In vitro and in vivo profile of 5-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-1H-indole-2-carboxylic acid benzylmethyl carbamoylamide (dirlotapide), a novel potent MTP inhibitor for obesity

Article abstract of DOI:10.1016/j.bmcl.2007.01.018

The synthesis of a novel gut selective MTP inhibitor, 5-[(4′-trifluoromethyl-biphenyl-2-carbonyl)-amino]-1H-indole-2-carboxylic acid benzylmethyl carbamoylamide (dirlotapide), and its in vitro and in vivo profile are described. Dirlotapide (3) demonstrate

Full text of DOI:10.1016/j.bmcl.2007.01.018

Bioorganic & Medicinal Chemistry Letters 17 (2007) 1996–1999
In vitro and in vivo profile of 5-[(40-trifluoromethyl-biphenyl-2-
carbonyl)-amino]-1H-indole-2-carboxylic acid benzylmethyl
carbamoylamide (dirlotapide), a novel potent
MTP inhibitor for obesity
Jin Li,* Brian S. Bronk, John P. Dirlam, Alan E. Blize, Peter Bertinato, Burton H. Jaynes,
Anne Hickman, Christine Miskell, Usha A. Pillai, Jay S. Tibbitts, Michelle L. Haven,
Nicole L. Kolosko, Chris J. Barry and Tara B. Manion
Pfizer Inc., Groton, CT 06340, USA
Received 18 November 2006; revised 5 January 2007; accepted 8 January 2007
Available online 17 January 2007
Abstract—The synthesis of a novel gut selective MTP inhibitor, 5-[(40-trifluoromethyl-biphenyl-2-carbonyl)-amino]-1H-indole-2-
carboxylic acid benzylmethyl carbamoylamide (dirlotapide), and its in vitro and in vivo profile are described. Dirlotapide (3) dem-
onstrated excellent potency against MTP enzyme in HepG2 cells and canine hepatocytes. This novel MTP inhibitor also showed
excellent efficacy when tested in a canine food intake model.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Obesity is a major health problem that has been associ-
ated with an increased incidence of type 2 diabetes mel-
litus, hypertension, and dyslipidemia.1,2 Currently,
obesity is the second leading cause of preventable death
in the United States, resulting in more than 300,000
deaths per year.3 In the US, more than 35% of the adult
population is overweight (BMI 25–29), and 26% of the
adult population is obese (BMI P30).4 Over the past
10 years, the prevalence of obesity in the United States
has increased by about 50% and a large component of
this increase has been in children.5,6 Weight loss is
expected to have a clinically significant impact on the
risk or selected co-morbidities (type 2 diabetes, cardio-
vascular disease, obesity associated quality of life issues,
etc.). Currently there are only two branded anti-obesity
agents on the market, orlistat (1, Roche, launched
1998)7 and sibutramine (2, Abbott, 1997).8,9 Microsom-
al triglyceride transfer protein (MTP)10 is involved in the
assembly of triglyceride-rich chylomicrons in entero-
cytes10–12 and very low density lipoproteins (VLDL) in
hepatocytes. MTP is located in intestinal and liver
tissues where it plays a role in lipid assembly and trans-
port.10 A gut selective MTP inhibitor will inhibit trans-
port of lipids within the endoplasmic reticulum without
significantly decreasing serum triglyceride levels.13 MTP
inhibitors represent a unique class of anti-obesity agents
that offer potent weight loss through appetite suppres-
sion14 and intestinal fat mal-absorption.15 In this paper,
we would like to disclose the discovery of a potent and
gut selective MTP inhibitor, dirlotapide (3), which is
currently in clinical trials as an anti-obesity agent
(Fig. 1).
The preparation of dirlotapide (3) is through a three-
step linear synthesis and outlined in Scheme 1. Trifluo-
romethyl biphenyl carboxylic acid (4) was coupled with
amino indole 516 using EDC/HOBT coupling conditions
to provide the ester 6. Hydrolysis of the ester function
under basic conditions furnished the acid 7 in high yield.
The acid 7 was then coupled with phenylglycine deriva-
tive 8 to provide dirlotapide 3.
As depicted in Table 1, dirlotapide (3) is a potent inhib-
itor of MTP as demonstrated by the inhibition of Apo B
secretion from human HepG2 cells (IC50 = 1.5 nM) and
in canine hepatocytes (IC50 = 2.9 nM).
Keywords: MTP; Obesity.
*
The selectivity of dirlotapide (3) for MTP inhibition,
relative to inhibition of other receptors in the body,
Corresponding author. Tel.: +1 860 715 3552; fax: +1 860 715
9259; e-mail: jin.li@pfizer.com
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.01.018

J. Li et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1996–1999
Table 1. In vitro MTP inhibition dataa
1997
H
N
OHC
Compound
Canine MTPi
IC50 (nM)
Human HepG2
MTPi IC50 (nM)
O
O
Dirlotapide (3)
2.9
1.5
a Values are averages of at least five determinations.
1. orlistat
CF3
Table 2. Caco-2 data for dirlotapide (3)
Concn (lM)
+/ꢀ Ca2+
pH
P(app)A–B
· 106 cm/s
P(app)B–A
· 106 cm/s
O
N
O
Me2N
1
1
+
ꢀ
+
6.5/7.4
6.5/7.4
6.5/7.4
0.6 0.0
0.9 0.3
0.4 0.0
0.4 0.1
—
0.3 0.1
HN
N
H
Cl
O
N
10
2. sibutramine
3. dirlotapide
Table 3. In vitro clearance for dirlotapide (3)
Figure 1. Selected anti-obesity drugs.
Species
Microsomal CLint Microsomal CLh Hepatocyte CLh
(mL/min/kg)
(mL/min/kg)
(mL/min/kg)
was demonstrated by a lack of activity (defined as
IC50 > 1 lM) when tested at 1 lM against a broad range
of selected peripheral and central receptors.17 Since
absorption by intestinal enterocytes is required for effi-
cacy, but systemic exposure is not required, the ADME
profile of dirlotapide 3 was assessed both in vitro and
in vivo. Caco-2 cell permeability data of dirlotapide
(3) in three-week-old cells are indicative of poor trans-
cellular absorption (P(app)A–B < 1 · 10ꢀ6 cm/s) with no
indication of efflux (Table 2).
Rat
Dog
645
1091
835
91
39
28
27
7
31.5
—
Monkey
Human
—
7.8
Table 4. Inhibition of turnover in human liver microsomes using
specific CYP inhibitors
Inhibitor
P450 Concn (lM) Concn (lM) T1/2
inhibited inhibitor
dirlotapide
(min)
Dirlotapide (3) has moderate turnover in rat, dog,
monkey, and human microsomes with projected hepat-
ic extraction (ER) of 55%, 80%, 68%, and 35%, respec-
tively. The microsomal and hepatocyte clearance are
listed in Table 3.
—
Ketoconzole
Sulfaphenazole
—
—
0.5
0.5
0.5
0.5
0.5
0.5
48
369
47
3A4
2C9
1
5
(S)-Mephenytoin 2C19
200
5
70
36
Quinidine
Furafylline
2D6
1A2
5
45
The mean IC50 values of dirlotapide (3) for the inhibi-
tion of CYP1A2, CYP2C9, CYP2C19, CYP2D6, and
CYP3A are all >30 lM. Microsomal incubations with
specific CYP inhibitors (Table 4) were conducted to gain
more quantitative information regarding the relative
contribution of each isoform to the overall CYP medi-
ated metabolism of dirlotapide (3). These studies indi-
cated that 3A is the predominant CYP mediated
metabolic pathway for this compound.
Dirlotapide (3) binds significantly to plasma compo-
nents with free fractions of 0.032%, 0.0088%, 0.026%,
and 0.0097% in rat, dog, monkey, and human plasma,
respectively (Table 5). Dirlotapide (3) has also been
shown to have low red blood cell partitioning with
blood-to-plasma concentration ratios of 0.64, 0.54,
CF3
CF3
N
CO2Et
O
N
O
a
O
+
OH
OR
H2N
N
H
6 R=Et
7 R=H
b
4
5
CF3
Table 5. Nonspecific binding and red blood cell partitioning for
dirlotapide (3)
O
c
N
O
+
fu(b)b (%)
BPRa
HN
N
N
Species
N
Concn (lM)
H2N
H
O
0.032 (0.022)c
0.0088 (0.0052)c
0.026 (0.08)c
0.64
0.54
NA
0.61
O
N
Rat
Dog
4
4
6
5
1.0
0.2
0.2
0.2
8
dirlotapide 3
Monkey
Human
0.0097 (0.0020)c
Scheme 1. Reagents and conditions: (a) EDC, HOBT, DIPEA, DCM,
rt, 85%; (b) LiOH, THF/H2O, reflux, 4 h, 98%; (c) PyBroP, DIPEA,
DCM, rt, 95%.
a Blood/plasma concentration ratio.
b Free fraction (blood).
c Standard deviation.

1998
J. Li et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1996–1999
Table 6. IV pharmacokinetic summary for dirlotapide (3)
Species
Rat
Dog
Monkey
Dose (mg/kg)
Sex
0.3
M
1
M
5
M
0.5
M
1
1a
M
0.5
M/F
M
3
N
3
4
2
14
3
4
3/3
CLplasma (mL/min/kg)
Vdss (L/kg)
T1/2 (h)
92 (11)
11 (4.6)
2.0 (0.6)
51 (5.8)
55 (5.9)
37 (17)
8.0 (3.9)
5.8 (4.1)
485 (196)
530 (235)
5.5 (1.3)
1.0 (0.21)
5
4.7 (0.9)
1.0 (0.20)
14 (1.0)
7.6 (4.2)
—
12 (2.0)
2.2 (0.5)
6.7 (0.6)
684 (118)
703 (121)
3.3
11.3
5612
5892
7.8 (3.4)
—
2825 (1663)
AUC(0–t) (ng h/mL)
AUC(0–1) (ng h/mL)
1494 (284)
1550 (316)
3258 (678)
3614 (683)
a Study conducted in portal cannulated dogs.
Table 7. PO (fed) pharmacokinetic summary for dirlopatide (3)
Species
Rat
Dog
Dose (mg/kg)
Sex
3
M
30
M
300
M
2
1c
M
4
0.5
M
3
5
M
3
N
3
4
Vehicle
T1/2 (h)
a
a
b
d
6.8 (1.9)
e
e
5.9 (1.8)
6.3 (3.5)
40.7 (8.4)
4.7 (1.5)
2.1 (0.0)
2.6
6.3 (6.3)
379 (152)
21.0 (11.7)
2.8 (1.5)
2.3
NC
NC
27
6
14.1 (5.0)
663 (607)
31.1 (18.2)
3.0 (1.0)
43 (39)
AUC(0–1) (ng h/mL)
Cmax (ng/mL)
Tmax (h)
289 (72.1)
26.8 (3.7)
6.3 (2.0)
1933 (197)
250 (81)
3.0 (1.0)
12 (2)
F (%)
1
10.6 (7.2)
(a) 80/20 PEG/saline; (b) PEG; (d) miglyol; and (e) miglyol/cremophor/water.
c Study conducted in portal cannulated dogs.
Table 8. Efficacy of dirlotapide (3) administered orally in obese Beagles for 3 months
Baseline
Treatment
Change (%)
Food intake (g)
Cholesterol (ng/mL)
Fecal fat (%)
Placebo
Dirlotapide (3)
282.2
305.4
282.3
139.6
ꢀ2
ꢀ54
10
Placebo
Dirlotapide (3)
219
259
240
127
ꢀ51
13
Placebo
Dirlotapide (3)
1.64
1.71
1.85
7.50
339
and 0.61 in rats, dogs, and humans, respectively
(Table 5).
distribution (3–11 L/kg). Following oral administration
of 3 mg/kg the mean plasma Cmax of 4.7 ng/mL was
achieved at 2.1 h post-dose and the bioavailability (F)
was 2.6%. At an oral dose of 30 mg/kg the resulting mean
Cmax was 21 ng/mL at 2.8 h post-dose and F was 2.3%.
Between 3 and 30 mg/kg the systemic exposure increased
in proportion to dose. However, increasing the dose to
300 mg/kg produced no further increase in systemic
exposure when dosed as a solution in PEG 400. In
male Beagle dogs the mean plasma clearance was
6.0 mL/min/kg and the mean volume of distribution
was 1.0 L/kg (Table 6). Following oral administration
to dogs dirlotapide (3) was moderately absorbed with
oral bioavailability ranging from 12% to 43% (Table 7).
Intestinal selectivity should reduce the risk for adverse
side effects. The in vivo selectivity of dirlotapide (3)
for intestinal compared to liver MTP inhibition has been
demonstrated in several animal models. In a murine
model16 investigating intestinal MTP inhibition, dirlo-
tapide (3) demonstrated potent inhibition of intestinal
fat absorption with an ED25 of 0.16 mg/kg. In contrast,
it was a poor inhibitor of hepatic MTP as demonstrated
in a murine model of triglyceride lowering, where it had
an ED25 of 6 mg/kg.
The iv and oral pharmacokinetics of dirlotapide (3) were
investigated in rats and dogs, and iv pharmacokinetics
were investigated in monkeys. All pharmacokinetic
parameters presented are derived from measurement of
plasma concentrations. Tables 6 and 7 summarize the
calculated pharmacokinetic parameters from the in vivo
studies. Male rats had plasma clearance values ranging
from 14 mL/min/kg at a dose of 5 mg/kg to 92 mL/min/
kg at a dose of 0.3 mg/kg and moderate/high volume of
Robust anti-obesity efficacy was demonstrated in several
canine studies with ad libitum feeding. In a 3-month effi-
cacy study using medium chain triglyceride oil based
solution, obese spayed and neutered beagles received
individual dosages of dirlotapide (3) ranging from 0.15
to 0.5 mg/kg (average dose ꢁ0.39 mg/kg). Doses were
adjusted to limit the degree of weight loss to ꢁ2%/week.
The dogs lost a mean of 18.8% of body weight (1.5% per

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K.; Lan, S.-J.; Maccagnan, T. J.; Ricci, B.; Yan, M.;
Young, D.; Chen, Y.; Fryszman, O. M.; Logan, J. V. H.;
Musial, C. L.; Poss, M. A.; Robl, J. A.; Simpkins, L. M.;
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week) compared to placebo animals that gained a mean
of 10.6% body weight. In addition, average food intake
was decreased by 54%, cholesterol decreased by 51%,
and fecal fat increased from 1.86% to 7.50%. Following
the weight loss phase, body weight was maintained in
dogs receiving dirlotapide (3) over a 28-day period at
dosages of 0.1–0.35 mg/kg (Table 8).
In conclusion, a very potent MTP inhibitor, dirlotapide
(3), has been discovered and profiled. The robust effica-
cy has been shown in animal models. The unique
ADME properties of dirlotapide (3) minimized the drug
exposure systemically; therefore a better safety profile is
expected in clinical trials.
14. Hickman, M. A.; Lundy, K. M.; Morgan, B. P.
EP1099438A2, 2001.
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B. P. EP1099439A2, 2001.
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