Subjects
Healthy male and female subjects were screened up to 21 days before check-in (day 0). Subjects were eligible for study participation if all inclusion/exclusion criteria were met. Subjects were 18–55 years of age with a body mass index (BMI) of 18–30 kg/m2. Subjects were required to be in good general health as determined by a standard physical exam, 12-lead electrocardiogram (ECG), and clinical laboratory testing. Subjects were excluded for any known exposure to OCA and any of its components, probe substrates, or any comparable or similar product. Additional exclusion criteria were specific to study.
Study Design
These five studies were conducted as single-center, open-label, fixed-sequence, parallel-design, inpatient studies in healthy volunteers. Each study was designed to evaluate the single-dose PK of the substrate probe alone and in combination with repeat doses (at least 14 days) of 10 and 25 mg OCA, the two highest OCA doses intended for commercialization across indications (Table 1). To reduce the total number of studies required to assess seven probe substrates, we combined the assessment of midazolam and caffeine into one study and dextromethorphan and omeprazole in another study, evaluating each probe substrate separately on different days. OCA and probe substrates were administered in the fasted state.
Table 1 Overview of clinical drug–drug interaction studies with oral OCA in healthy, fasted subjects
Eligible subjects were randomized in a 1:1 ratio to one of two OCA treatment arms (10 or 25 mg) receiving OCA once daily. All subjects remained at the clinic from day 0 to the final day of dosing which varied depending on the time needed to properly evaluate the PK of each probe substrate, which was dependent on the plasma half-life of the probe substrate. Safety analyses were conducted on all subjects who received at least one dose of study drug.
All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1964, as revised in 2013. All subjects provided informed consent before any study procedures or assessments were performed. The studies were not registered as this is not a requirement for phase 1 studies.
Study I: Specific Substrates for CYP1A2 (Caffeine) and for CYP3A (Midazolam)
Subjects were required to avoid meals and fluids containing caffeine, theobromine, and xanthine for at least 72 h before day 0 and during the inpatient portion of the study. Subjects received a single oral dose of 2 mg midazolam in syrup form on day 1 and day 19 and a single oral dose of 200 mg caffeine in caplet form on day 3 and day 21 under fasting conditions (10-h overnight fast). On day 5 through day 23, oral doses of 10 or 25 mg OCA were administered daily under fasting conditions. Blood samples were collected at predose and at 0.25, 0.5, 1, 1.25, 1.5, 2, 3, 4, 6, 8, 12, 24, 36, and 48 h after dosing of midazolam on day 1 and day 19 for the measurement of plasma concentrations of midazolam and its metabolite (1-hydroxymidazolam). Similarly, blood samples were collected at predose and at 0.25, 0.5, 1, 1.5, 2, 3, 4, 6, 8, 12, 24, 36, and 48 h after dosing of caffeine on day 3 and day 21 for the measurement of plasma concentrations of caffeine and its metabolite (paraxanthine).
Study II: Specific Substrates for CYP2C9 (S-warfarin) and for CYP1A2 and CYP3A (R-warfarin)
Subjects were excluded if they were a poor metabolizer of warfarin as determined by CYP2C9 genotype testing, if they were highly sensitive to warfarin as determined by genetic testing for vitamin K epoxide reductase complex, subunit 1 (VKORC1) or had a prothrombin time (PT), activated partial thromboplastin time (aPTT), international normalized ratio (INR), protein C, or protein S value(s) outside the normal ranges at screening for which the results were clinically significant.
Several published DDI studies used a racemic warfarin single dose of 25 mg to produce well-characterized PK and PD responses [11,12,13]. In this study, a 25-mg dose of racemic warfarin was expected to provide sufficient plasma concentrations of both enantiomers to characterize their plasma PK, and also produce measurable PD effects with no undue risk to the subjects. Subjects were administered a single oral dose of 25 mg racemic warfarin (R- and S-warfarin) in tablet form on day 1 and day 21 under fasting conditions. Daily oral administration of OCA at doses of 10 or 25 mg was initiated on day 8 and continuing through day 27. PK blood samples for measuring plasma concentrations of R- and S-warfarin were collected at predose and at 0.25, 0.5, 1, 2, 3, 4, 6, 8, 12, 24, 36, 48, 60, 72, 96, 120, 144, and 168 h (i.e., over a 7-day period) after dosing of racemic warfarin on day 1 and day 21. The PD of racemic warfarin using the serial blood samples (predose and 12, 24, 36, 48, 60, 72, 96, 120, 144, and 168 h after dosing on day 1 and day 21) was assessed by measuring coagulation parameters PT, aPTT, and INR.
Study III: Specific Substrates for CYP2D6 (Dextromethorphan) and for CYP2C19 (Omeprazole)
CYP2D6 is a genetically controlled polymorphic enzyme such that approximately 5–10% of people of white ethnicity express no functional enzyme and are characterized as poor metabolizers. Subjects were excluded if they were a poor metabolizer of dextromethorphan as determined by CYP2D6 genotyping or omeprazole as determined by CYP2C19 genotyping or screened positive for Helicobacter pylori (H. pylori).
Subjects received a single oral 30-mg dose of dextromethorphan in capsule form on day 1 and day 18. On the basis of the terminal half-life (t
1/2) of dextromethorphan (2–4 h), blood samples for the measurement of plasma concentrations of dextromethorphan and its metabolite (dextrorphan) were collected at predose and at 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 12, 24, 48, and 72 h after dosing of dextromethorphan on days 1 and 18.
A single 20-mg dose of omeprazole in capsule form was administered orally on day 4, and once-daily dosing was initiated on days 21 through 28 as a second test treatment. Blood samples for the measurement of plasma concentrations of omeprazole and its metabolite (5-hydroxyomeprazole) were collected at predose and at 0.25, 0.5, 1, 1.5, 2, 2.5, 3, 4, 5, 6, 8, 10, and 12 h after dosing of omeprazole on days 4 and 21, and at 3 h postdose on day 28. OCA was administered orally once daily, beginning on day 5 and continued through day 28.
Study IV: BCRP, OATP1B1, and OATP1B3 Substrate (Rosuvastatin)
Rosuvastatin is a recommended substrate for assessment of inhibition of transporters responsible for its distribution and elimination by BCRP, OATP1B1, and OATP1B3 [14]. Several reported drug interaction studies have used rosuvastatin single doses ranging from 10 to 80 mg [15,16,17,18,19]. On the basis of the terminal half-life of rosuvastatin (20 h) and the bioanalytical plasma assay sensitivity, a single dose of 20 mg rosuvastatin was selected to provide concentrations of rosuvastatin sufficient to characterize the PK while minimizing risk to the subjects.
Subjects were administered a single oral dose of 20 mg rosuvastatin in tablet form on day 1 and day 19 under fasting conditions. Daily administration of oral OCA (10 or 25 mg) was initiated on day 6 and continued for 18 days. Blood samples for the measurement of plasma concentrations of rosuvastatin and its metabolite (N-desmethyl rosuvastatin) were collected at predose and at 0.5, 1, 2, 3, 3.5, 4, 4.5, 5, 5.5, 6, 7, 8, 10, 12, 24, 36, 48, 72, 96, and 120 h after dosing of rosuvastatin on day 1 and day 19.
Study V: P-gp Substrate (Digoxin)
The subjects who were sensitive to digoxin or other digitalis preparations were excluded. Digoxin (immediate release formulation) is a sensitive and recommended substrate for the assessment of P-gp, and is eliminated as unchanged drug primarily via the kidney [20]. Subjects were administered a single oral dose of 0.25 mg digoxin in tablet form on day 1 and day 19 under fasting conditions. Daily administration of OCA was initiated on day 6 and continued for 18 days. On the basis of the half-life of digoxin (1.5–2 days), blood samples for the measurement of plasma concentrations of digoxin were collected at predose and at 0.25, 0.5, 0.75, 1, 1.5, 2, 2.5, 3, 4, 6, 8, 10, 12, 24, 36, 48, 60, 72, 96, and 120 h after dosing on day 1 and day 19.
In each study, blood samples for the measurement of plasma concentrations of OCA and its metabolites (glyco-OCA and tauro-OCA) were collected predose on the first day of OCA administration (reference treatment), periodically throughout the dosing period, and on days when OCA and probe drugs were administered concomitantly (test treatment).
Analytical Methods
The plasma concentrations of each probe substrate and of OCA and its metabolites were determined by high-performance liquid chromatography (HPLC) with tandem mass spectrometric detection (LC–MS-MS) using validated analytical methods.
Pharmacokinetic and Pharmacodynamic Parameters
For all studies, the reported PK parameters included the area under the plasma concentration–time curve (AUC) from time 0 to the last sampling time with measurable analyte concentration (AUC0−t
), AUC from time 0 to infinity (AUC∞), and maximum observed plasma concentration (C
max). All PK parameters were measured using Phoenix® WinNonlin® (Version 6.2 or later).
The PD parameters for PT, aPTT, and INR were calculated in study II (racemic warfarin), including the area under the effect curve from time zero to the last sampling collection time point (0–168 h) (AUEC), maximum observed effect (E
max), and time to reach E
max (TEmax).
Statistical Analysis
Statistical analyses were performed using SAS® Version 9.2. The primary PK endpoints (AUC0−t
, AUC∞, and C
max) were analyzed using a linear mixed effect model on the natural log (ln) transformed PK parameters of the different probe substrates. Geometric least square mean (GLSM) and ratios for each of the substrate probes were obtained from the ln-transformed AUC0−t
, AUC∞, and C
max. No apparent drug–drug interaction was concluded if the GLSM ratio was contained within the bounds (0.80–1.25) of the 90% confidence interval (CI) [14].
PD parameters AUEC and E
max for PT, aPTT, and INR were analyzed using the same statistical analyses performed on the PK parameters.