Organic Cation Transporter 1 and 3 Contribute to the High Accumulation of Dehydrocorydaline in the Heart.

Dehydrocorydaline (DHC), one of the main active components of Corydalis yanhusuo, is an important remedy for the treatment of coronary heart disease. Our previous study revealed a higher unbound concentration of DHC in the heart than plasma of mice after oral administration of C. yanhusuo extract or DHC, but the underlying uptake mechanism remains unelucidated. In our investigations, we studied the transport mechanism of DHC in transgenic cells, primary neonatal rat cardiomyocytes, and animal experiments. Using quantitative real-time polymerase chain reaction and Western blotting, we found that uptake transporters expressed in the mouse heart include organic cation transporter 1/3 (OCT1/3) and carnitine/organic cation transporter 1/2 (OCTN1/2). The accumulation experiments in transfected cells showed that DHC was a substrate of OCT1 and OCT3, with K m of 11.29 ± 3.3 and 8.96 ± 3.7 μM, respectively, but not a substrate of OCTN1/2. Additionally, a higher efflux level (1.71-fold of MDCK-mock) of DHC was observed in MDCK-MDR1 cells than in MDCK-mock cells. Therefore, DHC is a weak substrate for MDR1. Studies using primary neonatal rat cardiomyocytes showed that OCT1/3 inhibitors (quinidine, decynium-22, and levo-tetrahydropalmatine) prevented the accumulation of DHC, whereas OCTN2 inhibitors (mildronate and l-carnitine) did not affect its accumulation. Moreover, the coadministration of OCT1/3 inhibitors (levo-tetrahydropalmatine, THP) decreased the concentration of DHC in the mouse heart. Based on these findings, DHC may be accumulated partly by OCT1/3 transporters and excreted by MDR1 in the heart. THP could alter the distribution of DHC in the mouse heart. SIGNIFICANCE STATEMENT: We reported the cardiac transport mechanism of dehydrocorydaline, highly distributed to the heart after oral administration of Corydalis yanhusuo extract or dehydrocorydaline only. Dehydrocorydaline (an OCT1/3 and MDR1 substrate) accumulation in primary cardiomyocytes may be related to the transport activity of OCT1/3. This ability, hampered by selective inhibitors (levo-tetrahydropalmatine, an inhibitor of OCT1/3), causes a nearly 40% reduction in exposure of the heart to dehydrocorydaline. These results suggest that OCT1/3 may contribute to the uptake of dehydrocorydaline in the heart.

Chen Y ，Li C ，Yi Y ，Du W ，Jiang H ，Zeng S ，Zhou H ... -  《-》

The contribution of human OCT1, OCT3, and CYP3A4 to nitidine chloride-induced hepatocellular toxicity.

Nitidine chloride (NC), a quaternary ammonium alkaloid, has numerous pharmacological effects, such as anticancer activity. However, it was found that NC also has hepatocellular toxicity. Because organic cation transporters 1 and 3 (OCT1 and OCT3) might mediate the influx of NC into hepatocytes, multidrug and toxin extrusion 1 (MATE1) probably mediates the efflux of NC from hepatocytes, while cytochrome P450 (P450) enzymes might contribute to NC metabolism, the present study was to evaluate the contribution of OCT1, OCT3, MATE1, and P450 enzymes to NC-induced hepatocellular toxicity. Our results showed that the uptake of NC in Madin-Darby canine kidney (MDCK) cells expressing human (h) OCT1 and OCT3 (MDCK-hOCT1 and MDCK-hOCT3) was significantly higher than that in mock cells; the hOCT1- and hOCT3-mediated uptake followed typical Michaelis-Menten kinetics. Meanwhile, NC was also a substrate of hMATE1, although its transport capacity was much lower than that of OCT1 NC-induced cytotoxicity in MDCK-hOCT1 or MDCK-hOCT3 cells was obviously higher than that in mock cells. Quinidine and (+)-tetrahydropalmatine [(+)-THP], OCT1 and OCT3 inhibitors, significantly reduced the uptake of NC in MDCK-hOCT1 cells, MDCK-hOCT3 cells, and rat primary hepatocytes, but only (+)-THP markedly attenuated the NC-induced toxicity. In addition, P450 enzymes, such as CYP3A4, mediated the metabolism of NC, and NC-induced toxicity in MDCK-hOCT1/hCYP3A4 cells was lower than that in MDCK-hOCT1 cells. Our results indicated that NC is a substrate of hOCT1, hOCT3, and CYP3A4; that OCT1 and OCT3 mediate the uptake of NC in hepatocytes and subsequently cause hepatotoxicity; and that NC-induced toxicity could be attenuated by CYP3A4-mediated metabolism.

Li L ，Tu M ，Yang X ，Sun S ，Wu X ，Zhou H ，Zeng S ，Jiang H ... -  《-》

Utility of P-glycoprotein and organic cation transporter 1 double-transfected LLC-PK1 cells for studying the interaction of YM155 monobromide, novel small-molecule survivin suppressant, with P-glycoprotein.

Iwai M ，Minematsu T ，Li Q ，Iwatsubo T ，Usui T ... -  《-》

Differences in Metformin and Thiamine Uptake between Human and Mouse Organic Cation Transporter 1: Structural Determinants and Potential Consequences for Intrahepatic Concentrations.

The most commonly used oral antidiabetic drug, metformin, is a substrate of the hepatic uptake transporter OCT1 (gene name SLC22A1). However, OCT1 deficiency leads to more pronounced reductions of metformin concentrations in mouse than in human liver. Similarly, the effects of OCT1 deficiency on the pharmacokinetics of thiamine were reported to differ between human and mouse. Here, we compared the uptake characteristics of metformin and thiamine between human and mouse OCT1 using stably transfected human embryonic kidney 293 cells. The affinity for metformin was 4.9-fold lower in human than in mouse OCT1, resulting in a 6.5-fold lower intrinsic clearance. Therefore, the estimated liver-to-blood partition coefficient is only 3.34 in human compared with 14.4 in mouse and may contribute to higher intrahepatic concentrations in mice. Similarly, the affinity for thiamine was 9.5-fold lower in human than in mouse OCT1. Using human-mouse chimeric OCT1, we showed that simultaneous substitution of transmembrane helices TMH2 and TMH3 resulted in the reversal of affinity for metformin. Using homology modeling, we suggest several explanations, of which a different interaction of Leu155 (human TMH2) compared with Val156 (mouse TMH2) with residues in TMH3 had the strongest experimental support. In conclusion, the contribution of human OCT1 to the cellular uptake of thiamine and especially of metformin may be much lower than that of mouse OCT1. This may lead to an overestimation of the effects of OCT1 on hepatic concentrations in humans when using mouse as a model. In addition, comparative analyses of human and mouse orthologs may help reveal mechanisms of OCT1 transport. SIGNIFICANCE STATEMENT: OCT1 is a major hepatic uptake transporter of metformin and thiamine, but this study reports strong differences in the affinity for both compounds between human and mouse OCT1. Consequently, intrahepatic metformin concentrations could be much higher in mice than in humans, impacting metformin actions and representing a strong limitation of using rodent animal models for predictions of OCT1-related pharmacokinetics and efficacy in humans. Furthermore, OCT1 transmembrane helices TMH2 and TMH3 were identified to confer the observed species-specific differences in metformin affinity.

Meyer MJ ，Tuerkova A ，Römer S ，Wenzel C ，Seitz T ，Gaedcke J ，Oswald S ，Brockmöller J ，Zdrazil B ，Tzvetkov MV ... -  《-》

Interaction and Transport of Benzalkonium Chlorides by the Organic Cation and Multidrug and Toxin Extrusion Transporters.

Humans are chronically exposed to benzalkonium chlorides (BACs) from environmental sources. The U.S. Food and Drug Administration (FDA) has recently called for additional BAC safety data, as these compounds are cytotoxic and have great potential for biochemical interactions. Biodistribution studies revealed that BACs extensively distribute to many tissues and accumulate at high levels, especially in the kidneys, but the underlying mechanisms are unclear. In this study, we characterized the interactions of BACs of varying alkyl chain length (C8 to C14) with the human organic cation transporters (hOCT1-3) and multidrug and toxin extrusion proteins (hMATE1/2K) with the goal to identify transporters that could be involved in BAC disposition. Using transporter-expressing cell lines, we showed that all BACs are inhibitors of hOCT1-3 and hMATE1/2K (IC50 ranging 0.83-25.8 μM). Further, the short-chain BACs (C8 and C10) were identified as substrates of these transporters. Interestingly, although BAC C8 displayed typical Michaelis-Menten kinetics, C10 demonstrated a more complex substrate-inhibition profile. Transwell studies with transfected Madin-Darby canine kidney cells revealed that intracellular accumulation of basally applied BAC C8 and C10 was substantially higher (8.2- and 3.7-fold, respectively) in hOCT2/hMATE1 double-transfected cells in comparison with vector-transfected cells, supporting a role of these transporters in mediating renal accumulation of these compounds in vivo. Together, our results suggest that BACs interact with hOCT1-3 and hMATE1/2K as both inhibitors and substrates and that these transporters may play important roles in tissue-specific accumulation and potential toxicity of short-chain BACs. Our findings have important implications for understanding human exposure and susceptibility to BACs due to environmental exposure. SIGNIFICANCE STATEMENT: Humans are systemically exposed to benzalkonium chlorides (BACs). These compounds broadly distribute through tissues, and their safety has been questioned by the FDA. Our results demonstrate that hOCT2 and hMATE1 contribute to the renal accumulation of BAC C8 and C10 and that hOCT1 and hOCT3 may be involved in the tissue distribution of these compounds. These findings can improve our understanding of BAC disposition and toxicology in humans, as their accumulation could lead to biochemical interactions and deleterious effects.

Vieira LS ，Seguin RP ，Xu L ，Wang J ... -  《-》