New pyridazinone derivatives with vasorelaxant and platelet antiaggregatory activities

Article abstract of DOI:10.1016/j.bmcl.2010.09.031

New 6-substituted and 2,6-disubstituted pyridazinone derivatives were obtained starting from easily accessible alkyl furans by using oxidation with singlet oxygen to give 4-methoxy or 4-hydroxybutenolides, key intermediates of this synthetic strategy. The

Full text of DOI:10.1016/j.bmcl.2010.09.031

Bioorganic & Medicinal Chemistry Letters 20 (2010) 6624–6627
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
New pyridazinone derivatives with vasorelaxant and platelet
antiaggregatory activities q
Tamara Costas a, Pedro Besada a, Alessandro Piras a, Laura Acevedo b, Matilde Yañez b, Francisco Orallo b,
Reyes Laguna b, Carmen Terán a,
⇑
a Departamento de Química Orgánica, Universidade de Vigo, 36310 Vigo, Spain
b Departamento de Farmacología, Universidade de Santiago de Compostela, 15782 Santiago de Compostela, Spain
a r t i c l e i n f o
a b s t r a c t
Article history:
New 6-substituted and 2,6-disubstituted pyridazinone derivatives were obtained starting from easily
accessible alkyl furans by using oxidation with singlet oxygen to give 4-methoxy or 4-hydroxybuteno-
lides, key intermediates of this synthetic strategy. The new pyridazinone derivatives have been studied
as vasorelaxant and antiplatelet agents. Analysis of biological data revealed the silyl ethers (4a–i) and
N,O-dibenzyl derivatives (6g–i) as the most active compounds.
Received 2 August 2010
Revised 3 September 2010
Accepted 4 September 2010
Available online 15 September 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Pyridazinones
Butenolides
Vasorelaxant
Antiplatelet agents
The pyridazine nucleus represents a versatile scaffold to devel-
op new pharmacologically active compounds. This nitrogen het-
erocycle is included in chemicals with a wide range of biological
activities and can also be used to link other pharmacophoric
groups.1 For instance, the pyridazinone derivatives are known as
drugs with interesting effects in cardiovascular system due to their
platelet aggregation inhibition as well as their antihypertensive
activity and cardiotonic properties (Fig. 1).1e,2–7
In general, the 6-arylpyridazinone structure was considered
essential for the activity on cardiovascular system, especially to
ensure inhibition of some key targets as phosphodiesterase III
(PDE III),8,1e,2 and this can be the reason why most of pyridazinone
derivatives known include aryl residues at position 6. However, in
the last few years new pyridazinone derivatives with activity on
cardiovascular system were described in which the phenyl group
O
O
HN
N
HN
N
CO2Bn
CO2Bn
CH3
O
N
N
OCH3
OCHF2
CN
CN
Levosimendan
Bn
HN
N
Zardaverine
5-Alkylidenepyridazinone
Figure 1. Representative pyridazinone derivatives with activity on cardiovascular
system.
to derivatives with novel mechanisms of action we have designed
and synthesized a preliminary series of derivatives of structure I.
These compounds show two positions for structural diversity, C6
and N2 ( Fig. 2), since both positions seem very important to
modulate the effect of pyridazinone derivatives on cardiovascular
system.10,1e
at C6 was removed or replaced, such as a1-adrenoreceptor antago-
nists or platelet aggregation inhibitors.9,1e,2 Thus, replacement of
aryl by alkenyl fragments or their inclusion in other positions of
pyridazine ring gave rise to potent antiplatelet agents whose activ-
ity is not related with the inhibition of the PDE III.1e,2
Bearing in mind the good properties described by pyridazinone
based compounds for the control of cardiovascular diseases, in
particular as vasodilators and antiplatelet agents and looking for
new modifications on the pyridazinone nucleus that could give rise
OR6
n = 1, 2, 3
O
n
R6 = TBDPS, H, Bn
R2 = H, CH3, Bn
N N
R2
q
I
In memory of Professor Francisco Orallo.
⇑
Corresponding author. Tel.: +34 986812276; fax: +34 986812262.
E-mail address: mcteran@uvigo.es (C. Terán).
Figure 2. General structure of pyridazinone analogues synthesized.
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.09.031

T. Costas et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6624–6627
6625
The new pyridazinone analogues proposed will allow us to ex-
O
Bn
plore how vasorelaxant and antiplatelet activity is affected by the
presence at C6 of an alkyl chain of varying magnitude (1, 2 or 3 car-
bon atoms), functionalized with alcohol or ether groups, as well as
by inclusion of a methyl or a benzyl group at N2. The selection of
substituents at N2 was based on the activity data previously
known (Fig. 2).1e,2
N
N
n = 1-3
4g-i
a
n
O
R
OTBDPS
N
N
O
b
R2
O
R
The pyridazinone derivatives studied in this work were synthe-
sized as outlined in Schemes 1 and 2.
a
N
N
N
N
n
OTBDPS
According to Scheme 1, the pyridazinone core was obtained in
moderate yield in two steps starting from the adequate 2-alkylfu-
ran. Thus, oxidation of alkylfurans 1 with singlet oxygen11 gave an
appropriate butenolide suitable to react with hydrazine or methyl
hydrazine.12 The 2-alkylfuran 1a, precursor of 6-methyl deriva-
tives, was easily prepared by reaction of furfuryl alcohol with
tert-butyldiphenylsilyl chloride (TBDPSCl) and imidazol in DMF13
while its homologues, the alkylfurans 1b and 1c, were obtained fol-
lowing previously described synthetic approaches.11,14
n
n
OH
n = 1-3
4a-c R = H
OR6
n = 1-3
n = 1-3
4d-f R = CH3
5g-i R2 = Bn, R6 = H
5a-c R = H
5d-f R = CH3
6d-f R2 =CH3, R6 = Bn
6g-i R2 = R6 = Bn
Scheme 2. Reagents and conditions: (a) NaH, BnBr, Bu4NI, THF, rt; 68% (4g), 70%
(4h), 61% (4i), 45% (5g), 23% (5h), 49% (5i), 57% (6d), 99% (6e), 87% (6f), 40% (6g),
61% (6h), 30% (6i); (b) TBAF, THF, rt, 86% (5a), 94% (5b), 99% (5c), 99% (5d), 99% (5e),
98% (5f).
Once obtained the 2-alkylfurans 1a–c, two strategies are fol-
lowed to build the heterocyclic ring. The first one involves oxida-
tion of compounds
1 with singlet oxygen and subsequent
treatment with acetic anhydride, to give the 4-methoxybutenolide
intermediates 2 in moderate to good yields (56–89%).15,11,14 The
second one, involves oxidation of 1 with oxygen singlet in the pres-
ence of the Hünig’s base to afford the 4-hydroxybutenolides 3,16
which can be used in the following step without purification.17
However, to our surprise, the latter methodology has not been
adequate to provide the 4-hydroxybutenolide 3a, which has been
obtained in a yield significantly lower than the corresponding
4-methoxybutenolide 2a. These poor results are in contrast with
previous observations and could be related to the possible instabil-
ity of compound 3a.16
above indicated (Scheme 2).21 The acid character of the pyridazi-
none hydrazidic hydrogen makes possible a selective benzylation
at N2 for compounds 5a–c using one equivalent of benzyl bromide.
However, when excess benzyl halide was used, although the
N-benzyl derivative was initially obtained, increased reaction
times led to mixtures of N-benzyl and N,O-dibenzyl derivatives,
in varying proportions, whose separation was possible by column
chromatography.
The new pyridazinone derivatives synthesized, compounds 4–6
as well as the reference drug (milrinone) were screened for vasore-
laxant activity as reported previously,22 using intact rat aortic rings
Both intermediates, compounds 2 and 3, by reaction with
hydrazine or methyl hydrazine in ethanol afforded the 6- and
2,6-substituted pyridazinones respectively (4a–f, 11–45% yields).18
The low yield obtained in some cases could be explained taking
pre-contracted with noradrenaline (NA, 1
The cumulative addition of milrinone and the compounds 4–6
(1–100 M) caused a concentration-dependent relaxation of the
contractions induced by NA. The corresponding IC50 values are
shown in Table 1.
lM).
l
into account the high reactivity of the a
,b-unsaturated system.11,14
It is noteworthy that both types of butenolides are suitable to
Many of synthesized pyridazinones produced vasorelaxation in
provide the desired compounds, although the hydroxy derivatives,
micromolar range (32.5–78.25 lM), being less efficient than milri-
probably due to their equilibrium with the acyclic form (
acid), are more reactive, allowing milder reaction conditions and
better yields, in general.
c
-keto
none. The N,O-dibenzyl derivatives, compounds 6g–i, were the
most active compounds. The following effective compounds are
the silyl ethers 4, in particular the N-benzyl derivative 4i. On the
other hand, all hydroxyl derivatives (5a–i) were inactive, with
Treatment of 6-silyloxy derivatives 4a–c with benzyl bromide
at room temperature in the presence of NaH and Bu4NI catalytic
gave the expected N-benzyl derivatives 4g–i.19 In addition,
compounds 4a–f by treatment with TBAF provided the hydroxy
derivatives 5a–f in very good yields (86–99%),20 which have been
satisfactorily transformed into the N-benzyl-6-hydroxyalkyl
derivatives 5g–i, into the O-benzyl derivatives 6d–f, as well as into
the N,O-dibenzyl derivatives 6g–i using the standard procedure
IC50 values greater than 100 lM.
Platelet aggregation studies of compounds 4–6 were performed
in washed human platelets following the Born’s turbidimetric meth-
od,23 and using thrombine (0.25 U/mL) and collagen (2.5
lg/mL) as
platelet aggregation inductors.22b
Most of new pyridazinones selectively inhibited aggregation in-
duced by collagen in the low
lM range (1.80–69.6 lM), since none
OTBDPS
a
O
O
n
c
OCH3
2a-c
OTBDPS
n
OTBDPS
O
O
n
N N
b
d
R
1a-c
OTBDPS
4a-c R = H, n = 1-3
4d-f R = CH3, n = 1-3
O
O
n
OH
3a-c
Scheme 1. Reagents and conditions: (a) (1) O2, hv, rose bengal, MeOH, À78 °C; (2) Ac2O, Py, DMAP, rt, 56% (2a); (b) O2, hv, rose bengal, DIPEA, MeOH, À78 °C; (c) NH2NH2ÁH2O
or CH3NHNH2, EtOH, reflux, 14% (4a), 41% (4d); (d) NH2NH2ÁH2O or CH3NHNH2, EtOH, 0 °C to rt, 34% (4b), 45% (4c), 11% (4e), 31% (4f), yields for two steps.

6626
T. Costas et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6624–6627
Table 1
Since all tested compounds showed a selective effect towards
Vasorelaxant activity (IC50 in l
M)a of tested compounds
the aggregation induced by collagen, exploratory studies were per-
formed in order to evaluate the TXA2 pathway. In addition, biolog-
ical tests were carried out using purified enzymes according to a
previously reported protocol.27 Thus, preliminary studies per-
formed with compound 4e revealed moderate COX-1 inhibition
(results not shown) that can explain, at least in part, the antiplate-
let effect observed.
Compound
Noradrenaline (NA, 1 lM)
4a
4b
4c
4d
4e
62.61 (5.12)
56.2 (5.6)
78.25 (6.89)
58.4 (4.4)
47.0 (3.1)
>100
4f
In conclusion, 24 new pyridazinone derivatives were obtained
starting from easily accessible alkyl furans by using oxidation with
singlet oxygen to give 4-methoxy or 4-hydroxybutenolides, key
intermediates of this synthetic strategy. The new molecules have
been characterised as vasorelaxant and platelet antiaggregatory
agents. The target compounds could show a pharmacological pro-
file as antiplatelet drugs similar to that of aspirin. Further studies
are in progress to clarify the mechanisms by which the new pyrid-
azinone analogues produce their vasorelaxant and antiplatelet
effects.
4g
4h
4i
5a
5b
5c
5d
5e
5f
>100
>100
39.9 (8.6)
>100
>100
>100
>100
>100
>100
5g
5h
5i
>100
>100
>100
6d
6e
6f
6g
6h
6i
>100
>100
Acknowledgments
76.4 (7.1)
35.3 (5.7)
32.3 (2.5)
32.5 (4.3)
0.122 (0.008)
We acknowledge the Xunta de Galicia (PGIDIT07PXIB, INCI-
TE08ENA314019ES and INCITE09E1R314094ES) and Universidade
de Vigo for financial support. P.B. thanks the Xunta de Galicia for
an Isidro Parga Pondal contract.
Milrinone
a
Values are means of five experiments, standard
deviation is given in parentheses.
References and notes
of them inhibited thrombin-induced aggregation at concentrations
below 100
l
M. This selectivity was also developed by several chal-
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Polenzani, L.; dal Piaz, V. J. Med. Chem. 2009, 52, 7397.
cones,24 some zoanthamine-type alkaloids,25 and also by aspirin,26
used as standard drug for this study. Table 2 shows the antiplatelet
activity data against collagen (IC50 values) of the target compounds
and the reference drug, obtained from experiments conducted at
concentration intervals of 1.25–10
ing on the compound.
Analysis of these last biological data also revealed the silyl
ethers (4a–i) and N,O-dibenzyl derivatives (6g–i) as the most ac-
tive compounds of these series, being more potent than aspirin.
lM and of 25–100 lM, depend-
2. Meyers, C.; Yañez, M.; Elmaatougi, A.; Verhelst, T.; Coelho, A.; Fraiz, N.;
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Chem. Lett. 2003, 13, 171.
Table 2
Antiplatelet activity (IC50 in
l
M)a of tested compounds
Compound
Collagen (2.5 lg/mL)
4a
4b
4c
4d
4e
4f
4g
4h
4i
5a
5b
5c
5d
5e
5f
5g
5h
5i
6d
6e
6f
6g
6h
6i
1.80 (0.11)
3.87 (0.38)
2.13 (0.16)
4.55 (0.30)
3.17 (0.16)
2.34 (0.6)
4.6 (0.4)
4.8 (0.2)
3.9 (0.2)
>100
64.3 (5.2)
>100
69.6 (5.0)
60.4 (3.1)
>100
10. Manetti, F.; Corelli, F.; Strappaghetti, G.; Botta, M. Curr. Med. Chem. 2002, 9,
1303.
11. Pérez, M.; Canoa, P.; Gómez, G.; Terán, C.; Fall, Y. Tetrahedron Lett. 2004, 45,
5207.
12. (a) Escobar, C.; Fariña, F.; Sañudo, J. M. An. Quim. 1971, 67, 43; (b) Bourgignon, J.
J.; Wermuth, C. G. J. Org. Chem. 1981, 46, 4889; (c) Mylari, B. L.; Zembrowski, W.
J.; Beyer, T. A.; Aldinger, C. E.; Siegel, T. W. J. Med. Chem. 1992, 35, 2155.
13. Nicolau, K. C.; Aversa, R. J.; Jin, J.; Rivas, F. J. Am. Chem. Soc. 2010, 132, 6855.
14. Estévez, L.; Besada, P.; Fall, Y.; Teijeira, M.; Terán, C. Synthesis 2010, 425.
15. 5-(tert-Butyldiphenylsilyloxymethyl)-5-methoxy-5H-furan-2-one (2a): 1H NMR
(CDCl3) d (ppm): 7.62 (m, 4H), 7.40 (m, 6H), 7.09 (d, J = 7.5 Hz, 1H), 6.29 (d,
J = 5.7 Hz, 1H), 3.88 (s, 2H), 3.24 (s, 3H), 1.01 (s, 9H). 13C NMR (CDCl3) d (ppm):
169.8, 152.1, 135.6, 132.4, 129.9, 127.8, 126.1, 110.3, 65.2, 51.4, 26.7, 19.2.
16. Teijeira, M.; Suárez, P. L.; Gómez, G.; Terán, C.; Fall, Y. Tetrahedron Lett. 2005,
46, 5889.
>100
>100
4.7 (0.3)b
>100
>100
50.7 (2.3)
8.0 (0.4)
4.11 (0.38)
5.9 (0.5)
38.5 (2.9)
17. All hydroxybutenolides
3
exhibited satisfactory 1H NMR data. Selected
spectroscopic data for compound 3b: 1H NMR (CDCl3) d (ppm): 7.73 (m, 4H),
7.46 (m, 6H), 7.29 (d, J = 5.6 Hz, 1H), 6.09 (d, J = 5.6 Hz, 1H), 4.31 (m, 1H), 3.89
(m, 1H), 2.34 (m, 1H), 1.90 (m, 1H), 1.11 (s, 9H).
Aspirin
a
Values are means of five experiments, standard
deviation is given in parentheses.
18. Spectroscopic data for a representative compound 4b: 1H NMR (CDCl3) d (ppm):
7.58 (m, 4H), 7.40 (m, 6H), 7.18 (d, J = 9.7 Hz, 1H), 6.84 (d, J = 9.7 Hz, 1H), 3.92
b
Active in 50% of the studied plasmas.

T. Costas et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6624–6627
6627
(t, J = 6.1 Hz, 2H), 2.79 (t, J = 6.1 Hz, 2H), 1.00 (s, 9H). 13C NMR (CDCl3) d (ppm):
160.9, 147.0, 135.5, 134.9, 133.2, 129.8, 129.6, 127.7, 62.6, 37.5, 26.8, 19.1.
19. Spectroscopic data for a representative compound 4i: 1H NMR (CDCl3) d (ppm):
7.63 (m, 4H), 7.38 (m, 8H), 7.26 (m, 3H), 6.99 (d, J = 9.5 Hz, 1H), 6.82 (d,
J = 9.5 Hz, 1H), 5.24 (s, 2H), 3.70 (t, J = 6.0 Hz, 2H), 2.68 (t, J = 7.6 Hz, 2H), 1.87
(m, 2H), 1.05 (s, 9H). 13C NMR (CDCl3) d (ppm): 159.7, 147.6, 136.5, 135.5,
133.7, 132.6, 130.1, 129.6, 128.6, 128.5, 127.7, 127.6, 62.6, 55.1, 30.9, 30.8,
26.9, 19.2.
J = 6.2 Hz, 1H). 13C NMR (CDCl3) d (ppm): 159.8, 145.9, 138.0, 136.5, 133.2,
129.8, 128.6, 128.5, 128.4, 127.7, 127.6, 73.1, 68.4, 55.1, 35.1.
22. (a) Orallo, F.; Álvarez, E.; Camiña, M.; Leiro, J. M.; Gómez, E.; Fernández, P. Mol.
Pharmacol. 2002, 61, 294; (b) Vilar, S.; Quezada, E.; Santana, L.; Uriarte, E.;
Yañez, M.; Fraiz, N.; Alcaide, C.; Cano, E.; Orallo, F. Bioorg. Med. Chem. Lett. 2006,
16, 257.
23. Born, G. V. R. Nature 1962, 194, 927.
24. Lin, C. N.; Hsieh, H. K.; Ko, H. H.; Hsu, M. F.; Lin, H. C.; Chang, Y. L.; Chung, M. I.;
Kang, J. J.; Wang, J. P.; Teng, C. M. Drug Dev. Res. 2001, 53, 9.
25. Villar, R. M.; Gil-Longo, J.; Daranas, A. H.; Souto, M. L.; Fernández, J. J.; Peixinho,
S.; Barral, M. A.; Santafé, G.; Rodríguez, J.; Jiménez, C. Bioorg. Med. Chem. 2003,
11, 2301.
26. De Meyers, S. F.; Vanhoorelbeke, K.; Bross, K.; Salles, I. I.; Deckmyn, H. Br. J.
Haematol. 2008, 142, 515.
27. Alcaro, S.; Artese, A.; Botta, M.; Zizzari, A. T.; Orallo, F.; Ortuso, F.; Schenone, S.;
Brullo, C.; Yáñez, M. ChemMedChem 2010, 5, 1242.
20. Spectroscopic data for a representative compound 5d: 1H NMR (CD3OD) d (ppm):
7.55 (d, J = 9.5 Hz, 1H), 6.97 (d, J = 9.5 Hz, 1H), 4.47 (s, 2H), 3.73 (s, 3H). 13C
NMR (CD3OD) d (ppm): 162.6, 149.8, 133.5, 130.3, 63.5, 40.6.
21. Spectroscopic data for representative compound 5h: 1H NMR (CDCl3) d (ppm):
7.42 (m, 2H), 7.34 (m, 3H), 7.11 (d, J = 9.6 Hz, 1H), 6.92 (d, J = 9.6 Hz, 1H), 5.31
(s, 2H), 3.96 (t, J = 5.8 Hz, 2H), 2.84 (t, J = 5.8 Hz, 2H). 13C NMR (CDCl3) d (ppm):
159.6, 146.1, 136.3, 133.0, 130.5, 128.7, 128.8, 60.4, 54.9, 36.8. Compound 6h:
1H NMR (CDCl3) d (ppm): 7.42 (m, 2H), 7.31 (m, 8H), 7.21 (d, J = 9.5 Hz, 1H),
6.89 (d, J = 9.5 Hz, 1H), 5.31 (s, 2H), 4.52 (s, 2H), 3.77 (t, J = 6.2 Hz, 2H), 2.89 (t,