Synthesis and biological evaluation of homopiperazine derivatives with β-aminoacyl group as dipeptidyl peptidase IV inhibitors

Article abstract of DOI:10.1016/j.bmcl.2008.10.076

Compounds with homopiperazine skeleton are designed to find a potent DPP-IV inhibitor without inhibiting CYP. Thus a series of β-aminoacyl-containing homopiperazine derivatives was synthesized and evaluated. Compounds with acid moiety were found to be potent inhibitors of DPP-IV without inhibiting CYP 3A4. More specifically, compound 7m showed nanomolar activity with no inhibition towards five subtypes of CYPs, was considered as a prototype for further derivatization. Based on its X-ray co-crystal structure with human DPP-IV, we identified compounds 7s and 7t which showed good in vitro activity, no CYP inhibition, and good selectivity.

Full text of DOI:10.1016/j.bmcl.2008.10.076

Bioorganic & Medicinal Chemistry Letters 18 (2008) 6525–6529
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of homopiperazine derivatives
with b-aminoacyl group as dipeptidyl peptidase IV inhibitors
b
a
a
a
a
Jin Hee Ahn a, , Woul Seong Park , Mi Ae Jun , Mi Sik Shin , Seung Kyu Kang , Ki Young Kim ,
*
Sang Dal Rhee a, Myung Ae Bae a, Kwang Rok Kim a, Sung Gyu Kim c, Sun Young Kim c, Sang Kwon Sohn c,
Nam Sook Kang a, Jie Oh Lee d, Duck Hyung Lee b, Hyae Gyeong Cheon a, Sung Soo Kim a,
*
a Drug Discovery Division, Korea Research Institute of Chemical Technology, Yuseong-Gu, Daejeon 305-600, Republic of Korea
b Department of Chemistry, Sogang University, Seoul 121-742, Republic of Korea
c Namyang Research Center, Yungjin Pharm. Co., Ltd, Musong-dong, Kyungki-do, Republic of Korea
d Korea Advanced Institute of Science and Technology, Yuseong-gu, Daejeon 305-701, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Compounds with homopiperazine skeleton are designed to find a potent DPP-IV inhibitor without inhib-
iting CYP. Thus a series of b-aminoacyl-containing homopiperazine derivatives was synthesized and eval-
uated. Compounds with acid moiety were found to be potent inhibitors of DPP-IV without inhibiting CYP
3A4. More specifically, compound 7m showed nanomolar activity with no inhibition towards five sub-
types of CYPs, was considered as a prototype for further derivatization. Based on its X-ray co-crystal
structure with human DPP-IV, we identified compounds 7s and 7t which showed good in vitro activity,
no CYP inhibition, and good selectivity.
Received 3 August 2008
Revised 1 October 2008
Accepted 10 October 2008
Available online 21 October 2008
Keywords:
Dipeptidyl peptidase IV
Diabetes
Ó 2008 Elsevier Ltd. All rights reserved.
Inhibitor
Homopiperazine
A non-insulin dependent diabetes mellitus (NIDDM) is charac-
terized by chronic hyperglycemia, and belongs to a group of meta-
bolic disorders with multiple etiologies. It is very common and
may result from insulin resistance, inadequate secretion of insulin,
hepatic glucose overproduction or glucose intolerance.1
2006.8 Also, Merck has developed 1,4-diazepine-2-one derivative
(A) as a potential back-up candidate.9
F
F
H3PO4
NH2
CF3
O
F
F
O
NH2
A
O
Glucagon like peptide-1 (GLP-1)2 is released from L cells of the
small intestine in response to the digestion of food, and plays an
important role in secretion of insulin. Increased activity of GLP-1
will lead to sustained insulin secretion, which regulate an elevated
glucose level. It also retards gastric emptying, induction of satiety
and stimulation, regeneration and differentiation of islet b-cells.3 A
dipeptidyl peptidase IV (DPP-IV), a serine protease present in many
tissues and body fluids, exist either with membrane bound or sol-
uble enzyme. It degrades GLP-1 (GLP-1 [7-36] amide) into inactive
GLP [9-36] amide4,5 at N-terminus position. Inhibition of DPP-IV
increases the concentration of GLP-1, as a result increases insulin
secretion,6 which can ameliorate hyperglycemia in type 2 diabetes.
In recent past, many reports on use of small molecules as inhib-
itors of DPP-IV are available in the literauture.7 Merck described a
series of structurally novel b-amino amide derivatives, and among
them MK-0431 (sitagliptin) was launched into the market in
N
N
N
N
NH
F
N
F
CF3
Januvia (MK-0431)
In our earlier report, we have described the synthesis and bio-
logical evaluation of a series of b-aminoacyl group containing cyclic
hydrazine derivatives (Fig. 1).10
F
F
F
F
F
F
NH2
O
O
NH2
O
O
N
N
N
N
n
n = 1,2,3
I
* Corresponding authors. Tel.: +82 42 860 7076; fax: +82 42 860 7160.
E-mail address: jhahn@krict.re.kr (J.H. Ahn).
Figure 1. b-Aminoacyl-containing cyclic hydrazine derivatives.
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.10.076

6526
J. H. Ahn et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6525–6529
Table 1
In vitro activity against CYP.
we have further evaluated toxicity related test with this compound
(I), and found to inhibit CYP 3A4 subtype as shown in Table 1.
Cytochrome P450 (CYP) enzymes play a major role in metabo-
lizing drug molecules. Many lead candidate molecules in pharma-
ceutical development fail due to inhibition of one or more isozymic
forms of CYP enzymes. CYP 3A4, is one of the most important cyto-
chrome P450s present in human liver,11 has ability to metabolize
>50% of administered therapeutic agents. It accounts for the large
number of documented drug–drug interactions associated with
CYP 3A4 inhibition.12 To overcome CYP inhibition (particularly
CYP 3A4), we have synthesized over 100 compounds based on cyc-
lic hydrazine, however all the compounds found to show CYP 3A4
inhibition.
CYP subtype
Activity at 10 lM (%)
1A2
2C9
2C19
2D6
3A4
91.5
71.5
90.3
51.0
5.4
F
F
HCl
NH2
F
O
N
In our recent findings, we identified homopiperazine derivative
NH
5, which showed moderate DPP-IV inhibition (IC50 1.44
CYP 3A4 inhibition (93.3% at 10 M) (Fig. 2).
lM) and no
5
l
Figure 2. Homopiperazine derivative 5.
These observations prompted us to turn our attention from cyc-
lic hydrazine to homopiperazine skeleton.
F
F
F
Boc
F
NH
O
a
HCl
NH2
F
Cbz
N
HN
F
NH2
O
R
O
F
F
N
F
Boc
N Cbz
N
N
NH
O
F
N
1
3
N
R
F
OH
F
2
b
F
F
F
We now wish to report here the synthesis and biological evaluation
of a series of b-aminoacyl-containing homopiperazine derivatives
as DPP-IV inhibitors. The sequence of reaction steps involved in syn-
thesis is shown in Schemes 1 and 2. Cbz-protected homopiperazine
(1) was reacted with b-amino acid (2) in presence of EDCI to provide
the coupled product (3). It is on reduction with 10% palladium on
carbon in hydrogen atmosphere to provide a key intermediate 4,
followed by boc-deprotection using 4 M HCl to result corresponding
HCl salt (5). The compound 4 is also derivatized with diverse elec-
trophiles to obtain compounds 6, followed by deprotection to give
compound 7.
Similarly, Boc-protected homopiperazine (8) was reacted with
heteroaryl chloride to give the coupled compound 10, followed
by boc-deprotection to give product 11. The compounds 11 were
further coupled with b-amino acid to give 12, and on deprotection
with HCl to afford compounds 13.
HCl
F
F
Boc
NH2
O
NH
O
c
N
N
NH
NH
F
4
d
5
F
F
HCl
NH2
F
F
Boc
O
NH
O
c
N
N
N
R
N R
F
F
7
6
Scheme 1. Reagents and conditions: (a) (R)-3-BocNH-4-(2,4,5-trifluorophe-
nyl)butanoic acid (2), EDCI, Et3N, CH2Cl2, room temperature, 12 h; (b) 10% Pd/C,
H2 balloon, MeOH, room temperature, 12 h; (c) 4 M HCl, ethyl acetate, room
temperature, 12 h; (d) electrophile, CH2Cl2, TEA, room temperature.
The b-aminoacyl containing homopiperazine derivatives (5, 7
and 13) were evaluated in vitro for DPP-IV inhibition, and the re-
sults were summarized in Table 2. MK-0431 and compound I were
used as reference.
Among the cyclic hydrazine derivatives, compound I showed
good potency, selectivity and in vivo efficacy. In course of time,
Y
Y
X
X
a
b
Boc
NH
Boc
N
N
N
N
HN
N
N
Cl
X
Y
8
10
11
N
9
c
X, Y = C or N
F
F
F
HCl
NH2
F
Boc
F
NH
O
O
d
X
N
Y
X
N
Y
N
N
N
N
F
13
12
Scheme 2. Reagents and conditions: (a) 9, ethylene glycol, 130 oC, 2 h; (b) 4 M HCl, ethyl acetate, room temperature, 12 h; (c) (R)-3-BocNH-4-(2,4,5-trifluorophenyl)butanoic
acid (2), EDCI, Et3N, CH2Cl2, room temperature, 12 h; (d) 4 M HCl, ethyl acetate, room temperature, 12 h.

J. H. Ahn et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6525–6529
6527
The activity data of basic verses substituted homopiperazine
derivatives is compared. N-Aroyl substituted homopiperazines
such as 7a, 7c and 7d showed nearly 3-fold more potent than that
of the basic homopiperazine 5 (IC50 1440 nM), however activity of
acetyl derivative 7b is diminished. Carbamate 7e (IC50 490 nM),
heteroaryl 13a and 13b exhibited equal potency with that of aroyl
derivatives. Urea derivatives such as 7f and 7g demonstrated
weaker activities. N-Aralkyl substituted compound 7k (IC50
387 nM) showed better in vitro inhibitory activity than that of al-
kyl 7l (IC50 6400 nM). Tosyl derivative 7h (IC50 350 nM) found to
show highest activity, and is a potential candidate for further
derivatization.
Based on the above data, we chose three representative com-
pounds (7a, 7h and 13b) for CYP re-evaluation. Unfortunately, all
the compounds found to show CYP 3A4 inhibition (Table 3).
Our recent experience13 with acid moiety in a molecule pro-
moted good DPP-IV inhibitory activity with no CYP 3A4 inhibition.
Thus, we have synthesized number of compounds having acid or
acid equivalent group, and evaluated. Fortunately, acid derivatives
(7m–7o) showed no CYP 3A4 inhibition at 10 lM concentration as
Table 3
Table 2
In vitro CYP 3A4 activity of homopiperazine derivatives with b-aminoacyl group.
In vitro DPP-IV inhibitory activity of homopiperazine derivatives with b-aminoacyl
group.
F
HCl
F
F
HCl
F
NH2
O
NH2
O
N
N
R
F
N
N
R
F
Compound
R
CYP 3A4 activity at 10
2.1
lM (%)
a
Compound
R
IC50 (nM)
O
5
H
1440
510
7a
O
7a
7b
O
S
7h
1.2
O
O
O
O
3400
N
13b
16.9
89.0
N
MK-0431
O
O
7c
7d
7e
490
410
490
Table 4
In vitro DPP-IV and CYP 3A4 activity of homopiperazine derivatives with b-aminoacyl
N
O
group.
O
F
HCl
F
NH2
O
N
O
O
N
R
F
7f
7g
7h
7i
1392
1500
350
N
H
a
Entry
R
IC50 (nM)
223
CYP 3A4 activity at 10
74.6
lM (%)
O
N
N
7m
O
S
O
OH
O
O
S
O
OH
O
O
S
O
7n
7o
239
788
83.3
83.4
3100
405
O
O
S
O
7j
NO2
O
O
OH
N
H
7k
7l
387
6400
597
7p
7q
180
5.7
NH2
N
13a
O
N
N
O
N
288
87
36.6
86.1
13b
423
SO2NH2
MK-0431
I
87
70
MK-0431
a
a
IC50 values were determined from direct regression curve analysis.
IC50 values were determined from direct regression curve analysis.

6528
J. H. Ahn et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6525–6529
Table 6
shown in Table 4. Amide and sulfonamide derivatives (7p and 7q)
still inhibited CYP 3A4.
Compound 7m being not shown CYP 3A4 inhibition, was further
screened with other CYP subtypes and found to show no inhibition
(Table 5).
In vitro DPP-IV inhibitory activity and CYP 3A4 activity of homopiperazine derivatives
with b-aminoacyl group.
F
HCl
F
NH2
O
Therefore, we determined the crystal structure of human DPP-
IV complexed with 7m. As can be seen by viewing the binding site
region in the complex (Fig. 3), the b-aminoacyl group has a similar
conformation as that of MK-0431.8 Also, the carbonyl moiety in
amino acyl group showed water-bridged hydrogen bonding with
Y547. Whereas, carboxy or benzoyl part showed no significant
interaction, therefore we further derivatized.
We have derivatized homopiperazine skeleton with acid moi-
ety, and evaluated the inhibitory activities. The results are summa-
rized in Table 6. Substituents in meta position (7s and 7t) exhibited
better activity than para (7m and 7r) with IC50 value of 50–67 nM,
as well as no CYP 3A4 inhibition.
N
N
R
F
a
Entry
R
IC50 (nM)
CYP 3A4 activity at 10
74.6
lM (%)
O
O
7m
223
OH
O
7r
7s
7t
7u
150
50
55.5
66.1
74.0
OH
OH
OH
Compounds 7s and 7t were further investigated for their
N
selectivity towards
a variety of DPP-IV related peptidases,
O
O
and found to show good selectivity against DPP-2 and DPP-
8 (Table 7).
O
O
In conclusion, this investigation has led to the synthesis and
biological evaluation of
a series of b-aminoacyl-containing
homopiperazine derivatives. Several homopiperazine derivatives
O
67
Table 5
N
O
In vitro activity against five CYP subtypes.
F
O
HCl
F
O
260
87
100
NH2
O
OH
O
N
N
F
MK-0431
86.1
a
IC50 values were determined from direct regression curve analysis.
OH
O
Table 7
Selectivity of compounds 7s and 7t towards DPP-IV related enzymes.
CYP subtype
Activity at 10 lM (%)
1A2
2C9
2C19
2D6
3A4
97.3
103.2
96.0
89.1
74.6
Compound
DPP-IV IC50 a(nM)
DPP-2 IC50 a(nM)
DPP-8 IC50 a(nM)
7s
7t
50
67
46,770
99,434
4265
18,197
a
IC50 values were determined from direct regression curve analysis.
with acid moiety were found to be potent inhibitors of DPP-IV
with no CYP 3A4 inhibition. The compound 7m showed sub-
micromolar activity with no CYP inhibition towards five sub-
types, and is a prototype for further derivatization. Based on
the results, we identified 7s and 7t, which showed good
in vitro activity, no CYP inhibition, and good selectivity. Further
studies are underway to optimize this compound class for the
treatment of diabetes.
Acknowledgments
This research was supported by the Center for Biological
Modulators of the 21st Century Frontier R&D Program, Ministry
of Education, Science and Technology, Korea. We thank the
staff of the 4A and 6B beam lines of Pohang Accelerator
Laboratory.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at doi:10.1016/j.bmcl.2008.10.076.
Figure 3. X-ray co-crystal structure of 7m with DPP-IV (pdb code 3EIO).

J. H. Ahn et al. / Bioorg. Med. Chem. Lett. 18 (2008) 6525–6529
6529
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