Potent, selective, and orally active adenosine A2A receptor antagonists: Arylpiperazine derivatives of pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidines

Article abstract of DOI:10.1016/j.bmcl.2006.11.083

Antagonism of the adenosine A_2A receptor offers great promise in the treatment of Parkinson's disease. Employing the known pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine A_2A antagonist SCH 58261 as a starting point, we identified the potent and selective (vs. A1) antagonist 11 h, orally active in the rat haloperidol-induced catalepsy model. We further optimized this lead to the methoxyethoxyethyl ether 12a (SCH 420814), which shows broad selectivity, good pharmacokinetic properties, and excellent in vivo activity.

Full text of DOI:10.1016/j.bmcl.2006.11.083

Bioorganic & Medicinal Chemistry Letters 17 (2007) 1376–1380
Potent, selective, and orally active adenosine A2A receptor
antagonists: Arylpiperazine derivatives of pyrazolo[4,3-e]-
1,2,4-triazolo[1,5-c]pyrimidines
Bernard R. Neustadt,a,* Jinsong Hao,a Neil Lindo,a William J. Greenlee,a
Andrew W. Stamford,a Deen Tulshian,a Ennio Ongini,b John Hunter,a
Angela Monopoli,b Rosalia Bertorelli,b Carolyn Foster,a Leyla Arik,a
Jean Lachowicz,a Kwokei Nga and Kung-I Fenga
aDepartments of Chemical Research, CNS Pharmacology, Drug Metabolism and Pharmacokinetics,
Schering-Plough Research Institute, 2015 Galloping Hill Road, Kenilworth, NJ 07033, USA
bSchering-Plough Research Institute, San Raffaele Science Park, 20132 Milan, Italy
Received 11 October 2006; revised 20 November 2006; accepted 30 November 2006
Available online 3 December 2006
Abstract—Antagonism of the adenosine A2A receptor offers great promise in the treatment of Parkinson’s disease. Employing the
known pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine A2A antagonist SCH 58261 as a starting point, we identified the potent and
selective (vs. A1) antagonist 11 h, orally active in the rat haloperidol-induced catalepsy model. We further optimized this lead to
the methoxyethoxyethyl ether 12a (SCH 420814), which shows broad selectivity, good pharmacokinetic properties, and excellent
in vivo activity.
Ó 2006 Elsevier Ltd. All rights reserved.
Parkinson’s disease (PD) is a very serious neurological
disorder, and current methods of treatment fail to
achieve long-term control. Since adenosine A2A receptor
antagonists have been shown to restore the deficits
caused by degeneration of the striatonigral dopamine
system, which is compromised by the loss of striatal neu-
rons in this disease, A2A antagonism affords a possible
treatment for PD.1 The A2A antagonist KW-6002 (istra-
defylline) was shown to be effective in animal models of
PD, and recent clinical studies demonstrated efficacy in
alleviation of symptoms of the disease.2
good potency and selectivity.5 Beginning in 1994,
reports described a series of pyrazolo[4,3-e]-1,2,4-triaz-
olo[1,5-c]pyrimidines, some with good selectivity.6 The
parent compound of this tricyclic series is SCH
58261,6c
a
high-affinity A2A receptor antagonist
(Ki = 2 nM), which shows potent in vivo activity in ani-
mal models of PD. However, SCH 58261 is only moder-
ately selective for A2A receptors over A1 receptors,
possesses very poor solubility, and fails to show activity
upon oral administration. We report the discovery of
high-affinity A2A receptor antagonists with improved
selectivity and potent oral activity through modification
of the phenethyl side chain of SCH 58261.7
Adenosine A2A receptor antagonists of several structural
types have been described. The earliest described are
xanthines. Subsequently, non-xanthines CGS 159433
and CP 66,7134 were reported. These are non-selective
for the A2A receptor relative to the A1 receptor. KW-
6002 is a xanthine with moderate receptor selectivity.2
Several other bicyclic systems have been described with
For preparation of compounds 1 (Scheme 1) we initially
adopted the previously reported method,6b targeting key
intermediate 3 that could be alkylated at the 7-position.
However, this route behaved poorly in our hands, with 2
producing primarily 4, rather than the required 3. We
developed an alternative synthesis of 3 (Scheme 2) that
begins with commercially available 5.8 Dehydrative
cyclization of 7 with N,O-bis(trimethylsilyl)acetamide
(BSA) at reflux proceeded with concomitant Dimroth
rearrangement9 to afford the desired product 3.
Keywords: Adenosine A2A; Parkinson’s; Catalepsy; Pyrazolo[4,3-e]-
1,2,4-triazolo[1,5-c]pyrimidine.
*
Corresponding author. Tel.: +1 908 740 3495; fax: +1 908 740
7152; e-mail: bernard.neustadt@spcorp.com
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.11.083

B. R. Neustadt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1376–1380
1377
NH2
N
NH2
OMe
OMe
NH2
N
N
N
N
O
O
N
N
Me
N
N
N
N
O
N
N
EtN
N
N
O
N
N
Et
Cl
Cl
SCH 58261
CGS 5943
KW-6002
CP 66,713
SCH 58261 prepared by alkylation of 3 from the newly
developed route was identical by 1H and 13C NMR spec-
troscopy to an authentic sample prepared by the pub-
lished route,6b confirming that the required Dimroth
rearrangement had occurred. In addition, the structure
of SCH 58261 was confirmed independently by X-ray
crystallography,10 rigorously establishing the structural
assignments.
A variety of alkylated products were prepared from 3.
We found that arylpiperazine derivatives of type 11 were
of particular interest. These were produced by alkylation
of 3 (Scheme 3) with the bis-tosylate derived from ethyl-
ene glycol, which afforded the desired tosylate 10 as the
major product, along with minor amounts of the N8
regioisomer. Amination of 10 with N-arylpiperazines
and related amines proved very efficient and allowed
NH2
N
N
O
N
N
1
R
N
N
NH2
N
NH2
N
NH2
N
HCOOH
reflux
N
N
N
N
N
N
O
O
O
N
N
N
N
H N
t-Bu
N
3
2
N
N
N
4
t-Bu
Scheme 1. Desired targets and a critical step in the previously reported synthetic route to pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidines.
O
NH2
NH2
N
O
NH2
N
NH2
N
dehydrative
H2NHN
N2H4
rearrangement
N
N
N
N
O
O
N
N
N
O
O
BSA
O
N
H
6
N
H
Cl
Cl
Cl
5
HN
N
H
N
H
HN
CHO
CHO
N
7
3
N
NH2
N
NH2
N
R-X
N
N
N
O
+
O
N
N
N
N
R
N
9
N
8
N
R
Scheme 2. Newly developed synthetic route to pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidines.
NH2
N
NH2
N
NH2
N
OTs
N
NH
Ar
N
N
N
N
N
N
O
O
O
N
TsO
N
N
DMF, 80o
NaH, DMF
HN
N
N
10
TsO
N
N
N
N
11
Ar
N
3
Scheme 3. Convergent synthetic route from intermediate 10.

1378
B. R. Neustadt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1376–1380
the separation of N7- and N8-alkylated materials to be
carried out only once, at the stage of precursor 10. Req-
uisite N-arylpiperazines were typically prepared by
Buchwald–Hartwig amination11 of aryl bromides with
piperazine.
the p-hydroxyphenyl derivative 11b, which was orally
inactive at the 1.0 mg/kg dose. In an effort to identify
analogs with improved duration of activity, a number
of phenyl-substituted analogs 11c–11k that offered the
potential to block metabolism were profiled in the cata-
lepsy assay. Of these, the 2,4-difluorophenyl derivative
11h (SCH 412348) and the 2,4,6-trifluorophenyl deriva-
tive 11k displayed potent anti-cataleptic activity at a
dose of 1.0 mg/kg, and the activity was sustained at
the 4 h time point.
The majority of arylpiperazine derivatives that were pre-
pared afforded high A2A receptor binding affinity, with
high selectivity over A1 receptors12 (Table 1). Affinity
was assessed using radioligand competition assays as
previously described;12 SEM values were equal to or be-
low 15% of derived Ki values. Among these, the first to
demonstrate significant oral activity in the rat haloperi-
dol-induced catalepsy assay13 was the unsubstituted
phenylpiperazine 11a. Potent oral activity was observed
at 1.0 mg/kg at 1 h post-dose, but not at 4 h. This is con-
sistent with the observed metabolic conversion of 11a to
While SCH 412348 and 11k displayed potent oral activ-
ity, their solubility remained poor, and subsequent
efforts were directed toward improving solubility by fur-
ther modifying the phenylpiperazine substitution.
Accordingly, analogs incorporating ether-linked substit-
uents were prepared, exemplified by 12a–12g (Table 2).
Table 1. Activity of arylpiperazine compounds
NH2
N
N
N
N
11
Ar
N
N
O
N
N
a
Compound
Ar
A2A Kia, nM
A1/A2A
Rat catalepsy,
% inhibition at 1 mg/kgb
1 h
4 h
11a
11b
11c
11d
11e
11f
11g
1.6
1.3
1.4
2.1
1.0
0.5
0.6
343
186
848
571
101
1348
894
52
0
0
HO
EtO
NC
F
27
0
8
11
20
Cl
F
25
80
F
11h
11i
0.6
1.4
0.6
1.1
>1600
169
75
45
65
65
F
F
F
F
F
F
F
F
11j
1498
319
40
75
11k
F
a Average of duplicate determinations, human receptors.
b Average for n = 3. Maximum reduction attainable is 60–80%.

B. R. Neustadt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1376–1380
Table 2. 2-(Methoxyethoxy)phenyl and related substitution
1379
NH2
N
R'
N
N
N
12
R
N
N
O
N
N
a
Compound
R
R0
A2A Kia, nM
A1/A2A
Rat catalepsy,
% inhibition at 1 mg/kgb
1 h
77
4 h
70
MeO
MeO
HO
O
O
12a
12b
12c
12dc
12e
12fc
12g
12h
H
F
1.1
0.4
0.6
1.6
0.8
2.2
0.6
0.6
1340
1736
1525
539
60
15
55
15
30
8
70
25
20
10
5
O
H
H
H
H
H
H
MeO
Me
O
Me
O
O
427
Me
OS
O
523
MeO
O
1158
930
2
O
O
59
30
a See Table 1.
b See Table 1.
c Racemic compound.
In several cases these exhibited sub-nanomolar A2A
receptor binding affinity and retained excellent selectivi-
ty over A1 receptors. Of particular interest were the
methoxyethoxy derivatives 12a and 12b, which dis-
played robust oral anti-cataleptic activity that was main-
tained at 4 h, and 12a (SCH 420814) was subjected to
more extensive profiling.
The solubility of SCH 420814 as the crystalline free base
in water was 0.2 lM at native pH (5.1),17 and 2 mM in
0.01 N HCl. Pharmacokinetic properties of SCH
420814 in the rat are shown in Table 3. In this species
the compound demonstrated a relatively short half-life
(t1/2), moderate clearance (Cl), a high steady-state
100
80
SCH 420814 exhibits high affinity for both human and
rat A2A receptors, with Ki values of 1.1 and 2.5 nM,
respectively. In addition, the compound is more than
1000-fold selective for human A2A receptors over A1,14
15
A2B
,
and A3 receptors,14 with Ki values at human
60
A1, A2B, and A3 receptors of >1000 nM, >1700 nM,
and >1000 nM, respectively. In cell-based assays, SCH
420814 blocked adenylate cyclase activity stimulated
by the A2A agonist CGS 21680 with Kb values of
0.7 nM (rat) and 1.3 nM (human), confirming that it is
an antagonist of A2A receptors. The compound did
not show significant binding against a panel of 59 unre-
lated receptors, enzymes, and ion channels.16 In vivo,
SCH 420814 dose-dependently reversed haloperidol-in-
duced catalepsy in the rat with a MED of 0.3 mg/kg
1 h after oral administration (Fig. 1).
**
40
**
20
0
Veh
0.03
0.1
0.3
1 mg/kg, po
SCH420814
Figure 1. Dose–response of oral SCH 420814 in rat haloperidol-
induced catalepsy. **P < 0.01 versus vehicle, Dunnett’s t-test.

1380
B. R. Neustadt et al. / Bioorg. Med. Chem. Lett. 17 (2007) 1376–1380
Table 3. Rat pharmacokinetic properties of SCH 420814 HCl salta
po dose
(mg/kg)
iv dose
(mg/kg)
Cmax po
(nM)
Tmax, po
(h)
AUC po
(nM h)
t1/2 iv
(h)
Cl, iv
(mL minꢀ1 kgꢀ1
Vss, iv
(L kgꢀ1
)
Fpo
(%)
)
3
1
762
0.25
1560
2.1
37
2.6
57
a Mean values, n = 3 per route.
Pratt, R. M.; Revell, D.; Upton, R.; Dourish, C. T.
Neurology 2003, 61 (Suppl. 6), S101.
volume of distribution (Vss), and a brain-to-plasma ratio
of 1. When administered to rats as the hydrochloride
salt, the compound was well absorbed, with a measured
oral bioavailability of 57%. In several species, a major
metabolite was the O-desmethyl compound.
6. (a) Baraldi, P. G.; Manfredini, S.; Simoni, D.; Zapaterrra,
L.; Zocchi, C.; Dionisotti, S.; Ongini, E. Bioorg. Med.
Chem. Lett. 1994, 4, 2539; (b) Baraldi, P. G.; Cacciari, B.;
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Drug Dev. Res. 1997, 42, 63; (d) Baraldi, P. G.; Cacciari,
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7. This work was reported in part at the American Chemical
Society 231st National Meeting (March 2006), paper
MEDI 204.
In conclusion, novel, orally active phenylpiperazine
adenosine A2A receptor antagonists derived from SCH
58261 were identified, exemplified by SCH 412348. Opti-
mization of the phenylpiperazine substitution resulted in
identification of SCH 420814, which displays potent oral
anti-cataleptic activity and favorable pharmacokinetic
properties in rats. Further pharmacological character-
ization of SCH 412348 and SCH 420814 will be reported
in due course.
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