Download An integrated strategy to determine the absorption, distribution

An integrated strategy to determine the
absorption, distribution, metabolism, excretion,
and the absolute bioavailability of Bevenopran
(CB-5945) in healthy male subjects
A. Adedoyin1, R. Shibuya1, S. Bodie1, S. Mair2, I. Nisbet2, L. Stevens2, I. Shaw2, S. Wood3, L. Techner1,
Cubist Pharmaceuticals,1 Lexington, USA, Quotient Clinical Ltd,2 Nottingham, UK, Quotient Bioresearch Ltd3 Rushden, UK,
INTRODUCTION
Bevenopran (formerly LY2136231, ADL5945, and CB-5945), an orally bioavailable antagonist of μ
opioid receptors and δ opioid receptors, is currently under development by Cubist Pharmaceuticals,
for the treatment of opioid induced constipation in patients with chronic non-cancer pain.
This study was designed to evaluate the systemic availability of parent bevenopran and related
metabolites via determination of the absolute bioavailability of bevenopran and total radioactivity
following oral and IV administrations. Intravenous dosing was used to provide definitive information
on bevenopran PK which may be of value for subsequent PK modeling and simulation in support of
future clinical studies. This study also aimed to further understand the metabolism of bevenopran in
humans through determination of the routes of metabolism and excretion of [14C]-bevenopran, and
bile sampling using the Entero-test® (1). Radiochromatographic profiling of plasma, urine, and feces
followed by appropriate chemical structure analysis by mass spectrometry was to allow identification
and quantification of any ‘major’ bevenopran metabolites and their elimination routes.
MATERIALS AND METHODS
This was an open-label, nonrandomized,, 2-period, fixed-sequence study in 6 healthy male subjects.
The study was approved by the UK, MHRA and an independent ethics committee prior to consenting
and screening subjects.
Each enrolled subject received the following regimens in the order below:
Period 1: Single oral dose of 0.5 mg bevenopran solution followed after 1 hour by a 15 minute IV
infusion of 5 μg [14C]-bevenopran (containing not more than [NMT] 37 kBq [1000 nCi] 14C) The
Entero-Test™ bile string was administered after IV administration of radiolabelled drug in Period 1.
Period 2: Single oral dose of 0.5 mg [14C]-bevenopran solution (containing NMT 3 MBq [81 μCi] 14C)
For Period 1, blood samples were taken for the measurement of bevenopran, total radioactivity, and
[14C]-bevenopran concentrations. Urine and feces were collected for the measurement of total
radioactivity. Bile fluid was collected using an Entero-test capsule at specified time points after
dosing; the Entero-test strings were analyzed for total radioactivity and metabolites.
For Period 2, blood samples were taken after dosing for the measurement of total radioactivity (whole
blood and plasma) and bevenopran (plasma only), and for metabolite profiling and identification
(plasma). Urine and feces were collected for the measurement of total radioactivity and bevenopran
(urine only), and for metabolite profiling and identification (plasma, urine and feces).
Safety assessments (clinical laboratory tests, vital signs, electrocardiograms (ECGs), and physical
examinations) were performed at specified time points and adverse events (AEs) were monitored
throughout the study.
Analytical methods employed:
• Concentrations of bevenopran in plasma for oral doses in Periods 1 and 2 were determined using
LC-MS/MS.
• Concentrations of 14C-bevenopran after intravenous dosing were determined using LC-AMS
• Total radioactivity in plasma, bile string extract and excreta following intravenous microtracer
dosing in Period 1 were determined using AMS
• Total radioactivity concentrations in plasma and excreta following oral administration in Period 2
were determined using scintillation based methods
All analytical methods were validated prior to use.
RESULTS AND DISCUSSION
Number of subjects (planned and analyzed):
All 6 subjects were included in the safety population and PK populations for both Periods 1 and 2.
Five subjects were included in the mass balance population for Period 1; Subject 004 was excluded
due to a protocol deviation which resulted in a urine sample being discarded.
Five subjects were included in the mass balance population for Period 2; Subject 005 was excluded
due to an AE of constipation.
Mass balance and pharmacokinetics following administration of bevenopran (oral, 0.5mg
solution) and [14C]-bevenopran (5µg intravenous) in Period 1
The average mass balance recovery of total radioactivity in excreta for the IV dose was 65% of the
total radioactive dose recovered during the 72-hour sampling period with over half of the dose
recovered during the first 48 hours. Almost all of the recovery was via urine. Bile string data indicated
there was some elimination of total radioactivity in the bile. However, the recovery of radioactivity
observed in the feces was low (0.009%), indicating that total excretion via the bile was very low
and/or that any radiolabeled materials excreted via the bile were reabsorbed prior to defecation.
Table 1: PK parameters for bevenopran, [14C]-bevenopran, and total radioactivity for oral & IV
administration in period 1.
Pharmacokinetic
Parameters
tlag (h)1)
tmax (h)a
Cmax (pg/mL)2)
AUClast (pg∙h/mL)b
AUC∞ (pg∙h/mL)b
AUC%extrap (%)
t½ (h)
MRT∞ (h)
CL/F (mL/min)3)
CLR (mL/min)
Vz/F (mL)c
F (%)
Period 1
Bevenopran 0.5mg
Oral
Period 1
14C-bevenopran 5µg
IV
Period 1
14C-bevenopran 5µg
IV
Bevenopran
Mean ± SD
(N = 6)
0.0 (0.0–0.0)
3.0 (2.0–6.0)
728 ± 129
15600 ± 3510
17500 ± 2820 [n = 5]
4.77 ± 1.140 [n = 5]
16.9 ± 1.49 [n = 5]
22.4 ± 1.92 [n = 5]
486 ± 70.7 [n = 5]
NC
879000 ± 430000
70.63 ± 2.935
[14C]-bevenopran
Mean ± SD
(N = 6)
NC
0.2 (0.2–0.2)
92.0 ± 20.0
227 ± 38.5
237 ± 38.4
4.08 ± 1.815
16.8 ± 3.64
18.5 ± 2.36
360 ± 57.1
NC
530000 ± 186000
NC
Total Radioactivity
Mean ± SD
(N = 6)
NC
0.2 (0.2–0.2)
100 ± 26.6
402 ± 45.3
471 ± 59.7 [n = 5]
14.30 ± 2.339 [n = 5]
26.2 ± 1.92 [n = 5]
33.7 ± 3.19 [n = 5]
179 ± 21.5 [n = 5]
139 ± 19.3
412000 ± 49300
NC
+44 115 9749000 (UK)
+1-800-769-3518 (USA)
The total clearance of [14C]-bevenopran at 360 mL/min is lower than plasma flow to the liver
(812 mL/min). In the absence of any fecal excretion (<0.1%), it can be concluded that any drug
product excreted via the bile would have been reabsorbed, re-circulated, and eliminated via the urine.
The volume of distribution (Vz) at 530 L is considerably higher than extracellular fluid (18.2 L)
indicating extensive distribution into the tissues.
Absolute bioavailability of bevenopran was 70.63% suggesting bevenopran is well absorbed following
oral administration as a solution.
The mean Cmax and AUC∞ for plasma total radioactivity were 100 pg eq/mL and 471 pg eq∙h/mL,
respectively, with divergence of total radioactivity and [14C]-bevenopran concentrations after
approximately 0.42 hours, with AUC∞ for total radioactivity approximately 2-fold greater than that for
[14C]-bevenopran. This suggests formation of radiolabeled metabolites. The mean t½ for total
radioactivity was 26.2 hours, which was longer than that for [14C]-bevenopran, suggesting the
presence of metabolites that are cleared more slowly than parent drug.
The renal clearance of total radioactivity was 139 mL/min, which is considered similar to the
glomerular filtration rate (125 mL/min). Renal clearance represented approximately 78% of the
observed total clearance (179 mL/min for total radioactivity), which was consistent with the majority of
total radioactivity and parent drug being recovered from the urine.
Mass Balance and Pharmacokinetics Following Oral Administration of [14C]-bevenopran 0.5mg
solution in Period 2
The mass balance of total radioactivity for the oral dose was determined, with an average of 96% of
the total radioactive dose recovered in urine and feces. An average of 73% was recovered in the
urine and 23% in the feces during the 178-hour sampling period.
Comparison of the urinary recovery of total radioactivity after IV and oral dosing of [14C]-bevenopran
would give a direct estimation of the fraction absorbed for the oral solution. This calculation assumes
that the oral dose is stable within the gastrointestinal tract and that only parent drug is available for
absorption. The mass balance recovery from urine was over different time periods for the IV and oral
radiolabeled doses. To provide comparative data, the mean urinary recovery over 72 hours postdose
for those subjects with evaluable urinary excretion data for both routes was used (n = 5; data for
Subjects 004 and 005 were omitted for Periods 1 and 2, respectively).
Over the 72-hour period after dosing with each regimen, the recovery of total radioactivity in the urine
represented 66% after IV administration in Period 1 compared to 67% after oral administration in
Period 2. The ratio of these values indicates that approximately 100% of the oral dose was
absorbed. With the fraction absorbed at approximately 100% and the absolute bioavailability of
bevenopran at 71%, this suggests an approximate 30% loss of absorbed drug via pre-systemic first
pass loss via the gut wall and liver.
Table 2: PK parameters for oral administration of [14C]-bevenopran 0.5mg solution in period 2
Period 2
Bevenopran 0.5mg
solution
Pharmacokinetic
Parameters
tlag (h)1)
tmax (h)a
Cmax (pg/mL)2)
AUClast (pg∙h/mL)b
AUC∞ (pg∙h/mL)b
AUC%extrap (%)
t½ (h)
MRT∞ (h)
CL/F (mL/min)
CLR (mL/min)
Vz/F (mL)
Bevenopran
Plasma Total
Radioactivity
Whole Blood Total
Radioactivity
Mean ± SD
(N = 6)
0.0 (0.0–0.0)
6.0 (4.0–6.0)
878 ± 157
20200 ± 3500
20900 ± 3430
3.79 ± 1.066
19.9 ± 1.17
25.7 ± 2.36
407 ± 66.1
220 ± 37.9
704000 ± 138000
Mean ± SD
(N = 6)
0.0 (0.0–0.0)
3.5 (3.0–4.0)
1560 ± 476
42000 ± 12500
43900 ± NC [n = 2]
13.93 ± NC [n = 2]
30.4 ± NC [n = 2]
40.8 ± NC [n = 2]
NC
135 ± 22.8
NC
Mean ± SD
(N = 6)
0.0 (0.0–0.0)
6.0 (3.0–6.0)
1200 ± 288
12100 ± 4860
NC
NC
NC
NC
NC
NC
NC
Metabolite Profiling of Bile String Data Following Intravenous Administration of
[14C]-Bevenopran 0.5µg in Period 1
The profiling results showed that only a single major peak was observed in the reconstructed
radiochromatograms of bile string extract and glucuronidase treated bile string extract. This peak
appeared at a retention time consistent with bevenopran and constituted almost 80% of total
on-column total radioactivity analysis. The remaining 14C activity was observed in 1 region of the
pools, with no peak containing >10.5% of the total radioactivity analysis.
Safety and tolerability of oral and intravenous administration of bevenopran
Both oral solutions and the IV infusion of bevenopran were well tolerated and there were no serious
or severe AEs, and no subject was withdrawn as a result of an adverse event
CONCLUSIONS
The use of intravenous and oral tracers combined with therapeutic oral dosing with bevenopran has
enabled determination of the complete absorption, bioavailability and disposition profiling of the
drug in a single study. In addition to the standard mass balance recoveries and metabolite profiling
this study design has also provided the following pharmacokinetic information
• The absolute bioavailability of bevenopran was determined to be 71% .
• The ratio of radioactive dose recovered in urine following IV dosing in Period 1 and oral dosing in
Period 2 indicates that 100% of the oral dose was absorbed. Comparison of the fraction
absorbed and absolute bioavailability indicates a loss of approximately 30% on first-pass through
the gut wall and liver
• The bile string data showed biliary excretion in man with approximately 80% of total material in
the bile as unchanged bevenopran plus 10% of minor metabolites
ACKNOWLEDGMENTS
The authors acknowledge the support from Eckert & Ziegler Vitalea Science, Davis, CA for
providing AMS based analyses for this study.
Study was funded by Cubist Pharmaceuticals, Lexington, MA, USA.
REFERENCES
1. Vassit Johnson LS, Young MA, Stevens LA et al (2012): A microtracer study of GSK962040,
a motilin receptor agonist, to support dosing regimens in the critical care setting. Clin.
Pharm. Ther. 91: Suppl. 1 S80.
19th North American ISSX/29th JSSX Meeting
www.quotientclinical.com
[email protected]