Identification
of a selective inhibitor of organic anion transporting polypeptide
(OATP) 1B1 is critical in order to determine the contribution of OATP1B1-mediated
uptake of investigational drugs into human hepatocytes for successful
in vitro-to-in vivo extrapolation (IVIVE) of hepatic uptake and drug–drug
interaction (DDI). The following study examined the inhibitory effects
of estropipate (EPP) on major sinusoidal drug uptake transporters
and explored its utility regarding IVIVE of statin hepatic disposition.
EPP and its free-base form (i.e., estrone sulfate) showed a potent
and high degree of selectivity in inhibiting the OATP1B1-mediated
transport of rosuvastatin with an IC50 value averaging
0.05 ± 0.01 and 0.12 ± 0.07 μM for human and cynomolgus
monkey OATP1B1 (hOATP1B1 and cOATP1B1), respectively, whereas weak
inhibition was observed for human and monkey OATP1B3, OATP2B1, sodium-taurocholate
cotransporting polypeptide (NTCP), organic anion transporter 2, and
organic cation transporter 1 with IC50 values ranging from
8.6 to 64.0 μM. EPP, together with rifamycin SV, was subsequently
used to determine the fractions of hepatic uptake clearance (f
T) of statins, including rosuvastatin, pitavastatin,
and dehydropravastatin, which are reported to be mediated by OATP1B1,
OATP1B3, OATP2B1, and NTCP. Finally, the magnitudes of in vivo inhibition
of rosuvastatin clearance caused by EPP and rifampin in cynomolgus
monkeys were predicted by using individual transporter IC50 and f
T (AUC fold change 1.28 vs 1.21,
2.71 vs 1.75, and 3.35 vs 2.83, respectively). These results suggest
that EPP is an appropriate OATP1B1-selective inhibitor to establish
the relative contribution of OATP1B1 to hepatic uptake in vitro and
to discern the role of OATP1B1 in hepatic disposition in vivo.
As an expansion investigation of drug-drug interaction (DDI) from previous clinical trials (Shen et al., 2019), additional plasma endogenous metabolites were quantitated in the same subjects to further identify the potential biomarkers of OAT1/3 inhibition. In the single dose, open label, three-phase with fixed order of treatments study, 14 healthy human volunteers orally received 1000 mg probenecid alone, or 40 mg furosemide alone, or 40 mg furosemide at 1 h after receiving 1000 mg probenecid on Days 1, 8, and 15, respectively. Endogenous metabolites including kynurenic acid, xanthurenic acid, indo-3-acetic acid, pantothenic acid, p-cresol sulfate, and bile acids in the plasma were measured by LC-MS/MS. The Cmax of kynurenic acids was significantly increased about 3.3-and 3.7-fold over the baseline values at pre-dose followed by the treatment of probenecid alone or in combination with furosemide respectively. In comparison with the furosemide alone group, the Cmax and AUC0-12 of kynurenic acid were significantly increased about 2.4 and 2.5-fold by probenecid alone, and 2.7 and 2.9-fold by probenecid plus furosemide, respectively. The increases in Cmax and AUC of plasma kynurenic acid by probenecid are comparable to the increases of furosemide Cmax and AUC reported previously. Additionally, the plasma concentrations of xanthurenic acid, indo-3-acetic acid, pantothenic acid, and p-cresol sulfate, but not bile acids, were also significantly elevated by probenecid treatments. The magnitude of effect size analysis for known potential endogenous biomarkers demonstrated that kynurenic acid in the plasma offers promise as a superior addition for early DDI assessment involving OAT1/3 inhibition.
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