Fluorescent probes for the dual investigation of MRP2 and OATP1B1 function and drug interactions

Hepatic organic anion transporting polypeptides—OATP1B1, OATP1B3, and OATP2B1—are expressed at the basolateral membrane of hepatocytes, being responsible for the uptake of a wide range of natural substrates and structurally unrelated pharmaceuticals. Impaired function of hepatic OATPs has been linked to clinically relevant drug–drug interactions leading to altered pharmacokinetics of administered drugs. Therefore, understanding the commonalities and differences across the three transporters represents useful knowledge to guide the drug discovery process at an early stage. Unfortunately, such efforts remain challenging because of the lack of experimentally resolved protein structures for any member of the OATP family. In this study, we established a rigorous computational protocol to generate and validate structural models for hepatic OATPs. The multistep procedure is based on the systematic exploration of available protein structures with shared protein folding using normal-mode analysis, the calculation of multiple template backbones from elastic network models, the utilization of multiple template conformations to generate OATP structural models with various degrees of conformational flexibility, and the prioritization of models on the basis of enrichment docking. We employed the resulting OATP models of OATP1B1, OATP1B3, and OATP2B1 to elucidate binding modes of steroid analogs in the three transporters. Steroid conjugates have been recognized as endogenous substrates of these transporters. Thus, investigating this data set delivers insights into mechanisms of substrate recognition. In silico predictions were complemented with in vitro studies measuring the bioactivity of a compound set on OATP expressing cell lines. Important structural determinants conferring shared and distinct binding patterns of steroid analogs in the three transporters have been identified. Overall, this comparative study provides novel insights into hepatic OATP-ligand interactions and selectivity. Furthermore, the integrative computational workflow for structure-based modeling can be leveraged for other pharmaceutical targets of interest.

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“…The uptake of pyranine was measured on microplates based on the method developed by us previously. 60 , 61 …”

Section: Methodsmentioning

confidence: 99%

Data-Driven Ensemble Docking to Map Molecular Interactions of Steroid Analogs with Hepatic Organic Anion Transporting Polypeptides

Tuerkova

Ungvári

Laczkó-Rigó

et al. 2021

J. Chem. Inf. Model.

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“…A431 (ATCC), HEK‐293 (ATCC), or MDCKII (ATCC) cells overexpressing OATPs, 1A2, 1B1/3 or 2B1, or mock transfected controls were generated previously as described in Refs. 19,26,30. Cell lines were cultured in DMEM (Gibco, Thermo Fischer Scientific, Waltham, MA, US) supplemented with 10% fetal calf serum, 100 U/ml penicillin, and 100 μg/ml streptomycin at 37°C, 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

8‐acetoxy‐trisulfopyrene as the first activatable fluorogenic probe for add‐and‐read assessment of Organic anion‐transporting polypeptides, OATP1B1, OATP1B3, and OATP2B1

et al. 2021

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“…For the OATP1B1 and OATP1B3 transporter assays, the interaction between potential anti-COVID-19 agents and OATPs was investigated in a microplate-based indirect assay employing pyranine as described 53 . Briefly, A431 cells (ATCC) overexpressing the OATPs 53 and mock transfected A431 cells as control were seeded on 96-well plates in a density of 8x10 4…”

Section: Methodsmentioning

confidence: 99%

Interactions of Anti-COVID-19 Drug Candidates With Hepatic Transporters May Cause Liver Toxicity and Affect Drug Metabolism

Ambrus

Bakos

Sarkadi

et al. 2021

Preprint

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Transporters in the human liver play a major role in the metabolism of endo-and xenobiotics. Apical (canalicular) transporters extrude compounds to the bile, while basolateral hepatocyte transporters promote the uptake or expel various compounds into the venous blood stream. In the present work we have examined the in vitro interactions of some key repurposed drugs advocated to treat COVID-19 (lopinavir, ritonavir, ivermectin, remdesivir and favipiravir), with the relevant key transporters in the hepatocytes. These transporters included the ABCB11/BSEP, ABCC2/MRP2, and MATE1 in the canalicular membrane, as well as ABCC3/MRP3, ABCC4/MRP4, OCT1, OATP1B1, OATP1B3, and NTCP, residing in the basolateral membrane. Lopinavir and ritonavir in low micromolar concentrations inhibited the ABCB11/BSEP and MATE1 exporters, as well as the OATP1B1/1B3 uptake transporters. Ritonavir had a similar inhibitory pattern, also inhibiting OCT1. Remdesivir strongly inhibited ABCC4/MRP4, OATP1B1/1B3, MATE1 and OCT1. Thus, these agents may cause severe drug-drug interactions and drug-induced liver injury. Favipiravir had no significant effect on any of these transporters. Since both general drug metabolism and drug-induced liver toxicity are strongly dependent on the functioning of these transporters, the variable interactions reported here may have important clinical relevance in the drug treatment of this viral disease and the existing co-morbidities.

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Fluorescent probes for the dual investigation of MRP2 and OATP1B1 function and drug interactions

Cited by 19 publications

References 65 publications

Data-Driven Ensemble Docking to Map Molecular Interactions of Steroid Analogs with Hepatic Organic Anion Transporting Polypeptides

Data-Driven Ensemble Docking to Map Molecular Interactions of Steroid Analogs with Hepatic Organic Anion Transporting Polypeptides

8‐acetoxy‐trisulfopyrene as the first activatable fluorogenic probe for add‐and‐read assessment of Organic anion‐transporting polypeptides, OATP1B1, OATP1B3, and OATP2B1

Interactions of Anti-COVID-19 Drug Candidates With Hepatic Transporters May Cause Liver Toxicity and Affect Drug Metabolism

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