Synthesis and pharmacological evaluation of novel heterotricyclic acylhydrazone derivatives, designed as PAF antagonists

Article abstract of DOI:10.1016/S0928-0987(00)00102-0

This paper describes the synthesis and the antiplatelet properties of new heterotricyclic N-acylhydrazone derivatives (7a-e), structurally analogous to known hetrazepinic PAF antagonists, exploring molecular hybridization as a tool for molecular designing. The synthetic route employed to access compounds (7a-e) used, as starting material, the previously described methyl 3-hydroxy-8-methyl-6-phenyl-6H-pyrazolo[3,4-b]thieno[2,3-d]pyridine-2-carboxylate derivative. The results from inhibitory effects of these novel acylhydrazone derivatives (7a-e) upon PAF-induced platelet aggregation, indicated that all compounds present a significant antithrombotic profile. Copyright (C) 2000 Elsevier Science B.V.

Full text of DOI:10.1016/S0928-0987(00)00102-0

European Journal of Pharmaceutical Sciences 11 (2000) 285–290
www.elsevier.nl/locate/ejps
Synthesis and pharmacological evaluation of novel heterotricyclic
acylhydrazone derivatives, designed as PAF antagonistsq
Aline G.M. Fraga, Carlos R. Rodrigues, Ana L.P. de Miranda, Eliezer J. Barreiro,
*
Carlos A.M. Fraga
˜
´
´
ˆ
´
Laboratorio de Avaliac¸ao e Sıntese de Substancias Bioativas (LASSBio, http://www.ufrj.br/|pharma/lassbio), Faculdade de Farmacia,
Universidade Federal do Rio de Janeiro, P.O. Box 68006, 21944-970, Rio de Janeiro, RJ, Brazil
Received 25 November 1999; received in revised form 26 April 2000; accepted 27 April 2000
Abstract
This paper describes the synthesis and the antiplatelet properties of new heterotricyclic N-acylhydrazone derivatives (7a–e), structurally
analogous to known hetrazepinic PAF antagonists, exploring molecular hybridization as a tool for molecular designing. The synthetic
route employed to access compounds (7a–e) used, as starting material, the previously described methyl 3-hydroxy-8-methyl-6-phenyl-6H-
pyrazolo[3,4-b]thieno[2,3-d]pyridine-2-carboxylate derivative. The results from inhibitory effects of these novel acylhydrazone deriva-
tives (7a–e) upon PAF-induced platelet aggregation, indicated that all compounds present a significant antithrombotic profile.
Elsevier Science B.V. All rights reserved.
2000
Keywords: PAF antagonists; N-Acyl hydrazone; Pyrazolo[3,4-b]thieno[2,3-d]pyridine derivatives; Antithrombotic activity
1. Introduction
anti-aggregating activity in the PAF-induced rabbit platelet
aggregation model (Fig. 1) (Weber and Heuer, 1989).
The clinical application of the hetrazepine derivatives
(2) and (3) is still limited by its poor pharmacokinetic
profile. These compounds have a metabolic vulnerability
for the 1,4-diazepine methylene group to enzymatic hy-
droxylation in human (Garzone and Kroboth, 1989).
These results led us to explore a new heterocyclic
pattern developed in our laboratory (Carvalho et al.,
1996a), in the design of metabolically stable pyrazolo[3,4-
b]thieno[2,3-d]pyridine amide series (Carvalho et al.,
1996b), e.g., (4), using as rational basis, the pharma-
cophoric three-point model for PAF antagonist activity,
disclosed by Bures et al. (1994) (Fig. 1). Unfortunately,
the results of this previous study indicated a very poor
anti-aggregating activity in the PAF-induced model for
compounds (4) and (5) (Carvalho et al., 1996b), despite
the presence of all minimal structural requirements, antici-
pated by the Bure’s pharmacophoric model.
The platelet-activating factor (PAF, 1) is an endogenous
phospholipid mediator first described by Benveniste et al.
(1972), as the most potent platelet stimulator, also present-
ing hypotensive properties (Vargaftig et al., 1980). This
autacoid is now recognized as being directly or indirectly
involved in many pathophysiological responses, such as
ischemia, systemic anaphylaxis and inflammatory diseases
(Braquet et al., 1987, 1989). Since the PAF bioreceptor has
been recently isolated and sequenced (Chao and Olson,
1993), the design of new PAF receptor antagonists (PAF-
ant) consists of an important therapeutic strategy to
development of new antithrombotic agents (Ayscough and
Whittaker, 1995). Several different classes of structurally
related or heterocyclic PAFant have been described in the
literature (Braquet, 1991). Among these, the hetrazepinic
amides, such as the lead-compound WEB2086 (2) and the
conformationally restricted analogue WEB2170 (3), were
described as potent competitive PAFant agents, showing
This unexpected biological profile led us to design new
structurally related derivatives (7a–e) (Fig. 1), exploring
the molecular hybridization of the previously described
tricyclic amide derivatives (5) (Carvalho et al., 1996b) and
the potent platelet antiaggregating pyrazolyl N-acylhydra-
zone derivative (6) (Matheus et al., 1991; Soler, 1993;
qThis paper is the contribution [50 from LASSBio.
*Corresponding author. Tel.: 155-21-2609-192, ext. 220/223; fax:
155-21-260-2299.
E-mail address: cmfraga@pharma.ufrj.br (C.A.M. Fraga).
0928-0987/00/$ – see front matter
PII: S0928-0987(00)00102-0
2000 Elsevier Science B.V. All rights reserved.

286
A.G.M. Fraga et al. / European Journal of Pharmaceutical Sciences 11 (2000) 285–290
(1H NMR) spectra were determined in dimethylsulfoxide-
d6, using tetramethylsilane as an internal standard with a
Brucker AC 200 spectrometer. Splitting patterns were as
follows: s, singlet; d, doublet; t, triplet; m, multiplet; br,
broad. Infrared spectra (IR) were obtained with a Perkin-
Elmer 1600 spectrometer as KBr pellets. The mass spectra
(MS) were obtained on a GC/VG Micromass 12 at 70 eV.
Microanalysis data were obtained with a Perkin-Elmer 240
analyzer, using a Perkin-Elmer AD-4 balance.
The progress of all reactions was monitored by TLC
performed on 2.035.0-cm aluminum sheets precoated with
silica gel 60 (HF-254, Merck) to a thickness of 0.25 mm.
The developed chromatograms were visualized under
ultraviolet light (254–265 nm). Merck silica gel (70–230
mesh) was used for column chromatography. Solvents used
in reactions were dried, redistilled prior to use and stored
˚
over 3–4-A molecular sieves.
2.1.1. 3-Hydroxy-8-methyl-6-phenylpyrazolo[3,4-
b]thieno[2,3-d]pyridine-2-carbohydrazine (9) (Dias et
al., 1994; Ribeiro et al., 1998)
To a solution of 2.37 g (7.00 mmol) of hydroxy-ester
derivative (Carvalho et al., 1996a) (8) in 30 ml of ethanol,
was added 8.5 ml of 80% hydrazine monohydrate. The
reaction mixture was maintained under reflux for 25 h,
until the end of reaction was indicated by TLC. Then, the
media was poured on ice and the resulting precipitate was
filtered out, affording the title compound in 86% yield, as a
Fig. 1. Design concept of new tricyclic acylhydrazone derivatives (7a–e).
Miranda et al., 1994). Thus, we decided to include the acyl
group at C-2 of the pyrazolo[3,4-b]thieno[2,3-d]pyridine
system of (5) in the NAH framework present in (6),
furnishing the new hybrid derivatives (7a–e).
1
yellow light solid, m.p. .2508C. H NMR (200 MHz):
d510.95 (s, 1H, CONH–), 8.70 (5, 1H, Py-H), 8.12 (d,
9
9
2H, Ar-H2, J57.7 Hz), 7.51 (t, 2H, Ar-H3, J57.3 Hz),
9
In this new series, presented herein, the methoxy group
located at C-3 in the tricyclic system of (5) was replaced
by a hydroxyl group to induce an intramolecular hydrogen
bond between this donor substituent and the acceptor
present in acylhydrazone moiety. This was done in order to
mimic the conformational restriction introduced by annula-
tion of the side chain of (2), which previously was shown
to result in the more active cyclopentyl derivative
WEB2170 (3) (Fig. 1) (Weber and Heuer, 1989).
The nature of the para-substituent present at the phenyl
group (i.e., H, OMe, Br, NMe2, CN) in the new deriva-
tives (7a–e), which was elected to introduce an important
variation in sp-Hammett values (ranging from 20.83
(NMe2) to 10.66 (CN) (Hansch and Leo, 1979)), can be
useful to evidence any electronic contribution of this
structural sub-unit on the antithrombotic activity of this
series of compounds.
7.30 (t, 1H, Ar-H4, J57.1 Hz), 4.44 (br, 2H, –NH2), 2.62
(5, 1H, –CH3), 1.3 (br, 1H, OH) ppm; MS (70 eV) m/z
(relative abundance): 339 (M1, 37%), 308 (100%), 252
(18%), 77 (42%); IR (KBr) cm21: 3305 (n O–H), 3027 (n
N–H), 2923 (n N–H), 1651 (n C0), 1252 (n C–O).
Anal. Calc. for C16H13N5O2S: C, 56.63; H, 3.86; N,
20.64. Found: C, 56.68; H, 3.90; N, 20.59.
2.1.2. General procedure for preparation of tricyclic
acylhydrazones (Dias et al., 1994; Ribeiro et al., 1998)
(7a–e, Table 2)
To a solution of 0.21 g (0.62 mmol) of (9) in absolute
ethanol (ca. 10 ml) containing two drops of 37% hydro-
chloric acid, 0.62 mmol of corresponding benzaldehyde
derivative was added. The mixture was stirred at room
temperature for 45 min, until extensive precipitation was
visualized. Next, the mixture was poured in cold water,
neutralized with 10% aqueous sodium bicarbonate solution
and the precipitate formed was filtered out and dried.
2. Experimental procedures
2.1.3. Benzylidene 3-hydroxy-8-methyl-6-
phenylpyrazolo[3,4-b]thieno[2,3-d]pyridine-2-
carbohydrazide (7a)
2.1. Chemistry
Melting points were determined with a Quimis 340
apparatus and are uncorrected. Proton magnetic resonance
The derivative (7a) was obtained as a white solid, by
condensation of (9) with benzaldehyde (see Table 2); MS

A.G.M. Fraga et al. / European Journal of Pharmaceutical Sciences 11 (2000) 285–290
287
(70 eV) m/z (relative abundance): 427 (M1, 33%), 308
(100%), 252 (21%), 77 (58%), 63 (18%); IR (KBr) cm21
gregometer. PRP (400 ml) was incubated at 378C for 1 min
with continuous stirring at 900 rpm and then stimulated
with PAF (50 nM in distilled water).
:
3021 (n O–H), 2931 (n NH), 1636 (n C5O), 1270 (n
C–O).
Test compounds (7a–e) and the vehicle (0.5% DMSO, 2
ml) were incubated 5 min with PRP samples before
addition of the aggregating agent. The DMSO used as
vehicle did not have either pro- or antiplatelet aggregation
activity. WEB2170 (0.15 and 1 mM), a hetrazepinic PAF
antagonist, was used as standard.
The platelet aggregation was expressed in percentage
(%) and data were analyzed statistically by Student’s t-test
for a P value of ,0.05 and expressed as mean6S.D. for n
experiments in triplicate.
2.1.4. (49-Methoxybenzylidene) 3-hydroxy-8-methyl-6-
phenylpyrazolo[3,4-b]thieno[2,3-d]pyridine-2-
carbohydrazine (7b)
The derivative (7b) was obtained as a white solid, by
condensation of (9) with 4-methoxybenzaldehyde (see
Table 2); MS (70 eV) m/z (relative abundance): 457 (M1,
46%), 308 (52%), 252 (16%), 150 (100%), 77 (50%), 63
(13%); IR (KBr) cm21: 3025 (n O–H), 2929 (n N–H),
1631 (n C5O), 1252 (n C–O).
2.3. Computational methodology
2.1.5. (49-Bromobenzylidene) 3-hydroxy-8-methyl-6-
phenylpyrazolo[3,4-b]thieno[2,3-d]pyridine-2-
carbohydrazine (7c)
The derivative (7c) was obtained as a white solid, by
condensation of (9) with 4-bromobenzaldehyde (see Table
2); MS (70 eV) m/z (relative abundance): 506 (M1, 25%),
308 (100%), 252 (29%), 89 (45%), 77 (61%), 63 (26%);
IR (KBr) cm21: 3027 (n O–H), 2928 (n N–H), 1633 (n
C5O), 1249 (n C–O), 695 (n C–Br).
Geometry optimizations were performed at self con-
sistent field (SCF) level using the AM1 Hamiltonian,
within the MOPAC version 7.0 package (Stewart, 1993) on
an IBM RISC system/6000 workstation under the IBM
AIX version 3.0 operational system, and on a Pentium 100
MHz running under the FreeBSD Unix system. The
potential energy surface (PES) slices were calculated for
the torsion angles (see Fig. 2), which varied independently
between 0 and 3608 with a 308 increment.
2.1.6. (49-N,N-Dimethylaminobenzylidene) 3-hydroxy-8-
methyl-6-phenylpyrazolo[3,4-b]thieno[2,3-d]pyridine-2-
carbohydrazine (7d)
The derivative (7d) was obtained as an orange solid, by
condensation of (9) with 4-N,N-dimethylaminobenzal-
dehyde (see Table 2); MS (70 eV) m/z (relative abun-
dance): 470 (M1, 21%), 308 (100%), 252 (23%), 163
(87%), 77 (68%), 63 (13%); IR (KBr) cm21: 3022 (n
O–H), 2919 (n N–H), 1630 (n C5O), 1271 (n C–O).
Minimum energy structures were then reoptimized
adopting the keywords GNORM50.1, PRECISE and
MMOK to correct the peptide bonds, and were unequivo-
cally characterized by Hessian matrix analysis.
3. Results and discussion
3.1. Molecular modeling
2.1.7. (49-Cyanobenzylidene) 3-hydroxy-8-methyl-6-
phenylpyrazolo(3,4-b]thieno[2,3-d]pyridine-2-
carbohydrazine (7e)
The derivative (7e) was obtained as a white solid, by
condensation of (9) with 4-cyanobenzaldehyde (see Table
2); MS (70 eV) m/z (abundance relative): 452 (M1, 32%),
308 (100%), 252 (24%), 149 (82%), 77 (78%), 63 (48%);
IR (KBr) cm21: 3030 (n O–H), 2930 (n N–H), 2361 (n
C≡N), 1631 (n C=O), 1264 (n C–O).
Molecular modeling studies were carried out using the
AM1 method, in order to generate a set of representative
low-energy conformations for one representative N-acyl-
hydrazone derivative, i.e., (7a). Inspection of its structure
shows the possibility of geometric isomerism at iminic
C=N bond (Z- and E-diastereomers). In order to anticipate
the diastereomer relative stability, we modeled (Z)- and
(E)-isomers of compound (7a). Additionally, we have
investigated the conformational behaviour of the amide
framework (HNCO), considering two minimum-energy
conformers with syn and anti arrangements, corresponding
to 0 and 1808 dihedral angles, respectively (de Sant’Anna
et al., 1996). The difference observed in the heat of
formation DHf of (Z)- and (E)-isomers was ca. 3.0 kcal/
mol, favoring the (Z)-diastereomer (Fig. 2A,C; Table 1).
Subsequently, we investigated the syn and anti arrange-
ments for the most stable (Z)-isomer. The syn conformer
(Fig. 2A) was more stable by ca. 4.0 kcal/mol than anti
conformer (Fig. 2B) (Table 1). This difference can be
rationalized by the presence of favorable antiparallel
2.2. Pharmacological
2.2.1. Platelet aggregation
Blood was collected from rabbits by puncture of the
central ear artery into 3.8% sodium citrate (9:1, v/v).
Platelet-rich plasma (PRP) was prepared by centrifugation,
5003g for 9 min at room temperature. PRP was adjusted
to 53106 platelets/ml after counting.
Platelet aggregation was monitored by the turbidimetric
method (Born and Cross, 1963) in a Chrono-Log ag-

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A.G.M. Fraga et al. / European Journal of Pharmaceutical Sciences 11 (2000) 285–290
Fig. 2. Representation of the minimal energy conformations (A–D) of the tricyclic acylhydrazone (7a).
dipole–dipole interaction between carbonyl and N–H
groups of the peptidic bond.
(conformer D, Fig. 2), in spite of the more planar character
of the latter. These results confirm the conformational
analogy of the compounds of series (7) with cyclopentyl
hetrazepine (3).
Because of the greater stability of (Z)-diastereomer
identified in compound (7a), we investigated two plausible
conformers presenting intramolecular hydrogen bonds, in
order to evaluate the conformational restriction of the
pharmacophoric side chain anticipated in the design of this
new series. The results of this conformational studies have
showed that the hydrogen bond between C=O???HO in
conformer A (Fig. 2) is more stable than C=N???HO
3.2. Chemistry
The new target tricyclic acylhydrazone derivatives (7a–
e) were synthesized as depicted in Scheme 1. Our synthetic
approach to these new compounds identified the 8-methyl-
6-phenyl-6H-pyrazolo[3,4-b]thieno[2,3-d]pyridine-2-car-
bohydrazide (9) as the key intermediate (Scheme 1). This
compound was prepared, in 86% yield, by treatment of the
methyl ester derivative (8) with hydrazine hydrate in
ethanol at reflux (Dias et al., 1994; Ribeiro et al., 1998).
The infrared spectrum of (9) indicated the acylhydrazine
moiety by the absorptions at 2923 and 2853 cm21, typical
Table 1
Heat of formation (DHf), dipole moment (m) and hydrogen bond distance
˚
(HB, A) for the conformers of (7a) obtained by semi-empirical AM1
method
Conformera
Diastereomer
DHf
(kcal/mol)
m
HB
(A)
˚
(D)
for –NH–NH2 of hydrazine group, and at 1651 cm21
referent to amide carbonyl group.
,
A
B
C
D
Z
Z
E
Z
130.30
134.51
133.49
139.55
3.8
4.3
3.6
3.8
HO???O
(2.01)
HO???O
(2.05)
HO???O
(2.03)
HO???N
(1.98)
The synthesis of the novel pyrazolo[3,4-b]thieno[2,3-
d]pyridine acylhydrazone derivatives (7a–e) was con-
cluded in good yields (Table 2), by acid catalyzed con-
densation of the tricyclic acylhydrazide derivative (9) with
benzaldehyde, 4-methoxybenzaldehyde, 4-bromobenzal-
dehyde, 4-dimethylaminobenzaldehyde, 4-cyanobenzal-
a See Fig. 2.

A.G.M. Fraga et al. / European Journal of Pharmaceutical Sciences 11 (2000) 285–290
289
compared to the minor one, indicating the (Z)-configura-
tion (Table 2). This behavior is in agreement with our
molecular modeling studies as well as with previous results
disclosed by Karabatsos for the relative configuration of
hydrazones and related compounds (Karabatsos et al.,
1962, 1963; Karabatsos and Taller, 1963).
3.3. Pharmacology
The antithrombotic activity of these novel tricyclic
acylhydrazone derivatives (7a–e) was evaluated by their
ability to inhibit platelet aggregation of rabbit platelet-rich
plasma (PRP) induced by PAF at 50 nM. The results of the
anti-platelet profile of (7a–e) are displayed in Table 3.
All compounds studied, at a dose of 100 mM, were able
to inhibit by ca. 30%, the platelet aggregation induced by
PAF, showing a better biological profile than precedent
amide series (4) and (5) (Carvalho et al., 1996b).
Compounds (7a) and (7c) were also evaluated at 10 mM,
presenting, respectively, 10.4 and 13.6% of inhibition of
the PAF-induced platelet aggregation (Table 3). Addition-
ally, compound (7a) presented a poor platelet anti-ag-
gregating profile when compared with WEB2170 (3) used
as standard, at the same molar concentration, i.e., 1 mM
(Table 3).
Scheme 1.
dehyde, respectively, in ethanol at room temperature, as
described previously (Dias et al., 1994; Ribeiro et al.,
1998) (Scheme 1).
Finally, we determined the relative configuration of the
N=C bond in acylhydrazone derivatives (7a–e), in order to
assure the diastereomeric ratio, essential to understand the
1
biological profile. Analysis of the H NMR spectra of the
compounds (7a) and (7c–e) (Table 2) revealed the pres-
ence of only one hydrogen signal referent to imino bond.
In contrast, only compound (7b) was obtained as a 3:1
diastereomeric mixture. The imino hydrogen signal of the
major diastereomer is downfielded by 0.2–0.3 ppm when
The similar PAF antagonist activity of compounds (7a–
e), could be due to the phenyl group of the hydrazone
moiety and its hydrophobic interactions with PAF
bioreceptor. Considering the major hydrophobic contribu-
tion of phenyl ring (p51.96 (Hansch and Leo, 1979)) in
Table 2
Yields, physical and spectroscopic data of pyrazolo[3,4-b]thieno[2,3-d]pyridine acylhydrazone derivatives (7a–e)
Compound
7a
W
Molecular
formulaa
Yield
(%)
M.p.
Ratio
1H NMRc
(8C)
(Z:E)
d (ppm)
H
C24H17N5O2S
95
84
.250b
100:0
75:25
9.04 (s, 1H, Py-H); 8.33 (d, 2H, Ar-H-29, J58.3 Hz); 8.27(s, 1H,
N5CH (Z)); 7.87 (d, 2H, Ar-H299 , J57.4 Hz), 7.71 (t, 2H, Ar-H3
7.65–7.58 (m, 2H, Ar-H4 and Ar-H499 ); 7.51 (t, 2H, Ar-H399 );
3.10 (br, 2H, NH1OH); 2.91 (s, 3H, ArCH3)
9, J58.1 Hz);
9
7b
OCH3
C24H19N5O3S
.250b
9.13 (s, 1H, Py-H); 8.59 (s, 1H, N5CH (E)); 8.35 (d, 2H, Ar-H-29,
J58.5 Hz); 8.32 (s, 1H, N5CH (Z)); 7.88 (t, 1H, Ar-H929 , J58.7 Hz);
7.64 (t, 2H, Ar-H3, J57.7 Hz); 7.44 (t, 1H, Ar-H49, J57.2 Hz);
7.15 (d, 2H, Ar-H939 , J58.8 Hz); 3.98 (s, 3H, OCH3); 3.22 (br, 2H,
NH1OH); 2.89 (s, 3H, ArCH3)
7c
7d
7e
Br
C23H16BrN5O2S
C25H22N6O2S
C24H16N6O2S
89
81
87
.250b
.250b
.250b
100:0
100:0
100:0
9.15 (s, 1H, Py-H); 8.35 (d, 2H, Ar-H-29, J57.5 Hz); 8.24 (s, 1H,
N5CH (Z));7.98–7.79 (m, 4H, Ar-H929 and Ar-H399 ); 7.68 (t, 2H, Ar-H3
J57.8 Hz);7.47 (t, 1H, Ar-H49, J57.5 Hz); 3.35 (br, 2H, NH1OH);
2.85 (s, 3H, ArCH3)
9,
N(CH3)2
8.98 (s, 1H, Py-H); 8.22 (d, 2H, Ar-H29, J57.5 Hz); 8.06 (s, 1H,
N5CH (Z)); 7.64 (d, 2H, Ar-H20, J58.6 Hz); 7.52 (t, 2H, Ar-H93, J57.2 Hz);
7.31 (t, 1H, Ar-H49, J57.3 Hz); 6.78 (d, 2H, Ar-H939 , J58.8 Hz);
2.99 (s, 6H, N(CH3)2); 2.95 (br, 2H, NH1OH); 2.7 (s, 3H, ArCH3)
9.01 (s, 1H, Py-H); 8.37 (d, 2H, Ar-H-29, J57.4 Hz); 8.21 (s, 1H,
N5CH (Z)); 8.07 (d, 2H, Ar-H929 , J58.2 Hz), 7.87 (d, 2H, Ar-H399 , J58.5 Hz);
CN
7.62 (t, 2H, Ar-H93, J57.5 Hz); 7.43 (t, 1H, Ar-H49, J57.6 Hz);
3.01 (br, 2H, NH1OH); 2.78 (s, 3H, ArCH3)
a The analytical results for C, H, N were within 10.4% of calculated values.
b Recrystallized from ethanol/water. Obtained from KBr plates.
c Recorded at 200 MHz, using DMSO-d6 as solvent.

290
A.G.M. Fraga et al. / European Journal of Pharmaceutical Sciences 11 (2000) 285–290
Table 3
dent histamine release from rabbit platelets: the role for IgE, basophilis
and a platelet activating factor. J. Exp. Med. 136, 1356–1377.
Born, G.V.R., Cross, M.J., 1963. The aggregation of blood platelets. J.
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Antiplatelet evaluation of novel pyrazolo[3,4-b]thieno[2,3-d]pyridine
acylhydrazone derivatives (7a–e) in a model induced by PAF at 50 nM
Compound
W
C (mM)a
nb
Aggregationc
(%)
Inhibition
(%)
Control
–
–
7
44.160.9
–
Braquet, P., Touqui, L., Shen, T.Y., Vargaftig, B.B., 1987. Perspectives in
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Braquet, P., 1991. In: Handbook of PAF and PAF Antagonists. CRC
Press, Boca Raton, FL.
WEB2170
WEB2170
–
–
1
0.15
3
6
2.860.2
28.562.9
93.6*
35.3*
7a
7b
7c
7d
7e
H
OMe
Br
N(CH3)2
CN
100
100
100
100
100
8
8
8
8
8
29.261.7
32.461.9
28.562.0
30.561.4
31.661.5
33.7*
26.5*
35.3*
30.8*
28.3*
Bures, M.G., Danaher, E., DeLazzer, J., Martin, Y.C., 1994. New
molecular modeling tools using three-dimensional chemical substruc-
tures. J. Chem. Inf. Comput. Sci. 34, 218–223.
Carvalho, A.S., Fraga, C.A.M., Barreiro, E.J., 1996a. Synthesis of
condensed tricyclic pyrazolo[3,4-b]thieno[2,3-d]pyridine and related
isostere derivatives. J. Heterocyclic Chem. 33, 309–313.
Carvalho, A.S., Fraga, C.A.M., Silva, K.C.M., Miranda, A.L.P., Barreiro,
E.J., 1996b. Synthesis and anti-platelet evaluation of new tricyclic PAF
antagonists, designed as structurally related to hetrazepine class-
WEB2086. J. Braz. Chem. Soc. 5, 247–256.
7a
7c
H
Br
10
10
3
3
39.560.8
38.160.8
10.4*
13.6*
7a
H
1
3
42.661.8
3.4 ns
Chao, W., Olson, M.S., 1993. Platelet activating factor: receptors and
signal transduction. Biochem. J. 292, 617–629.
a C, final concentration.
b n, number of experiments carried out in triplicate.
de Sant’Anna, C.M.R., de Alencastro, R.B., Rodrigues, C.R., Barreiro,
G., Barreiro, E.J., Neto, J.D.M., Freitas, A.C.C., 1996. A semiempiri-
cal study of pyrazole acylhydrazones as potential antimalarial agents.
Int. J. Quantum Chem.: Quantum Biol. Symp. 23, 111–119.
Dias, L.R.S., Alvim, M.J.F., Freitas, A.C.C., Barreiro, E.J., Miranda,
A.L.P., 1994. Synthesis and analgesic properties of 5-acyl-
arylhydrazone 1-H pyrazolo[3,4-b]pyridine derivatives. Pharm. Acta
Helv. 69, 163–169.
c The values of platelet aggregation represent the means6S.D.
* P,0.05 (Student t-test); ns, not significant.
comparison to that of the W substituent (p520.33 to
10.19 (Hansch and Leo, 1979)), no significant differences
in the platelet aggregation inhibitory profile of the novel
compounds (7a–e) could be observed.
Garzone, P.D., Kroboth, P.D., 1989. Pharmacokinetics of the newer
benzodiazepines. Clin. Pharmacokinet. 16, 337–364.
Hansch, C., Leo, A., 1979. In: Substituent Constants for Correlation
Analysis in Chemistry and Biology. Wiley, New York.
Karabatsos, G.J., Graham, J.D., Vane, F.M., 1962. Syn-anti isomer
determination of 2,4-dinitrophenylhydrazones and semicarbazones by
N.M.R. J. Am. Chem. Soc. 84, 753–755.
Karabatsos, G.J., Taller, R.A., 1963. Structural studies by nuclear
magnetic resonance. V. Phenylhydrazones. J. Am. Chem. Soc. 85,
3624–3629.
Karabatsos, G.J., Shapiro, B.L., Vane, F.M., Fleming, J.S., Ratka, J.S.,
1963. Structural studies by nuclear magnetic resonance. II. Aldehyde
2,4-dinitrophenylhydrazones. J. Am. Chem. Soc. 85, 2784–2788.
Matheus, M.E., Oliveira, L.F., Freitas, A.C.C., Carvalho, A.M.A.S.P.,
Barreiro, E.J., 1991. Antinociceptive property of new 4-acyl-
arylhydrazone pyrazole compounds. Braz. J. Med. Biol. Res. 24,
1219–1222.
4. Conclusions
As concluding remarks, the synthetic route described
herein represents an useful, efficient and high yield meth-
od, exploring the heterocyclic synthon (8) as starting
material to access new tricyclic acylhydrazone derivatives
(7a–e), structurally planned as hybrid of tricyclic amides
(5) and acylhydrazone derivative (6). The pharmacological
results confirmed the anticipated improvement of anti-PAF
activity in comparison with previous amide series (5) and
indicated that these heterotricyclic acylhydrazone deriva-
tives could be considered as a new lead-series of antit-
hrombotic agents, acting at the PAF receptor level.
Miranda, A.L.P., Soler, O., Freitas, A.C.C., Barreiro, E., 1994. A new
series of platelet aggregation inhibitors: 4-acyl-pyrazolylhydrazone
derivatives. Can. J. Physiol. Pharmacol. 72 (Suppl. 1), 210.
Ribeiro, I.G., da Silva, K.C.M., Parrini, S.C., Miranda, A.L.P., Fraga,
C.A.M., Barreiro, E.J., 1998. Synthesis and antinociceptive properties
Acknowledgements
of
new
structurally
planned
imidazo[1,2-a]pyridine
3-
acylarylhydrazone derivatives. Eur. J. Med. Chem. 33, 225–235.
We are grateful to CNPq (BR, grant [52.0033/96-5),
FUJB (BR) and FAPERJ (BR) for financial support and
fellowships (AGMF, ALPM, CAMF and EJB).
´
Soler, O., 1993. MSc. Thesis, Faculdade de Farmacia, UFRJ, Rio de
Janeiro, Brazil.
Stewart, J.J.P., 1993. MOPAC 7.0, F.J. Seiler Research Lab., US Air
Force Academy, Colorado Springs.
Vargaftig, B.B., Lefort, J., Chignard, M., Benveniste, J., 1980. Platelet
activating factor induces
unrelated to the formation of prostaglandin derivatives. Eur. J.
Pharmacol. 65, 185–192.
a platelet-dependent bronchoconstriction
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