used to analyze the relationship of the uptake of oxaliplatin by prostate cancer cells with inhibitory rate of oxaliplatin to prostate cancer cells and 505916443@qq.com mRNA expressions of OCTN2. RESULTS OCTN2 was located on the membrane of cancer cells， and the uptake of zjdtztougao@163.com oxaliplatin by cancer cells was 0.283±0.264 （n＝12）mRNA and protein expression of OCTN2 varied significantly among different cancer cells. The sensitivity of cancer cells with high expression of OCTN2 to oxaliplatin （IC50 of 4.61 μmol/L） was higher than that of cancer cells with lower expression of OCTN2 （IC50 of 26.23 μmol/L）. The inhibitory rate of oxaliplatin to cancer cells was （25.4±10.8）% （n＝12）. There was a correlation between the uptake of oxaliplatin by prostate cancer cells and the inhibition rate of oxaliplatin to prostate cancer cells and mRNA expression of OCTN2 （P＜0.05）. CONCLUSIONS High-expressed OCTN2 may promote the uptake of oxaliplatin by prostate cancer cells， and its expression can serve as a reference for predicting the sensitivity of prostate cancer cells to oxaliplatin chemotherapy.
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Kidney injury and decreased chemosensitivity of tumor cells are obstacles with cisplatin (CDDP) chemotherapy. Down-regulation of the organic cation transporter 2 (OCT2) and multidrug resistance-associated protein 2 (MRP2) is a key means to alleviate CDDP-induced kidney injury and increase chemosensitivity. Astragaloside IV (AS IV) is obtained from the well-known traditional Chinese herb Astragalus membranaceus. This study explored the role of AS IV in preventing kidney injury and enhancing the antitumor effect of CDDP by suppressing OCT2 expression in kidney and MRP2 in tumors. This project was reviewed and approved by the Animal Ethics Committee of the First Hospital of Jilin University. The effects of AS IV on CDDP inhibition of tumor growth and promotion of apoptosis were assessed in Lewis lung tumor (LLC)-bearing mice by H&E and TUNEL staining. Kidney injury was assessed by serum biochemical parameters and H&E staining. We used Western blotting and immunohistochemistry assays to detect OCT2 and MRP2 expression in kidney and tumor. The concentration of CDDP in kidney and tumor was measured by HPLC-MS/MS. AS IV enhanced CDDP chemosensitivity by increasing tumor cell apoptosis and slowing tumor growth, and decreased kidney injury as evidenced by lower blood creatinine (Cr) and blood urea nitrogen (BUN). Co-administration of AS IV suppressed MRP2 overexpression induced by CDDP in tumor tissues and may be an important mechanism for enhancing CDDP chemosensitivity. Moreover, AS IV reduced CDDP-induced kidney injury in mice along with suppression of OCT2 expression in kidney. The concentration of CDDP was increased in tumor but decreased in kidney. In total, AS IV not only enhanced the antitumor effect of CDDP by suppressing MRP2 expression in tumor cells, but also decreased kidney injury induced by CDDP. The results provide new insight into the combined use of a chemotherapy drug and natural ingredients to treat cancer.
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Human organic cation transporter 1 (hOCT1) is a transmembrane influx transporter protein encoded by the SLC22A1gene. hOCT1 plays a pivotal role in the hepatocellular and renal uptake of several xenobiotics and endogenoussubstrates. The human SLC22A1 gene is highly polymorphic. Non-synonymous single nucleotide polymorphisms(SNPs) of the human SLC22A1 gene tend to impair the transmembrane conductance of substrates by hOCT1. Herein,we describe the effect of 1022C>T and 1222A>G variations in the human SLC22A1 gene on hOCT1 structure andsubstrate binding. The three-dimensional (3D) structures of hOCT1 variants were ab initio models using the iTASSERserver, and drug-binding residues of the transmembrane domain were predicted using the Prankweb server. Substratebinding was analyzed by molecular docking using AutoDock 4.2.6. Amino acid residues, crucial for substrate bindingand transport, were altered in Met408Val and Pro341Leu variants and were suggestive of conformational changeinduced by 1022C>T and 1222A>G SNPs. Moreover, a statistically significant difference was observed betweenthe binding affinities of substrates to wild and mutant variants. Therefore, it is evident that 1022C>T and 1222A>Gnon-synonymous SNPs impair the drug uptake process of hOCT1, and hence patients with the former variants need tobe closely monitored for idiosyncratic adverse drug reactions or sub-therapeutic responses while being initiated intotherapy with hOCT1 substrates.
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Nitidine chloride (NC) is a compound with prominent anti-tumor activity. To determine potential cardiotoxicity of NC, this study was designed to investigate the distribution of NC in rat heart and the underlying mechanism. The animal studies were approved by Institutional Animal Care and Use Committee of Zhejiang University Medical Center (2015-380-01) and complied with the standards of animal welfare in China. At 0.25, 0.5 and 2 h after a single intravenous injection (iv) of 5 mg·kg-1 NC, the concentrations of NC in rat heart were 47.7, 71.1 and 63.2 μg·g-1 respectively, which were 576, 1 352 and 1 212 folds of that in plasma. This study also revealed that the NC concentration in heart was 458.5 μg·g-1 (7 336 folds of that in plasma) at 2 h after the last dose in rats, after daily iv administration of NC at 5 mg·kg-1·day-1 for successive 20 days. Further studies showed that the accumulations of NC in MDCK-hOCT1 and MDCK-hOCT3 cells were 16.1 and 4.99 folds higher than that of the mock cells, respectively. There is no significant difference between the accumulations of NC in MDCK cells transfected with hOCTN1, hOCTN2 or hPMAT and the mock cells. Additionally, quinidine, L-tetrahydropalmatine and Decynium 22, the inhibitors of OCTs, clearly reduced the accumulations of NC in primary cardiomyocytes and cardiac fibroblasts from neonatal rats. MTT assay showed that the LC50 of NC on cardiomyocytes and cardiac fibroblasts were 10.9 and 10.4 μmol·L-1, respectively. Moreover, treatment of the primary cardiomyocytes and cardiac fibroblasts with NC (1～15 μmol·L-1) for 48 h resulted in significantly increased LDH enzyme leakage. These results indicated that NC can be highly accumulated in the heart, and accumulation is mediated by OCT1 and OCT3, but not by OCTN1, OCTN2 and PMAT. The accumulated NC has potential cytotoxicity as shown in the results from primary cardiomyocytes and cardiac fibroblasts.
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Current study systematically investigated the interaction of two alkaloids, anisodine and monocrotaline, with organic cation transporter OCT1, 2, 3, MATE1 and MATE2-K by using in vitro stably transfected HEK293 cells. Both anisodine and monocrotaline inhibited the OCTs and MATE transporters. The lowest IC was 12.9 µmol·L of anisodine on OCT1 and the highest was 1.8 mmol·L of monocrotaline on OCT2. Anisodine was a substrate of OCT2 (K = 13.3 ± 2.6 µmol·L and V = 286.8 ± 53.6 pmol/mg protein/min). Monocrotaline was determined to be a substrate of both OCT1 (K = 109.1 ± 17.8 µmol·L, V = 576.5 ± 87.5 pmol/mg protein/min) and OCT2 (K = 64.7 ± 14.8 µmol·L, V = 180.7 ± 22.0 pmol/mg protein/min), other than OCT3 and MATE transporters. The results indicated that OCT2 may be important for renal elimination of anisodine and OCT1 was responsible for monocrotaline uptake into liver. However neither MATE1 nor MATE2-K could facilitate transcellular transport of anisodine and monocrotaline. Accumulation of these drugs in the organs with high OCT1 expression (liver) and OCT2 expression (kidney) may be expected.
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Drug transporters play vital roles in absorption, distribution and excretion of drugs. Understanding the transport activity can improve the effectiveness and safety of drugs and guide clinical rational use of drugs. Metformin is a first-line drug in the treatment of type 2 diabetes mellitus, of which the pharmacokinetics involves several transporters. The changes in expression and function of these transporters affect directly the pharmacokinetics/pharmacodynamics of metformin. This paper reviews the research progress of pharmacokinetics of metformin based on transporters, and these transporters are organic cation transporters (OCTs), multidrug and toxin extrusion proteins (MATE), plasma membrane monoamine transporter protein (PMAT), serotonin reuptake transporter (SERT), thiamine transporter 2 (THTR-2), and carnitine/organic cation 1 (OCTN1).
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Human carnitine/organic cation transporter 1 and 2(hOCTN1 and hOCTN2) mediate transport of endogenous and exogenous compounds. The present study aimed to establish cell models with stable expression of hOCTN1 or hOCTN2 to study interactions with compounds and transporters. MDCK cells were transfected with pcDNA3.1(+) plasmid vector containing hOCTN1 or hOCTN2(pcDNA3.1(+)-hOCTN1/2), several stable transfected clones were obtained after G418 screening. hOCTN1 and hOCTN2 clones were screened with ergothioneine and mildronate respectively as substrates to identify the best candidates. We explored interactions of endogenous substances, alkaloids, flavonoids and ACEIs with hOCTN1/2. As a result, the cellular accumulation of ergothioneine in MDCK-hOCTN1 or mildronate in MDCK-hOCTN2 was 122 and 108 folds of the control cells, respectively. The kinetic parameters, Km and Vmax of ergothioneine, mediated by MDCK-hOCTN1, were 8.19±0.61 μmol·L-1 and 1427±49 pmol·mg-1(protein)·min-1; while Km and Vmax of mildronate by MDCKhOCTN2 were 52.3±4.3 μmol·L-1 and 2454±64 pmol·mg-1(protein)·min-1. Dopamine, glutamine, piperine, berberine, nuciferine, lisinopril and fosinopril could inhibit ergothioneine or mildronate uptake by MDCKhOCTN1/2. In conclusion, cell models with good stable hOCTN1 and hOCTN2 functions have been established successfully, which can be applied to the study of interactions between compounds and transporters of hOCTN1 and hOCTN2.
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In recent years, a new type of organic cation transporter has been found to transport organic cation drugs like pyrilamine, diphenhydramine and oxycodone in brain capillary endothelial cells, Caco-2 cells and other cells. Its transport activity can not be inhibited by typical organic cation transporter substrates or inhibitors, and its transport characteristics are different from those reported for the organic cation transporters, such as organic cation transporters (OCTs), organic cation/carnitine transporters (OCTNs), multidrug and toxin extrusion transporters (MATE) and the plasma membrane monoamine transporter (PMAT). It is a novel organic cation transporter, called pyrilamine-sensitive H+/OC antiporter. This review will present a comprehensive summary to elaborate the transport characteristics, structure of the substrates, tissue expression and the differences with other organic cation transporters.
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OBJECTIVE:To systematically review the effect of organic cation transporter 2 [(OCT2)808G>T] gene polymor-phism on the metformin hydrochloride pharmacokinetics in vivo,and to provide evidence-based reference for clinical medication. METHODS:Retrieved from PubMed,EMBase,Foreign Medical Journey Service,CJFD,VIP database and Wanfang database,re-lated studies about the effect of (OCT2)808G>T gene polymorphism on the metformin hydrochloride pharmacokinetics in vivo were collected,and Meta-analysis was performed by using Rev Man 5.1 statistics software. RESULTS:A total of 5 retrospective studies were included,involving 172 patients. The result of gene type was type GT,type TT and type GG. Results of Meta-analysis showed,compared with type GT volunteers,type TT could prolong the half-time period of metformin hydrochloride;compared with type TT,type GG could increase the peak concentration. However,(OCT2)808G>T gene polymorphism had no effects on the renal clearance rate,creatinine clearance rate and area under the drug-time curve. CONCLUSIONS:(OCT2)808G>T gene poly-morphism has certain effect on the half-time period and peak concentration of metformin hydrochloride in vivo of health volunteer, and has no effect on the renal clearance rate,creatinine clearance rate and area under the drug-time curve. Due to the limit of re-search methodological quality,large-scale and high quality studies are required for further validation of the conclusions.
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Objective To investigate the associations between polymorphisms of organic cation transporter OCTN1/2 (organic cation transporter 1/2) and the susceptibility of Crohn' s disease (CD) through a meta-analysis.Methods Databases of PubMed,EMBASE,MedLine,and CNKI (Chinese),Wanfang (Chinese) were searched for published case control studies on the association between polymorphisms of OCTN1/2 gene and the susceptibility of CD which were published before September 2012.The meta-analysis was applied with Review Manager 4.2 software and Stata 10.0 software.Results Nineteen eligible studies,including 14 from Europeans,3 from Asians,1 from Oceania,and 1 from the US were included in the meta-analysis.In total,significant associations were found between OCTN1/2 polymorphisms and the susceptibility of CD for all genetic models.In subgroup analyses,significant associations were found in the European population for OCTN1/2.Associations were not significant in the non-European population for OCTN1 (TT vs.CT:OR=1.25,95％CI:0.75-1.98,P=0.34; TT vs.CC+CT:OR=l.48,95％CI:0.95-2.29,P=0.08) and for OCTN2 (CC vs.GC:OR=1.03,95％CI:0.68-1.56,P=0.89; CC vs.GG + GC:OR=1.23,95％CI:0.83-1.82,P=0.31).However,there were significant associations found between OCTN1/2(TT vs.CC,TT+CT vs.CC,CC vs.GG,CC + GC vs.GG) polymorphisms and the susceptibility of CD found in the nonEuropean population.Conclusion Results from this meta-analysis suggested that OCTN1/2 polymorphisms were associated with the susceptibility of CD in the European population.Associations between OCTN1/2 polymorphisms and the susceptibility of CD in the non-European population required searching for large samples to confirm the findings.
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The organic cation transport systems were initially recognized in studies of renal elimination.A variety of endogenous and exogenous compounds that are harmful to the body are eliminated from the kidney via glomerular filtration and/or active tubular secretion.Among those compounds are polar organic cations that require a transporter-mediated process to cross cellular membrane and be eventually excreted into the urine.The organic cation transport systems in the kidney have been recognized for several decades.Following the insight gained from the kidney,organic cation transporters (OCTs) have also been characterized in other tissues.In particular,the OCTs in the liver and in the intestine have been proposed as important determinants of drug absorption and disposition.Over the past several decades,the mechanisms of organic cation transport have been extensively characterized using in vivo models,ex vivo organ perfusions,in vitro tissue preparations and cell lines.In particular,the molecular cloning and characterization of OCTs have remarkably increased our knowledge of this important subfamily of solute carriers.In this article,we first review the molecular cloning and characterization of OCTs.We will then describe their transport mechanisms,tissue distribution and localization,and their specificity of interaction with organic cations.Regulatory mechanisms and genetic animal models will be reviewed as they provide the most important insight on this class of transporters in recent years.Finally,we will summarize genetic variation in human OCT genes and raise clinical perspectives on these important transporters.
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A function of the kidney is elimination of a variety of xenobiotics ingested and wasted endogenous compounds from the body. Organic anion and cation transport systems play important roles to protect the body from harmful substances. The renal proximal tubule is the primary site of carrier-mediated transport from blood into urine. During the last decade, molecular cloning has identified several families of multispecific organic anion and cation transporters, such as organic anion transporter (OAT), organic cation transporter (OCT), and organic anion-transporting polypeptide (oatp). Additional findings also suggested ATP-dependent organic ion transporters such as MDR1/P-glycoprotein and the multidrug resistance-associated protein (MRP) as efflux pump. The substrate specificity of these transporters is multispecific. These transporters also play an important role as drug transporters. Studies on their functional properties and localization provide information in renal handling of drugs. This review summarizes the latest knowledge on molecular properties and pharmacological significance of renal organic ion transporters.