Our results demonstrate the selectivity of CPI and CPIII towards the OATP1B/MRP pathway, and the herein reported data further underline the potential of CPI and CPIII as selective and sensitive clinical biomarkers to quantify OATP1B-mediated DDIs.

Organic anion-transporting polypeptides (OATP) 1B1 and OATP1B3 are drug transporters mediating the active hepatic uptake of their substrates. Because they exhibit overlapping substrate specificities, the contribution of each isoform to the net hepatic uptake needs to be considered when predicting drug-drug interactions. The relative contribution of OATP1B1-and OATP1B3-mediated uptake of statins into hepatocytes was estimated based on either relative transporter protein expression data or relative activity data. Therefore, kinetics of eight statins and OATP1B1-and OATP1B3-specific reference substrates was determined in OATP1B1-and OATP1B3-expressing human embryonic kidney 293 cells and in human cryopreserved hepatocytes. Absolute OATP1B1 and OATP1B3 protein abundance was determined by liquid chromatography-tandem mass spectrometry in all expression systems. Transporter activity data generated in recombinant cell lines were extrapolated to hepatocyte values using relative transporter expression factors (REF) or relative activity factors (RAF). Our results showed a pronounced OATP1B1 and comparatively low OATP1B3 protein expression in the investigated hepatocyte lot. Based on REF scaling, we demonstrated that the active hepatic uptake clearances of reference substrates, atorvastatin, pravastatin, rosuvastatin, and simvastatin were well predicted within twofold error, demonstrating that OATP1B1 and OATP1B3 were major contributors. For other statins, the net hepatic uptake clearance was underpredicted, suggesting the involvement of other hepatic uptake transporters. Summarized, we showed that REF-and RAF-based predictions were highly similar, indicating a direct transporter expressionactivity relationship. Moreover, we demonstrated that the REF-scaling method provided a powerful tool to quantitatively assess the transporterspecific contributions to the net uptake clearance of statins in hepatocytes.

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Introduction Endogenous biomarkers are promising tools to assess transporter-mediated drug-drug interactions (DDI) early in humans.Methods We evaluated on a common and validated in vitro system the selectivity of 4-pyridoxic acid (PDA), homovanillic acid (HVA), glycochenodeoxycholate-3-sulfate (GCDCA-S) and taurine towards different renal transporters, including multidrug resistance-associated protein (MRP), and assessed the in vivo biomarker sensitivity towards the strong organic anion transporters (OAT) inhibitor probenecid at 500mg every 6h to reach close to complete OAT inhibition.Results PDA and HVA were substrates of the OAT1/2/3, OAT4 (PDA only) and MRP4; GCDCA-S was more selective, having affinity only towards OAT3 and MRP2. Taurine was not a substrate of any of the investigated transporters under the in vitro conditions tested. Plasma exposure of PDA and HVA significantly increased and the renal clearance of GCDCA-S, PDA and HVA decreased; the magnitude of these changes was comparable to the ones of known clinical OAT probe substrates. PDA and GCDCA-S were the most promising endogenous biomarkers of the OAT pathway activity: PDA plasma exposure was the most sensitive to probenecid inhibition, and, in contrast, GCDCA-S was the most sensitive OAT biomarker based on renal clearance, with higher selectivity towards the OAT3 transporter. ConclusionThe current findings illustrate a clear benefit of measuring PDA plasma exposure during Phase 1 studies when a clinical drug candidate is suspected to be an OAT inhibitor based on in vitro data.Subsequently, combined monitoring of PDA and GCDCA-S in both urine and plasma is recommended in order to tease out the involvement of OAT1/3 in the inhibition interaction.

Hepatic elimination is a function of the interplay between different processes