Introduction
Glucagon-like peptide-1 (GLP-1) is an incretin hormone secreted from L-cells in the
gastrointestinal tract in response to food intake [1]
[2]. GLP-1 regulates glucose homeostasis, stimulation of insulin release, and inhibition
of glucagon secretion from pancreatic β-cells through GLP-1 receptor signals. Food
intake and gastric emptying are also regulated through a GLP-1 receptor agonism effect
[3]
[4]
[5]. GLP-1 has been shown to preserve or increase pancreatic β-cell mass through the
inhibition of apoptosis, and stimulation of proliferation and differentiation. Therefore,
GLP-1 receptor agonists (GLP-1 RAs) have the potential to improve diabetes and obesity.
However, once GLP-1 is secreted into systemic circulation, it is rapidly degraded
by dipeptidyl peptidase IV (DPP-4) and neutral endopeptidase 24.11 (NEP) [6]
[7]. Consequently, the biological half-life of native GLP-1 is less than 2 min [8].
Recently, several synthetic GLP-1R agonists have been approved and made available
for the treatment of type 2 diabetes (T2DM). These GLP-1 RAs are resistant to degradation
by DPP-4 or NEP, and exhibit extended biological half-lives, a long-lasting glucose
lowering effect, and a delay in the rate of gastric emptying [9]
[10]
[11]
[12]. Among these RAs, liraglutide has been well investigated for its pharmacological
effects and pharmacokinetics in animal and clinical studies. The mechanism of biological
stability of liraglutide was obtained from macromolecularization, with strong binding
of albumin via acylated side chains [13]. The pharmacokinetic profiles of liraglutide revealed a long half-life and good
subcutaneous bioavailability in humans [14]. Therefore, these features are suitable for once-daily administration.
In our previous studies, we designed and characterized a novel GLP-1 RA, SKL-18287.
This RA consists of only natural L-amino acid, and is believed to exist in an oligomer
form in systemic circulation, with a molecular weight of around 40,000 daltons [15]. This unique feature of self-association may result in high biological stability
and a long-lasting glucose-lowering effect in T2DM treatment.
Interestingly, SKL-18287 shows a long-lasting glucose lowering effect in rodent diabetes
models, and displays more potent therapeutic effects than liraglutide in a non-obese
T2DM model (Goto-Kakizaki rats). SKL-18287 also displays a lower effect on gastric
emptying in the early treatment period in normal rats [16].
The purpose of the present study is to characterize the pharmacokinetic profiles of
SKL-18287 in various animal species, and to predict its biological half-life in humans.
We also demonstrate the tissue distribution of radioactivity after administration
of [3H]-SKL-18287 in rats. These results might be useful for clinical study design for
SKL-18287.
Materials and Methods
Materials
SKL-18287 (purity>95%, [Fig. 1]) and an internal standard (peptide 2, HAEGTFTSDVSSYLEGQAAKEFIAWLVKGKALIDEILAA-NH2) were prepared by solid-phase peptide synthesis. Tritium-labeled SKL-18287 was synthesized
by Sekisui Medical Co., Ltd (Tokyo, Japan) using tritium tyrosine, as described previously
[15]. Formic acid, acetonitrile, MeOH, toluene, NH4OH, and EDTA-2K were purchased from Wako Pure Chemical Corporation (Osaka, Japan).
Fig. 1 Primary structure of SKL-18287. *Position of the [3H] label. The C-terminal appendage is derived from the C-terminal helix 3 of the albumin
binding domain 3 of protein G, from the bacterial Streptococcal G strain 148. Ref.
[15].
Animals
Seven-week old male Sprague-Dawley (SD) rats were purchased from Charles River Laboratories
Japan, Inc. (Kanagawa, Japan). Eighteen-month old male cynomolgus monkeys were housed
and studied at Japan Laboratory Animals, Inc. (Tokyo, Japan). Forty-month old mini-pigs
were housed and studied at Nihon Bioresearch Center, Inc. (Gifu, Japan).
All animals were housed under standard conditions, with a 12-h light/dark cycle and
free access to water and a commercial diet (CRF-1 for rats, PS-A for monkeys, and
MP-A for mini-pigs; Oriental Yeast, Japan) for at least 5 days. All animal procedures
were approved by the Sanwa Kagaku Kenkyusho Institutional Animal Care and Use Committee,
and conducted in accordance with the institutional guidelines for the Conduct of Animal
Experiments in Research Institutions under the Jurisdiction of the Ministry of Health,
Labor, and Welfare (2006).
Pharmacokinetic study in rats, monkeys, and mini-pigs
SKL-18287 was suspended in phosphate-buffered saline (PBS) and administered intravenously
or subcutaneously to male SD rats, male cynomolgus monkeys, and male mini-pigs. The
doses for rats were 10 µg/kg for intravenous administration, and 10, 30, and 100 µg/kg
for subcutaneous administration. The doses for monkeys and pigs were 17.4 µg/kg (4
nmol/kg) for both administration routes. Blood samples were collected via the jugular
vein for rats, the cephalic vein for monkeys, and the jugular venous or femoral venous
catheter for mini-pigs, using a heparinized syringe at 0.083, 0.25, 0.5, 1, 2, 4,
6, 8, 10, 12, 24, 48, and 72 h for intravenous and 0.25, 0.5, 1, 2, 4, 6, 8, 10, 12,
24, 48, and 72 h for subcutaneous administration. Plasma was prepared by centrifugation
(1,500 × g, 4°C, 10 min) of the blood samples and stored at−70 °C until analysis.
Plasma sample preparation and LC-MS/MS conditions
The plasma samples and peptide 2 were loaded onto an OASIS MCX 96-well plate that
had been washed with 200 µL of MeOH and equilibrated with 200 µL of water. The wells
were subsequently washed with 400 µL of formic acid/water/acetonitrile (1/95/5, v/v/v)
and 400 µL of acetonitrile. SKL-18287 and peptide 2 were eluted from the wells with
30 µL of 28% NH4OH/water/acetonitrile (5/50/50, v/v/v), and the eluents were diluted with 70 µL of
formic acid/water/acetonitrile (5/50/50, v/v/v) for the analysis.
LC-MS/MS analyses were performed on an SC-10A HPLC system (Shimazu Corporation, Kyoto,
Japan) coupled to an API 4000 tandem mass spectrometer (Sciex, Shimazu Corporation,
Kyoto, Japan). Eluents were separated on the YMC Triart C8 (2.0 × 50 mm, 5 μm, YMC
Co., Ltd.) and kept at 50 °C using gradient flow of the mobile phase, consisting of
buffer A (formic acid/water=0.5 /100, v/v) and buffer B (formic acid/acetonitrile=0.5
/100, v/v). SKL-18287 was determined in the multiple reaction monitoring mode with
positive ionization, using peptide 2 as an internal standard.
Pharmacokinetic analysis
The pharmacokinetic parameters were calculated using a non-compartmental analysis
(WinNonlin Professional, Version 5.2.1; Pharsight Corporation, CA, USA). Bioavailability
was calculated by dividing AUC0-∞ after subcutaneous administration by AUC 0-∞ after intravenous administration.
Prediction of the human pharmacokinetic parameters of SKL-18287
The allometric scaling method was used to predict the human pharmacokinetic parameters
of SKL-18287 [17]. Body weight, plasma clearance, and volume of distribution after the intravenous
administration of SKL-18287 were plotted between body weight and plasma clearance,
and body weight and volume of distribution. The following equations were used for
scaling of the plasma clearance and volume of distribution. The allometric coefficients
(a, c) and allometric exponents (b, d) were calculated using data from the tested
species.
CLp (mL/h)=a·Wb
Vdss (mL)=c·Wd
W: body weight (kg)
a, c: allometric coefficients
b, d: allometric exponents
Human plasma clearance and volume of distribution were estimated with the body weight
set at 70 kg. Elimination half-life was calculated by the following equation:
Predicted t
1/2 =0.693·predicted human Vdss/Predicted human CLp
Tissue distribution of radioactivity in rats
Male SD rats (n=18; 3 per time point) were administered a single subcutaneous dose
of 30 µg eq. /kg of [3H]-SKL-18287 (414 MBq/mg), and the tissue concentration of radioactivity was determined
at 0.5, 2, 6, 12, 24, and 48 h after administration. At each time point, blood was
collected using an EDTA-2K-containing syringe from the abdominal aorta, under anesthesia
with isoflurane inhalation. Rats were sacrificed by exsanguination and the designated
tissues were excised. Plasma was prepared by centrifugation (1,500×g, 4°C, 10 min)
of the blood samples. Tissues collected from the SD rats were cerebrum, cerebellum,
pituitary gland, spinal cord, eyeball, harderian gland, parotid gland, sublingual
gland, mandibular gland, thyroid gland, trachea, mandibular lymph node, thymus, heart,
lung, liver, kidney, adrenal gland, spleen, pancreas, aorta, fat, brown fat, skeletal
muscle, dorsal skin, skin, thigh bone, bone marrow, testis, epididymis, seminal vesicle,
prostate gland, urinary bladder, stomach, small intestine, cecum, and large intestine.
Plasma and blood were each collected in a filter paper cup for combustion, as a sample
for radioactivity assay. The other tissues were washed with saline, wiped with filter
paper to remove saline, and weighed. The liver, pancreas, and kidney were each placed
in a plastic container, mixed with the same volume of ice-cold acetonitrile, and minced.
An equal weight of saline was then added, followed by homogenized with a polytron
homogenizer. Approximately 0.5 g (liver) and approximately 0.3 g (pancreas and kidney)
aliquots of each prepared homogenate were collected in a filter paper cup. The entire
dorsal skin, including the dosing site, was weighed, dissolved in 100 mL of a 2 mol/L
sodium hydroxide solution and 10 mL of toluene with heating, and diluted with water
to 200 mL. A 1 mL aliquot of the diluted solution was collected in a filter paper
cup. The gastrointestinal tissues were weighed, dissolved in 10 (stomach, cecum, and
large intestine) or 20 (small intestine) mL of 0.5 mol/L sodium hydroxide solution
with heating, and diluted with distilled water to 20 (stomach, cecum, and large intestine)
or 40 (small intestine) mL. A 1 mL aliquot of each diluted solution was collected
in a filter paper cup. The gastrointestinal contents collected at washing of the gastrointestinal
tract were placed in PP tubes previously rinsed with 1% Tween 80 and dried, for each
region, mixed with distilled water to approximately 100 g, and homogenized with a
polytron homogenizer. An approximately 0.5 g aliquot of the homogenate was collected
in a filter paper cup. The other tissues were collected entirely or in 150-mg aliquots
of tissues in a filter paper cup. Each sample, in a filter paper cup for combustion,
was dried at 40°C for more than 24 h in an incubator, and combusted using an automatic
combustion system. Generated 3H2O was absorbed in 12 mL of the scintillator Monophase S (PerkinElmer, USA), and subjected
to the radioactivity assay. The measured values from the combustion method were corrected
with the recovery, which was calculated using the 3H standard sample (Spec-Chec-3H, PerkinElmer, USA) for the sample oxidizer. The concentrations of radioactivity
in blood, plasma, and tissues were measured by 2700 TR liquid scintillation counter
(PerkinElmer, USA), and expressed as nmol eq./g or mL.
Results
Pharmacokinetics study in rats
The plasma concentration time profiles of SKL-18287 in male SD rats after intravenous
(10 µg/kg) and subcutaneous administration (10, 30, and 100 µg/kg) are shown in [Fig. 2]. The PK parameters of SKL-18287 in rats are shown in [Table 1]. For intravenous administration, SKL-18287 was disposed in a biphasic manner, and
the elimination half-life was 5.4±0.3 h. The plasma clearance (CLp) and apparent volume of distribution (Vdss) were 11.8±0.3 mL/h/kg and 75.6±1.7 mL/kg, respectively. The area under the plasma
concentration curve (AUC) of SKL-18287 was 851±23 ng·h/mL. For subcutaneous administration,
AUC and Cmax revealed a dose-dependent increase (10–100 µg/kg). The mean Tmax and bioavailability (BA) were from 3.2 to 4.0 h, and 64.3 to 68.4%, respectively.
The mean elimination half-life (5.2 to 5.8 h) and mean MRT (8.8– 9.7 h) changed little
among the 3 dose groups.
Fig. 2 Plasma concentration vs. the time profile of SKL-18287 in male SD rats, after single
intravenous (IV) and subcutaneous (SC) administration of SKL-18287 at a doses of 10
and 10 to 100 µg/kg, respectively. Data are expressed as mean±SD (n=5).
Table 1 Pharmacokinetic parameters of SKL-18287 administered to male SD rats.
|
Parameters
|
IV
|
SC
|
|
10 µg/kg
|
10 µg/kg
|
30 µg/kg
|
100 µg/kg
|
|
Cmax (ng/mL)
|
–
|
43.0±5.4
|
145±34
|
507±132
|
|
Tmax (h)
|
–
|
4.0±2.0
|
3.2±1.8
|
3.2±1.8
|
|
AUC 0-t (ng · h/mL)
|
820±19
|
510±58
|
1560±140
|
5810±920
|
|
AUC 0-∞ (ng· h/mL)
|
851±23
|
548±66
|
1640±150
|
5820±930
|
|
CLp (mL/h/kg)
|
11.8±0.3
|
–
|
–
|
–
|
|
Vdss (mL/kg)
|
75.6±1.7
|
–
|
–
|
–
|
|
t1/2 (h)
|
5.4±0.3
|
5.8±0.4
|
5.4±0.3
|
5.2±0.2
|
|
MRT (h)
|
6.4±0.2
|
9.7±0.7
|
8.8±1.0
|
9.0±0.1
|
|
BA (%)
|
–
|
64.3±7.7
|
64.4±5.7
|
68.4±10.9
|
Data are expressed as the mean±SD (n=3). IV; intravenous administration, SC; subcutaneous
administration, BA; bioavailability
Pharmacokinetics study in monkeys and mini-pigs
The plasma concentration time profiles of SKL-18287 in cynomolgus monkeys and mini-pigs
after intravenous and subcutaneous administration (17.4 µ/kg) are shown in [Fig. 3]. The pharmacokinetic parameters of SKL-18287 in monkeys and pigs are summarized
in [Table 2]. For intravenous administration, SKL-18287 was disposed in a biphasic manner in
monkeys. A bimodal distribution was observed in mini-pigs. The elimination half-life
for monkeys and pigs was 9.0±1.8 h and 12.9±1.0 h, respectively. The CLp and Vdss were similarly low for both species, at 2.89 mL/h/kg and 34.0 mL/kg, respectively
for monkeys, and 3.68 mL/h/kg and 67.0 mL/kg, respectively for pigs. For subcutaneous
administration, AUC and Cmax were similar for both species at 3800±1310 ng·h/mL and
151±37.2 ng/mL, respectively for monkeys, and 3960 ±943 ng·h/L and 130±30 ng·h/mL,
respectively for pigs. The tmax was 6.7 h for monkeys and 8.7 h for pigs. The mean
bioavailability was 58.1% for monkeys and 87.6% for mini-pigs.
Fig. 3 Plasma concentrations vs. the time profile of SKL-18287 in monkeys and mini-pigs,
after single intravenous (IV) and subcutaneous (SC) administration of SKL-18287 at
a dose of 17.4 μg/kg: a monkeys, b mini-pigs. Data are expressed as mean±SD (n=3–5).
Table 2 Pharmacokinetic parameters of SKL-18287 administered to monkeys and mini-pigs.
|
parameters
|
Monkey
|
Mini-pig
|
|
Route
|
IV
|
SC
|
IV
|
SC
|
|
Cmax (ng/mL)
|
–
|
151±37.2
|
–
|
130±30
|
|
Tmax (h)
|
–
|
6.7±1.2
|
–
|
8.7±3.1
|
|
AUC 0-t (ng · h/mL)
|
6350±2330
|
3660±1360
|
4960±477
|
3770±912
|
|
AUC 0-∞ (ng · h/mL)
|
6400±2340
|
3800±1310
|
5090±519
|
3960±943
|
|
CLp (mL/h/kg)
|
2.89±0.89
|
–
|
3.68±0.36
|
–
|
|
Vdss (mL/kg)
|
34.0±4.6
|
–
|
67.0±4.3
|
–
|
|
t
1/2 (h)
|
9.0±1.8
|
11.4±1.0
|
12.9±1.0
|
15.4±0.3
|
|
MRT (h)
|
12.3±2.6
|
18.6±1.5
|
18.2±0.7
|
25.5±0.7
|
|
BA (%)
|
–
|
58.1±20.0
|
–
|
87.6±13.6
|
Data are expressed as the mean±SD (n=3). IV; intravenous administration, SC; subcutaneous
administration, BA; bioavailability
Prediction of human pharmacokinetic parameters
The interspecies relationships for the clearance and volume of distribution taken
from all animal data are shown in [Fig. 4]. All data were fitted using allometric equations, and allometric coefficients were
calculated. A good correlation was observed between body weight and clearance, and
between body weight and volume of distribution. The clearance and volume of distribution
of SKL-18287 in humans were then predicted using allometric equations. The calculated
parameters in humans are shown in [Table 3]. When the human weight was set at 70 kg, the plasma clearance and volume of distribution
were calculated at 148 mL/h and 3170 mL, respectively. The estimated half-life in
humans was 14.8 h.
Fig. 4 Allometric relationships between body weight and a plasma clearance and b volume of distribution of SKL-18287. The linear regression for the logarithmic values
was calculated using the least-squares method (a: y=7.17 ×0.71, r2=0.966; b: y=60.6 ×0.93, r2=0.969). Extrapolated human values, based on a body weight of 70 kg (open circles)
and values from individual animals (black circles) are also shown.
Table 3 Predicted human pharmacokinetic parameters for SKL-18287.
|
W (kg)
|
CLp (mL/h)
|
Vdss (mL)
|
t
1/2
(h)
|
|
Rat
|
0.25
|
2.95
|
18.9
|
4.44
|
|
Monkey
|
6.9
|
19.8
|
233
|
8.2
|
|
Mini-pig
|
25.0
|
92
|
1675
|
12.6
|
|
Human
|
70.0
|
148
|
3170
|
14.8
|
Tissue distribution of radioactivity in rats
Following a single subcutaneous administration of 30 µg eq. /kg of [3H]-SKL-18287 to rats, radioactivity was highest in most tissues at 6 h post-dose ([Table 4]), indicating relatively slow distribution to tissues. The radioactivity in tissues
at 6 h after dosing was highest in plasma, followed by pancreas, blood, adrenal grand,
bone marrow, and liver. Low radioactivity levels were found in the fat, cerebrum,
cerebellum, and spinal cord during the study period. The tissue concentration vs.
the time profiles of radioactivity and tissue to plasma concentration ratios after
administration of [3H]-SKL-18287 is shown in [Fig. 5]. The radioactivity concentrations of pancreas and liver were relatively higher than
those of other tissues until 12 h post dose, and disappeared as the plasma concentration
decreased. The tissue concentrations of radioactivity in heart, kidney, and lung showed
relatively slow absorption and disappearance, compared to plasma. The tissue concentrations
of radioactivity in cerebrum, cerebellum, thyroid gland, stomach, small intestine,
and large intestine showed slow absorption, and then reached a steady state after
24 h post-dose.
Fig. 5 Tissue radioactivity concentrations and tissue to plasma concentration ratios in
male SD rats after a single subcutaneous administration of [3H]-SKL-18287 at a dose of 30 µg eq. /kg: a radioactivity concentrations in plasma, heart, lung, liver kidney, and pancreas;
b radioactivity concentrations in plasma, cerebrum, cerebellum, thyroid gland, stomach,
small intestine, and large intestine; and c tissue to plasma concentration ratios of radioactivity in major tissues, at 2 h and
6 h post-dose. Data are expressed as mean±SD or mean value (n=3).
Table 4 Concentrations of the radioactivity in the tissues of SD rats after a single subcutaneous
administration of [3H]-SKL-18287, at a dose of 30 µg eq. /kg.
|
Tissue
|
Radioactivity concentration (ng eq. of SKL-18287/g or mL)
|
|
0.5 h
|
2 h
|
6 h
|
12 h
|
24 h
|
48 h
|
|
Plasma
|
98±14.2
|
214±36
|
191±1
|
85.9±14.4
|
29.5±4.9
|
12.6±0.5
|
|
Blood
|
59.4±7.3
|
133±20
|
115±3
|
54.9±9.1
|
22±3.1
|
13.4±0.3
|
|
Cerebrum
|
1.79±0.49
|
4.73±0.13
|
7.05±0.04
|
8.24±0.28
|
7.86±0.18
|
8.27±0.27
|
|
Cerebellum
|
2.06±0.89
|
5.19±0.05
|
7.81±0.06
|
8.67±0.54
|
8.15±0.39
|
8.66±0.28
|
|
Pituitary gland
|
12.2±4.4
|
31.3±4.6
|
41.7±8.8
|
52.7±3.7
|
38±4.8
|
26.9±0.3
|
|
Spinal cord
|
2.01±0.93
|
3.6±0.25
|
6.44±0.78
|
6.58±0.12
|
5.81±0.39
|
5.98±0.15
|
|
Eyeball
|
1.43±0.42
|
4.97±0.33
|
8.08±0.83
|
7.75±0.23
|
7.05±0.03
|
6.8±0.29
|
|
Harderian gland
|
3.65±0.87
|
11.4±1.2
|
20.4±0.5
|
29.1±0.8
|
25.1±1.5
|
23.8±1.1
|
|
Parotid gland
|
7.81±1.45
|
35.1±1.5
|
70±9.7
|
54.2±12.6
|
31.4±6.2
|
20.6±2.6
|
|
Sublingual gland
|
7.06±1.45
|
29±4.2
|
43.6±2.6
|
34.7±3.3
|
25.5±3
|
19.5±0.8
|
|
Mandibular gland
|
7.81±2.25
|
29.8±4.1
|
49.7±5.6
|
39.9±1.7
|
26.1±3.1
|
22.8±1.2
|
|
Thyroid gland
|
13.4±4.2
|
30.1±7.3
|
38.9±8.2
|
40.8±1.3
|
24.2±4.6
|
24±4.5
|
|
Trachea
|
4.66±0.93
|
11.2±5
|
26.9±8.1
|
20.9±3.4
|
16.4±5.6
|
13.4±0.8
|
|
Mandibular lymph node
|
4.73±0.86
|
22.3±1.6
|
33.8±4.9
|
36.3±0.8
|
26.1±3.4
|
25.9±1.6
|
|
Thymus
|
3.31±1.41
|
12.3±1
|
23.6±1.7
|
31.2±2.1
|
31.2±2.6
|
29.7±0.1
|
|
Heart
|
7.92±1.75
|
21.8±2.4
|
29.5±0.3
|
23.2±1.3
|
18.7±0.6
|
17±0.1
|
|
Lung
|
11.1±1.1
|
31±2.6
|
58.7±1.9
|
64.3±6
|
64.6±12.3
|
53.8±11.9
|
|
Liver
|
25.2±6.3
|
93.6±28.4
|
78.9±3.8
|
69.6±7.2
|
35.7±1.9
|
26±0.4
|
|
Kidney
|
23±3.7
|
55.7±4.9
|
54.6±5.4
|
55.7±1.4
|
42.8±0.7
|
34.7±0.3
|
|
Adrenal gland
|
32.8±7.2
|
76.9±16.6
|
90.9±19.7
|
66.3±6.8
|
44.9±6.1
|
31.2±3.3
|
|
Spleen
|
8.59±1.64
|
31.6±7.3
|
42.4±2
|
48.5±3.7
|
42±5.2
|
29.8±2.4
|
|
Pancreas
|
20.1±6.6
|
93.5±14.3
|
132±8
|
96.4±21.3
|
36.6±2.7
|
22.5±1.5
|
|
Aorta
|
4.49±1.06
|
12.4±0.7
|
21.7±7.8
|
21.6±3.2
|
17.9±5.2
|
15.3±3.3
|
|
Fat
|
0.935±0.2
|
2.71±0.28
|
3.27±0.65
|
3.28±0.48
|
3.99±0.22
|
3.46±0.27
|
|
Brown fat
|
4.42±1.35
|
11.2±1
|
26.1±6.6
|
19.6±0.8
|
16.7±3.3
|
19.2±2.6
|
|
Skeletal muscle
|
2.09±0.43
|
7.07±0.97
|
11.9±0.5
|
11.3±1.3
|
13.3±0.5
|
12.4±1.7
|
|
Dorsal skin*
|
382±64
|
130±36
|
51.8±19.4
|
37±4.3
|
26.4±1.8
|
22.2±5.4
|
|
Skin
|
3.77±0.71
|
16.8±0.7
|
28.7±2.3
|
30.9±3.7
|
38.5±5.4
|
30.1±5.3
|
|
Thigh bone
|
5.37±1.1
|
15.3±5.2
|
20.4±4.5
|
14.1±4.3
|
12.5±3.8
|
10.4±1.4
|
|
Bone marrow
|
10.5±1.6
|
51.2±16.3
|
79.3±15.4
|
108±13
|
96.3±15.3
|
64.7±7.7
|
|
Testis
|
1.92±0.28
|
10.6±2.3
|
20.2±4.4
|
17.7±1.1
|
15.5±1.6
|
12.6±0.6
|
|
Epididymis
|
2.3±0.63
|
10.4±2.1
|
20.6±0.8
|
22.1±1.4
|
17.9±1.1
|
14.9±1.4
|
|
Seminal vesicle
|
4.66±1.59
|
14.8±0.8
|
34.1±1.2
|
39.2±4
|
37.5±1.7
|
42.4±8.5
|
|
Prostate gland
|
5.67±1.62
|
26.8±9.4
|
39.9±9
|
47.1±5.5
|
41.9±3.8
|
34.9±4
|
|
Urinary bladder
|
3.93±0.64
|
15.8±4.2
|
27.3±2.2
|
23.7±1.7
|
16.2±0.6
|
14.5±1.7
|
|
Stomach
|
6.35±1.33
|
23.4±3.7
|
43.2±5.2
|
44.8±2.5
|
28.8±5.6
|
23.9±3.2
|
|
Small intestine
|
5.64±0.59
|
21.8±4.9
|
37.2±3.5
|
48.9±6.5
|
37.8±3.7
|
23.2±4.5
|
|
Cecum
|
5.13±0.66
|
17.4±4
|
30.3±2.9
|
40.9±15.2
|
26.4±2.8
|
22.9±0.7
|
|
Large intestine
|
4.26±0.54
|
13.9±3.6
|
23.2±3.2
|
24.9±1.4
|
22.5±0.8
|
18.9±1.4
|
Data are expressed as the mean±SD (n=3), *, including dosing site.
Discussion
In this present study, we investigated the pharmacokinetic properties of SKL-18287,
a novel long acting GLP-1 RA, in rats, monkeys, and mini-pigs. After intravenous administration
of SKL-18287, the apparent volume of distribution was similar to the plasma volume,
in all animal species. These results indicate that a large fraction of SKL-18287 is
localized to systemic circulation. After subcutaneous administration, SKL-18287 was
slowly absorbed into systemic circulation, reaching a maximum concentration at around
3.2 to 4 h in rats, and 6.7 and 8.7 h in monkeys and mini-pigs, respectively. The
elimination half-life after intravenous and subcutaneous administration of SKL-18287
was shorter in rats (IV, 5.4 h; SC, 5.2 to 5.8 h) than in monkeys (IV, 9.0 h; SC,
11.4 h) and in mini-pigs (IV, 12.9 h; SC, 15.4 h). Since SKL-18287 is highly stable
in rat and human plasma [15], the elimination rate of SKL-18287 from systemic circulation may depend on the order
of the plasma flow rate in the kidneys [18]. SKL-18287 exhibited good bioavailability in all animal species, indicating the
SKL-18287 was almost fully absorbed from the subcutaneous tissues. Based on the studies
of Steensgaard, et al. [19] and Wang, et al. [20], liraglutide forms an oligomer (heptamer, octamer, or dodecamer) in solution, a
feature considered to result in slow absorption from the dosing site (subcutaneous
tissue). SKL-18287 also forms an oligomer in solution, and similar phenomena (i. e.,
slow absorption) were observed in all animal species.
A number of studies on protein and peptide drugs have shown that allometric scaling
is effective in predicting human clearance and volume of distribution [21]
[22]
[23]. The relationship between plasma clearance and body weight, and between volume of
distribution and body weight revealed high correlations, and human pharmacokinetic
parameters were predicted by a simple allometric scaling equation [17]. The estimated half-life of SKL-18287 in humans was 14.8 h, a result that was similar
to those of liraglutide in healthy volunteers (range 11–15 h) [14].
Tissue distribution of radioactivity was not as extensive after subcutaneous administration
of [3H]-SKL-18287, and remained mainly at the injection site at the early time points.
Interestingly, the radioactivity concentration in the pancreas was higher than in
other tissues, until 12 h post-dose. The pancreas is a major target of GLP-1 RA, and
GLP-1 receptors are highly expressed in this tissue. Regarding other GLP-1 receptor
expressing tissues, such as cerebrum, cerebellum, lung, heart, kidney, small intestine,
and large intestine, the concentration of radioactivity in each tissue was not so
high. Although we do not know the reason for the distribution of SKL-18287 to the
pancreas, it is possible that SKL-18287 multimer formation may be affecting the tissue
distribution to the pancreas. Additional work is necessary to elucidate the distribution
properties of SKL-18287.
In our previous study, SKL-18287 had more potent therapeutic effects than liraglutide
in Goto-Kakizaki rats. Moreover, SKL-18287 showed a lower effect on gastric emptying
in the early treatment period in normal rats [16]. The present tissue distribution results suggest that higher pancreas selectivity
influences insulin secretion, leading to a potent glucose lowering effect.
