Xanthine mimetics as potent dipeptidyl peptidase IV inhibitors

Article abstract of DOI:10.1016/j.bmcl.2006.09.024

A series of xanthine mimetics containing 5,5 and 5,6 heterocycle fused imidazoles were synthesized as dipeptidyl peptidase IV inhibitors. Compound 7 is potent (h-DPPIV K_i = 2 nM) and exhibits excellent selectivity and no species specificity aga

Full text of DOI:10.1016/j.bmcl.2006.09.024

Bioorganic & Medicinal Chemistry Letters 16 (2006) 6226–6230
Xanthine mimetics as potent dipeptidyl peptidase IV inhibitors
Ravi Kurukulasuriya,* Jeffrey J. Rohde, Bruce G. Szczepankiewicz, Fatima Basha,
Chunqui Lai, Hwan-Soo Jae, Martin Winn, Kent D. Stewart, Kenton L. Longenecker,
Thomas W. Lubben, Stephen J. Ballaron, Hing L. Sham and Thomas W. von Geldern
Metabolic Disease Research, Global Pharmaceutical Research and Development, Abbott Laboratories, 100 Abbott Park Road,
Abbott Park, IL 60064-6098, USA
Received 20 July 2006; revised 7 September 2006; accepted 8 September 2006
Available online 28 September 2006
Abstract—A series of xanthine mimetics containing 5,5 and 5,6 heterocycle fused imidazoles were synthesized as dipeptidyl peptidase
IV inhibitors. Compound 7 is potent (h-DPPIV Ki = 2 nM) and exhibits excellent selectivity and no species specificity against rat and
human enzymes. The X-ray structure confirms that the binding mode of 7 to rat DPPIV is similar to the parent xanthines.
Ó 2006 Elsevier Ltd. All rights reserved.
The incretin hormones glucagon like peptide-1 (GLP-1)
and glucose dependent insulinotropic polypeptide (GIP)
play an important role in glucose homeostasis with
effects on the pancreas, gastrointestinal tract, muscle tis-
sue, and brain.1,2 GLP-1 enhances glucose-stimulated
insulin secretion from the b-cells of the pancreas,3 pro-
motes insulin biosynthesis,4 and inhibits postprandial
glucagon secretion.5 Administration of GLP-1 reduces
the rate of gastric emptying,6 suppresses appetite and,
importantly, promotes b-cell mass.7 Exenatide, a potent
GLP-1 mimetic administered subcutaneously, is current-
ly approved for treatment of type 2 diabetes.8 Under
physiological conditions, the incretin effect is brief
because of the short half-life of the incretin hormones
due to the action of dipeptidyl peptidase IV (DPPIV)
enzyme. DPPIV is a serine protease that cleaves a dipep-
tide from the N-terminus of the active form of GLP-1,
GIP, neuropeptides, and chemokines, and renders them
inactive. This discovery has led to the development of
DPPIV inhibitors to increase the half-life of circulating
incretin hormones and normalize glucose homeostasis.9
DPPIV has now become a validated target with several
small molecule inhibitors in late stage clinical trials for
the treatment of type 2 diabetes.10
DPPIV inhibitors can be broadly categorized into pep-
tidic and non-peptidic series, with a majority of the
inhibitors that have successfully advanced to late stage
trials being of peptidyl origin.10 DPPIV inhibitors relat-
ed to xanthines include purine, uracil, imidazole, pyrim-
idine, and pyridine analogs.11 Each of these subclasses
serves as a core to direct key peripheral groups in a
direction important to maintain several points of con-
tact needed for potent DPPIV inhibition. Takeda San
Diego (formerly Syrrx) recently disclosed SYR-322
which bears a central uracil moiety.12 SYR-322 was
reported to be in phase III clinical trials making it likely
that compounds in this class will play an important role
in DPPIV inhibition and type 2 diabetes therapy
(Fig. 1).
A high throughput screen of the Abbott library of com-
pounds for potential DPPIV inhibitors revealed several
xanthine analogs exhibiting low micromolar potency
O
CN
N
N
NH2
O
N
Syr-322
DPPIV IC50 = 4 nM
Keywords: Dipeptidyl peptidase IV (DPPIV); Xanthines; SAR; X-ray
structure; Type 2 diabetes.
*
Corresponding author. Tel.: +1 847 935 5699; e-mail:
ravi.kurukulasuriya@abbott.com
Figure 1. Lead DPPIV inhibitor in late stage clinical trials.
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.09.024

R. Kurukulasuriya et al. / Bioorg. Med. Chem. Lett. 16 (2006) 6226–6230
6227
for the enzyme. Preliminary SAR on the xanthine
revealed that N-7 alkylation of the xanthine moiety
was needed, preferably with a (2-cyano) benzyl group.
Substitution at the 8-position was also required with 3-
amino piperidine being optimal. N-1 substitution was
tolerated but not required. A protein X-ray crystallo-
graphic study of the most potent screening lead 6,13
bound within the active site of rat DPPIV, was in accor-
dance with these early SAR observations.14 Groups at
the 7- and 8-positions of the xanthine ring system made
specific protein contacts. In addition, a hydrogen bond
between the 6-carbonyl oxygen and the backbone N–H
of Tyr 632 of the protein was observed. Positions 1 and
2 were in close proximity to the protein but had no
direct interaction, while N-3 and N-9 appeared to be
in a large solvent pocket (Fig. 2).
ation at N-7 and deprotection provided DPPIV inhibi-
tor 6. Related fused imidazole compounds were
synthesized by similar procedures.
The in vitro activity of the compounds was tested
against human and rat DPPIV enzyme. Data for select-
ed compounds are shown in Table 1. Xanthine 6 with an
ortho-cyano benzyl group off the imidazole (N-7) nitro-
gen was extremely potent (h-DPPIV Ki = 2 nM). The
potency is retained when the A ring of the xanthine is re-
placed with a methyl maleimide as in 7 (h-DPPIV
Ki = 2 nM) or methyl pyridazinone as in 8 (h-DPPIV
Ki = 5 nM). The ortho-cyano group can be substituted
with small electron-withdrawing groups such as a chloro
as in 9 (h-DPPV Ki = 9 nM) or trifluoromethyl as in 10
(h-DPPIV Ki = 10 nM) with a 5-fold loss in potency.
Larger groups such as a trifluoromethoxy as in 11
(h-DPPIV Ki = 22 nM) are less desirable with a 10-fold
loss in DPPIV activity compared to 7. Notably human
DPPIV inhibition is dramatically weakened when the
ortho-cyano group is moved to the meta-position as in
12 (h-DPPIV Ki = 1200 nM). Modeling of meta-substit-
uents indicated that a meta acetonitrile as in 13 would
allow the cyano group to reorient to be similar to 8.
As modeling predicted, potency was re-established with
13 (h-DPPIV Ki = 11 nM). The SAR indicates that both
the benzylic moiety and a small electron-withdrawing
group (EWG) are important for potent DPPIV inhibi-
tion and the optimum orientation for the protein–
inhibitor interaction is when the small EWG is at the
ortho-position of the aryl group. Extended groups at
the A ring are less desirable. For example, benzyl malei-
mide as in 14 (h-DPPIV Ki = 29 nM) was less potent
than methyl maleimide 7. Similarly naphthoquinone as
in 15 (h-DPPIV Ki = 25 nM) was less potent than 7.
Not all heterocyclic substitutions at the A ring were
tolerated. Substitution of the C-6 carbonyl of the uracil
moiety of 6 with a fluoro group as in 16 (h-DPPIV
Ki = 57 nM) diminished potency while a pyridine N-ox-
ide as in 17 (h-DPPIV Ki = 300 nM) was weakly active.
The location of the C-6 carbonyl group relative to the
benzyl moiety off the imidazole is also critical. For
example when the ortho-cyano benzyl group is attached
to the imidazole N-9 nitrogen distal to the carbonyl of
the pyridazinone as in 18 (h-DPPIV Ki = 260 nM) the
potency decreased sharply.
Based on the X-ray data, we embarked on a synthetic
program which targeted xanthine analogs that retain
the imidazole and 6-carbonyl moieties but possess alter-
native A-ring heterocycles. We envisioned that such a
change would still enable the core to orient the vital
N-7 benzylic and the C-8 (3-amino)-piperidine pharma-
cophores in a similar direction as the xanthine to main-
tain potent DPPIV inhibition while providing the
flexibility to modulate the pharmacokinetic properties
of the compounds. To aid the medicinal chemistry strat-
egy, a convergent procedure was developed to obtain 5,5
and 5,6 ring fused xanthine mimetics. A Cu1 mediated
cyclization utilizing the acyclic guanidine derived from
3-amino piperidine was the key step (Scheme 1).15 Alkyl-
Figure 2. Preliminary SAR on the xanthine hit.
O
O
NHBoc
+
Br
Br
O
N
NH2
N
a
N
N
NHBoc
O
N
N
O
N
H2N
NH2 Cl
4
2
3
CN
O
O
NHBoc
NH3
H
b
c, d
N
N
N
N
N
N
N
N
O
N
O
N
5
6
Scheme 1. Reagents and conditions: (a) NaOMe, MeOH, 80 °C, 64%; (b) NaH, CuI, THF, 75 °C, 77%; (c) BnBr, K2CO3, DMF, 23 °C, 50%;
(d) TFA, CH2Cl2, 23 °C, 95%.

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R. Kurukulasuriya et al. / Bioorg. Med. Chem. Lett. 16 (2006) 6226–6230
Table 1. Human DPPIV binding for selected compounds
Cl
NH2
N
R
N
Cl
NH3
L
NHBoc
H2N
+
Het
N
Het
L
N
Compound
Het
Product
h-DPPIV Ki (nM)
CN
O
O
N
Br
O
NH3
N
O
6
2
N
N
N
N
N
O
CN
O
Br
NH3
O
N
N
O
7
8
9
2
5
9
N
N
Br
N
O
CN
O
Br
Br
O
NH3
N
N
N
N
N
N
N
Cl
O
O
O
Br
Br
NH3
O
N
N
N
N
N
O
O
CF3
N
Br
Br
NH3
O
N
O
10
11
12
13
14
10
N
N
N
O
OCF3
Br
NH3
O
N
O
22
N
N
N
N
Br
O
CN
O
Br
Br
N
1200
O
NH3
N
N
N
N
N
N
CN
O
Br
Br
N
N
11
O
NH3
N
N
N
N
N
CN
O
Br
NH3
O
N
O
29
N
N
N
N
Br
O
(continued on next page)

R. Kurukulasuriya et al. / Bioorg. Med. Chem. Lett. 16 (2006) 6226–6230
6229
Table 1 (continued)
Compound
Het
Product
h-DPPIV Ki (nM)
CF3
N
O
O
O
Br
Br
NH3
15
16
17
18
25
57
N
N
O
CF3
F
NH2
MeO
N
N
N
N
Cl
Br
O
300
260
NH3
N
N
N
N
O
NH3
N
N
O
N
N
N
Br
Br
N
N
CN
The X-ray structure of 7 bound to rat DPPIV (pdb code
2I3Z) assisted in further rationalizing the SAR by
revealing critical binding features necessary for potent
DPPIV inhibition (Fig. 3). Analysis of the crystal struc-
ture revealed that the 3-amino piperidine was in an ideal
position to interact with Glu 203 and Glu 204. The
maleimide was oriented to form a p stacking interaction
with Tyr 548. The ortho-cyano benzyl group is in the
vicinity of Ser 631 (the catalytically active serine in
DPPIV). Unlike the cyano pyrrolidine DPPIV inhibitors
that have been reported in the literature14,16 there was
no direct interaction between the nitrile of 7 and the
hydroxyl group of Ser 631. However, the nitrile is being
directed to a shallow hydrophobic pocket in P114
increasing the inhibitor–protein interaction. This result
possibly explains the similar potencies observed when
the nitrile was replaced with a chloro or trifluoromethyl
group as in 9 and 10 (Table 1). The crystal structure de-
picts the benzyl group to be orthogonal to the plane of
the heterocycle. It is likely that steric and electronic
properties of the adjacent carbonyl of the maleimide
are contributing to the orthogonality of the benzyl
group as predicted by AM1 modeling. Methyl malei-
mide, 7, seems to be optimum and larger groups or
extensions may be in the way of the protein, as demon-
strated by the diminished potency of 14 and 15.
Compound 7 was unstable in rat plasma and showed a
poor pharmacokinetic (PK) profile in Sprague–Dawley
CN
CN
O
NH3
p, q
NHBoc
O
N
N
N
N
N
H
O
N
N
N
π -stacking
with Tyr 548
NH
O
19
x, y, z
CN
N
No direct
interaction
with Ser 631
O
O
NH3
N
N
N
N
Electrostatic
interaction with
Glu 203 and Glu 204
20
Scheme 2. Reagents and conditions: (p) Methylamine, ACN, 80 °C,
90%; (q) TFA, CH2Cl2, 23 °C, 95%. (x) N,N0-dimethylhydrazine,
ACN, 80 °C, 90%; (y) 6N HCl, MeOH, H2O, 80 °C, 80%; (z) TFA,
CH2Cl2, 23 °C, 95%.
Figure 3. X-ray structure of 7 bound to rat DPPIV.

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R. Kurukulasuriya et al. / Bioorg. Med. Chem. Lett. 16 (2006) 6226–6230
Table 2. Human and rat DPPIV binding
fused imidazoles. Compound 7 was found to be a potent
and selective DPPIV inhibitor, but it showed a signifi-
cant loss of activity in the presence of plasma. Com-
pound 20 showed minimal potency changes in the
presence of plasma. Hence the shift in potency of 7
was attributed to plasma instability and not protein
binding. X-ray analysis of 7 bound to DPPIV indicates
that the binding mode is similar to the parent xanthine
6. Xanthine mimetics allowed us to unravel the key
structural features important for potent DPPIV activity
while providing good selectivity and no species specific-
ity enabling them to be studied in rodent models of dia-
betes. These attributes make them excellent compounds
to study DPPIV inhibition in vivo as well as assist in
designing future DPPIV inhibitors based on this impor-
tant class of heterocycles.
Compound
h-DPPIV
(0% plasma)
Ki (nM)
r-DPPIV
(0% plasma)
Ki (nM)
r-DPPIV
(18% plasma)
Ki (nM)
6
7
2
2
3
2
7
600
ND
24
19
20
450
11
ND
17
Table 3. Inhibition of other peptidases for selected compounds
(Ki = nM)
Compound
DPP8
DPP9
POP
6
7
>30,000
>3000
>3000
>3000
>30,000
>3000
ND
>30,000
>30,000
>30,000
>30,000
ND
>3000
ND
ND
8
13
15
20
>3000
ND
References and notes
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ity. We reasoned that the low plasma stability was partly
due to the electrophilicity of the carbonyl groups which
made the maleimide susceptible to nucleophilic attack.
In an attempt to make more stable DPPIV inhibitors,
7 was reacted with either N-methyl amine to convert
the A ring to the ring opened diamide 19 or reacted with
N,N0-dimethylhydrazine to prepare the corresponding
pyridazine dione 20 (Scheme 2).
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Compounds 7, 19, and 20 were tested against human
and rat DPPIV enzyme in the presence of 2% and 18%
rat plasma to study the effects of plasma stability on
DPPIV inhibition (Table 2). Xanthine 6, maleimide 7,
and pyridazinedione 20 did not show species specificity
against human and rat DPPIV enzymes in the absence
of plasma. Xanthine 6 is stable in rat plasma with a mere
2-fold shift in potency in the presence of 18% rat plasma
(r-DPPIV Ki = 3–7 nM). Confirming the PK and plasma
instability result, a significant shift in potency was ob-
served for 7 against DPPIV in the absence of plasma
(r-DPPIV Ki = 2 nM) compared to 18% rat plasma (r-
DPPIV Ki = 600 nM). The ring opened analog 19 was
weak against h-DPPIV. The ring expanded product 20
was 5-fold less potent than 7 (h-DPPIV Ki = 11 nM).
However, the compound was stable in rat plasma with-
out a significant shift in potency in the presence of plas-
ma (r-DPPIV Ki = 17–24 nM in 18% plasma).
Human peptidases including DPP7, DPP8, DPP9, and
POP are structurally or functionally related to DPPIV
and most importantly inhibition of DPP8 and/or
DPP9 is associated with significant toxicity.17 The com-
pounds were screened and found to be inactive against
these isozymes (Table 3).
16. Engel, M.; Hoffman, T.; Wagner, L.; Wermann, M.;
Heiser, U.; Kiefersauer, R.; Huber, R.; Bode, W.;
Demuth, H. U.; Brandstetter, H. PANS 2003, 100, 5063.
17. Lankas, G.; Leiting, B.; Roy, R. S. Diabetes 2004, 53,
A2-OR.
In summary, we have discovered novel, potent, and
selective xanthine mimetics based on 5,5 and 5,6 ring