RESUMEN
Oleanane pentacyclic triterpenoids have been widely used in clinical practice. However, studies on their interactions with hepatic transporters remain limited. In this study, we systematically investigated the inhibitory effects of 14 oleanane pentacyclic triterpenoids on organic anion transporting polypeptide 1B1 and 1B3 (OATP1B1 and OATP1B3), two liver-specific uptake transporters. Through fluorescence-based cellular uptake assays, we identified three potent OATP1B1 inhibitors (saikosaponin B1, saikosaponin A and 18ß-glycyrrhetinic acid) and five potent OATP1B3 inhibitors (echinocystic acid, 3-oxo-16α-hydroxy-olean-12-en-28ß-oic acid, chikusetsu saponin IVa, saikosaponin B1 and 18ß-glycyrrhetinic acid). Structural analysis revealed that free oleanane triterpenoids inhibited OATP1B1/1B3 more potently than triterpene glycosides. Despite their similar structures, 18ß-glycyrrhetinic acid exhibited much stronger inhibition on OATP1B1/1B3 than 18α-glycyrrhetinic acid, while both were substrates of OATP1B3. Interestingly, OATP1B3 overexpression significantly increased reactive oxygen species (ROS) levels in HepG2 cells after treatment with 18ß-glycyrrhetinic acid. To conclude, this study highlights the potential interactions of oleanane pentacyclic triterpenoids with OATP1B1/1B3, and provides novel insights into the anti-cancer activity of 18ß-glycyrrhetinic acid.
Asunto(s)
Transportador 1 de Anión Orgánico Específico del Hígado , Ácido Oleanólico , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Humanos , Ácido Oleanólico/análogos & derivados , Ácido Oleanólico/farmacología , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Células HEK293 , Células Hep G2 , Saponinas/farmacología , Ácido Glicirretínico/farmacología , Ácido Glicirretínico/análogos & derivadosRESUMEN
Telmisartan, a selective inhibitor of angiotensin II receptor type 1 (AT1), demonstrates nonlinear pharmacokinetics (PK) when orally administered in ascending doses to healthy volunteers, but the underlying mechanisms remain unclear. This study presents a physiologically based pharmacokinetic model integrated with target-mediated drug disposition (TMDD-PBPK model) to explore the mechanism of its nonlinear PK. We employed the Cluster-Gauss Newton method for top-down analysis, estimating the in vivo Km,OATP1B3 (Michaelis-Menten constant for telmisartan hepatic uptake via Organic Anion Transporting Polypeptide 1B3) to be 2.0-5.7 nM. This range is significantly lower than the reported in vitro value of 810 nM, obtained in 0.3% human serum albumin (HSA) conditions. Further validation was achieved through in vitro assessment in plated human hepatocytes with 4.5% HSA, showing a Km of 4.5 nM. These results underscore the importance of albumin-mediated uptake effect for the hepatic uptake of telmisartan. Our TMDD-PBPK model, developed through a "middle-out" approach, underwent sensitivity analysis to identify key factors in the nonlinear PK of telmisartan. We found that the nonlinearity in the area under the concentration-time curve (AUC) and/or maximum concentration (Cmax) of telmisartan is sensitive to Km,OATP1B3 across all dosages. Additionally, the dissociation constant (Kd) for telmisartan binding to the AT1 receptor, along with its receptor abundance, notably influences PK at lower doses (below 20 mg). In conclusion, the nonlinear PK of telmisartan appears primarily driven by hepatic uptake saturation across all dose ranges and by AT1-receptor binding saturation, notably at lower doses.
Asunto(s)
Hepatocitos , Modelos Biológicos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Telmisartán , Telmisartán/farmacocinética , Telmisartán/administración & dosificación , Humanos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/antagonistas & inhibidores , Hepatocitos/metabolismo , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacocinética , Bloqueadores del Receptor Tipo 1 de Angiotensina II/administración & dosificación , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Hígado/metabolismo , Dinámicas no Lineales , Bencimidazoles/farmacocinética , Bencimidazoles/administración & dosificación , Benzoatos/farmacocinética , Benzoatos/administración & dosificación , Voluntarios Sanos , Administración OralRESUMEN
OBJECTIVE: To explore the characteristics of SLCO1B1/SLCO1B3 gene variants among children with Rotor syndrome (RS). METHODS: Four children who were admitted to the Department of Hepatology of Hunan Children's Hospital between January 2019 and January 2022 were selected as the study subjects. Trio-whole exome sequencing was carried out for the four families, and gel electrophoresis was used to verify an insertional variant of long-interspersed element-1 (LINE-1). RESULTS: Genetic testing has identified three variants of the SLCO1B1 gene, including c.1738C>T (p.R580*), c.757C>T (p.R253*) and c.1622A>C (p.Q541P), and two variants of the SLCO1B3 gene, including c.481+22insLINE-1 and c.1747+1G>A among the children. Three of them were found to harbor homozygous variants of the SLCO1B1/SLCO1B3 genes, and one has harbored compound heterozygous variants. Sanger sequencing confirmed the existence of all variants, and gel electrophoresis has confirmed the existence of the LINE-1 insertional variant of about 6 kb within intron 6 of the SLCO1B3 gene in all children. CONCLUSION: The pathogenesis of the RS among the four children may be attributed to the variants of the SLCO1B1/SLCO1B3 genes. The LINE-1 insertion variant of the SLCO1B3 gene may be common among Chinese RS patients.
Asunto(s)
Pruebas Genéticas , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Humanos , Masculino , Femenino , Niño , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Preescolar , Pruebas Genéticas/métodos , Transportador 1 de Anión Orgánico Específico del Hígado/genética , Secuenciación del Exoma , Lactante , MutaciónRESUMEN
BACKGROUND AND OBJECTIVE: The prediction of pharmacokinetic parameters for drugs metabolised by cytochrome P450 enzymes has been the subject of active research for many years, while the application of in vitro-in vivo extrapolation (IVIVE) techniques for non-cytochrome P450 enzymes has not been thoroughly evaluated. There is still no established quantitative method for predicting hepatic clearance of drugs metabolised by uridine 5'-diphospho-glucuronosyltransferases (UGTs), not to mention those which undergo hepatic uptake. The objective of the study was to predict the human hepatic clearance for telmisartan based on in vitro metabolic stability and hepatic uptake results. METHODS: Telmisartan was examined in liver systems, allowing to estimate intrinsic clearance (CLint, in vitro) based on the substrate disappearance rate with the use of liquid chromatography tandem mass spectrometry (LC-MS/MS) technique. Obtained CLint, in vitro values were corrected for corresponding unbound fractions. Prediction of human hepatic clearance was made from scaled unbound CLint, in vitro data with the use of the well-stirred model, and finally referenced to the literature value of observed clearance in humans, allowing determination of the essential scaling factors. RESULTS: The in vitro scaled CLint, in vitro by UGT1A3 was assessed using three systems, human hepatocytes, liver microsomes, and recombinant enzymes. Obtained values were scaled and hepatic metabolism clearance was predicted, resulting in significant clearance underprediction. Utilization of the extended clearance concept (ECC) and hepatic uptake improved prediction of hepatic metabolism clearance. The scaling factors for hepatocytes, assessing the in vitro-in vivo difference, changed from sixfold difference to only twofold difference with the application of the ECC. CONCLUSIONS: The study showed that taking into consideration hepatic uptake of a drug allows us to obtain satisfactory scaling factors, hence enabling the prediction of in vivo hepatic glucuronidation from in vitro data.
Asunto(s)
Glucurónidos , Glucuronosiltransferasa , Microsomas Hepáticos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Telmisartán , Glucuronosiltransferasa/metabolismo , Telmisartán/farmacocinética , Telmisartán/metabolismo , Humanos , Microsomas Hepáticos/metabolismo , Glucurónidos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Hígado/metabolismo , Hígado/enzimología , Tasa de Depuración Metabólica , Espectrometría de Masas en Tándem/métodos , Hepatocitos/metabolismo , Modelos Biológicos , Cromatografía Liquida/métodos , Benzoatos/farmacocinética , Benzoatos/metabolismoRESUMEN
BACKGROUND: Refractoriness to surgical resection and chemotherapy makes intrahepatic cholangiocarcinoma (ICC) a fatal cancer of the digestive system with high mortality and poor prognosis. Important function invests circRNAs with tremendous potential in biomarkers and therapeutic targets. Nevertheless, it is still unknown how circRNAs contribute to the evolution of ICC. METHODS: CircRNAs in paired ICC and adjacent tissues were screened by circRNAs sequencing. To explore the impact of circRNAs on ICC development, experiments involving gain and loss of function were conducted. Various experimental techniques, including quantitative real-time PCR (qPCR), western blotting, RNA immunoprecipitation (RIP), luciferase reporter assays, RNA pull-down, chromatin immunoprecipitation (ChIP), ubiquitination assays and so on were employed to identify the molecular regulatory role of circRNAs. RESULTS: Herein, we reported a new circRNA, which originates from exon 9 to exon 15 of the SLCO1B3 gene (named circSLCO1B3), orchestrated ICC progression by promoting tumor proliferation, metastasis and immune evasion. We found that the circSLCO1B3 gene was highly overexpressed in ICC tissues and related to lymphatic metastasis, tumor sizes, and tumor differentiation. Mechanically, circSLCO1B3 not only promoted ICC proliferation and metastasis via miR-502-5p/HOXC8/SMAD3 axis, but also eradicated anti-tumor immunity via suppressing ubiquitin-proteasome-dependent degradation of PD-L1 by E3 ubiquitin ligase SPOP. We further found that methyltransferase like 3 (METTL3) mediated the m6A methylation of circSLCO1B3 and stabilizes its expression. Our findings indicate that circSLCO1B3 is a potential prognostic marker and therapeutic target in ICC patients. CONCLUSIONS: Taken together, m6A-modified circSLCO1B3 was correlated with poor prognosis in ICC and promoted ICC progression not only by enhancing proliferation and metastasis via potentiating HOXC8 expression, but also by inducing immune evasion via antagonizing PD-L1 degradation. These results suggest that circSLCO1B3 is a potential prognostic marker and therapeutic target for ICC.
Asunto(s)
Neoplasias de los Conductos Biliares , Colangiocarcinoma , Metiltransferasas , ARN Circular , Humanos , Antígeno B7-H1/genética , Antígeno B7-H1/metabolismo , Neoplasias de los Conductos Biliares/genética , Neoplasias de los Conductos Biliares/metabolismo , Neoplasias de los Conductos Biliares/patología , Línea Celular Tumoral , Proliferación Celular/genética , Colangiocarcinoma/genética , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patología , Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Proteínas Nucleares/metabolismo , Pronóstico , Proteínas Represoras/metabolismo , ARN/metabolismo , ARN Circular/genética , ARN Circular/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genéticaRESUMEN
Amanita phalloides is one of the deadliest mushrooms worldwide, causing most fatal cases of mushroom poisoning. Among the poisonous substances of Amanita phalloides, amanitins are the most lethal toxins to humans. Currently, there are no specific antidotes available for managing amanitin poisoning and treatments are lack of efficacy. Amanitin mainly causes severe injuries to specific organs, such as the liver, stomach, and kidney, whereas the lung, heart, and brain are hardly affected. However, the molecular mechanism of this phenomenon remains not understood. To explore the possible mechanism of organ specificity of amanitin-induced toxicity, eight human cell lines derived from different organs were exposed to α, ß, and γ-amanitin at concentrations ranging from 0.3 to 100 µM. We found that the cytotoxicity of amanitin differs greatly in various cell lines, among which liver-derived HepG2, stomach-derived BGC-823, and kidney-derived HEK-293 cells are most sensitive. Further mechanistic study revealed that the variable cytotoxicity is mainly dependent on the different expression levels of the organic anion transporting polypeptide 1B3 (OATP1B3), which facilitates the internalization of amanitin into cells. Besides, knockdown of OATP1B3 in HepG2 cells prevented α-amanitin-induced cytotoxicity. These results indicated that OATP1B3 may be a crucial therapeutic target against amanitin-induced organ failure.
Asunto(s)
Amanitinas , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Humanos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Amanitinas/toxicidad , Células HEK293 , Línea Celular , Supervivencia Celular/efectos de los fármacos , Alfa-Amanitina/toxicidad , Células Hep G2RESUMEN
Organic anion-transporting polypeptides (OATP) 1B1 and OATP1B3, encoded by the SLCO gene family of the solute carrier superfamily, are involved in the disposition of many exogenous and endogenous compounds. Preclinical rodent models help assess risks of pharmacokinetic interactions, but interspecies differences in transporter orthologs and expression limit direct clinical translation. An OATP1B transgenic mouse model comprising a rodent Slco1a/1b gene cluster knockout and human SLCO1B1 and SLCO1B3 gene insertions provides a potential physiologically relevant preclinical tool to predict pharmacokinetic interactions. Pharmacokinetics of exogenous probe substrates, pitavastatin and pravastatin, and endogenous OATP1B biomarkers, coproporphyrin-I and coproporphyrin-III, were determined in the presence and absence of known OATP/Oatp inhibitors, rifampin or silymarin (an extract of milk thistle [Silybum marianum]), in wild-type FVB mice and humanized OATP1B mice. Rifampin increased exposure of pitavastatin (4.6- and 2.8-fold), pravastatin (3.6- and 2.2-fold), and coproporphyrin-III (1.6- and 2.1-fold) in FVB and OATP1B mice, respectively, but increased coproporphyrin-I AUC0-24h only (1.8-fold) in the OATP1B mice. Silymarin did not significantly affect substrate AUC, likely because the silymarin flavonolignan concentrations were at or below their reported IC50 values for the relevant OATPs/Oatps. Silymarin increased the Cmax of pitavastatin 2.7-fold and pravastatin 1.9-fold in the OATP1B mice. The data of the OATP1B mice were similar to those of the pitavastatin and pravastatin clinical data; however, the FVB mice data more closely recapitulated pitavastatin clinical data than the data of the OATP1B mice, suggesting that the OATP1B mice are a reasonable, though costly, preclinical strain for predicting pharmacokinetic interactions when doses are optimized to achieve clinically relevant plasma concentrations.
Asunto(s)
Interacciones Farmacológicas , Transportador 1 de Anión Orgánico Específico del Hígado , Ratones Transgénicos , Pravastatina , Rifampin , Silimarina , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Animales , Rifampin/farmacocinética , Ratones , Transportador 1 de Anión Orgánico Específico del Hígado/genética , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Humanos , Silimarina/farmacocinética , Pravastatina/farmacocinética , Pravastatina/administración & dosificación , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Quinolinas/farmacocinética , Coproporfirinas/metabolismo , Masculino , Transportadores de Anión Orgánico/genética , Transportadores de Anión Orgánico/metabolismoRESUMEN
The development of genetic reporters for magnetic resonance imaging (MRI) is essential for investigating biological functions inâ vivo. However, current MRI reporters have low sensitivity, making it challenging to create significant contrast against the tissue background, especially when only a small fraction of cells express the reporter. To overcome this limitation, we developed an approach for amplifying the sensitivity of molecular MRI by combining a chemogenetic contrast mechanism with a biophysical approach to increase water diffusion through the co-expression of a dual-gene construct comprising an organic anion transporting polypeptide, Oatp1b3, and a water channel, Aqp1. We first show that the expression of Aqp1 amplifies MRI contrast in cultured cells engineered to express Oatp1b3. We demonstrate that the contrast amplification is caused by Aqp1-driven increase in water exchange, which provides the gadolinium ions internalized by Oatp1b3-expressing cells with access to a larger water pool compared with exchange-limited conditions. We further show that our methodology allows cells to be detected using approximately 10-fold lower concentrations of gadolinium than that in the Aqp1-free scenario. Finally, we show that our approach enables the imaging of mixed-cell cultures containing a low fraction of Oatp1b3-labeled cells that are undetectable on the basis of Oatp1b3 expression alone.
Asunto(s)
Acuaporina 1 , Genes Reporteros , Imagen por Resonancia Magnética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Agua , Agua/química , Humanos , Imagen por Resonancia Magnética/métodos , Acuaporina 1/metabolismo , Acuaporina 1/genética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Gadolinio/química , Medios de Contraste/química , Medios de Contraste/metabolismo , Células HEK293 , AnimalesRESUMEN
The addition of darolutamide, an androgen receptor signalling inhibitor, to therapy with docetaxel has recently been approved as a strategy to treat metastatic prostate cancer. OATP1B3 is an SLC transporter that is highly expressed in prostate cancer and is responsible for the accumulation of substrates, including docetaxel, into tumours. Given that darolutamide inhibits OATP1B3 in vitro, we sought to characterise the impact of darolutamide on docetaxel pharmacokinetics. We investigated the influence of darolutamide on OATP1B3 transport using in vitro and in vivo models. We assessed the impact of darolutamide on the tumour accumulation of docetaxel in a patient-derived xenograft (PDX) model and on an OATP1B biomarker in patients. Darolutamide inhibited OATP1B3 in vitro at concentrations higher than the reported Cmax. Consistent with these findings, in vivo studies revealed that darolutamide does not influence the pharmacokinetics of Oatp1b substrates, including docetaxel. Docetaxel accumulation in PDX tumours was not decreased in the presence of darolutamide. Metastatic prostate cancer patients had similar levels of OATP1B biomarkers, regardless of treatment with darolutamide. Consistent with a low potential to inhibit OATP1B3-mediated transport in vitro, darolutamide does not significantly impede the transport of Oatp1b substrates in vivo or in patients. Our findings support combined treatment with docetaxel and darolutamide, as no OATP1B3 transporter based drug-drug interaction was identified.
Asunto(s)
Docetaxel , Neoplasias de la Próstata , Pirazoles , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Ensayos Antitumor por Modelo de Xenoinjerto , Humanos , Masculino , Docetaxel/farmacología , Docetaxel/farmacocinética , Animales , Ratones , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Pirazoles/farmacología , Pirazoles/farmacocinética , Interacciones Farmacológicas , Línea Celular Tumoral , Células HEK293RESUMEN
BACKGROUND: In predictions about hepatic clearance (CLH), a number of studies explored the role of albumin and transporters in drug uptake by liver cells, challenging the traditional free-drug theory. It was proposed that liver uptake can occur for transporter substrate compounds not only from the drug's unbound form but also directly from the drug-albumin complex, a phenomenon known as uptake facilitated by albumin. In contrast to albumin, dextran does not exhibit binding properties for compounds. However, as a result of its inherent capacity for stabilization, it is widely used to mimic conditions within cells. METHODS: The uptake of eight known substrates of the organic anion-transporting polypeptide 1B3 (OATP1B3) was assessed using a human embryonic kidney cell line (HEK293), which stably overexpresses this transporter. An inert polymer, dextran, was used to simulate cellular conditions, and the results were compared with experiments involving human plasma and human serum albumin (HSA). RESULTS: This study is the first to demonstrate that dextran increases compound uptake in cells with overexpression of the OATP1B3 transporter. Contrary to the common theory that highly protein-bound ligands interact with hepatocytes to increase drug uptake, the results indicate that dextran's interaction with test compounds does not significantly increase concentrations near the cell membrane surface. CONCLUSIONS: We evaluated the effect of dextran on the uptake of known substrates using OATP1B3 overexpressed in the HEK293 cell line, and we suggest that its impact on drug concentrations in liver cells may differ from the traditional role of plasma proteins and albumin.
Asunto(s)
Dextranos , Transportadores de Anión Orgánico , Humanos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/farmacología , Transportador 1 de Anión Orgánico Específico del Hígado/genética , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/farmacología , Células HEK293 , Transportadores de Anión Orgánico/genética , Transportadores de Anión Orgánico/metabolismo , Hepatocitos/metabolismo , Hígado , Proteínas de Transporte de Membrana/metabolismo , Albúminas , Transportadores de Anión Orgánico Sodio-Independiente/genética , Transportadores de Anión Orgánico Sodio-Independiente/metabolismoRESUMEN
Organic anion transporting polypeptide (OATP) 1B1 and OATP1B3 (collectively, OATP1B) transporters encoded by the solute carrier organic anion transporter (SLCO) genes mediate uptake of multiple pharmaceutical compounds. Nonalcoholic steatohepatitis (NASH), a severe form of nonalcoholic fatty liver disease (NAFLD), decreases OATP1B abundance. This research characterized the pathologic and pharmacokinetics effects of three diet- and one chemical-induced NAFLD model in male and female humanized OATP1B mice, which comprises knock-out of rodent Oatp orthologs and insertion of human SLCO1B1 and SLCO1B3. Histopathology scoring demonstrated elevated steatosis and inflammation scores for all NAFLD-treatment groups. Female mice had minor changes in SLCO1B1 expression in two of the four NAFLD treatment groups, and pitavastatin (PIT) area under the concentration-time curve (AUC) increased in female mice in only one of the diet-induced models. OATP1B3 expression decreased in male and female mice in the chemical-induced NAFLD model, with a coinciding increase in PIT AUC, indicating the chemical-induced model may better replicate changes in OATP1B3 expression and OATP substrate disposition observed in NASH patients. This research also tested a reported multifactorial pharmacokinetic interaction between NAFLD and silymarin, an extract from milk thistle seeds with notable OATP-inhibitory effects. Males showed no change in PIT AUC, whereas female PIT AUC increased 1.55-fold from the diet alone and the 1.88-fold from the combination of diet with silymarin, suggesting that female mice are more sensitive to pharmacokinetic changes than male mice. Overall, the humanized OATP1B model should be used with caution for modeling NAFLD and multifactorial pharmacokinetic interactions. SIGNIFICANCE STATEMENT: Advanced stages of NAFLD cause decreased hepatic OATP1B abundance and increase systemic exposure to OATP substrates in human patients. The humanized OATP1B mouse strain may provide a clinically relevant model to recapitulate these observations and predict pharmacokinetic interactions in NAFLD. This research characterized three diet-induced and one drug-induced NAFLD model in a humanized OATP1B mouse model. Additionally, a multifactorial pharmacokinetic interaction was observed between silymarin and NAFLD.
Asunto(s)
Enfermedad del Hígado Graso no Alcohólico , Transportadores de Anión Orgánico , Silimarina , Humanos , Masculino , Femenino , Ratones , Animales , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Ratones Transgénicos , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Hígado/metabolismo , Transportadores de Anión Orgánico/metabolismo , Proteínas de Transporte de Membrana/metabolismo , Silimarina/metabolismo , Interacciones FarmacológicasRESUMEN
Daprodustat is the first oral hypoxia-inducible factor prolyl hydroxylase inhibitor approved recently for the treatment of anemia caused by chronic kidney disease (CKD) in adults receiving dialysis. We evaluated the role of organic anion transporting polypeptide (OATP)1B-mediated hepatic uptake transport in the pharmacokinetics (PKs) of daprodustat using in vitro and in vivo studies, and physiologically-based PK (PBPK) modeling of its drug-drug interactions (DDIs) with inhibitor drugs. In vitro, daprodustat showed specific transport by OATP1B1/1B3 in the transfected cell systems and primary human and monkey hepatocytes. A single-dose oral rifampin (OATP1B inhibitor) reduced daprodustat intravenous clearance by a notable 9.9 ± 1.2-fold (P < 0.05) in cynomolgus monkeys. Correspondingly, volume of distribution at steady-state was also reduced by 5.0 ± 1.1-fold, whereas the half-life change was minimal (1.5-fold), corroborating daprodustat hepatic uptake inhibition by rifampin. A PBPK model accounting for OATP1B-CYP2C8 interplay was developed, which well described daprodustat PK and DDIs with gemfibrozil (CYP2C8 and OATP1B inhibitor) and trimethoprim (weak CYP2C8 inhibitor) within 25% error of the observed data in healthy subjects. About 18-fold increase in daprodustat area under the curve (AUC) following gemfibrozil treatment was found to be associated with strong CYP2C8 inhibition and moderate OATP1B inhibition. Moreover, PK modulation in hepatic dysfunction and subjects with CKD, in comparison to healthy control, was well-captured by the model. CYP2C8 and/or OATP1B inhibitor drugs (e.g., gemfibrozil, clopidogrel, rifampin, and cyclosporine) were predicted to perpetrate moderate-to-strong DDIs in healthy subjects, as well as, in target CKD population. Daprodustat can be used as a sensitive CYP2C8 index substrate in the absence of OATP1B modulation.
Asunto(s)
Citocromo P-450 CYP2C8 , Interacciones Farmacológicas , Hepatocitos , Transportador 1 de Anión Orgánico Específico del Hígado , Insuficiencia Renal Crónica , Rifampin , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Adulto , Animales , Femenino , Humanos , Masculino , Citocromo P-450 CYP2C8/metabolismo , Inhibidores del Citocromo P-450 CYP2C8/farmacocinética , Glicina/análogos & derivados , Glicina/farmacocinética , Células HEK293 , Hepatocitos/metabolismo , Hepatocitos/efectos de los fármacos , Hepatopatías/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/antagonistas & inhibidores , Macaca fascicularis , Insuficiencia Renal Crónica/metabolismo , Rifampin/farmacología , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/antagonistas & inhibidoresRESUMEN
Sotorasib is a small molecule that irreversibly inhibits the Kirsten rat sarcoma viral oncogene homolog (KRAS) protein with a G12C amino acid substitution mutant protein. The impact of cytochrome P450 (CYP) 3A4 inhibition and induction on sotorasib pharmacokinetics (PKs) was evaluated in 2 separate studies in healthy volunteers (N = 14/study). The impact of CYP3A4 inhibition was interrogated utilizing repeat doses of 200 mg of itraconazole, a strong CYP3A4 inhibitor, on 360 mg of sotorasib PKs. The impact of CYP3A4 induction was interrogated utilizing multiple doses of 600 mg of rifampin, a strong CYP3A4 inducer. Additionally, the impact of organic anion transporting polypeptide (OATP) 1B1/3 inhibition on 960 mg of sotorasib PKs was interrogated after a single dose of 600 mg of rifampin. CYP3A4 inhibition did not significantly impact sotorasib Cmax but did lead to a 26% increase in sotorasib AUCinf. CYP3A4 induction decreased sotorasib Cmax by 35% and AUCinf by 51%. OATP1B1/3 inhibition decreased sotorasib Cmax and AUCinf by 16% and 23%, respectively. These results support that sotorasib can be given together with strong CYP3A4 and OATP1B1/3 inhibitors but the co-administration of sotorasib and strong CYP3A4 inducers should be avoided.
Asunto(s)
Inductores del Citocromo P-450 CYP3A , Inhibidores del Citocromo P-450 CYP3A , Citocromo P-450 CYP3A , Interacciones Farmacológicas , Transportador 1 de Anión Orgánico Específico del Hígado , Rifampin , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos , Humanos , Transportador 1 de Anión Orgánico Específico del Hígado/antagonistas & inhibidores , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Masculino , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/antagonistas & inhibidores , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Adulto , Inhibidores del Citocromo P-450 CYP3A/farmacología , Inhibidores del Citocromo P-450 CYP3A/administración & dosificación , Citocromo P-450 CYP3A/metabolismo , Rifampin/farmacología , Rifampin/administración & dosificación , Inductores del Citocromo P-450 CYP3A/farmacología , Femenino , Adulto Joven , Administración Oral , Persona de Mediana Edad , Voluntarios Sanos , Área Bajo la Curva , Itraconazol/farmacología , Itraconazol/administración & dosificación , Itraconazol/farmacocinética , Compuestos de Espiro/farmacocinética , Compuestos de Espiro/administración & dosificación , Compuestos de Espiro/farmacologíaRESUMEN
Intravenously administered chemotherapeutic cabazitaxel is used for palliative treatment of prostate cancer. An oral formulation would be more patient-friendly and reduce the need for hospitalization. We therefore study determinants of the oral pharmacokinetics of cabazitaxel in a ritonavir-boosted setting, which reduces the CYP3A-mediated first-pass metabolism of cabazitaxel. We here assessed the role of organic anion-transporting polypeptides (OATPs) in the disposition of orally boosted cabazitaxel and its active metabolites, using the Oatp1a/b-knockout and the OATP1B1/1B3-transgenic mice. These transporters may substantially affect plasma clearance and hepatic and intestinal drug disposition. The pharmacokinetics of cabazitaxel and DM2 were not significantly affected by Oatp1a/b and OATP1B1/1B3 activity. In contrast, the plasma AUC0-120 min of DM1 in Oatp1a/b-/- was 1.9-fold (p < 0.05) higher than that in wild-type mice, and that of docetaxel was 2.4-fold (p < 0.05) higher. We further observed impaired hepatic uptake and intestinal disposition for DM1 and docetaxel in the Oatp-ablated strains. None of these parameters showed rescue by the OATP1B1 or -1B3 transporters in the humanized mouse strains, suggesting a minimal role of OATP1B1/1B3. Ritonavir itself was also a potent substrate for mOatp1a/b, showing a 2.9-fold (p < 0.0001) increased plasma AUC0-120 min and 3.5-fold (p < 0.0001) decreased liver-to-plasma ratio in Oatp1a/b-/- compared to those in wild-type mice. Furthermore, we observed the tight binding of cabazitaxel and its active metabolites, including docetaxel, to plasma carboxylesterase (Ces1c) in mice, which may complicate the interpretation of pharmacokinetic and pharmacodynamic mouse studies. Collectively, these results will help to further optimize (pre)clinical research into the safety and efficacy of orally applied cabazitaxel.
Asunto(s)
Transportadores de Anión Orgánico Sodio-Independiente , Transportadores de Anión Orgánico , Taxoides , Animales , Humanos , Masculino , Ratones , Carboxilesterasa/metabolismo , Docetaxel , Hígado/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Ratones Transgénicos , Transportadores de Anión Orgánico/metabolismo , Transportadores de Anión Orgánico Sodio-Independiente/metabolismo , Ritonavir , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismoRESUMEN
The organic anion-transporting polypeptide 1B3 and P-glycoprotein (P-gp) provide efficient directional transport (OATP1B3-P-gp) from the blood to the bile that serves as a key determinant of hepatic disposition of the drug. Unfortunately, there is still a lack of effective means to evaluate the disposal ability mediated by transporters. The present study was designed to identify a suitable endogenous biomarker for the assessment of OATP1B3-P-gp function in the liver. We established stably transfected HEK293T-OATP1B3 and HEK293T-P-gp cell lines. Results showed that azelaic acid (AzA) was an endogenous substrate for OATP1B3 and P-gp using serum pharmacology combined with metabolomics. There is a good correlation between the serum concentration of AzA and probe drugs of rOATP1B3 and rP-gp when rats were treated with their inhibitors. Importantly, after 5-fluorouracil-induced rat liver injury, the relative mRNA level and expression of rOATP1B3 and rP-gp were markedly down-regulated in the liver, and the serum concentration of AzA was significantly increased. These observations suggest that AzA is an endogenous substrate of both OATP1B3 and P-gp, and may serve as a potential endogenous biomarker for the assessment of the function of OATP1B3-P-gp for the prediction of changes in the pharmacokinetics of drugs transported by OATP1B3-P-gp in liver disease states.
Asunto(s)
Ácidos Dicarboxílicos , Hígado , Metabolómica , Animales , Humanos , Ratas , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Biomarcadores , Células HEK293 , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones OrgánicosRESUMEN
The protein phosphatase inhibitor microcystin-LR (MC-LR), a hepatocyte-selective cyanotoxin, induces phenotypic changes in HEK293 OATP1B3-expressing (HEK293-OATP1B3) cells, which include cytoskeletal reorganization (HEK293-OATP1B3-AD) and anoikis resistance (HEK293-OATP1B3-FL) transformed cells, respectively. These cells acquire resistance to MC-LR and partial epithelial-mesenchymal transition (EMT) characteristics. In cancer cells, EMT is generally involved in multi-drug resistance. Here, we focused on the multi-drug resistance of HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. The MTT assay and immunoblotting were conducted to examine the responses of HEK293-OATP1B3, HEK293-OATP1B3-AD, and HEK293-OATP1B3-FL cells to multiple toxins and drugs that function as substrates for OATP1B3, including MC-LR, nodularin (Nod), okadaic acid (OA), and cisplatin (CDDP). HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells were more resistant to MC-LR, Nod, and OA than HEK293-OATP1B3 cells. Conversely, the three cell types were equivalently sensitive to CDDP. By using protein phosphatase assay, the reduction of the inhibitory effect of MC-LR and Nod on phosphatase activity might be one reason for the resistance to MC-LR and Nod in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. Furthermore, the parental HEK293-OATP1B3 cells showed enhanced p53 phosphorylation and stabilization after MC-LR exposure, while p53 phosphorylation was attenuated in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells. Moreover, in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells, AKT phosphorylation was higher than that of the parental HEK293-OATP1B3 cell line. These results suggest that the multi-toxin resistance observed in HEK293-OATP1B3-AD and HEK293-OATP1B3-FL cells is associated with AKT activation and p53 inactivation.
Asunto(s)
Toxinas Marinas , Transportadores de Anión Orgánico Sodio-Independiente , Proteínas Proto-Oncogénicas c-akt , Humanos , Transportadores de Anión Orgánico Sodio-Independiente/metabolismo , Transportadores de Anión Orgánico Sodio-Independiente/farmacología , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Células HEK293 , Microcistinas/metabolismo , Ácido Ocadaico/toxicidad , Transición Epitelial-Mesenquimal , Fosfoproteínas FosfatasasRESUMEN
BACKGROUND: Severe neonatal hyperbilirubinemia could lead to kernicterus and neonatal death. This study aimed to analyze the association between single nucleotide polymorphisms in genes involved in bilirubin metabolism and the incidence of severe hyperbilirubinemia. METHODS: A total of 144 neonates with severe hyperbilirubinemia and 50 neonates without or mild hyperbilirubinemia were enrolled in 3 institutions between 2019 and 2020. Twelve polymorphisms of 5 genes (UGT1A1, SLCO1B1, SLCO1B3, BLVRA, and HMOX1) were analyzed by PCR amplification of genomic DNA. Genotyping was performed using an improved multiplex ligation detection reaction technique based on ligase detection reaction. RESULTS: The frequencies of the A allele in UGT1A1-rs4148323 and the C allele in SLCO1B3-rs2417940 in the severe hyperbilirubinemia group (30.2% and 90.6%, respectively) were significantly higher than those in the controls (30.2% vs.13.0%, 90.6% vs. 78.0%, respectively, both p < 0.05). Haplotype analysis showed the ACG haplotype of UGT1A1 were associated with an increased hyperbilirubinemia risk (OR 3.122, p = 0.001), whereas the GCG haplotype was related to a reduced risk (OR 0.523, p = 0.018). CONCLUSION: The frequencies of the A allele in rs4148323 and the C allele in rs2417940 are highly associated with the incidence of severe hyperbilirubinemia in Chinese Han neonates. TRIAL REGISTRATION: Trial registration number:ChiCTR1800020424; Date of registration:2018-12-29.
Asunto(s)
Hiperbilirrubinemia Neonatal , Polimorfismo de Nucleótido Simple , Recién Nacido , Humanos , Transportador 1 de Anión Orgánico Específico del Hígado/genética , Alelos , Hiperbilirrubinemia Neonatal/genética , Glucuronosiltransferasa/genética , China/epidemiología , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/genética , Hemo-Oxigenasa 1/genética , Hemo-Oxigenasa 1/metabolismoRESUMEN
Organic anion-transporting polypeptides (OATPs) 1B1 and 1B3 are two highly homologous transport proteins. However, OATP1B1- and 1B3-mediated estradiol-17ß-glucuronide (E17ßG) uptake can be differentially affected by clotrimazole. In this study, by functional characterization on chimeric transporters and single mutants, we find that G45 in transmembrane domain 1 (TM1) and V386 in TM8 are critical for the activation of OATP1B3-mediated E17ßG uptake by clotrimazole. However, the effect of clotrimazole on the function of OATP1B3 is substrate-dependent as clotrimazole does not stimulate OATP1B3-mediated uptake of 4',5'-dibromofluorescein (DBF) and rosuvastatin. In addition, clotrimazole is not transported by OATP1B3, but it can efficiently permeate the plasma membrane due to its lipophilic properties. Homology modeling and molecular docking indicate that E17ßG binds in a substrate binding pocket of OATP1B3 through hydrogen bonding and hydrophobic interactions, among which its sterol scaffold forms hydrophobic contacts with V386. In addition, a flexible glycine residue at position 45 is essential for the activation of OATP1B3. Finally, clotrimazole is predicted to bind at an allosteric site, which mainly consists of hydrophobic residues located at the cytoplasmic halves of TMs 4, 5, 10, and 11.
Asunto(s)
Estradiol/análogos & derivados , Transportadores de Anión Orgánico Sodio-Independiente , Transportadores de Anión Orgánico , Transportadores de Anión Orgánico Sodio-Independiente/metabolismo , Clotrimazol/farmacología , Miembro 1B3 de la Familia de los Transportadores de Solutos de Aniones Orgánicos/metabolismo , Transportador 1 de Anión Orgánico Específico del Hígado/metabolismo , Simulación del Acoplamiento Molecular , Transportadores de Anión Orgánico/metabolismo , Transporte BiológicoRESUMEN
Solute carrier organic anion (SLCO) transporters (OATP transporters) are involved in cellular uptake of drugs and hormones. Germline variants in SLCO1B3 and SLCO2B1 have been implicated in prostate cancer progression and therapy response, including to androgen deprivation and statin medications, but results have appeared heterogeneous. We conducted a cohort study of five single-nucleotide polymorphisms (SNPs) in SLCO1B3 and SLCO2B1 with prior evidence among 3208 men with prostate cancer who participated in the Health Professionals Follow-up Study or the Physicians' Health Study, following participants prospectively after diagnosis over 32 years (median, 14 years) for development of metastases and cancer-specific death (lethal disease, 382 events). Results were suggestive of, but not conclusive for, associations between some SNPs and lethal disease and differences by androgen deprivation and statin use. All candidate SNPs were associated with SLCO mRNA expression in tumor-adjacent prostate tissue. We also conducted a systematic review and harmonized estimates for a dose-response meta-analysis of all available data, including 9 further studies, for a total of 5598 patients and 1473 clinical events. The A allele of the exonic SNP rs12422149 (14% prevalence), which leads to lower cellular testosterone precursor uptake via SLCO2B1, was associated with lower rates of prostate cancer progression (hazard ratio per A allele, 0.80; 95% confidence interval, 0.69-0.93), with little heterogeneity between studies (I2, 0.27). Collectively, the totality of evidence suggests a strong association between inherited genetic variation in SLCO2B1 and prostate cancer prognosis, with potential clinical use in risk stratification related to androgen deprivation therapy.