4-Amino-3(2H)-pyridazinones bearing arylpiperazinylalkyl groups and related compounds: Synthesis and antinociceptive activity

Article abstract of DOI:10.1016/S0014-827X(03)00162-9

A series of 4-amino-3(2H)-pyridazinones substituted at position 2 with arylpiperazinylalkyl groups and analogues were synthesized and their antinociceptive effect was evaluated in the mouse abdominal constriction model. Preliminary SARs studies were performed. Several of the novel compounds dosed at 100 mg/kg s.c. significantly reduced the number of writhes induced by the noxious stimulus. Compound 12e showed 100% inhibition of writhes and was able to protect all the treated animals from the effect of the chemical stimulus. Subsequent dose-response studies revealed 12e to be almost 40-fold more potent than the structurally related Emorfazone.

Full text of DOI:10.1016/S0014-827X(03)00162-9

Il Farmaco 58 (2003) 1063Á1071
/
www.elsevier.com/locate/farmac
4-Amino-3(2H)-pyridazinones bearing arylpiperazinylalkyl groups
and related compounds: synthesis and antinociceptive activity
Vittorio Dal Piaz a,*, Claudia Vergelli a, Maria Paola Giovannoni a,
Mark A. Scheideler b, Giuseppe Petrone b, Paola Zaratin b
a Dipartimento di Scienze Farmaceutiche, Universita` di Firenze, via G. Capponi 9, 50121 Firenze, Italy
b Neurobiology Research, GlaxoSmithKline Pharmaceuticals, via Zambeletti, 20021, Baranzate di Bollate, Milan, Italy
Received 3 January 2003; accepted 22 April 2003
Abstract
A series of 4-amino-3(2H)-pyridazinones substituted at position 2 with arylpiperazinylalkyl groups and analogues were
synthesized and their antinociceptive effect was evaluated in the mouse abdominal constriction model. Preliminary SARs studies
were performed. Several of the novel compounds dosed at 100 mg/kg s.c. significantly reduced the number of writhes induced by the
noxious stimulus. Compound 12e showed 100% inhibition of writhes and was able to protect all the treated animals from the effect
of the chemical stimulus. Subsequent doseÁ
/
response studies revealed 12e to be almost 40-fold more potent than the structurally
related Emorfazone.
´
# 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
Keywords: Arylpiperazinylalkylpyridazinones; Synthesis; Antinociceptive activity; Emorfazone; SARs studies
1. Introduction
antinociceptive agents remains an important challenge
for medicinal chemists.
Pain relief represents a major unmet medical need and
millions of people worldwide have to use drugs for
treatment of different levels of pain. Although non
steroidal antiinflammatory agents (NSAIDs) and
opioids have been the cornerstone of pain therapy for
a long time, both classes are limited by severe side
effects. NSAIDs, which are mainly used to treat mild to
moderate inflammatory pain, induce gastric irritation
and nephrotoxicity [1]. Opioids display an array of
adverse reactions, such as respiratory depression, seda-
tion, constipation [2]. Moreover, repeated administra-
tions of these drugs induce tolerance to the analgesic
effects and physical dependence. Neuropathic pain,
which is a particular form of pain, can be controlled
only with antidepressant and anticonvulsant drugs [3].
For all these reasons, the search for more safe novel
Among the pyridazine derivatives endowed with
antinociceptive effects, Ag 246 1 [4] and Emorfazone 2
[5] have emerged (Chart 1); the latter, which was
launched in Japan, displays an interesting profile
because its activity is not mediated by interaction with
the prostaglandins system or by affinity for opioids
receptors [6,7]. The literature reports many examples of
antinociceptive agents bearing an arylpiperazinylalkyl
moiety linked both to the 2-nitrogen of a pyridazine ring
(compounds 3 and 4) [8,9] and to a different lactamic
system (compound 5) [10].
Our studies in this field have allowed the identifica-
tion of compound 6 (ED50ꢀ14.9 mg/kg, s.c.) as a
/
potent antinociceptive agent in writhing test [9,11]. On
this basis, we decided to combine the arylpiperazinyl
moiety and the amino and vinyl functions (which are
essential structural requirements for the activity of
compound 6) in the same phenylpyridazinone backbone,
in order to verify if the contemporary presence of these
structural elements is associated with better activity.
* Corresponding author.
E-mail address: vittorio.dalpiaz@unifi.it (V.D. Piaz).
´
0014-827X/03/$ - see front matter # 2003 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
doi:10.1016/S0014-827X(03)00162-9

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V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
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Chart 1.
2. Chemistry
Mannich condensation between 2-methoxyphenylpi-
perazine and compound 18 [16], which in turn was
obtained from 13 through the usual reduction [17] and
dehydratation sequence, afforded the final 19 (Scheme
3).
The synthetic pathway affording the final 5-vinylpyr-
idazinones 12aÁi is depicted in Scheme 1. Compounds
/
8a,b were obtained by treatment of the previously
described isoxazolo[3,4-d]pyridazinones 7a,b [12,13]
with dibromoethane in standard conditions. The alkyla-
tion takes place regioselectivity at pyridazine-2-nitrogen
in agreement with a variety of literature reports [14,15].
Displacement of the bromine in 8a,b with the appro-
The chemical and physical data of all compounds are
reported in Tables 1Á4.
/
priate cycloalkylamine afforded 9aÁ
were transformed into the corresponding 5-acetyl-4-
amino derivatives 10aÁi by reductive cleavage with
hydrazine hydrate and 10% Pd/C. The final compounds
12aÁi were obtained by treatment of 10aÁi with sodium
borohydride, followed by dehydratation of the second-
ary alcohols 11aÁi with poliphosphoric acid (PPA).
Scheme 2 shows the synthesis of the analogues 16aÁ
/
i, which, in turn,
3. Experimental procedures
/
3.1. Chemistry
/
/
All melting points were determined on a Buchi
¨
1
apparatus and are uncorrected. H NMR spectra were
/
/
c
recorded with Varian Gemini 200 instruments. Chemical
shifts are reported in ppm, using the solvent as internal
standard. Extracts were dried over Na2SO4 and the
solvents were removed under reduced pressure. E.
Merck F-254 commercial plates were used for analytical
TLC to follow the course of reaction. Silica gel 60
bearing a three methylenic spacer. The introduction of
the appropriate 1-aryl-4-(3-bromopropyl)piperazine
moiety was carried out in standard conditions on
compound 13 [12]; the intermediates 14aÁ/c were trans-
formed into 16aÁc following the two steps procedure
/
above. Again any trace of oxygen-alkylated derivative
was detected in the reaction mixture.
(Merck 70Á/230 mesh) was used for column chromato-
graphy.

V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
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1065
Scheme 1. Conditions: (a) Br(CH2)2Br, K2CO3, DMF; (b) cycloalkylamine, EtOH; (c) hydrazine hydrate, EtOH; (d) NaBH4, MeOH; (e) PPA.
3.1.1. General procedure for compounds 8a,b
mixture cooled, water (12 ml) was added and the final
products were recovered by suction.
A mixture of the appropriate isoxazolopyridazinone
7a,b (0.6 mmol), dibromoethane (1.2 mmol) and potas-
sium carbonate (4 mmol) in anhydrous DMF (3 ml) was
3.1.1.1. 6-(2-Bromoethyl)-3,4-dimethylisoxazolo[3,4-
d]pyridazin-7(6H)-one (8b). 1H NMR (CDCl3): d 2.50
heated at 60 8C under stirring for 1Á/3 h. After the
Scheme 2. Conditions: (a) Br-(CH2)2-
, K2CO3, DMF; (b) NaBH4, MeOH; (c) PPA.

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V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á1071
/
Scheme 3. Conditions: (a) NaBH4, MeOH; (b) PPA; (c) HCHO,
, EtOH.
(s, 3H, 4-CH3), 2.90 (s, 3H, 3-CH3); 3.70 (t, Jꢀ
/8.1 Hz,
exception of compounds 9b and 9h for which it was
necessary extraction with CHCl3 (3ꢁ15 ml) and
evaporation of the solvent.
2H, CH2Br); 4.50 (t, Jꢀ8.1 Hz, 2H, NCH2).
/
/
3.1.2. General procedure for compounds 9aÁi
/
A mixture of the appropriate 2-bromoethyl derivative
8a,b (0.3 mmol), the appropriate cycloalkylamine (1
mmol), potassium carbonate (0.8 mmol) in anhydrous
3.1.2.1. 3-Methyl-4-phenyl-6-[(4-phenylpiperazin-1-
yl)ethyl]-isoxazolo[3,4-d]pyridazin-7(6H)-one (9a).
1H NMR (CDCl3): d 2.50 (s, 3H, CH3), 2.85Á
3.50 (m,
8H, piperazine); 3.40 (d, Jꢀ7.9 Hz, 2H, NCH2CH2Ã
piperazine); 4.45 (d, Jꢀ8.0 Hz, 2H, NCH2CH2Ãpiper-
azine); 6.80Á7.00 (m, 5H, Ar); 7.54 (s, 5H, Ar).
/
DMF (2 ml) was heated at 50Á
/
110 8C for 1Á/5 h. After
/
/
the mixture cooled and cold water (10 ml) was added;
the crude precipitate was recovered by suction with the
/
/
/
Table 1
Physical and chemical data of compounds 8a,b, 9aÁ
/i, 21 and 22
a
Comp.
X
R
Yield (%)
M.p. (8C)
Formula b
8a
8b
9a
9b
9c
9d
9e
9f
Ph
Me
Ph
Ph
Ph
Ph
Me
Ph
Ph
Ph
Ph
82
60
72
93
62
97
69
64
62
78
69
157Á
127Á
158Á
Oil
76Á
145Á
117Á
102Á
90Á
Oil
157Á
/
159
130
160
C14H12BrN3O2
C9H10BrN3O2
C24H25N5O2
C26H29N5O3
C25H27N5O2
C24H24N5O2Cl
C20H25N5O2
C19H22N4O2
C20H24N4O2
C26H28N4O2
C18H20N4O3
/
NC6H5
/
NC6H4OC2H5(o)
NC6H4CH3(p)
NC6H4Cl(o)
NC6H4CH3(p)
CH2
/
78
/
147
119
104
/
/
9g
9h
9i
CHCH3
CHCH2C6H5
O
/
92
/158
a
All compounds were recrystallized from EtOH with the exception of compounds 9b and 9h which were purified by column chromatography
using as eluent cyclohexane/ethyl acetate 1:2 and CHCl3/MeOH 9:1, respectively.
b
The elemental analyses are within 90.4% of the theoretical values for C, H, N.
/

V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
/1071
1067
Table 2
Physical and chemical data of compounds 10aÁ
/
i and 14aÁc
/
a
Comp.
N
X
R
Yield (%)
M.p. (8C)
Formula b
10a
10b
10c
10d
10e
10f
10g
10h
10i
2
2
2
2
2
2
2
2
2
3
3
3
NC6H5
Ph
Ph
Ph
Ph
Me
Ph
Ph
Ph
Ph
Ph
Ph
Ph
72
98
86
62
80
72
83
95
90
98
98
97
154Á
105Á
149Á
149Á
124Á
132Á
97Á
Oil
152Á
145Á
139Á
85Á
/
156
108
151
151
126
133
C24H27N5O2
C26H31N5O3
C25H29N5O2
C24H26N5O2Cl
C20H27N5O2
C19H24N4O2
C20H26N4O2
C26H30N4O2
C18H22N4O3
C25H29N5O2
C26H31N5O2
C27H33N5O3
NC6H4OC2H5(o)
NC6H4CH3(p)
NC6H4Cl(o)
NC6H4CH3(p)
CH2
/
/
/
/
/
CHCH3
/
100 dec.
CHCH2C6H5
O
/
155
147
141
14a
14b
14c
NC6H5
NC6H4CH3(p)
NC6H4OC2H5(o)
/
/
/
87 dec.
a
Compounds were recrystallized from EtOH with the exception of 10b which was recrystallized from cyclohexane and 10h which was purified by
column chromatography using as eluent CHCl3/MeOH 9:1.
b
The elemental analyses are within 90.4% of the theoretical values for C, H, N.
/
Table 3
Physical and chemical data of compounds 11aÁ
/
i and 15aÁc
/
Comp.
N
X
R
Yield (%)
M.p. (8C)
Recryst. solvent
Formula c
11a
11b
11c
11d
11e
11f
11g
11h
11i
2
2
2
2
2
2
2
2
2
3
3
3
NC6H5
Ph
Ph
Ph
Ph
Me
Ph
Ph
Ph
Ph
Ph
Ph
Ph
64
85
71
91
71
72
80
68
87
50
75
50
80Á
70Á
86Á
82Á
/
82
75
90
84
EtOH
C24H29N5O2
C26H33N5O3
C25H31N5O2
C24H28N5O2Cl
C20H29N5O2
C19H26N4O2
C20H28N4O2
C26H32N4O2
C18H24N4O3
C25H31N5O2
C26H33N5O3
C27H35N5O3
NC6H4OC2H5(o)
NC6H4CH3(p)
NC6H4Cl(o)
NC6H5CH3(p)
CH2
/
Cyclohexane
EtOH
EtOH
/
/
166Á
Oil a
73Á76
65Á70
Oil a
70Á72
152Á154
84Á87
/168
Acetone
CHCH3
/
A b
CHCH2C6H5
O
/
EtOH
15a
15b
15c
NC6H5
NC6H4CH3(p)
NC6H4OC2H5(o)
/
Cyclohexane
EtOH
Cyclohexane
/
/
a
Purified by column chromatography using as CHCl3/MeOH 7:3 for 11f and CHCl3/MeOH 9:1 for 11i.
Cyclohexane/ethyl acetate 1:1.
b
c
The elemental analyses are within 90.4% of the theoretical values for C, H, N.
/

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V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
/1071
Table 4
Physical and chemical data of compounds 12aÁ
/
i, 16aÁc and 19
/
a
Comp.
N
X
R
Yield (%)
M.p. (8C)
Formula b
12a
12b
12c
12d
12e
12f
12g
12h
12i
2
2
2
2
2
2
2
2
2
3
3
3
1
NC6H5
Ph
Ph
Ph
Ph
Me
Ph
Ph
Ph
Ph
Ph
Ph
Ph
Ph
56
40
84
35
50
57
66
57
42
76
31
17
19
120Á
59Á61
129Á
138Á
100Á
Oil
118Á
/
125
C24H27N5O
C25H29N5O
C26H31N5O2
C24H26N5OCl
C20H27N5O
C19H24N4O
C20H26N4O
C26H30N4O
C18H22N4O2
C25H29N5O
C26H31N5O
C27H33N5O2
C24H27N5O2
NC6H4OC2H5(o)
NC6H4CH3(p)
NC6H4Cl(o)
NC6H4CH3(p)
CH2
/
/
131
140
101
/
/
CHCH3
/121
CHCH2C6H5
O
118Á/120
Oil
16a
16b
16c
19
NC6H5
115Á
130Á
102Á
159Á
/
118
133
104
161
NC6H4CH3(p)
NC6H4OC2H5(p)
NC6H4OCH3(o)
/
/
/
a
All compounds were recrystallized from EtOH, with exception of 12f and 12i which were purified by column chromatography using as CHCl3/
MeOH 7:3 and CHCl3/MeOH 9:1, respectively.
b
The elemental analyses are within 90.4% of the theoretical values for C, H, N.
/
1
3.1.3. General procedure for compounds 10aÁ
The appropriate 2-substituted isoxazolo[3,4-d]pyrida-
zinones (0.5 mmol) 9aÁi were suspended in EtOH (10
ml). Hydrazine hydrate (1.5 mmol) and 10% Pd/C (40
mg) were added and the mixture was refluxed for 1 h.
Then, after cooling the catalyst was filtered off and the
/
i
(15a). H NMR (CDCl3): d 1.45 (d, Jꢀ
CH3); 2.25Á2.40 (m, 2H, CH2CH2CH2); 2.90Á
10H, 8H piperazine and CH2CH2CH2Ãpiperazine);
4.20Á4.40 (m, 2H, NCH2CH2CH2Ãpiperazine); 4.70Á
4.85 (m, 1H, CHOH); 5.95 (exch. br s, 1H, OH);
6.85Á6.95 (m, 3H, Ar); 7.20Á7.50 (m, 7H, Ar).
/
7.8 Hz, 3H,
/
/
3.35 (m,
/
/
/
/
/
/
/
solvent was evaporated in vacuo affording 10aÁi as
/
residue.
3.1.5. General procedure for compounds 12aÁ
The appropriate 5-hydroxyethylpyridazinones 11aÁ
15aÁc (0.4 mmol) were reacted with PPA (40 mmol) for
2Á24 h. For compounds 12e and 16aÁc the reaction was
carried out at 50 8C, for all others at rt. After treatment
with ice water the mixture was neutralized with 1 N
NaOH and compounds were recovered by suction (12a,
/
i and 16aÁ
/
c
3.1.3.1. 5-Acetyl-4-amino-6-phenyl-2-{[4(4-
methyl)phenylpiperazin-1-yl]ethyl}-pyridazin-3(2H)-
/i,
/
1
one (10c). H NMR (CDCl3): d 1.80 (s, 3H, COCH3),
/
/
2.30 (s, 3H, CÃ
3.25 (t, Jꢀ7.8 Hz, 2H, CH2CH2Ã
Jꢀ7.8 Hz, 2H, CH2CH2Ãpiperazine); 6.85 (m, 2H, Ar);
7.05 (m, 2H, Ar); 7.45 (s, 5H, Ar).
/
CH3); 2.80Á/3.15 (m, 8H, piperazine),
/
/
piperazine); 4.45 (t,
/
/
12cÁ
20 mmol) (12b, 12f, 12i and 16aÁ
the solvent.
/
e and 12g,h) or after extraction with CH2Cl2 (3ꢁ
/
/c) and evaporation of
3.1.4. General procedure for compounds 11aÁ
/
i and 15aÁc
/
Sodium borohydride (2.8 mmol) was added portion-
wise to a stirred solution of the appropriate 4-amino-5-
acetyl derivatives 10aÁ
/
i or 14aÁ
/
c in MeOH (5Á/16 ml).
3.1.5.1. 4-Amino-6-phenyl-2[(4-phenylpiperazin-1-
The reaction mixture was stirred for 20Á
/
120 min at rt
1
and, after concentration in vacuo, it was diluted with
cold water. Compounds were recovered by suction, with
the exception of 11b, 11f,g and 11i which were obtained
yl)propyl]5-vinylpyridazin-3(2H)-one (16a). H NMR
(CDCl3): d 2.40Á2.60 (m, 2H, CH2CH2CH2); 2.95Á3.20
(m, 6H, CH2CH2CH2Ãpiperazine and 4H piperazine);
3.50Á3.65 (m, 4H, piperazine); 4.35 (t, Jꢀ7.8 Hz, 2H,
CONCH2); 5.50 (d, 1H, Jꢀ18.3 Hz, CHÄCH2); 5.60 (d,
1H, Jꢀ12.4 Hz, CHÄCH2); 6.25 (dd, 1H, Jꢀ18.3 Hz,
Jꢀ12.4 Hz, CH ÄCH2); 6.85Á7.00 (m, 3H, Ar); 7.25Á
7.35 (m, 2H, Ar); 7.45 (s, 5H, Ar).
/
/
/
after extraction with CH2Cl2 (3ꢁ
/
20 ml) and evapora-
/
/
tion in vacuo.
/
/
/
/
/
/
/
/
/
3.1.4.1. 4-Amino-5-hydroxyethyl-6-phenyl-2-[(4-
phenylpiperazin-1-yl)propyl]-pyridazin-3(2H)-one

V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
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1069
3.1.6. General procedure for compounds 14aÁ
/
c
Software Limited, Horley, UK). The antinociceptive
activity of the tested compounds was also evaluated as
A mixture of the appropriate 1-aryl-4-(3-bromopro-
pyl)piperazine (0.75 mmol), 5-acetyl-4-aminopyridazi-
none (13) (0.5 mmol) and potassium carbonate (4.0
mmol) in acetone (DMF for 14b) was heated at 70 8C
quantal
protection,
using
protected
treated
the
formula:/
% quantal protectionꢀ
ꢁ100; where ‘pro-
for 1Á3 h under stirring. After cooling, cold water was
/
tected’ means the number of animals completely pro-
tected from the effects of PPQ and ‘treated’ means the
number of animals treated in each group.
added and products were recovered by suction.
3.1.6.1. 5-Acetyl-4-amino-6-phenyl-2-[(4-
phenylpiperazin-1-yl)propyl]-pyridazin-3(2H)-one
1
(14a). H NMR (CDCl3): d 1.80 (s, 3H, CH3), 2.10Á
/
2-
2.80 (m, 6H,
piperazine and 4H piperazine); 3.20Á
3.35 (m, 4H, piperazine); 4.30 (t, Jꢀ7.9 Hz, 2H,
CONCH2); 6.85Á6.95 (m, 3H, Ar); 7.20Á7.35 (m, 2H,
Ar); 7.45 (s, 5H, Ar).
4. Results and discussion
30 (m, 2H, CH2CH2CH2), 2.60Á
/
CH2CH2CH2Ã
/
/
All the final compounds were evaluated as antinoci-
ceptive agents at the dose of 100 mg/kg s.c. in writhing
test induced by PPQ and mouse abdominal constrictions
were calculated. The obtained results are reported in
Table 6.
/
/
/
3.1.7. Synthesis of 4-amino-6-phenyl-2-{[4-(2-
methoxy)phenylpiperazin-1yl]methyl}-5-vinylpyridazin-
3(2H)-one (19)
To a suspension of compound 18 (0.05 mmol) in
EtOH (3.5 ml), 2-methoxyphenylpiperazine (0.1 mmol)
and 37% CH2O (0.1 mmol) were added. The mixture
was refluxed for 2 h under stirring, the solvent was
evaporated in vacuo and the residue crystallized from
EtOH.
In order to define some preliminary structureÁactivity
/
relationships, the synthesized compounds can be exam-
ined considering three different chemical series:
a) 4-amino-5-vinylpyridazinones with piperidinyl(mor-
pholinyl) ethyl chains (compounds 12fÁ
b) 4-amino-5-vinylpyridazinones with arylpiperaziny-
lalkyl chains (compounds 12aÁe, 16aÁc and 19)
/
i)
/
/
c) 4-amino-5-acetylpyridazinones 10c, 10f and 10g.
1H NMR (CDCl3): d 3.00Á
3.85 (s, 3H, OCH3); 3.90 (s, 2H, NCH2N); 5.50Á
4H, CHÄCH2 and NH2); 6.30 (dd, Jꢀ15.0, 9.0 Hz, 1H,
CH ÄCH2); 6.80Á7.05 (m, 4H, Ar); 7.35Á7.50 (m, 5H,
Ar).
/
3.15 (m, 8H, piperazine);
5.65 (m,
/
In the first series, compound 12f, bearing an unsub-
stituted piperidine was found completely devoid of
activity; isosteric replacement of the CH2 with oxygen
(12i) resulted in a slight improvement of activity,
whereas homologation of 12f led to a strong increase
of activity (compound 12g, 63% inhibition); this com-
pound, which was also able to protect 20% of the treated
animal from the noxious stimulus, is one of the most
active among the compounds described in this study. On
the contrary, the phenylpiperidinyl analogue 12h proved
to be less active. These data seem to indicate that small
lipophilic groups are the best arranged at position 4 of
the cycloalkylamine.
/
/
/
/
/
3.2. Pharmacology
The antinociceptive effect of the compounds reported
in Table 5 was evaluated by the p-phenylquinone-
induced abdominal constriction test according the
procedure described by Siegmund et al. [18] and
modified by Milne [19]. Male CD-1 mice (21Á
/
35)
maintained at 2391 8C, were injected intraperitoneally
/
with p-phenylquinone (PPQ) (2 mg/kg using a 0.02%
solution in 5% ethanol/95% distilled water, 20 min after
subcutaneous administration of the test compound).
Groups of 5 mice were used for each dose tested. The
number of characteristic abdominal constrictions for
each mouse was counted for a period of 8 min after the
PPQ injection. The mean number of abdominal con-
strictions was compared for each treatment group with
the mean number in the vehicle-treated control group.
The percentage inhibition was then calculated for each
compound-treated group. All compounds were dis-
solved in DMSO and administered subcutaneous in a
volume of 5 ml/kg. The same volume of DMSO was
administered to controls. The ED50 and related 95%
confidence intervals were determined by the nonlinear
fitting analysis using the software GraFit (Eriacus
Among the compounds belonging to the second
series, the 4-phenyl derivative 12a (nꢀ2) proved to be
/
weak active (12% inhibition). Introduction of a chlorine
(12d) or an ethoxy group (12b) in the ortho position left
almost unchanged the activity, whereas introduction of
a CH3 in the para position (12c) was associated with a
good level of activity (53% inhibition and 20% of
quantal protection). When this same fragment was
introduced in the substrate bearing a methyl group at
position 6 of the pyridazine (12e) a dramatic increase of
activity was observed (100% inhibition and 100%
quantal protection). DoseÁ
very high potency (ED50ꢀ2.5 mg/kg) (Fig. 1).
Elongation of the methylenic spacer to nꢀ
/
response studies revealed
/
/
3 gave
interesting results, the 4-unsubstituted phenyl derivative
16a displaying a significant level of activity (51%

1070
V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á1071
/
Table 5
Analgesic activity of pyridazinone derivatives determined by p-phenylquinone-induced (PPQ) mouse abdominal constriction test
a
Comp.
No. of abdominal constrictions
Inhibition (%)
Quantal protection (%)
12f
12g
18.209
7.40 *9
12.40 *9
14.409
16.609
12.409
8.809
13.209
0.00 *9
9.60 *9
12.009
4.609
7.80 *9
14.009
19.409
10.009
18.409
0.80 *9
7.00 *9
/
1.50
3.26
0.87
2.87
4.30
4.76
3.57
3.46
0.00
2.68
2.90
1.89
3.25
5.26
1.72
4.46
1.86
0.80
2.02
7
63
38
27
12
34
53
30
100
51
36
76
60
30
1
0
20
0
/
12h
/
12i
/
0
12a
/
0
12b
12c
/
20
20
0
/
12d
/
12e
16a
/
100
0
/
16b
/
0
16c
19
/
40
20
20
0
/
10c
/
10f
/
10g
/
50
40
Saline
Morphine b
Emorfazone b
/
/
96
64
80
20
/
a
Compounds were dissolved in DMSO 100% and were administered (100 mg/kg, s.c.) 20 min before PPQ (2 mg/kg, i.p.).
Compounds were dissolved in saline; morphine was administered at 1 mg/kg s.c. and Emorfazone at 100 mg/kg s.c..
b
* P B
/
0.05 vs. appropriate vehicle treated group (nꢀ5).
/
Table 6
Elemental analyses for the final compounds 12aÁ
/
i, 16aÁ
/
c and 19
Comp.
Calculated (%)
Found (%)
C
H
N
C
H
N
12a
12b
12c
12d
12e
12f
12g
12h
12i
71.78
72.25
70.07
66.11
67.95
70.33
70.96
75.32
66.22
72.25
72.68
70.55
69.03
6.79
7.05
7.03
6.02
7.71
7.47
7.76
7.31
6.81
7.05
7.29
7.25
6.53
17.44
16.86
15.72
16.07
19.82
17.27
16.56
13.52
17.17
16.86
16.31
15.24
16.78
72.03
72.50
70.35
65.90
68.15
70.50
71.02
75.05
65.96
72.39
73.00
70.81
68.85
6.80
7.03
7.00
6.00
7.74
7.44
7.74
7.44
6.83
7.02
7.31
7.22
6.50
17.51
16.92
15.78
16.01
19.75
17.34
16.50
13.50
17.10
16.80
16.37
15.30
16.84
16a
16b
16c
19
Fig. 1.
was completely inactive, the p-tolyl derivative 10c
showed an intermediate value of inhibition (30%).
Taken together these results suggest that in the
compounds examined in this study the presence of small
lipophilic groups at position 4 of the cycloalkylamine
and the nature of the substituent at position 6 of the
pyridazine are important requirements for antinocicep-
tive activity in writhing test. On the contrary, the nature
of the function at position 5 and the length of the
methylenic spacer seem to play only a marginal role.
This study allowed to identify compound 12e as a very
promising lead, useful for further chemical development.
This novel agent, compared with the structurally related
inhibition) and the para ethoxy phenyl analogue 16c
being even more active (76% inhibition, 40% quantal
protection). Finally compound 19, which is the only
characterized by a mono methylenic spacer, was proved
endowed with a good level of activity (60% inhibition,
20% quantal protection).
In the series of 4-amino-5-acetylpyridazinones, the
structural requirements of the cycloalkylamino moiety
at the end of the carbon chain demonstrated to be very
similar to that of the 4-amino-5-vinyl analogues: in fact
the best arranged system was the 4-methylpiperidine
(compound 10g, 50% inhibition and 40% quantal
protection); the unsubstituted piperidine derivative 10f

V.D. Piaz et al. / Il Farmaco 58 (2003) 1063Á
/1071
1071
[6] M. Sato, Y. Ishizuka, A. Yamaguchi, Pharmacological investiga-
tion of 4-ethoxy-2-methyl-5-morpholino-3(2H)-pyridazinone,
Emorfazone 2 was over 40-fold more potent in the
mouse abdominal constriction test (Fig. 1). A significant
improvement in comparison with the previous lead 6
Arzneim. Forsch. 31 (1981) 1738Á
[7] M. Sato, A. Yamaguchi, Studies on mechanism of action of
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/
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[8] C. Rubat, P. Coudert, E. Albuisson, J. Bastide, J. Couquelet, P.
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