Evaluation of the Selectivity of Several Organic Anion Transporting Polypeptide 1B Biomarkers Using Relative Activity Factor Method.

In recent years, some endogenous substrates of organic anion transporting polypeptide 1B (OATP1B) have been identified and characterized as potential biomarkers to assess OATP1B-mediated clinical drug-drug interactions (DDIs). However, quantitative determination of their selectivity to OATP1B is still limited. In this study, we developed a relative activity factor (RAF) method to determine the relative contribution of hepatic uptake transporters OATP1B1, OATP1B3, OATP2B1, and sodium-taurocholate co-transporting polypeptide (NTCP) on hepatic uptake of several OATP1B biomarkers, including coproporphyrin I (CPI), coproporphyrin I CPIII, and sulfate conjugates of bile acids: glycochenodeoxycholic acid sulfate (GCDCA-S), glycodeoxycholic acid sulfate (GDCA-S), and taurochenodeoxycholic acid sulfate (TCDCA-S). RAF values for OATP1B1, OATP1B3, OATP2B1, and NTCP were determined in cryopreserved human hepatocytes and transporter transfected cells using pitavastatin, cholecystokinin, resveratrol-3-O-ß-D-glucuronide, and taurocholic acid (TCA) as reference compounds, respectively. OATP1B1-specific pitavastatin uptake in hepatocytes was measured in the absence and presence of 1 µM estropipate, whereas NTCP-specific TCA uptake was measured in the presence of 10 µM rifampin. Our studies suggested that CPI was a more selective biomarker for OATP1B1 than CPIII, whereas GCDCA-S and TCDCA-S were more selective to OATP1B3. OATP1B1 and OATP1B3 equally contributed to hepatic uptake of GDCA-S. The mechanistic static model, incorporating the fraction transported of CPI/III estimated by RAF and in vivo elimination data, predicted several perpetrator interactions with CPI/III. Overall, RAF method combined with pharmacogenomic and DDI studies is a useful tool to determine the selectivity of transporter biomarkers and facilitate the selection of appropriate biomarkers for DDI evaluation. SIGNIFICANCE STATEMENT: The authors developed a new relative activity factor (RAF) method to quantify the contribution of hepatic uptake transporters organic anion transporting polypeptide (OATP)1B1, OATP1B3, OATP2B1, and sodium taurocholate co-transporting polypeptide (NTCP) on several OATP1B biomarkers and evaluated their predictive value on drug-drug interactions (DDI). These studies suggest that the RAF method is a useful tool to determine the selectivity of transporter biomarkers. This method combined with pharmacogenomic and DDI studies will mechanistically facilitate the selection of appropriate biomarkers for DDI prediction.

In Vitro Assessment of Transporter Mediated Perpetrator DDIs for Several Hepatitis C Virus Direct-Acting Antiviral Drugs and Prediction of DDIs with Statins Using Static Models.

Inhibitory effects of asunaprevir, daclatasvir, grazoprevir, paritaprevir, simeprevir, and voxilaprevir, direct-acting antiviral (DAA) drugs for the treatment of chronic hepatitis C virus (HCV) infection, were evaluated in vitro against a range of clinically important drug transporters. In vitro inhibition studies were conducted using transporter transfected cells and membrane vesicles. The risk of clinical drug-drug interactions (DDIs) was assessed using simplified static models recommended by regulatory agencies. Furthermore, we refined and developed static models to predict complex DDIs with several statins (pitavastatin, rosuvastatin, atorvastatin, and pravastatin) by mechanistically assessing differential inhibitory effects of perpetrator drugs on multiple transporters, such as organic anion transporting polypeptides (OATP1B), breast cancer resistance protein (BCRP), multidrug resistance protein 2 (MRP2), organic anion transporter 3 (OAT3), and cytochrome P450 CYP3A enzyme, as they are known to contribute to absorption, distribution, metabolism and excretion (ADME) of above statins. These models successfully predicted a total of 46 statin DDIs, including above DAA drugs and their fix-dose combination regimens. Predicted plasma area under curve ratio (AUCR) with and without perpetrator drugs was within ~ 2-fold of observed values. In contrast, simplified static R-value model resulted in increased false negative and false positive predictions when different prediction cut-off values were applied. Our studies suggest that mechanistic static model is a promising and useful tool to provide more accurate prediction of the risk and magnitude of DDIs with statins in early drug development and may help to improve the management of clinical DDIs for HCV drugs to ensure effective and safe HCV therapy. GRAPHICAL ABSTRACT.

In Vitro Evaluation of the Drug Interaction Potential of Doravirine.

Doravirine is a novel nonnucleoside reverse transcriptase inhibitor for the treatment of human immunodeficiency virus type 1 infection. In vitro studies were conducted to assess the potential for drug interactions with doravirine via major drug-metabolizing enzymes and transporters. Kinetic studies confirmed that cytochrome P450 3A (CYP3A) plays a major role in the metabolism of doravirine, with ∼20-fold-higher catalytic efficiency for CYP3A4 versus CYP3A5. Doravirine was not a substrate of breast cancer resistance protein (BCRP) and likely not a substrate of organic anion transporting polypeptide 1B1 (OATP1B1) or OATP1B3. Doravirine was not a reversible inhibitor of major CYP enzymes (CYP1A2, -2B6, -2C8, -2C9, -2C19, -2D6, and -3A4) or of UGT1A1, nor was it a time-dependent inhibitor of CYP3A4. No induction of CYP1A2 or -2B6 was observed in cultured human hepatocytes; small increases in CYP3A4 mRNA (≤20%) were reported at doravirine concentrations of ≥10 µM but with no corresponding increase in enzyme activity. In vitro transport studies indicated a low potential for interactions with substrates of BCRP, P-glycoprotein, OATP1B1 and OATP1B3, the bile salt extrusion pump (BSEP), organic anion transporter 1 (OAT1) and OAT3, organic cation transporter 2 (OCT2), and multidrug and toxin extrusion 1 (MATE1) and MATE2K proteins. In summary, these in vitro findings indicate that CYP3A4 and CYP3A5 mediate the metabolism of doravirine, although with different catalytic efficiencies. Clinical trials reported elsewhere confirm that doravirine is subject to drug-drug interactions (DDIs) via CYP3A inhibitors and inducers, but they support the notion that DDIs (either direction) are unlikely via other major drug-metabolizing enzymes and transporters.

Identification of Endogenous Biomarkers to Predict the Propensity of Drug Candidates to Cause Hepatic or Renal Transporter-Mediated Drug-Drug Interactions.

Drug transporters expressed in liver and kidney play a critical role in the elimination of a wide range of drugs and xenobiotics and inhibition of these transporters may therefore cause clinically significant drug-drug interactions (DDIs). Currently, in vitro transporter inhibition data are used to assess the risk that a drug candidate may act as an inhibitor of a transporter in patients at clinically relevant exposures. However, this approach is hampered by low confidence in in vitro to in vivo extrapolations, and large inter-system and inter-laboratory variability in in vitro data. Several endogenous compounds have been identified as substrates of drug transporters. Determining the impact of perpetrator drugs on the plasma or urinary exposure of these potential endogenous biomarkers in humans is being explored as an alternative approach to assess the DDI liability of drug candidates, especially in early drug development. In this review, we provide an overview of recently identified biomarkers used to study the inhibition of hepatic and renal transporters; summarize the methods and strategies employed to identify biomarkers; and discuss the utility, limitation, and future direction of biomarker approaches to predict transporter-mediated DDIs.

The Complexities of Interpreting Reversible Elevated Serum Creatinine Levels in Drug Development: Does a Correlation with Inhibition of Renal Transporters Exist?

In humans, creatinine is formed by a multistep process in liver and muscle and eliminated via the kidney by a combination of glomerular filtration and active transport. Based on current evidence, creatinine can be taken up into renal proximal tubule cells by the basolaterally localized organic cation transporter 2 (OCT2) and the organic anion transporter 2, and effluxed into the urine by the apically localized multidrug and toxin extrusion protein 1 (MATE1) and MATE2K. Drug-induced elevation of serum creatinine (SCr) and/or reduced creatinine renal clearance is routinely used as a marker for acute kidney injury. Interpretation of elevated SCr can be complex, because such increases can be reversible and explained by inhibition of renal transporters involved in active secretion of creatinine or other secondary factors, such as diet and disease state. Distinction between these possibilities is important from a drug development perspective, as increases in SCr can result in the termination of otherwise efficacious drug candidates. In this review, we discuss the challenges associated with using creatinine as a marker for kidney damage. Furthermore, to evaluate whether reversible changes in SCr can be predicted prospectively based on in vitro transporter inhibition data, an in-depth in vitro-in vivo correlation (IVIVC) analysis was conducted for 16 drugs with in-house and literature in vitro transporter inhibition data for OCT2, MATE1, and MATE2K, as well as total and unbound maximum plasma concentration (Cmax and Cmax,u) data measured in the clinic.

Raltegravir has a low propensity to cause clinical drug interactions through inhibition of major drug transporters: an in vitro evaluation.

Raltegravir (RAL) is a human immunodeficiency virus type 1 (HIV-1) integrase inhibitor approved to treat HIV infection in adults in combination with other antiretrovirals. The potential of RAL to cause transporter-related drug-drug interactions (DDIs) as an inhibitor has not been well described to date. In this study, a series of in vitro experiments were conducted to assess the inhibitory effects of RAL on major human drug transporters known to be involved in clinically relevant drug interactions, including hepatic and renal uptake transporters and efflux transporters. For hepatic uptake transporters, RAL showed no inhibition of organic anion-transporting polypeptide 1B1 (OATP1B1), weak inhibition of OATP1B3 (40% inhibition at 100 µM), and no inhibition of organic cation transporter 1 (OCT1). Studies of renal uptake transporters showed that RAL inhibited organic anion transporters 1 and 3 (OAT1 and OAT3) with 50% inhibitory concentrations (IC50s) (108 µM and 18.8 µM, respectively) well above the maximum concentration of drug in plasma (Cmax) at the clinical 400-mg dose and did not inhibit organic cation transporter 2 (OCT2). As for efflux transporters, RAL did not inhibit breast cancer resistance protein (BCRP) and showed weak inhibition of multidrug and toxin extrusion protein 1 (MATE1) (52% inhibition at 100 µM) and MATE2-K (29% inhibition at 100 µM). These studies indicate that at clinically relevant exposures, RAL does not inhibit or only weakly inhibits hepatic uptake transporters OATP1B1, OATP1B3, and OCT1, renal uptake transporters OCT2, OAT1, and OAT3, as well as efflux transporters BCRP, MATE1, and MATE2-K. The propensity for RAL to cause DDIs via inhibition of these transporters is therefore considered low.

Transport of the dipeptidyl peptidase-4 inhibitor sitagliptin by human organic anion transporter 3, organic anion transporting polypeptide 4C1, and multidrug resistance P-glycoprotein.

Sitagliptin, a selective dipeptidyl peptidase 4 inhibitor recently approved for the treatment of type 2 diabetes, is excreted into the urine via active tubular secretion and glomerular filtration in humans. In this report, we demonstrate that sitagliptin is transported by human organic anion transporter hOAT3 (Km=162 microM), organic anion transporting polypeptide OATP4C1, and multidrug resistance (MDR) P-glycoprotein (Pgp), but not by human organic cation transporter 2 hOCT2, hOAT1, oligopeptide transporter hPEPT1, OATP2B1, and the multidrug resistance proteins MRP2 and MRP4. Our studies suggested that hOAT3, OATP4C1, and MDR1 Pgp might play a role in transporting sitagliptin into and out of renal proximal tubule cells, respectively. Sitagliptin did not inhibit hOAT1-mediated cidofovir uptake, but it showed weak inhibition of hOAT3-mediated cimetidine uptake (IC50=160 microM). hOAT3-mediated sitagliptin uptake was inhibited by probenecid, ibuprofen, furosemide, fenofibric acid, quinapril, indapamide, and cimetidine with IC50 values of 5.6, 3.7, 1.7, 2.2, 6.2, 11, and 79 microM, respectively. Sitagliptin did not inhibit Pgp-mediated transport of digoxin, verapamil, ritonavir, quinidine, and vinblastine. Cyclosporine A significantly inhibited Pgp-mediated transport of sitagliptin (IC50=1 microM). Our data indicate that sitagliptin is unlikely to be a perpetrator of drug-drug interactions with Pgp, hOAT1, or hOAT3 substrates at clinically relevant concentrations. Renal secretion of sitagliptin could be inhibited if coadministered with OAT3 inhibitors such as probenecid. However, the magnitude of interactions should be low, and the effects may not be clinically meaningful, due to the high safety margin of sitagliptin.