Maturation of metabolizing capacities occurred postnatally, as described in man.

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.

P-glycoprotein inhibitors, like zosuquidar, have widely been used to study the role of P-glycoprotein in oral absorption. Still, systematic studies on the inhibitor dose-response relationship on intestinal drug permeation are lacking. In the present study, we investigated the effect of 0.79 nM-2.5 μM zosuquidar on etoposide permeability across Caco-2 cell monolayers. We also investigated etoposide pharmacokinetics after oral or IV administration to Sprague Dawley rats with co-administration of 0.063–63 mg/kg zosuquidar, as well as the pharmacokinetics of zosuquidar itself. Oral zosuquidar bioavailability was 2.6–4.2%, while oral etoposide bioavailability was 5.5 ± 0.9%, which increased with increasing zosuquidar doses to 35 ± 5%. The intestinal zosuquidar concentration required to induce a half-maximal increase in bioavailability was estimated to 180 μM. In contrast, the IC 50 of zosuquidar on etoposide permeability in vitro was only 5–10 nM, and a substantial in vitro-in vivo discrepancy of at least four orders of magnitude was thereby identified. Overall, the present study provides valuable insights for future formulation development that applies fixed dose combinations of P-glycoprotein inhibitors to increase the absorption of poorly permeable P-glycoprotein substrate drugs.

It has been suggested that oral absorption of low-permeable P-glycoprotein (P-gp) substrates can be increased through saturation of P-gp. For BCS class IV drug substances, saturating P-gp is challenging due to low aqueous solubility. The present study investigated if the BCS IV drug substance etoposide could be solubilized to a concentration saturating P-gp after oral administration.A formulation consisting of 10% (w/v) of pluronic ® F-127 and polyvinylpyrrolidone/vinyl acetate (PVP/VA), and 57% (v/v) ethanol enhanced etoposide's solubility approximately 100 times (16 mg mL -1 ) compared to its aqueous solubility. In vitro, this formulation was stable upon dilution in simulated intestinal fluid. In male Sprague-Dawley rats, oral administration of increasing solubilized etoposide doses using the formulation matrix increased the AUC0-∞ of etoposide dose-proportionally but resulted in a lower absolute oral bioavailability (F) and rate of absorption as compared to control.At the highest investigated dose (100 mg kg -1 ), AUC0-∞ and Cmax were significantly increased by 2.9and 1.4-fold, respectively, compared to control dosed at 20 mg kg -1 . A single oral dose of 20 mg kg -1 zosuquidar followed by 20 mg kg -1 oral etoposide increased F 8.6-fold. In conclusion, a stable formulation with improved etoposide solubility was developed, yet the formulation did not result in increased oral bioavailability of etoposide.

P‑glycoprotein (P‑gp) limits the oral absorption of drug substances. Potent small molecule P‑gp inhibitors (e.g., zosuquidar) and nonionic surfactants (e.g., polysorbate 20) inhibit P‑gp by proposedly different mechanisms. Therefore, it was hypothesised that a combination of zosuquidar and polysorbate 20 may potentiate inhibition of P‑gp-mediated efflux. P‑gp inhibition by zosuquidar and polysorbate 20 in combination was assessed in a calcein‑AM assay and in a transcellular etoposide permeability study in MDCKII‑MDR1 and Caco‑2 cells. Furthermore, solutions of etoposide, zosuquidar, and polysorbate 20 were orally administered to Sprague Dawley rats. Zosuquidar elicited a high level of nonspecific adsorption to various labware, which significantly affected the outcomes of the in vitro studies. Still, at certain zosuquidar and polysorbate 20 concentrations, additive P‑gp inhibition was observed in vitro. In vivo, however, oral etoposide bioavailability decreased by coadministration of both zosuquidar and polysorbate 20 when compared to coadministration of etoposide with zosuquidar alone. For future formulation development, the present study provided important and novel knowledge about nonspecific zosuquidar adsorption, as well as insights into combinational P‑gp inhibition by a third-generation P‑gp inhibitor and a P‑gp-inhibiting nonionic surfactant.