New antithrombotic 1-Phthalazinamines with Serotonin Antagonistic Properties

Article abstract of DOI:10.1002/ardp.200300775

We report nineteen 4-aryl- and 4-arylalkyl-1-phthalazinamines (5-8) which we prepared and tested for antithrombotic properties. All compounds were assayed for their antiplatelet activity in the "Born test" with collagen as inducer of the aggregation. N-[4-(1H-1,2,4-triazol-1-yl)butyl]-4-phenyl-1-phthalazin-amine (7c) was the most potent compound, having an IC₅₀ of 8 μM. When 5-HT (Serotonin) was used to start aggregation the N-(furan-2-yl-methyl)-4-phenyl-1-pthtalazinamine (8a) had an IC₅₀ of 2 μM. In vivo potencies were highly significant. N-[5-(1H-1,2,4-triazol-1-yl)pentyl]-4-phenyl-1-phthalazinamine (7d) inhibited thrombus formation by 12% (P < 0.002) in arterioles and 7% (P < 0.01) in venoles as tested with our laser thrombosis model. For compound 8 a we surprisingly found an antagonism to the 5HT_2A receptor, which is most likely the mechanism of the inhibition of aggregation by this compound.

Full text of DOI:10.1002/ardp.200300775

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597 Antithrombotic 1-Phthalazinamines with Serotonin Antagonistic Properties 591
Matthias Johnsena,*,
New antithrombotic 1-Phthalazinamines with
Serotonin Antagonistic Properties
Klaus Rehsea,
Heinz Pertza,
Johannes Peter Staschb,
Erwin Bischoffb
We report nineteen 4-aryl- and 4-arylalkyl-1-phthalazinamines (5–8) which we pre-
pared and tested for antithrombotic properties. All compounds were assayed for
their antiplatelet activity in the “Born test” with collagen as inducer of the aggrega-
tion. N-[4-(1H-1,2,4-triazol-1-yl)butyl]-4-phenyl-1-phthalazin-amine (7 c) was the
most potent compound, having an IC50 of 8 µM.When 5-HT (Serotonin) was used to
start aggregation the N-(furan-2-yl-methyl)-4-phenyl-1-pthtalazinamine (8 a) had
an IC50 of 2 µM. In vivo potencies were highly significant. N-[5-(1H-1,2,4-triazol-1-
yl)pentyl]-4-phenyl-1-phthalazinamine (7 d) inhibited thrombus formation by 12 %
(P < 0.002) in arterioles and 7 % (P < 0.01) in venoles as tested with our laser throm-
bosis model. For compound 8 a we surprisingly found an antagonism to the 5HT2A
receptor, which is most likely the mechanism of the inhibition of aggregation by this
compound.
a
Institut für Pharmazie,
Freie Universität Berlin,
Berlin, Germany
Institute of Cardiovascular
b
Research,
Bayer AG, Wuppertal,
Germany
Keywords: 1-Phthalazinamine; Antithrombotic properties; Born test; Laser throm-
bosis model; Serotonin antagonism
Received: January 23, 2003; Accepted March 4, 2003 [FP775]
DOI 10.1002/ardp.200300775
Introduction
In 1984, Ko et al.[1] described 3-(5-hydroxymethyl-2-fur-
yl)-1-phenylmethyl-indazole (YC-1) as the first drug
stimulating the soluble guanylatecyclase (sGC) inde-
pendent of NO. Activation of this enzyme in thrombo-
cytes leads via an increase in the concentration of cyclic
guanosinemonophosphate (cGMP) to a reduced re-
sponse to aggregation inducing agents. Later it was
found [2], that YC-1 is also capable of inhibiting phos-
phodiesterases, which degrade cGMP to guanosine-
monophosphate (GMP).YC-1 shows no specifity for any
isoform of the enzyme.In screening the literature for sub-
stances which are capable of activating sGC as well as
inhibiting PDE 5 we found an inhibitor of PDE 5 which
shows a certain degree of similarity toYC-1.These simi-
larities e.g., an electron-rich heterocyclus and lipophilic
moieties of YC-1 and 1-(3-chloroanilino)-4-phenyl-
phthalazine (MY 5445) prompted us to synthesize fur-
ther phthalazines on that basis to get compounds with
activities to the enzymes above (Figure 1).
Figure 1. Similarities of YC-1 and MY 5445.
Chemistry
N-substituted 1-phthalazinamines were prepared ac-
cording to Figure 2.
Subsequently, we report a series of 4-(aryl)- and 4-(aryl-
alkyl)-1-phthalazinamines with new substituents at the
exocyclic nitrogen.
Common synthons for the 4-aryl- and 4-arylalkyl-1-
phthalazineamines 5–8 are the 4-aryl- and 4-arylalkyl-
phthalazin-1(2H)-ones 2.These were prepared by heat-
ing an ethanolic solution of either 2-arylcarbonyl-ben-
* Part of the M. Johnsons PhD thesis, FU Berlin, Berlin (Germany)
2002.
zoic acids
1 or 3-(aryl-methylene)-1(3H)-benzo[c]-
furanones 3 with one equivalent of hydrazine hydrate.
Treatment of the phthalazinones 2 with phosphorous
oxichloride yields the corresponding 1-chloro-4-aryl-
and 1-chloro-4-arylmethyl-phthalazines 4.These can be
Correspondence: Klaus Rehse, Institut für Pharmazie, Freie
Universität Berlin, Königin-Luise-Straße 2+4, D-14195 Berlin,
Germany.
Phone:
+49
30
838-53290/64,
Fax:
+ 49 30 838-53251; e-mail: Rehiwer@zedat.fu-berlin.de
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

592 Johnsen et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597
Figure 2. Synthetic route to 1-phthalazineamines.
reacted with primary amines [3] to give the desired N-
substituted 1-phthalazinamines 5.
model [7]. The compounds were administered orally to
male Wistar rats (60 mg/kg). After 2 h the formation of
thrombi in mesenteric vessels was induced by the beam
of an argon laser through a microscope (35 mW, 50 ms).
The number of exposures (“shots”) necessary to form a
thrombus of defined size is counted. From the average
number of shots the percentage of inhibition of thrombo-
sis is calculated [8].
The 2-arylcarbonyl-benzoic acids were either commer-
cially available or prepared by Friedel-Crafts acylation of
phthalic anhydride.Compounds of type 3 were prepared
by a method described by Gabriel [4].
Biology
Born test
Inhibition of phosphodiesterases
All compounds were tested for antiaggregative proper-
ties with a modified [5] Born test [6].For that purpose, the
substance was dissolved in a small quantity of DMSO
and added to approximately 100 µl of freshly prepared
human platelet rich plasma (PRP). This stock solution
was added in calculated volumes to PRP, giving a final
volume of 220 µl of defined concentrations of the test
compound. Aggregation was induced using either colla-
gen or serotonin (5-HT). The IC50 values were deter-
mined from a semilogarithmic plot of the concentration
versus the percentage of inhibition of the aggregation.
According to their low IC50 values compounds 6 b, 6 c,
8 a, and 8 d were tested for their inhibitory action on sev-
eral isoforms of the enzyme by the Institute of Cardiovas-
cular Research of the Bayer AG, Wuppertal [9]. Briefly, a
3H-cAMP or a 3H-cGMP-SPA-Assay from Amersham
was used observing the instructions of the manufacturer
(Amersham, Braunschweig, Germany). The isoforms
tested were PDE 1, PDE 2, PDE 3, PDE 5, and PDE 9.All
enzymes, except PDE 9 which was of murine origin,
were recombinant human enzymes expressed in a bac-
ulovirus SF9 system.All determinations were performed
in duplicate and were repeated at least three times.The
IC50-values were determined from sigmoidal curves, fit-
ted to plots of enzyme activity versus log compound con-
centration using a GraphPad (San Diego, CA, USA)
curve fitting program.
Laser thrombosis model
The influence of selected test compounds on the forma-
tion of thrombi in vivo was assayed in a laser thrombosis
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597 Antithrombotic 1-Phthalazinamines with Serotonin Antagonistic Properties 593
Activation of sGC
Results and discussion
Compounds 6 b, 6 c, 8 a, and 8 d were also tested for
their activation of human recombinant sGC.The test was
performed at the Institute of Cardiovascular Research of
the Bayer AG, Wuppertal [10]. Briefly, the activity of the
enzyme was measured by determining the yield of
[α-32P]-cGMP 10 min after incubation with the test com-
pound.The test compound was added to a final concen-
tration of 10 or 100 µM.To investigate the mechanism of
the sGC activation a second determination was per-
formed in the presence of 0.1 µM of the NO releasing
agent 2-(N,N-diethylamino)-diazenolate-2-oxide (DEA/
NO).
The results of the determination of the antiaggregative
properties in vitro and the inhibition of thrombus forma-
tion in vivo are summarized in Table 1.
Inhibition of platelet aggregation
a) Collagen as inducing agent
All compounds were tested with collagen as aggregation
starter.
In type 5 compounds the 4-phenyl derivatives 5 a and 5 b
are more effective than the benzyl derivatives 5 c and 5 d
Changing the basic center in the substituent in 4-position
from dimethylamino (pKB ~ 5) to 1-imidazolyl (pKB ~ 7)
the type 6 compounds are obtained. Here again, the
4-phenyl derivatives 6 a–d have a lower IC50 than the
corresponding benzyl compounds 6 e or 6 f, respective-
ly.In total the less basic type 6 compounds are more effi-
cient than the type 5 compoundsThe lowest IC50 = 18 µM
is found for 6 c Interestingly, an obvious dependence of
Antagonism at the 5-HT2A receptor
Compound 8 a was tested for antagonistic action at the
5-HT2A receptor in a rat tail artery model [11].The effect
of 8 a on the contractive response of endothelium denud-
ed caudal artery cylinders to 5-HT was measured as
given in ref. 11.
Table 1. Antiaggregative properties (inducer collagen or serotonin) of type 5–8 compounds and acetylsalicylic acid
(asa) (general formula, see Figure) in vitro and inhibition of thrombus formation in vivo.– not tested.n.s.not significant;
1)
2)
3)
P Յ 0.002 P Յ 0.01 P Յ 0.05.
Nr
n
m
R
RЈ
IC50
(Coll)
[µM]
IC50
(5-HT)
[µM]
Inhibition of thrombus formation
(%)
arterioles
venoles
5 a
5 b
5 c
5 d
6 a
6 b
6 c
6 d
6 e
6 f
7 a
7 b
7 c
7 d
7 e
8 a
8 b
8 c
8 d
asa
0
0
1
1
0
0
0
0
1
1
0
0
0
0
1
0
0
0
1
–
2
3
2
3
2
3
4
5
2
3
2
3
4
5
3
1
1
1
1
–
Dimethylamino
Dimethylamino
Dimethylamino
Dimethylamino
Imidazol-1-yl
Imidazol-1-yl
Imidazol-1-yl
Imidazol-1-yl
Imidazol-1-yl
Imidazol-1-yl
1,2,4-Triazol-1-yl
1,2,4-Triazol-1-yl
1,2,4-Triazol-1-yl
1,2,4-Triazol-1-yl
1,2,4-Triazol-1-yl
Furan-2-yl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
76
100
160
160
80
45
18
24
170
70
140
60
8
31
210
28
51
51
68
–
–
–
–
–
12
5
37
–
15
–
–
12
–
–
2
–
7
6
22)
23)
n.s.
n.s.
–
–
n.s.
12)
12)
n.s
n.s.
7
7
4
7
13)
n.s.
n.s.
n.s.
–
n.s.
–
9
22)
n.s.
n.s.
10 22)
12 11)
12)
–
n.s.
n.s.
Furan-2-yl
Furan-2-yl
Furan-2-yl
–
Cl
OCH3
H
–
–
–
–
17
n.s.
n.s.
48 101)
20 51)
2)
3)
– not tested; n.s. not significant; 1) P Յ 0.002 P Յ 0.01 P Յ 0.05.
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

594 Johnsen et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597
the IC50 from the chainlength in the imidazolylalkyl sub-
stituent is observed. The peak efficacy is found for the
imidazolylbutyl derivative 6 c, while shorter or longer
alkyl chains show higher IC50 values indicating a smaller
antiplatelet effect (see 6 a–d). The same is true for the
even less basic 1,2,4-triazol-1-ylalkyl derivatives of the
type 7.Here again, the 4-phenyl derivative 7 b is more ef-
fective than the benzyl compound 7 e.Again the triazolyl-
butyl compound 7 c has the lowest IC50 i.e. 8 µM. Supris-
ingly the neutral furanylmethyl derivative 8 a showed a
remarkable antiplatelet activity. Comparison of 8 b–8 d
shows that the electronic properties of R' do not in-
fluence this activity.
Activation of sGC
Compound 8 d was the only one showing an effect on the
activity of sGC. When present in a concentration of
10 µM the activity of the enzyme was increased 2-fold.In
a concentration of 100 µM the increase was 5-fold.In the
presence of 0.1 µM DEA/NO the activity of the enzyme
was increased by factor 47.When, in addition, 100 µM of
8 d are present the enzyme shows a 83-fold increase of
the basal activity.
Using the same conditions 100 µM ofYC-1 lead to a 93-
fold activation of sGC, increased to a 550-fold activation
when further 0.1 µM DEA/NO are present.
In conclusion, the antiplatelet activity of 8 d should not be
due to the small activation of sGC.
b) Serotonin as inducing agent
For the further eludication of the mechanism of the an-
tiaggreggating activities other inducers of platelet aggre-
gation – especially serotonin – were used for the most
active compounds.The IC50 values observed are mostly
lower than those found when collagen was used to in-
duce aggregation.This is most evident with 6 c (IC50 = 18
versus 5 µM) and 8 a (IC50 = 28 versus 2 µM). This
prompted us to investigate the latter compound in an in
vitro 5-HT2A receptor model (see below.).
Antagonism of 8 a at the 5-HT2A receptor
When present in a concentration of 10 µM 8 a shows a
significant shift of the concentration/effect-curve to the
right (Figure 3).Using the formula of Furchgott [12] pA2 =
5.78 0.08. As there is also a depression of the maxi-
mum effect of 5-HT to 70 % the antagonism is non-com-
petitive. The combination of the shift of the concentra-
tion-effect-curve to the right and depression of the maxi-
mum effect resembles the mechanism of ritanserin,
which is an insurmountable antagonist of the 5-HT2A re-
ceptor [11].
Antithrombotic properties
The antithrombotic properties obtained from an in vivo
thrombosis model are summarized in the last two col-
umns of Table 1. In all types of compounds there are ob-
served small antithrombotic properties. It is obvious
however that in type 6 and type 7 derivatives with low
IC50 values also show antithrombotic properties. The
peak effects are seen in 7 b, 7 c, and 7 d. On the other
hand, in the type 8 compounds 8 a and 8 d such a corre-
lation is missing.Whether this difference has pharmaco-
dynamic (different mechanism) or pharmacokinetic rea-
sons cannot be decided presently. Compared to acetyl-
salicylic acid (asa) which inhibits thrombus formation un-
der the same conditions by 48 10 % (P Յ 0.002) in arte-
rioles and 20 5 % (P Յ 0.002) in venoles, these effects
are rather low.
The concentration in which a remarkable 5-HT antago-
nizing effect is found is nearly the same as the IC50 of
this compound for the inhibition of 5-HT-induced platelet
aggregation in vitro. This strongly indicates that com-
Inhibition of phosphodiesterases
Only the furan-2-ylmethyl derivatives 8 a and 8 d inhibit
PDE 5 with an IC50 of 10 µM. The IC50 for the other iso-
forms of the enzyme were out of the range of determina-
tion. Compounds 6 b and 6 c did not inhibit any of these
enzymes. We conclude that type 8 and type 6 com-
Figure 3. Concentration/effect curves of 5-HT on the
contraction of rat arterial smooth muscle in absence
(open circles) or presence of 0.3 µM (triangle up),1 µM
(triangle down), or 1 µM (filled circles) of 8 a. For further
detail see reference [11].
pounds have
a different profile concerning their
antiplatelet effect.
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597 Antithrombotic 1-Phthalazinamines with Serotonin Antagonistic Properties 595
pound 8 a inhibits platelet aggregation via a 5-HT2A re-
ceptor antagonism.
53 °C, yield 0.40 g (70 %). – Anal. C19H22N4 (306.413). –
1H-NMR / 400 MHz (CDCl3): δ (ppm) = 2.36 (s, 6 H, N(CH3)2),
2.74 (t, J = 5.7 Hz, 2 H, CH2CH2NMe2), 3.79 (m, 2 H,
HNCH2CH2NMe2), 4.55 (s, 2 H, CH2Ph), 6.16 (br. s, 1 H, D2O
exchange, NH), 7.14–7.18 (m, 1H, ph-4H), 7.22-7.34 (m, 4H,
Experimental
ph-2H-3H-5H-6H), 7.65-7.74 (m, 2H, ar-6H-7H), 7.87-7.92 (m,
Chemistry
᭹+
2H, ar-5H-8H). – MS (70 eV, 50 °C): m/z (%) = 248 (20) [M
-
H2CNMe2], 236 (10) [M -(CH2)2NMe2 + 2 H], 91 (13) [C7H+7], 71
᭹+
The full set of data is given in the PhD thesis of M. Johnsen,
Freie Universität Berlin, 2002.
(71) [H2C=CHNMe+2], 58 (100) [H2C=NMe+2].
Mp (uncorr.), Linström. – Elemental analysis: Elementar vario
EL. – NMR: Bruker DPX 400 (Bruker, Rheinstetten, Germany).
– MS:CH-7A-Varian-MAT (70 eV) (Varian, Braunschweig, Ger-
many).
N,N-Dimethyl-NЈ-(4-phenylmethyl-phthalazin-1-yl)-propane-
1,3-diamine (5 d)
From 0.55g (2.2 mmol) 4 e and 2.4 g N1,N1-dimethyl-1,3-pro-
panediamine, 3 h at 120 °C. Yellowish crystals (diethylether/
acetone), mp 48–50 °C, yield 0.49 g (70 %). – Anal C20H24N4
(320.440). – 1H-NMR / 400 MHz (CDCl3): δ (ppm) = 1.93 (tt, J =
5.8/5.8 Hz, 2 H, HNCH2CH2CH2NMe2), 2.37 (s, 6 H, N(CH3)2),
2.59 (t, J = 5.8 Hz, 2 H, HNCH2CH2CH2NMe2), 3.82 (t, J =
5.8 Hz, 2 H, HNCH2CH2CH2NMe2), 4.53 (s, 2 H, CH2Ph),
7.14–7.17 (m, 1 H, Ph-4H), 7.22–7.34 (m, 4 H, ph-2H-3H-5H-
6H)), 7.62–7.68 (m, 2 H, ar-6H-7H), 7.70 (d, 3J = 8.3 Hz, 1 H, ar-
5H), 7.78 (br. s, 1 H, D2O exchange, NH), 7.85 (d, 3J = 8.2 Hz,
The synthesis of 5 b has been reported [3] previously. All other
compounds were prepared for the first time.
General procedure for the synthesis of type 5–8 compounds
Approx. 0.5 g of the 1-chloro- compound 4 were dissolved in
2–4 ml of the corresponding amine and heated to 130 °C for
1–3 h until no chloro compound was detectable by TLC any-
more. After cooling to 40 °C the mixture was dissolved in 5 ml
methanol and then added dropwise to 100 ml of 1/2-conc brine.
In case a substance precipitated, it was collected, washed with
brine, and recrystallized form the solvent(s) stated. If the prod-
uct did not crystallize it was extracted with ethylacetate several
times, the combined extracts were washed with water, dried
over Na2SO4 and the solvent removed in vacuo.The residue was
recrystallized from the solvent(s) stated. If no crystalline product
was obtainable in this fashion, the compound was dissolved in
ethylacetate and the hydrochloride was precipitated by bubbling
gaseous HCl through the solution. The hydrochlorides were re-
crystallized from methanol unless otherwise stated.
᭹
+
1 H, ar-8H).– MS (70 eV, 130 °C):m/z (%) = 320 (11) [M ], 276
᭹
+
᭹
+
(7) [M᭹+-NMe2], 262 (35) [M -CH2NMe2], 248 (100) [M
-
(CH2)2NMe2], 236 (12) [M -(CH2)3NMe2 + 2 H], 91 (26) [C7H+7],
᭹
+
85 (24) [(CH2)2NMe+2], 58 (53) [H2C=NMe+2].
N-[2-(Imidazol-1-yl)ethyl]-4-phenyl-1-phthalazinamine (6 a)
From 0.5 g (2.1 mmol) 4 a and 1.3 g 2-imidazol-1-yl-ethylamine,
1.5 h at 130 °C. Brownish crystals, mp 164–166 °C, yield 0.55 g
(80 %). – Anal. C19H17N5 (315.380). – 1H-NMR / 400 MHz
(CDCl3): δ (ppm) = 4.10 (t, J = 5.2 Hz, 2 H, HNCH2CH2N-im),
4.48 (t, J = 5.2 Hz, 2 H, HNCH2CH2N-im), 5.66 (br. s, D2O ex-
change, 1 H, NH), 6.98 (s, 1 H, im-5H), 7.07 (s, 1 H, im-4H),
7.50–7.56 (m, 4 H, ph-3H-4H-5H + im-2H) 7.66–7.72 (m, 2 H,
ph-2H-6H), 7.75–7.79 (m, 3 H, ar-5H-6H-7H), 7.99–8.01 (m,
1 H, ar-8H). – MS (70 eV, 180 °C): m/z (%) = 315 (9) [M᭹+], 234
N,N-Dimethyl-NЈ-(4-phenyl-phthalazin-1-yl)-ethane-1,2-di-
amine dihydrochloride (5 a)
From 0.7 g (2.9 mmol) 4 a and 2.5 ml N1,N1-dimethyl-1,2-ethan-
ediamine, 1.5 h at 130 °C. Crystals (MeOH), mp 245 °C, yield
0.70 g (65 %). – Anal. C18H22N4Cl2 (365.302) – 1H-NMR /
400 MHz (CD3OD):δ (ppm) = 3.05 (s, 6 H, N(CH3)2), 3.63 (t, J =
5.9 Hz, 2 H, HNCH2CH2NMe2), 4.11 (t, J = 5.9 Hz, 2 H,
HNCH2CH2NMe2), 7.72–7.78 (m, 5 H, ph-2H-6H), 8.15–8.30
(m, 3 H, ar-5H-6H-7H), 8.66 (d, J = 8.2 Hz, 1 H, ar-8H).– MS (70
᭹
+
᭹
+
(23) [M -CH2-im], 221 (100) [M -(CH2)2-im], 81 (5) [CH2-
im+], 77 (40).
N-[3-(Imidazol-1-yl)propyl]-4-phenyl-1-phthalazinamine (6 b)
From 0.8 g (3.3 mmol) 4 a and 3 ml 3-imidazol-1-yl-pro-
pylamine, 1.5 h at 130 °C. Yellowish crystals, mp 188–190 °C,
yield 0.68 g (60 %). – Anal. C20H19N5 (329.407). – 1H-NMR /
400 MHz (CDCl3/D2O): δ (ppm) = 2.35 (tt, J = 6.8 /6.8 Hz, 2 H,
HNCH2CH2CH2im), 3.80 (t, J = 6.7 Hz, 2 H, HNCH2(CH2)2im),
4.15 (t, J = 6.9 Hz, 2 H, HN(CH2)2CH2Im), 7.01 (s, 1 H, im-5H),
7.08 (s, 1 H, im-4H), 7.46–7.54 (m, 4H, ph-3H-4H-5H + im-2H),
7.68-7.71 (m, 2 H, ph-2H-6H), 7.73–7.82 (m, 3 H, ar-5H-6H-
7H), 7.98 (d, 3J = 8.0 Hz, 1 H, ar-8H). – MS (70 eV, 50 °C): m/z
(%) = 329 (13) [M+᭹], 261 (58) [M+᭹imH], 248 (60) [M+᭹-CH2-im],
eV, 175 °C): m/z (%) = 246 (2) [M+᭹-HNMe2], 234 (18) [M+᭹
CH2NMe2], 71 (57) [CH2CH2NMe+2], 58 (100) [H2C=NMe2+].
-
N,N-Dimethyl-NЈ-(4-phenyl-phthalazin-1-yl)-propane-1,3-di-
amine dihydrochloride monohydrate (5 b)
From 0.75 g (3.1 mmol) 4 a and 2 ml N1,N1-dimethyl-1,3-pro-
panediamine, 1.5 h at 130 °C. Crystals (EtOH/H2O), mp 238–
242 °C, yield 0.55 g (45 %). – Anal. C19H22N4 · 2 HCl · H2O
(397.344). – 1H-NMR / 400 MHz (CD3OD):δ (ppm) = 2.27–2.35
(m, 2 H, NCH2CH2CH2NMe2), 2.95 (s, 6 H, N(CH3)2), 3.35 (t, J =
7.8 Hz, 2 H, NCH2CH2CH2NMe2), 3.82 (t, J = 6.8 Hz, 2 H,
NCH2CH2CH2NMe2), 7.66–7.69 (m, 5 H, phH), 8.08 (d, J =
7.7 Hz, ar-5H), 8.14 (ddd, J = 8.1/8.0/0.9 Hz, 1 H, ar-6H or
-7H), 8.20 (dd, J = 8.1/7.4 Hz, 1 H, ar-6H or -7H), 8.68 (d, J =
7.8 Hz, 1 H, arH-8). – MS (70 eV, 200 °C): m/z (%) = 306 (13)
235 (100) [M+᭹-CH2CH2-im + H], 234 (79), 220 (47) [M+᭹
(CH2)3-im], 165 (25), 81 (11) [CH2-im+], 77 (61) [C6H5+].
-
N-[4-(Imidazol-1-yl)butyl]-4-phenyl-1-phthalazinamine (6c)
From 0.65 g (2.7 mmol) 4a and 0.8 g 4-imidazol-1-yl-
butylamine. 2.5 h at 130 °C. Yellowish crystals (ethylacetate),
mp 134-135 °C, yield 0.38 g (40 %). - Anal. C21H21N5
(343.434). – 1H-NMR / 400 MHz (DMSO-D6): d (ppm) = 1.67
(tt, J = 6.9/7.5 Hz, 2H, NCH2CH2(CH2)2im), 1.84 (tt, J = 7.5/6.9
Hz, 2H, N(CH2)2CH2CH2im), 3.61-3.66 (m, 2H,
NCH2(CH2)3im), 4.04 (t, J = 7.9 Hz, 2H, N(CH2)3CH2im), 6.87
(s, 1H, im-5H), 7.19 (s, 1H, im-4H), 7.48-7.62 (m, 5H, phH),
7.64 (s, 1H, im-2H), 7.78 (d, J = 8.1 Hz, 1H, ar-5H), 7.82–7.91
(m, 2 H, ar-6-7 H), 8.34 (d, J = 8.1 Hz, 1 H, ar-8H). – MS (70 eV,
50 °C): m/z (%) = 343 (8) [M+᭹], 274 (64) [M+᭹-imH], 262 (38)
[M+᭹], 261 (28) [M᭹+-HNMe2], 248 (54) [M -CH2NMe2], 235
᭹
+
᭹
+
(100) [M -CH2CH2NMe2 + H], 234 (68), 205 (35), 165 (12), 84
(36), 58 (95) [H2C=NMe+2].
N,N-Dimethyl-NЈ-(4-phenylmethyl-phthalazin-1-yl)-ethane-1,2-
diamine (5 c)
From 0.5 g (1.9 mmol) 4 e and 2.5 g N1,N1-dimethyl-ethane-1,2-
diamine, 3 h at 120 °C. Yellowish crystals (acetone/H2O), mp
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

596 Johnsen et al.
Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597
[M+᭹-CH2-im], 248 (100) [M+᭹-CH2CH2-im], 234 (41) [M+᭹
-
N-[3-(1H-1,2,4-Triazol-1-yl)propyl]-4-phenyl-1-phthalazinamine
(7 b)
(CH2)3-im], 220 (88) [M+᭹-(CH2)4-im], 190 (17), 165 (31), 122
(40), 82 (25) [CH2im+ + H], 77 (70) [C6H5+], 68 (19) [imH+ + H].
From 0.45 g (1.9 mmol) 4 a and 0.9 g 3-triazol-1-yl-pro-
pylamine, 2.5 h at 120 °C.Yellow crystals, mp 123–124 °C, yield
0.47 g (75 %).– Anal.C19H18N6 (330.395).– 1H-NMR / 400 MHz
N-[5-(Imidazol-1-yl)pentyl]-4-phenyl-1-phthalazinamine (6 d)
From 0.5
g (2.1 mmol) 4 a and 0.9 g 5-imidazol-1-yl-
(CDCl3): δ (ppm)
= 2.45 (tt, J = 6.4 Hz/6.3 Hz, 2 H,
pentylamine, 2.5 h at 130 °C, opalising crystals (acetone/n-
hexane), mp 128–129 °C, yield 0.34 g (45 %), – Anal. C22H23N5
(357.461). – 1H-NMR / 400 MHz (CDCl3): δ (ppm) = 1.42–1.50
(m, 2 H, HN(CH2)2CH2(CH2)2-Im), 1.82–1.92 (m, 4 H,
HNCH2CH2CH2CH2CH2-im), 3.76–3.81 (m, 2 H, NCH2(CH2)4-
im), 3.97 (t, J = 7.0 Hz, 2 H, N(CH2)4CH2-im), 5.61 (br. s, D2O
exchange, 1 H, NH), 6.91 (s, 1 H, Im-5H), 7.04 (s, 1 H, Im-4H),
7.46–7.54 (m, 4 H, Ph-3H-4H-5H + im-2H), 7.68–7.70 (m, 2 H,
ph-2H-6H), 7.73–7.83 (m, 2 H, ar-6H-7H), 7.92–7.99 (m, 2 H,
ar-5H-8H). – MS (70 eV, 120 °C): m/z (%) = 357 (32) [M+᭹], 276
HNCH2CH2CH2tri), 3.88–3.90 (m, 2 H, HNCH2(CH2)2tri), 4.39
(t, J = 6.5 Hz, 2 H, HN(CH2)2CH2tri), 7.47–7.54 (m, 3 H, ph-3H-
4H-5H), 7.67–7.70 (m, 3 H, Ph-2H-6H + ar-5H), 7.77–7.81 (m,
1 H, ar-6H or -7H), 7.84–7.88 (m, 1 H, ar-6H or -7H), 7.98 (d, J =
8.1 Hz, 1 H, ar-8H(, 8.02 *s, 1 H, tri-5H), 8.06 (br. s, D2O ex-
change, 1 H, NH), 817 (s, 1 H, tri-3H). MS (70 eV, 180 °C): m/z
(%) = 330 (18) [M+᭹], 261 (6) [M+᭹-tri], 248 (100) [M+᭹-CH2-tri],
234 (29) [M+᭹-CH2CH2-tri], 220 (22) [M+᭹-(CH2)3-tri], 205 (17),
165 (10), 82 (2) [CH2-tri ] 77 (22) [C6H+5].
+
(25) [M+᭹-CH2-im], 262 (11) [M+᭹-(CH2)2-im], 248 (48) [M+᭹
-
N-[4-(1H-1,2,4-Triazol-1-yl)butyl]-4-phenyl-1-phthalazinamine
(7 c)
(CH2)3-im], 234 (100) [M+᭹-(CH2)4-im], 220 (67) [M+᭹-(CH2)5-
im], 205 (34), 165 (25), 137 (48), 124 (82), 96 (15) [(CH2)2-im+ +
H], 77 (55) [C6H+5].
From 0.4 g (1.7 mmol) 4 a and 0.7 g 4-triazol-1-yl-butylamine,
2 h at 120 °C. Yellow crystals (acetone/n-hexane); (acetone/
H2O), mp 101–103 °C, yield 0.14 g (25 %). – Anal C20H20N6
(344.422). – 1H-NMR / 400 MHz (CDCl3): δ (ppm) = 1.92 (tt, J =
7.0/6.8 Hz, 2 H, HN(CH2)2CH2CH2tri), 2.07 (tt, J = 6.8/6.7 Hz,
2 H, HNCH2CH2(CH2)2tri), 3.79 (m, 2 H, HNCH2 (CH2)3tri), 4.36
(t, J = 6.7 Hz, 2 H, HN(CH2)3CH2tri), 7.47–7.55 (m, 3 H, Ph-3H-
4H-5H), 7.68–7.70 (m, 2 H, Ph-2H-6H), 7.74–7.84 (m, 2 H, ar-
6H-7H), 7.97–7.99 (m, 3 H, ar-5H-8H + tri-3H), 8.14 (s, 1 H, tri-
5H). MS (70 eV, 200 °C): m/z (%) = 344 (26) [M+᭹], 262 (34)
[M+᭹-CH2-tri], 248 (97) [M+᭹-(CH2)2-tri], 234 (67) [M+᭹-(CH2)3-
tri], 221 (100) [M+᭹-(CH2)4-tri + H], 220 (82), 207 (62), 205 (48),
165 (37), 77 (60) [C6H+5].
N-[2-(Imidazol-1-yl)ethyl]-4-phenylmethyl-1-phthalazinamine
(6 e)
From 0.6 g (2.3 mmol) 4 e and 1.3 g 2-imidazol-1-yl-ethylamine,
3 h at 130 °C.Yellow crystals, mp 55 °C, yield 0.41 g (55 %), –
Anal. C20H19N5 (329.407). – 1H-NMR / 400 MHz (CDCl3) δ
(ppm) = 4.10 (t, J = 5.3 Hz, 2 H, HNCH2CH2im), 4.49 (t, J =
5.3 Hz, 2 H, HNCH2CH2im), 4.57 (s, 2 H, CH2Ph), 5.80 (br. s,
D2O exchange, 1 H, NH), 6.99 (s, 1 H, Im-5), 7.07 (s, 1 H, Im-
4H), 7.17–7.21 (m, 1 H, Ph-4H), 7.25–7.34 (m, 4 H, Ph-2H-3H-
5H-6H), 7.64 (s, 1HH, Im-2H), 7.69–7.73 (m, 2 H, ar-6H-7H),
7.81–7.83 (m, 1 H, ar-5H), 7.92–7.95 (m, 1 H, ar-8H). – MS
(70 eV, 50 °C): m/z (%) = 329 (18) [M+᭹], 261 (12) [M+᭹-im],248
(27) [M+᭹ -CH2im], 234 (100) [M+᭹ -(CH2)2im], 190 (3), 165 (2),
129 (6), 95 (31) [CH2im+], 91 (46) [C7H7+].
N-[5-(1H-1,2,4-Triazol-1-yl)pentyl]-4-phenyl-1-phthalazinamine
(7 d)
From 0.5 g (2.1 mmol) 4 a and 1.3 g 5-triazol-1-yl-pentylamine,
2 h at 120 °C.Yellow crystals (acetone/H2O), mp 117–120 °C,
yield 0.44 g (60 %). – Anal. C21H22N6 (358,449). – 1H-NMR /
400 MHz (CDCl3) δ (ppm) = 1.48 (tt, J = 7.3/7.5 Hz, 2 H,
N-[3-(Imidazol-1-yl)propyl]-4-phenylmethyl-1-phthalazinamine
(6 f)
From 0.7 g (2.7 mmol) 4 e and 4 g 3-imidazol-1-yl-propylamine,
4 h at 130 °C. Crystals, mp 180–181 °C, yield 0.68 g (75 %). –
Anal. C21H21N5 (343.434). - 1H-NMR / 400 MHz (CDCl3): δ
(ppm) = 2.34 (tt, J = 6.7 Hz/6.8 Hz, 2 H, HNCH2CH2CH2im),
3.76 (m , 2 H, HNCH2(CH2)2im), 4.14 (t, J = 6.9 Hz, 2 H,
HN(CH2)CH2im), 4.55 (s, 2 H, CH2Ph), 536 (br. s, D2O ex-
change, 1 H, NH), 7.00 (s, 1 H, im-5H), 7.08 (s, 1 H, im-4H),
7.15–7.19 (m, 1 H, ph-4H), 7.23–7.27 (m, 2 H, ph-3H-5H),
7.31–7.34 (m, 2 H, ph-2H-6H), 7.57 (s, 1 H, Im-2H), 7.66–7.76
(m, 3 H, ar-5H-6H-7H), 7.91–7.93 (m, 1 H, ar-H8). – MS (70 eV,
180 °C): m/z (%) = 343 (25) [M+᭹], 275 (39) [M+᭹-im], 262 (36)
[M+᭹ -CH2im], 248 (100) [M+᭹ -(CH2)2im], 234 (33) [M+᭹
-(CH2)3im], 109 (19), 91 (52) [C7H+7], 81 (9) [CH2im+].
HN(CH2)2CH2(CH2)2R), 1.87 (tt,
J = 7.4/7.5 Hz, 2 H,
HN(CH2)3CH2CH2tri), 1.98 (tt, J = 7.2/7.7 Hz, 2 H, HNCH2CH2
(CH2)3tri), 3.79 (m, 2 H, HNCH2 (CH2)4tri), 4.20 (t, J = 7.0 Hz,
2 H, HN(CH2)4CH2tri), 7.45–7.53 (m, 3 H, ph-3H-4H-5H),
7.65–7.69 (m, 2 H, ph-2H-6H), 7.76 (m, 1 H, ar-6H or -7H),
7.80–7.85 (m, 1 H, ar-6H or -7H), 7.92 (s, 1 H, tri-5H), 7.97 (d, 3J
= 8.2 Hz, 1 H, ar-5H), 8.02 (d, J = 7.8 Hz, 1 H, ar-8H). 8.07 (s,
1 H, tri-3H). – MS (70 eV, 85 °C): m/z (%) = 358 (26), [M+᭹], 276
(33) [M+᭹-CH2tri], 248 (61) [M+᭹-(CH2)3tri], 234 (65) [M+᭹
-
(CH2)4tri], 220 (100) [M+᭹-(CH2)5tri], 208 (50), 165 (26), 82 (5)
[CH2-tri+], 77 (52) [C6H+5].
N-[2-(1H-1,2,4-Triazol-1-yl)ethyl]-4-phenyl-1-phthalazinamine
(7 a)
N-[3-(1H-1,2,4-Triazol-1-yl)propyl]-4-phenylmethyl-1-phthal-
azinamine (7 e)
From 0.4 g (1.7 mmol) 4 a and 0.9 g 2-triazol-1-yl-ethylamine.
1.5 h at 120 °C.Yellow crystals (acetone/H2O); (acetone/n-hex-
ane), mp 158 °C, yield 0.35 g (65 %). – Anal. C18H16N6
(316.368). – 1H-NMR / 400 MHz (CDCl3): δ (ppm) = 4.29 (t, J =
5.3 Hz, 2 H, HNCH2CH2tri), 4.73 (t, 3J = 5.3 Hz, 2 H,
HNCH2CH2tri), 7.49–7.58 (m, 3 H, ph-3H-4H-5H), 7.67–7.69
(m, 2 H, Ph-2H-6H), 7.80–7.89 (m, 2 H, ar-6H-7H), 7.99 (d, J =
From 0.5 g (1.9 mmol) 4 e and 1.1 g 3-triazol-1-yl-propylamine,
1.5 h at 120 °C. Yellow crystals, mp 166–167 °C, yield 0.52 g
(80 %). – Anal. C20H20N6 (344.422). – 1H-NMR / 400 MHz
(CDCl3):
δ (ppm) = 2.42 (tt, J = 6.5/6.5 Hz, 2 H,
HNCH2CH2CH2tri), 3.83 (m, 2 H, HNCH2(CH2)2tri), 4.38 (t, J =
6.5 Hz, 2 H, HN(CH2)2CH2tri), 4.5 (s, 2 H, CH2Ph), 6.16 (br. s,
1 H, D2O exchange, NH), 7.15–7.21 (m, 1 H, ph-4H), 722–7.34
(m, 4 H, Ph-2H-3H-5H-6H), 7.69–7.77 (m, 2 H, ar-6H-7H),
7.91–7.96 (m, 2 H, ar-5H-8H), 8.01 (s, 1 H, tri-5H), 8.16 (s, 1 H,
tri-3H). – MS (70 eV, 195 °C): m/z (%) = 344 (22) [M+᭹], 262
3
8.3 Hz, 1 H, ar-5H), 8.03 (s, 1 H, tri-5H), 8.08 (d, J = 8.0 Hz,
1 H, ar-8H), 8.11 (s, 1 H, tri-3H). – MS (70 eV, 50 °C): m/z (%) =
316 (16) [M+᭹], 247 (26) [M+᭹-(1H)-tri], 234 (100) [M+᭹-CH2-trit],
221 (68) [M+᭹-CH2CH2-tri + H], 205 (23), 165 (13), 77 (38)
[C6H+5].
(100) [M+᭹-CH2tri], 248 (31) [M+᭹-(CH2)2tri], 234 (19) [M+᭹
(CH2)3tri], 184 (16), 91 (36) [C7H+7], 82 (3) [CH2-tri+].
-
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Pharm. Med. Chem. 2003, 336, 591–597 Antithrombotic 1-Phthalazinamines with Serotonin Antagonistic Properties 597
1
N-(Furan-2-yl-methyl)-4-phenyl-1-pthtalazinamine Hydrochlo-
ride (8 a)
(55 %). – Anal. C20H18ClN3O (351.834) – H-NMR / 400 MHz
(CD3OD):d (ppm) = 4.58 (s, 2 H, CH2ph), 4.98 (s, 2 H, NCH2fur.;
integral th H2O for 3,8 Protons), 6.43–645 (m, 1 H, fur-4H), 6.53
(d, J = 1.3 Hz, 1 H, fur-3H), 7.21–7.26 (m, 1 H, phH-4), 7.28–
7.35 (m, 4 H, Ph-2H-3H-5H-6H), 7.54 (d, J = 1.3 Hz, 1 H, furH-
5), 8.05–8.10 (m, 2 H, ar-6H-7H), 8.27–8.31 (m, 1 H, ar-5H),
8.52–8.55 (m, 1 H, ar-8H). – MS (70 eV, 200 °C): m/z (%) = 315
(100) [M+᭹], 286 (30) [M+᭹ – HC=O], 238 (40) [M+᭹-C6H5], 224
(62) [M+᭹-C7H7], 91 (35) [C7H+7].
From 0.6 g (2.5 mmol) 4 a and 2.5 ml furan-2-yl-methylamine,
2 h at 100 °C. Crystals (EtOH), mp 230 °C, yield 0.3 g (35 %). –
Anal.C19H16ClN3O (331.807).– 1H-NMR / 400 MHz (CD3OD):δ
(ppm) = 4.91 (s, 2 H, HNCH2-fur), 6.43–6.45 (m, 1 H, fur-4H),
6.54 (d, J = 3.1 Hz, fur-3H), 7.54 (d, J = 5.3 Hz, 1 H, fur-5H),
7.63–7.71 (m, 5 H, phH), 8.06–8.19 (m, 3 H, ar-5H-6H-7H),
8.61 (dd, J = 7.3/1.2 Hz, 1 H, ar-8H). – MS (70 eV, 165 °C): m/z
(%) = 301 (100) [M+᭹], 272 (62) [M+᭹ – HC=O], 246 (11), 220 (9)
[M+᭹-CH2-fur], 170 (27), 165 (15), 81 (39) [CH2-fur+], 77 ( 16)
[C6H+5].
References
[1] F.-N. Ko, C.-C. Wu, S. C. Kuo, F.Y. Lee, C. M. Teng, Blood
1994, 84, 4226–4233.
N-(Furan-2-ylmethyl)-4-(4-chlorophenyl)-1-phthalazinamine
(8 b)
[2] J. Galle et al., Br. J. Pharmacol. 1999, 127, 195–203.
[3] H. M. Holava Jr., R. A. Partyka, J. Med. Chem. 1969, 12,
From 0.5 g (1.9 mmol) 4 c and 2.1 g furan-2-yl-methylamine, 2 h
555–557.
at 100 °C. Light brown crystals, mp 190–192 °C, yield 0.55 g
1
(85 %). – Anal. C19H15ClN3O (336.799). – H-NMR / 400 MHz
[4] S. Gabriel, A. Michael, Ber. 1878, 11, 1007–1021.
[5] F. Seuter, Haemostasis 1976, 5, 85–95.
(CDCl3): δ (ppm) = 5.00 (s, 2 H, NCH2fur.), 6.37–6.38 (m, 1 H,
fur-4H), 6.46 (d, J = 2.9 Hz, 1 H, fur-3H), 7.41–7.42 (m, 1 H, fur-
5H), 7.50 (AAЈBBЈ, J = 8.4 Hz, 2H, Ph-3H-5H), 7.64 (AAЈBBЈ, J
= 8.4 Hz, 2 H, Ph-2H-6H), 7.78–7.87 (m, 2 H, ar-6H-7H), 7.92–
7.94 (m, 1 H, ar-5H), 8.00 (br. d, 1 H, ar-8H). – MS (70 eV,
250 °C): m/z (%) = 337 (18) [(37Cl)M+᭹], 335 (56) [M+᭹], 306 (32)
[M+᭹-CHO], 280 (9), 254 (10) [M+᭹-CH2fur.], 170 (30), 96 (30),
81 (100) [CH2fur.+].
[6] G. V. R. Born, Nature (London) 1962, 194, 927–929.
[7] K. Rehse, T. Ciborski, V. Schein, M. Kämpfe, B. Rose, E.
Unsöld, Arch. Pharm. 1991, 324, 301–305.
[8] K. Rehse, T. Ciborski, Arch. Pharm. 1995, 328, 71–75.
[9] I. Saenz de Tejada, J. Anguo, P. Cuevas, A. Fernández, I.
Moncada, A. Allona, E. Lledó, H. G. Körschen, U. Niewöh-
ner, H. Haning, E. Pages, E. Bischoff, Int. J. Impot. Res.
2001, 13, 282–290.
N-(Furan-2-ylmethyl)-4-(4-methoxyphenyl)-1-phthalazinamine
(8 c)
From 0.7 g (2.5 mmol) 4 d and 2.0 g furan-2-yl-methylamine, 2 h
at 100 °C. Crystals (acetone/H2O), mp 194 °C, yield 0.75 g
(90 %). – Anal. C20H17N3O2 (331.375). – 1H-NMR / 400 MHz
(CDCl3): δ (ppm) = 3.89 (s, 3 H, OCH3), 4.98 (s, 2 H, NCH2fur.),
5.63 (br. s, 1 H, D2O exchange, NH), 6.36–643 (m, 2 H, fur-3H-
4H), 7.05 (AAЈBBЈ, J = 8.8 Hz, 2 H, Ph-3H-5H), 7.64 (AAЈBBЈ, J
= 8.8 Hz, 2 H, Ph-2H-6H), 7.40–7.41 (m, 1 H, fur-5H), 7.73–
7.82 (m, 2 H, ar-6H-7H), 7.90 (d, J = 7.9 Hz, 1 H, ar-5H), 8.01
(dd, J = 7.8/1.3 Hz, 1 H, ar-8H). – MS (70 eV, 80 °C): m/z (%) =
[10] M. Hoenika, E.-M. Becker, H. Apeler, T. Sirichoke, H.
Schröder, R. Gerzer, J.-P. Stasch, J. Mol. Med. 1999, 77,
14–23.
[11] H. Pertz, S. Elz, J. Pharm. Pharmacol. 1995, 47, 310–316.
[12] R. F. Furchgott, In: Blaschko, H., Muscholl, E. (Eds.) Cate-
cholamines, Handbook of Experimental Pharmacology
33,. Springer Verlag, Berlin 1972, S. 283–335.
331 (100) [M+᭹], 302 (42) [M+᭹-CHO], 276 (10), 250 (6) [M+᭹
CH2fur.], 170 (29), 96 (19), 81 (35) [CH2fur.+].
-
N-(Furan-2-yl-methyl)-4-phenylmethyl-1-pthtalazinamine Hy-
drochloride (8 d)
From 0.6 g (2.3 mmol) 4 e and 2.5 g furan-2-yl-methylamine, 2 h
at 100 °C. Crystals (MeOH), mp 220–223 °C, yield 0.44 g
© 2003 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim