Establishment of In Vitro P-Glycoprotein Inhibition Assay and Its Exclusion Criteria to Assess the Risk Of Drug–Drug Interaction at the Drug Discovery Stage

Abstract. Antimalarials are widely used in African and Southeast Asian countries, where they are combined with other drugs for the treatment of concurrent ailments. The potential for P-glycoprotein (P-gp)-mediated drug-drug interactions (DDIs) between antimalarials and P-gp substrates was examined using a Caco-2 cell-based model. Selected antimalarials were initially screened for their interaction with P-gp based on the inhibition of rhodamine-123 (Rho-123) transport in Caco-2 cells. Verapamil (100 μM) and quinidine (1 μM) were used as positive inhibition controls. Lumefantrine, amodiaquin, and artesunate all showed blockade of Rho-123 transport. Subsequently, the inhibitory effect of these antimalarials on the bi-directional passage of digoxin (DIG) was examined. All of the drugs decreased basal-toapical (B-A) P-gp-mediated DIG transport at concentrations of 100 μM and 1 mM. These concentrations may reflect therapeutic doses for amodiaquin and artesunate. Therefore, clinically relevant DDIs may occur between certain antimalarials and P-gp substrates in general.

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“…The K i values were generated from ER 12 and calculations made using Microsoft Excel software (Redmond, WA).…”

Section: Methodsmentioning

confidence: 99%

“…9 Several methods for predicting the likelihood of intestinal enzyme and transporter-mediated DDIs have been suggested. [10][11][12] This study used the druginteraction number (DIN, the ratio of inhibitor dose/inhibition constant) as an index for predicting the occurrence of clinically relevant intestinal DDIs.…”

Section: Introductionmentioning

confidence: 99%

Potential P-Glycoprotein-Mediated Drug-Drug Interactions of Antimalarial Agents in Caco-2 cells

Oga

Sekine²,

Shitara³

et al. 2012

The American Society of Tropical Medicine and Hygiene

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“…both in the A-B and B-A direction), thus yielding more potent IC50 values. [187,191,192] Balimane et al [187] used the same experimental data sets to calculate % inhibition and/or IC50 values for more than 50 compounds using either A-B transport data, B-A transport data, the net secretory flux or the efflux ratio. [187] They found that only the IC50 values from B-A transport data and net secretory flux data correlated.…”

Section: Transcellular Transport Studiesmentioning

confidence: 99%

“…[213] The cut-off value of 5 was determined from Caco-2 cells both when net secretory flux data (corresponding to equation 8) and when efflux ratios (corresponding to equation 10) were used respectively to calculate the IC50 values. [191] Most recently, Sugimoto et al [192] proposed a cut-off value of 30 based on IC50 values determined from efflux ratios in P-gp-transfected LLC-PK1 cells and on changes in digoxin AUC from clinical studies. Cook et al [191] also collected IC50 values from the literature including several laboratories, cell lines and calculation methods.…”

Section: Determination Of the 'Cut-off' Valuementioning

confidence: 99%

“…For all the deducted cut-off values, the Ki/IC50 for P-gp substrates were determined with basis in the extracellular concentration, an approach that is dependent on the influx permeability of the drug substance and the expression level of the transporter in the particular in-vitro system. [191,192,212,213] Thus, differences in the in-vivo expression level between efflux transporters may imply that different cut-off values should be used. Furthermore, there may be general differences in the permeability properties of substrates/inhibitors for one efflux transporter compared with substrates/inhibitors for another transporter (e.g.…”

Section: Limitations Of the Cut-off Value For Intestinal Transportersmentioning

confidence: 99%

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Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug–drug interactions

Grandvuinet

Vestergaard

Rapin³

et al. 2012

Journal of Pharmacy and Pharmacology

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Objectives This review provides an overview of intestinal human transporters for organic anions and stresses the need for standardization of the various in‐vitro methods presently employed in drug–drug interaction (DDI) investigations. Key findings Current knowledge on the intestinal expression of the apical sodium‐dependent bile acid transporter (ASBT), the breast cancer resistance protein (BCRP), the monocarboxylate transporters (MCT) 1, MCT3‐5, the multidrug resistance associated proteins (MRP) 1–6, the organic anion transporting polypetides (OATP) 2B1, 1A2, 3A1 and 4A1, and the organic solute transporter α/β (OSTα/β) has been covered along with an overview of their substrates and inhibitors. Furthermore, the many challenges in predicting clinically relevant DDIs from in‐vitro studies have been discussed with focus on intestinal transporters and the various methods for deducting in‐vitro parameters for transporters (Km/Ki/IC50, efflux ratio). The applicability of using a cut‐off value (estimated based on the intestinal drug concentration divided by the Ki or IC50) has also been considered. Summary A re‐evaluation of the current approaches for the prediction of DDIs is necessary when considering the involvement of other transporters than P‐glycoprotein. Moreover, the interplay between various processes that a drug is subject to in‐vivo such as translocation by several transporters and dissolution should be considered.

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In Vitro Assays to Predict In Vivo Interactions of P ‐Glycoprotein with Drugs

Volpe

Balimane

2017

Encyclopedia of Drug Metabolism and Interactions

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P‐glycoprotein (P‐gp) is an efflux transporter that pumps endogenous and exogenous compounds out of cells. P‐gp limits drug absorption from the intestines, aids in drug excretion from the liver and kidneys, and prevents entry of drugs into the brain. Given that interactions of a drug with P‐gp can play an important part in its pharmacokinetic properties, there are efforts in early discovery and development to identify compounds as P‐gp substrates or inhibitors. In vitro assays are based on cell membranes (ATPase, vesicles) or intact cells (accumulation, bidirectional) expressing the P‐gp transporter. This chapter reviews these in vitro assays along with their advantages and limitations. In determining whether a new compound is a substrate or inhibitor of P‐gp during drug discovery and development, a tier approach provides a scheme where higher throughput assays lead to those with greater complexity. An important aspect of using in vitro P‐gp assays during drug discovery and development is that they need to be optimized and validated in order to have confidence in their outcomes in determining whether a new compound is a substrate or inhibitor of the efflux transporter. The use of acceptance criteria and negative and positive control substrates and inhibitors is discussed along with how different assay parameters can effect outcomes. In conclusion, well‐established in vitro P‐gp interaction assays are reasonably predictive of the in vivo P‐gp effects in the clinic.

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Establishment of In Vitro P-Glycoprotein Inhibition Assay and Its Exclusion Criteria to Assess the Risk Of Drug–Drug Interaction at the Drug Discovery Stage

Cited by 38 publications

References 37 publications

Potential P-Glycoprotein-Mediated Drug-Drug Interactions of Antimalarial Agents in Caco-2 cells

Potential P-Glycoprotein-Mediated Drug-Drug Interactions of Antimalarial Agents in Caco-2 cells

Intestinal transporters for endogenic and pharmaceutical organic anions: the challenges of deriving in-vitro kinetic parameters for the prediction of clinically relevant drug–drug interactions

In Vitro Assays to Predict In Vivo Interactions of P ‐Glycoprotein with Drugs

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