Synthesis and biological evaluation of azobicyclo[3.3.0] octane derivatives as dipeptidyl peptidase 4 inhibitors for the treatment of type 2 diabetes

Article abstract of DOI:10.1016/j.bmcl.2010.04.120

A series of novel azobicyclo[3.3.0]octane derivatives were synthesized and evaluated as dipeptidyl peptidase 4 (DPP-4) inhibitors. The effort resulted in the discovery of inhibitor 2a, which exhibited excellent efficacies in an oral glucose tolerance test. Introduction of methyl group (2j) could prolong the inhibition of serum DPP-4 activity.

Full text of DOI:10.1016/j.bmcl.2010.04.120

Bioorganic & Medicinal Chemistry Letters 20 (2010) 3565–3568
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of azobicyclo[3.3.0] octane derivatives
as dipeptidyl peptidase 4 inhibitors for the treatment of type 2 diabetes
a
a
a
a
a
Tang Peng Cho a, , Yang Fang Long , Lin Zhi Gang , Wang Yang , Lu He Jun , Shen Guang Yuan ,
Fu Jian Hong a, Wang Lin a, Guan Dong Liang a, Zhang Lei a, Luo Jing Jing a, Gong Ai Shen a, She Gao Hong a,
Wang Dan a, Feng Ying b, Yan Pang Ke a,b, Leng Ying b, Feng Jun a, Mong Xian Tai a
*
a Shanghai Hengrui Pharmaceuticals Co., Ltd, 279 Wenjing Road, Shanghai 200245, China
b State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, 555 Zu Chong Zhi Road, Shanghai 201203, China
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel azobicyclo[3.3.0]octane derivatives were synthesized and evaluated as dipeptidyl pep-
tidase 4 (DPP-4) inhibitors. The effort resulted in the discovery of inhibitor 2a, which exhibited excellent
efficacies in an oral glucose tolerance test. Introduction of methyl group (2j) could prolong the inhibition
of serum DPP-4 activity.
Received 10 March 2010
Revised 21 April 2010
Accepted 27 April 2010
Available online 18 May 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Azobicyclo[3.3.0]octane derivatives
Dipeptidyl peptidase 4
Oral glucose tolerance test
Type 2 diabetes mellitus
Inhibition of dipeptidyl peptidase (DPP-4) has been a promising
new approach to the treatment of type 2 diabetes mellitus (T2DM)
since three drugs have been approved, Sitagliptin (JanuviaÒ, MK-
0431),1 Vildagliptin (GlavusÒ, LAF-237),2 and Saxagliptin (Ong-
The 5
2. Ketone 4a was stereoselectively reduced to 5b-alcohol 6, which
was mesylated and inverted to 5 -amine 9 by employing potas-
a isomer of urea 2a was synthesized as shown in Scheme
a
sium phthalimide displacement followed by hydrolysis. Compound
lyza™, BMS-477118).3–5 (Fig. 1). DPP-4 is
a
key regulatory
9 was further substituted by (S)-1-(2-chloroacetyl)pyrrolidine-2-
enzyme and a signaling factor of insulin-stimulating hormones,
glucagon-like peptide (GLP-1), and glucose-dependent insulino-
tropic polypeptide (GIP).6–8 Due to rapid degradation of the both
incretin hormones by DPP-4, the half-lives of active GLP-1 and
GIP are extremely short.9 Inhibition of plasma DPP-4 enzyme leads
to prolong the actions of endogenous GLP-1 and GIP, which ulti-
mately decreases blood glucose levels and glucagon levels, and im-
proves glucose homeostasis with a low risk of hypoglycemia and
potential for disease modification.10
Bicyclo[3.3.0]octane derivative 1 was previously reported to be
a potent DPP-4 inhibitor,11 we now report the synthesis, biological
evaluation and SAR of this series of azobicyclo[3.3.0]octane com-
pounds (2) which possess less stereocenters and can be synthe-
sized more conveniently.
carbonitrile (10) to afford the desired 5a
-substituted isomer 2h.14
In order to discourage intramolecular cyclization of amine to
nitrile so as to improve stability of compounds 2a and 2h,15
5-methyl derivatives were also synthesized (Schemes 3 and 4).
Reaction of ketone 4a with tosylmethyl isocyanide gave a mixture
of diastereomeric carbonitriles 11, which was selectively methyl-
ated to give 5
nitrile 12 followed by Curtis rearrangement afforded the
corresponding 5 -methyl-5b-amine 14, which was substituted
with 10 to give 5 -methyl isomer 2i. In order to obtain 5b-methyl
a-methyl derivative 12. Acid hydrolysis of the carbo-
a
a
isomer 2j, another synthetic approach was applied. Wittig methyl-
enation of ketone 4a gave methylene 1512, which was treated with
AgClO4 and trimethylsilyl cyanide followed by hydrolysis to give
desired tertiary 5b-methyl isocyanide 16 in a yield of 18%.16 Isocy-
anide 16 was hydrogenated and alkylated to give 5b-methyl
isomer 2j. The stereochemistry of 5-methyl groups for 2i and 2j
was determined by 2D NOESY.
Enzymatic inhibitions of DPP-4, DPP-8, and DPP-9 of azobicy-
clo[3.3.0]octane derivatives were outlined in Table 1.17 The selectiv-
ity of DPP-4 against DPP-8 and DPP-9 is critical as the inhibition of
these two enzymes may be associated with profound toxicities.18
Analog2a showedan IC50 of 9 nM against DPP-4. The selectivityratio
As shown in Scheme 1, Boc-protected cyclopenta[c]pyrrol-
5(1H)-one 312 was first deprotected and then treated with various
acyl chlorides to afford key intermediates 4a–g, which underwent
reductive amination with (S)-1-(2-aminoacetyl)-pyrrolidine-2-car-
bonitrile (5) to give 5b substituted compounds 2a–g.13
* Corresponding author. Tel.: +86 21 54752877; fax: +86 21 54759072.
E-mail address: tangpc@shhrp.com (T.P. Cho).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.04.120

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T. P. Cho et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3565–3568
F
F
HO
NH2
O
HO
N
N
N
H
N
N
CN
O
H2N
N
F
N
CN
O
CF3
Vildagliptin
LAF-237
Saxagliptin
BMS-477118
Sitagliptin
MK-0431
N
HN
5
N
R1
6
HN
CN
O
5
2
4
CN
O
6
4
H
H
H
1
H
3
1
3
N
2
O
R2
O
1
2
Figure 1. Structures of selected DPP-4 inhibitors and design of new azobicyclo[3.3.0]octane derivatives as DPP-4 inhibitors.
O
N
O
N
N
HN
N
R2
a N(CH3)2
b NHCH(CH3)2
c N(CH2)4
d N(CH2)5
e N(CH2CH2)2O
CN
O
a, b
c
H
H
H
H
H
H
5
R2
O
O
O
R2
2a-g
f
OCH3
O
3
4a-g
g CH2OH
Scheme 1. Reagents and conditions: (a) HCl, Et2O, rt; (b) R2COCl, K2CO3, CH3CN, rt, 40–77%, over two steps; (c) (S)-1-(2-aminoacetyl)-pyrrolidine-2-carbonitrile TFA salt (5),
NaBH(OAc)3, THF, 15–43%.
N
O
O
O
N
OH
N
OMs
N
NH2
HN
N
CN
O
a
b
d
c
e
H
H
H
H
H
H
H
H
H
H
H
H
N
N
N
N
N
O
N
O
N
O
O
N
O
O
N
7
6
8
9
2h
4a
Scheme 2. Reagents and conditions: (a) LiAl(t-BuO)3H, THF, À30 °C, 90%; (b) MsCl, Et3N, CH2Cl2, 0 °C to rt, 75%; (c) phthalimide potassium, DMF, 60 °C, 90%; (d) N2H4, EtOH,
reflux, 50%; (e) (S)-1-(2-chloroacetyl)pyrrolidine-2-carbonitrile (10), K2CO3, KI, CH2Cl2, rt, 45%.
O
H
CN
O
N
CN
COOH
H
N
NH2
H
N
a
c
b
d,e
f
H
H
H
H
H
H
H
H
H
H
N
N
N
N
N
N
N
N
O
O
N
N
N
N
O
O
O
O
14
2i
13
4a
11
12
Scheme 3. Reagents and conditions: (a) tosylmethyl isocyanide, t-BuOK, 51%; (b) CH3I, LHMDS, THF, rt; (c) concd HCl solution, 50 °C, 92%; (d) ClCOOC2H5, NaN3, À5 °C;
(e) toluene, reflux, then 8 N HCl solution; (f) (S)-1-(2-chloroacetyl)pyrrolidine-2-carbonitrile (10), K2CO3, KI, CH2Cl2, rt, 55%.
(SR) of 2a for DPP-4 versus DPP-8 was 1931-fold. Previous work from
our laboratories indicated that increasing the steric bulk at 2-posi-
tion of bicyclo[3.3.0]octane derivatives reduced the potency.11 This
trend was also observed in the series of derivatives2 with compound
2b (IC50 0.12
lM), 2c (IC50 0.039 lM), 2d (IC50 0.069 lM), and 2e
(IC50 0.050 M). When compared to compound 2a, carbamate 2f
l
showed a threefold decrease and amide 2g exhibited twofold
decrease in DPP-4 inhibitory potency. The inverted isomer 2h was

T. P. Cho et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3565–3568
3567
O
H
O
N
N
NH2
H
NC
N
a
b
c
d
H
H
H
H
H
H
H
H
H
N
N
N
N
N
N
N
N
N
N
O
O
O
O
O
4a
16
15
17
2j
Scheme 4. Reagents and conditions: (a) methyltriphenylphosphonium iodide (CH3IP(C6H5)3), t-BuOK, 80%; (b) TMSCN, AgClO4, CH2Cl2, rt, 18%; (c) 6 N HCl solution, rt, 76%;
(d) 10, K2CO3, KI, CH2Cl2, rt, 87%.
The rat pharmacokinetic studies of selected compounds, as
shown in Table 2, revealed that 5 -methyl derivative 2i exhibited
Table 1
Potency and selectivities of azobicyclo[3.3.0]octane derivatives
a
a,b
best pharmacokinetic profiles. Introduction of methyl group in 5-
position has increased the half life in buffered solution (pH 7.2)
from 25.7 h to about 50 days.19
Compd
R2
R1
IC50
(lM)
DPP-4
DPP-8
DPP-9
2a
2b
2c
2d
2e
2f
2g
2h
2i
N(CH3)2
NHCH(CH3)2
N(CH2)4
N(CH2)5
N(CH2CH2)2O
OCH3
H(
H(
H(
H(
H(
H(
H(
H(b)
CH3(
CH3(b)
a
a
a
a
a
a
a
)
)
)
)
)
)
)
0.009
0.120
0.039
0.069
0.050
0.024
0.014
0.083
0.013
0.013
17.38
ND
ND
ND
ND
18.24
ND
ND
4.89
38.48
5.70
ND
ND
ND
ND
5.04
ND
ND
16.90
2.43
In pharmacodynamic studies, oral glucose tolerance tests
(OGTT) in lean mice (ICR mice) were conducted to determine the
efficacy of 2a.11 Oral administration of 2a with indicated doses to
ICR mice 0.5 h before an oral glucose challenge produced signifi-
cant decrease in glucose excursion. Compared to LAF-237, which
reduced the AUC0–120 min values 35.8%, 41.7%, and 51.4% at the dose
of 1, 3, and 10 mg/kg, compound 2a showed comparable effect
with the decrease rate of 39.7%, 42%, and 45%, respectively (Fig. 2).
At the same time, the efficacy of 2a, 2i, and 2j on the inhibition
of serum DPP-4 activity was examined in cynomolgus monkey
(Macaca mulatta). Oral dosing of 20 mg/kg 2j inhibited serum
DPP-4 activity by >75% within 9 h post-dose, whereas inhibition
of >75% persisted only for 5 h post-dose by the treatment of 2a
and 2i with the same dose. Moreover, at 12 h post-dose, 20 mg/
CH2OH
N(CH3)2
N(CH3)2
N(CH3)2
a)
2j
a
Average values (at least two experiments).
In vitro activities of LAF-2372, DPP-4 IC50 = 0.009
DPP-9 IC50 = 0.23
b
l
M; DPP-8 IC50 = 3.82
lM; selectivity ratio (SR, DPP-8 IC50/DPP-4 IC50) = 424.
l
M;
ND, not determined.
Table 2
2a-20mg/kg
120
Pharmacokinetic properties of selected DPP-4 inhibitors
2i-20mg/kg
2j-20mg/kg
100
Compd
CLz/F (L/h/kg)
t1/2 (h)
AUC0–t (ng h/mL)
Cmax (ng/mL)
80
60
40
20
0
2a
2i
2j
11.40 3.40
50.87 26.19
33.81 20.13
0.94 0.40
3.25 1.65
0.90 0.41
68.7 20.2
266.3 59.2
120.5 72.0
39.4 18.9
164.7 91.3
146.4 101.3
CLz/F, apparent total plasma clearance; t1/2, half life; AUC, area under the plasma
concentration–time; Cmax peak plasma concentration. Oral administrations of
3.0 mg/kg to Sprague-Dawley rats (n = 6).
,
0
4
8
12
16
20
24
Time (hour)
ninefold less potent than compound 2a in DPP-4 inhibitory potency.
Introduction of methyl group in 5-position (2i or 2j) displayed
slightly reduction in DPP-4 inhibitory potency.
Figure 3. Effect of 2a, 2i or 2j on serum DPP-4 activity in cynomolgus monkey. Data
are represented as mean SEM (n = 4).
Control
A
B
2a-1mg/kg
2a-3mg/kg
2a-10mg/kg
LAF237-1mg/kg
16
14
10
8
LAF237-3mg/kg
LAF237-10mg/kg
12
10
8
*
*
*
*
6
4
2
0
**
**
6
4
2
0
0
30
60
90
120
Control 1
3
10
1
3
10
Time (min)
2a (mg/kg)
LAF237 (mg/kg)
Figure 2. Glucose responses (A) and AUC0–120 min change rate (B) during an oral glucose tolerance test (OGTT) in ICR mice following treatment with 2a. Data are represented
as mean SEM (n = 5). *P <0.05, **P <0.01 versus control.

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T. P. Cho et al. / Bioorg. Med. Chem. Lett. 20 (2010) 3565–3568
8. (a) Nauck, M. A.; Wollschlager, D.; Werner, J.; Holst, J. J.; Orskov, C.; Creutzfeldt,
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1998, 35, 117; (c) Gautier, J. F.; Fetita, S.; Sobngwi, E.; Salaün-Martin, C. Diabetes
Metab. 2005, 31, 233; (d) Miller, S. A.; St. Onge, E. L.; Taylor, J. R. Formulary
2008, 43, 122; (e) Gautier, J. F.; Choukem, S. P.; Girard, J. Diabetes Metab. 2008,
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kg 2j and 2i inhibited serum DPP-4 activity by 51% and 38%, sepa-
rately, whereas the same dose of 2a showed much lower inhibition
rate with 24%, which also suggested that introduction of methyl
group could prolong the inhibition of serum DPP-4 activity. (Fig. 3)
In summary, we have identified a series of novel azobicy-
clo[3.3.0]octane derivatives as potent DPP-4 inhibitors. Compound
2a possesses good DPP-4 activity, high selectivity over other re-
lated enzymes, moderate pharmacokinetic profiles, and excellent
in vivo efficacy in an OGTT in lean mice. Its 5a-methyl analog 2i
showed better pharmacokinetic profiles and 5b-methyl analog 2j
showed better inhibition activity of serum DPP-4. Further studies
of 5-methyl azobicyclo[3.3.0]octane derivatives are in process.
13. All new compounds were characterized by 1H NMR and MS. Where noted,
compounds for evaluation were determined to be >95% pure by analytical
reverse-phase HPLC. Data for compound 2a (HCl salt): 1H NMR (CD3OD,
400 MHz, ppm) d 4.82 (m, 1H), 4.02 (m, 2H), 3.62–3.25 (m, 7H), 2.76 (s, 6H),
2.51–1.49 (m, 10H). MS (ESI) m/z: 334.5 [M+1]+. Data for compound 2i: 1H
NMR (DMSO-d6, 400 MHz, ppm) d 4.82 (m, 1H), 3.97 (s, 2H), 3.79 (m, 1H), 3.49
(m, 2H), 3.21 (m, 4H), 2.75 (s, 6H), 2.62 (m, 2H), 2.19 (m, 2H), 2.06 (m, 2H), 1.92
(m, 2H), 1.61 (m, 2H), 1.20 (s, 3H). MS (ESI) m/z: 348.2 [M+1]+. Data for
compound 2j: 1H NMR (DMSO-d6, 400 MHz, ppm) d 4.82 (m, 1H), 4.07 (s, 2H),
3.80 (m, 1H), 3.67 (m, 1H), 3.50 (m, 1H), 3.21 (m, 4H), 2.76 (s, 6H), 2.63 (m, 2H),
2.23–2.18 (m, 2H), 2.08–2.00 (m, 2H), 1.96–1.92 (m, 2H), 1.68–1.63 (m, 2H),
1.24 (s, 3H). MS (ESI) m/z: 348.2 [M+1]+.
Acknowledgments
The authors would like to thank Mr. Sun Piao Yang for his vision
and support in new drug discovery in China, and Analytical Chemis-
try Group of Shanghai Hengrui Pharmaceuticals Co., Ltd for NMR and
mass spectroscopy services.
14. Villhauer, E. B.; Brinkman, J. A.; Naderi, G. B.; Dunning, B. E.; Mangold, B. L.;
Mone, M. D.; Russell, M. E.; Weldon, S. C.; Hughes, T. E. J. Med. Chem. 2002, 45,
2362.
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