Amino-terminal region of human organic anion transporting polypeptide 1B1 dictates transporter stability and substrate interaction

Wang, Xuyang; Chen, Jie; Xu, Shaopeng; Ni, Chunxue; Fang, Zihui

doi:10.1016/j.taap.2019.114642

Cited by 10 publications

(12 citation statements)

References 23 publications

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“…In experimental studies, LYS41 and ARG580 have been shown to play a role in transport activity in both OATP1B1 and OATP1B3. 29 , 68 , 69 OATP1B1 active steroids are mainly showing TMH2 (ASN77), TMH4 (ASN178 and ARG181), TMH7 (THR345) and TMH10 (GLY552) involvement in our study (see Tables 2 and Table S9 ). ARG181 was found to be a functionally important residue in OATP1B1, indicated by site-directed mutagenesis studies.…”

Section: Resultssupporting

confidence: 61%

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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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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Section: Resultssupporting

confidence: 61%

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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“…This site also contains relevant residues for the substrate transport of OATP1A2, such as E172 [ 37 ], R168 [ 38 ], and other highly conserved residues, such as Arg556 (conserved in all OATPs; see Supporting information, Figure S3B ) and Lys33 (within OATP1 family, Figure 5 B; also see Supporting information, Figure S3A ). These positively charged residues, corresponding to Arg580 and Lys41 in OATP1B1 [ 39 ] and OATP1B3 [ 40 ], play a role in the uptake of substrates. Given the relevance of the predicted pocket, the potential binding mode of synthetic compounds ( 1–11 ) was suggested by docking along with the reference substrate Estrone-3-Sulfate (E3S) [ 38 ].…”

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

“…The reference substrate E3S (efficacy = 10.21, K m = 6.5 µM, V max = 66 ± 8.5 [ 39 ]) was evaluated as a model substrate for the steroidal derivatives, using docking and a total of 4 µs MD simulations. E3S docking pose was selected based on its interactions with previously described relevant residues for the binding of this substrate [ 39 , 40 , 41 , 42 , 43 ], namely Lys33, Arg556, Arg168, and Glu172. E3S initial binding mode ( Figure 5 C,E) displays its sulfate group oriented toward the extracellular region, interacting with Lys33, Arg556, and its carbonyl group oriented toward the intracellular side of the pocket, interacting with His551.…”

Section: Resultsmentioning

confidence: 99%

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Neurosteroids: Structure-Uptake Relationships and Computational Modeling of Organic Anion Transporting Polypeptides (OATP)1A2

et al. 2021

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In this study, we investigated the delivery of synthetic neurosteroids into MCF-7 human breast adenocarcinoma cells via Organic Anionic Transporting Polypeptides (OATPs) (pH 7.4 and 5.5) to identify the structural components required for OATP-mediated cellular uptake and to get insight into brain drug delivery. Then, we identified structure-uptake relationships using in-house developed OATP1A2 homology model to predict binding sites and modes for the ligands. These binding modes were studied by molecular dynamics simulations to rationalize the experimental results. Our results show that carboxylic acid needs to be at least at 3 carbon-carbon bonds distance from amide bond at the C-3 position of the androstane skeleton and have an amino group to avoid efflux transport. Replacement of hydroxyl group at C-3 with any of the 3, 4, and 5-carbon chained terminal carboxylic groups improved the affinity. We attribute this to polar interactions between carboxylic acid and side-chains of Lys33 and Arg556. The additional amine group showed interactions with Glu172 and Glu200. Based on transporter capacities and efficacies, it could be speculated that the functionalization of acetyl group at the C-17 position of the steroidal skeleton might be explored further to enable OAT1A2-mediated delivery of neurosteroids into the cells and also across the blood-brain barrier.

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“…Estradiol-17β-glucuronide and cholecystokinin-octapeptide (CCK8) can be used as selective probe substrates for evaluating OATP 1B1 [ 298 , 299 , 300 ] and OATP 1B3 [ 101 , 301 ] transporter activities, respectively. Measurement of OATP1B1/1B3 inhibition can also rely on the uptake of other specific substrates, such as zombie violet, live/dead green, cascade blue hydrazide, Alexa Fluor 405 succinimidyl ester [ 302 ], estrone-3-sulfate [ 101 , 303 ], statins [ 304 ] and sodium-fluorescein [ 305 ]. Optimisation of fluorescence-based in vitro transporter assays using sodium-fluorescein as probe substrate has allowed for HTP screening of the inhibitory potential of chemicals [ 305 ].…”

Section: Liver-specific Toxicity In Vitro Methodsmentioning

confidence: 99%

In Vitro Liver Toxicity Testing of Chemicals: A Pragmatic Approach

Tabernilla

Rodrigues

Pieters

et al. 2021

IJMS

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The liver is among the most frequently targeted organs by noxious chemicals of diverse nature. Liver toxicity testing using laboratory animals not only raises serious ethical questions, but is also rather poorly predictive of human safety towards chemicals. Increasing attention is, therefore, being paid to the development of non-animal and human-based testing schemes, which rely to a great extent on in vitro methodology. The present paper proposes a rationalized tiered in vitro testing strategy to detect liver toxicity triggered by chemicals, in which the first tier is focused on assessing general cytotoxicity, while the second tier is aimed at identifying liver-specific toxicity as such. A state-of-the-art overview is provided of the most commonly used in vitro assays that can be used in both tiers. Advantages and disadvantages of each assay as well as overall practical considerations are discussed.

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Amino-terminal region of human organic anion transporting polypeptide 1B1 dictates transporter stability and substrate interaction

Cited by 10 publications

References 23 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

Neurosteroids: Structure-Uptake Relationships and Computational Modeling of Organic Anion Transporting Polypeptides (OATP)1A2

In Vitro Liver Toxicity Testing of Chemicals: A Pragmatic Approach

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