5-Lipoxygenase inhibitors: Synthesis and structure-activity relationships of a series of 1-aryl-2H,4H-tetrahydro-1,2,4-triazin-3-ones

Article abstract of DOI:10.1021/jm960372b

Synthetic routes were developed to access a variety of novel 1-aryl- 2H,4H-tetrahydro-1,2,4-triazin-3-one analogs which were evaluated as 5- lipoxygenase (5-LO) inhibitors. The parent structure, 1-phenylperhydro- 1,2,4-triazin-3-one (4), was found to be a

Full text of DOI:10.1021/jm960372b

3938
J . Med. Chem. 1996, 39, 3938-3950
5-Lip oxygen a se In h ibitor s: Syn th esis a n d Str u ctu r e-Activity Rela tion sh ip s of
a Ser ies of 1-Ar yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on es
Pramila A. Bhatia, Clint D. W. Brooks,*,† Anwer Basha, J ames D. Ratajczyk, Bruce P. Gunn,
J ennifer B. Bouska, Carmine Lanni, Patrick R. Young, Randy L. Bell, and George W. Carter
Abbott Laboratories, Immunoscience Research Area, D-47K, AP10, 100 Abbott Park Road, Abbott Park, Illinois 60064-3500
Received May 23, 1996
Synthetic routes were developed to access a variety of novel 1-aryl-2H,4H-tetrahydro-1,2,4-
triazin-3-one analogs which were evaluated as 5-lipoxygenase (5-LO) inhibitors. The parent
structure, 1-phenylperhydro-1,2,4-triazin-3-one (4), was found to be a selective inhibitor of 5-LO
in broken cell, intact cell, and human blood assays with IC50 values of 5-21 µM. In a rat
anaphylaxis model, 4 blocked leukotriene formation with an ED50 ) 7 mg/kg when administered
orally. Compound 4 exhibited selectivity for inhibition of 5-LO with little activity against related
enzymes: 12-LO from human platelets, 15-LO from soybean, and cyclooxygenase (COX) from
sheep seminal vesicle. In pilot subacute toxicity testing, 4 did not produce methemoglobinemia
in rats (400 mg/kg po daily for 9 days) or in dogs (200 mg/kg po daily for 28 days). These
results indicated that the triazinone structure provided a 5-LO inhibitor template devoid of
the toxicity problems observed in the related phenidone (1) and pyridazinone (3) classes of
5-LO inhibitors. The parent compound 4 is a selective, orally bioavailable 5-LO inhibitor which
can serve as a useful reference standard for in vivo pharmacological studies involving
leukotriene-mediated phenonmena.
In tr od u ction
Selective inhibition of 5-lipoxygenase (5-LO) provides
a definitive means to limit the pathological effects of
all leukotrienes and their metabolites.1 The 5-LO in-
hibitor, zileuton,2 has demonstrated therapeutic bene-
fit in the treatment of asthma.3 Over the past decade,
we have investigated several structural classes of
analogs was reduced.15 In contrast to phenidone, the
5-LO inhibitors with the objective of identifying suit-
homolog 3 did not convert hemoglobin to methemoglobin
in vitro in whole blood from various species including
humans. However, in pilot toxicity studies, repeated
400 mg/kg daily oral doses of 3 resulted in the appear-
ance of Heinz bodies in red blood cells and progressive
methemoglobinemia. These unexpected results sug-
gested that a metabolite of 3 might be responsible for
the toxicity. None of the metabolites identified in the
rat, accounting for about 92% of the radioactivity,
converted hemoglobin to methemoglobin in vitro up to
400 µM. We therefore were left to rationalize that in
vivo hydrolysis of 3 might form a reactive phenylhy-
drazide intermediate 3b (Scheme 1), which was not
isolated as it may have formed adducts with macromol-
ecules. A reactive phenylhydrazine intermediate like
3b might be a likely source of the observed methemo-
globinemia since it is known that phenylhydrazine
rapidly converts human oxyhemoglobin to methemoglo-
bin.16 To test this hypothesis, we synthesized the novel
heterocycle, 1-phenyl-2H,4H-tetrahydro-1,2,4-triazin-3-
one (4), which was predicted to have increased stability
toward in vivo metabolic hydrolysis due to its cyclic urea
functionality. The biological properties and structure-
activity relationships of this new series of triazinone
5-LO inhibitors were investigated.
able orally active 5-LO inhibitors for clinical study to
evaluate the role of leukotrienes in human diseases.4
This report describes a series of 1-aryl-2H,4H-tetrahy-
dro-1,2,4-triazin-3-ones with oral activity as selective
5-LO inhibitors which will be referred to simply as
triazinones.
Phenidone (1)5 was one of the earliest orally active
inhibitors of 5-LO to be discovered. Animal studies
revealed that 1 and several analogs thereof were found
to be toxic due to methemoglobin formation upon
repeated dosing.6,7 The related aminopyrazoline series
of 5-LO inhibitors, represented by BW-755C (2),8 was
also found to cause methemoglobinemia.9 The Sterling
group also investigated 1-phenyl-3-pyrazolidinones and
found hematologic toxicity to be a limiting factor.10 The
redox properties of 1-phenyl-3-pyrazolidinones were
likely linked to the propensity for causing methemoglo-
binemia.11 We tested the hypothesis that the inhibition
of 5-LO by 1-phenyl-3-pyrazolidinones was not entirely
due to a nonspecific redox interaction by examining the
corresponding six-membered analog, 1-phenyl-2H-tet-
rahydropyridazin-3-one (A-53612, 3).12
The cyclic hydrazide, 3 was found to be a selective
orally active inhibitor of 5-LO.13,14 The increased
inhibitory selectivity of 3 for 5-LO compared to pheni-
done supported the hypothesis that by ring expansion
the inhibitory component due to nonspecific antioxidant
activity observed for the five-membered phenidone
Ch em istr y
Several synthetic routes were developed to access a
variety of novel 1-aryl-2H,4H-tetrahydro-1,2,4-triazin-
3-one analogs. The first synthesis of the parent com-
pound, 1-phenyl-2H,4H-tetrahydro-1,2,4-triazin-3-one
† Formerly known as Dee W. Brooks.
Abstract published in Advance ACS Abstracts, September 1, 1996.
S0022-2623(96)00372-X CCC: $12.00 © 1996 American Chemical Society

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3939
Sch em e 1. Hypothetical in Vivo Hydrolysis of
Pyridazinones to Toxic Phenylhydrazine Intermediates
An alternative synthesis from amino acid derivatives
was attempted as shown in Scheme 5. For example,
the synthesis of 5-(hydroxymethyl)-1-phenyl-2H,4H-
tetrahydro-1,2,4-triazin-3-one (32) was accomplished by
reaction of D,L-serine methyl ester with aniline to
provide the amide 32a , which was reduced with LiAlH4,
and the primary amino group was selectively converted
to the corresponding ethyl carbamate intermediate 32b.
The nitrosation and reduction steps provided the cor-
responding hydrazine intermediate which was difficult
to cyclize to the desired compound 32 (less than 1%
yield). The unprotected hydroxyl group was likely
responsible for the poor yield. Since 32 was found to
be a weak 5-LO inhibitor no further attempts to improve
this method were made.
Sch em e 2. Method A: Synthesis of Triazinones 4-9,
11, 12, and 14-17 from 1-Amino-2-haloethanesa
The heteroarylphenyl-substituted analogs 51-59 were
prepared as shown in Scheme 6. The (3-bromophenyl)-
triazinone 12 or 29 was used in a Pd-catalyzed coupling
reaction18 with the requisite trimethylstannyl hetero-
cycle, prepared by n-butyllithium-mediated metalation
of the heterocycle followed by quenching with trimeth-
ylstannyl chloride.19 For example, compound 54 was
prepared in 17% yield by the coupling of 29 with
2-(trimethylstannyl)thiophene.
a
Reagents: (a) ClCO2Et, Et3N, CH2Cl2, 0 °C, 1 h; (b) EtN(iPr)2,
NH2-R1, toluene, 80 °C, 24 h; (c) NaNO2, concentrated HCl, H2O,
5 °C, 2 h; (d) Zn dust, H2O, CH3CO2H, 5-15 °C, 1 h then room
temperature for 1 h; (e) EtMgBr, CH2Cl2, -40 °C, 0.5 h then 40
°C, 48 h.
The 4-N-substituted analogs 60-62 were prepared as
shown in Scheme 7. The 4-N substituent was deter-
mined by the acyl chloride used to react with N-
phenylethylenediamine. For example, the synthesis of
4-(phenylmethyl)-1-phenyl-2H,4H-tetrahydro-1,2,4-tri-
azin-3-one (62) was accomplished by reaction of N-
phenylethylenediamine with benzoyl chloride to provide
the intermediate amide 62a . Reduction of 62a with
LiAlH4 provided the intermediate amine which was
directly converted to the methyl carbamate 62b. The
nitrosation, reduction, and cyclization methods of Scheme
1 were applied to 62b to afford 62 in 24% overall yield.
The 4-N-hydroxy analogs 63-65 were prepared as
shown in Scheme 8. For example, the carbamate 63a
was reacted with chloroacetone oxime in the presence
of Na2CO3 to provide the adduct 63b. The oxime group
was reduced with borane-pyridine in the presence of 6
N ethanolic HCl to provide the intermediate hydroxy-
lamine, which was cyclized by treating with potassium
tert-butoxide to provide the 4-N-hydroxytriazinone ana-
log 63 in 20% overall yield.
Synthesis of 1-phenyl-3H-trihydro-1,3,4-oxadiazin-2-
one (67) was accomplished by the procedures of Scheme
9. 1-Acetyl-2-phenylhydrazine was reacted with 2-bro-
moethanol, and the intermediate was subjected to
hydrolysis with 6 N HCl at reflux to provide 1-(2-
hydroxyethyl)-1-phenylhydrazine which was treated
with carbonyldiimidazole to provide 67 in 6% overall
yield.
Synthesis of 1-phenyl-2H,4H-tetrahydro-1,2,4-triazin-
3-thione (68) in 42% yield was accomplished by treat-
ment of 4 with Lawesson’s reagent in toluene at 80 °C
for 15 h.
(4), was accomplished by the sequence of reactions
illustrated in Scheme 2. Bromoethylamine was con-
verted to the corresponding ethoxycarbamate 4a which
was then reacted with aniline to provide the intermedi-
ate ethyl N-[(phenylamino)ethyl]carbamate 4b. Stan-
dard procedures were used for nitrosation of 4b followed
by reduction, to provide the substituted phenylhydra-
zine intermediate 4c. The intramolecular cyclization
of the hydrazine, 4c, proved to be problematic until we
applied the urea method reported by Basha.17 Treat-
ment of 4c initially at -40 °C with slightly more than
1 equiv of ethylmagnesium bromide followed by warm-
ing at 40 °C for 2 days provided the desired phenyltri-
azinone 4 in 16% overall yield. The procedures shown
in Scheme 2 were used for the synthesis of analogs 5-9,
11, 12, and 14-17 by applying the requisite substituted
aniline (Table 1).
A more efficient synthesis of the triazinone hetero-
cycle was devised by changing the order of construction
of the ring system as shown in Scheme 3. For example,
the synthesis of 10 was accomplished by reaction of
2-chloroethyl isocyanate with (3-fluorophenyl)hydrazine
to provide the intermediate 10a which was cyclized by
heating in DMF at 75 °C with 1 equiv of NaI for 2 days
to provide 10 in 40% overall yield. The procedures
shown in Scheme 3 were used for the preparation of
analogs 10, 13, 19, 27, 29-31, and 33-50 (Table 1).
The synthesis of analogs 21-26 and 28 from amino
alcohols was devised as shown in Scheme 4. For
example, the synthesis of 5-methyl-1-phenyl-2H,4H-
tetrahydro-1,2,4-triazin-3-one (21) was accomplished by
conversion of D,L-2-amino-1-propanol to the correspond-
ing ethyl carbamate followed by a Swern oxidation to
provide the aldehyde intermediate 21a , which was
reacted with aniline under conditions of reductive ami-
nation with NaBH3CN to provide the ethyl [(phenyl-
amino)ethyl]carbamate 21b. This intermediate was
then converted to the desired triazinone analog by the
procedures of nitrosation, reduction, and cyclization
previously described in Scheme 2 to provide 21 in 6%
overall yield.
Resu lts a n d Discu ssion
Biologica l Testin g. The biological methods used to
provide the critical data for inhibitor optimization are
described as follows.2 Direct evaluation of inhibition of
5-LO catalysis was conducted using a crude supernatant
from sonicated rat basophilic leukemia cells and mea-
suring 5-HETE product formation. Compounds of in-

3940 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
Ta ble 1. Structure-Activity Relationships for Selected Triazinones
Bhatia et al.
in vivo ratc
ED50, mg/kg, or
(% I at 200 µmol/kg)
rat pod
Tmax, h;
Cmax, µM
IC50, µM
broken cella
21
(synth
meth)
compd
R1
R5
HWBLb
5.8
4
5
6
7
8
(A)
(A)
(A)
(A)
(A)
(A)
(B)
(A)
(A)
(B)
(A)
(A)
(A)
(A)
phenyl
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
7
1; 130
2.1; 78
1.1; 98
1.1; 105
4; 20
1.1; 32
1; 108
2; 59
4.2; 35
2; 64
4.1; 24
1.2; 72
1; 26
2-methylphenyl
3-methylphenyl
4-methylphenyl
3,5-dimethylphenyl
3-ethylphenyl
3-fluorophenyl
3-chlorophenyl
3-bromophenyl
3,5-difluorophenyl
3-(trifluoromethyl)phenyl
3-methoxyphenyl
3-(methylthio)phenyl
3-(phenylmethoxy)phenyl
3-hydroxyphenyl
3-nitrophenyl
386
NSe
18
15
2.0
NS (300 µM)e
12
13
19
5.3
4.9
8.9
17
47
14
(46%)
24
(38%)
18
(46%)
(59%)
(40%)
(44%)
(73%)
(14%)
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
2.1
4.6
2.2
0.2
1.1
NS (32 µM)e
49
1; 2.5
(B)
NSe
2; 62
3-aminophenyl
phenyl
phenyl
phenyl
phenyl
H
178
11
1.1; 95
2; 141
2.2; 20
1.2; 59
2; 48
(C)
(C)
(C)
(C)
(C)
(C)
(B)
(C)
(B)
(B)
(B)
(D)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(B)
(E)
(E)
(E)
(E)
(E)
(E)
(E)
(E)
(E)
CH3
(CH3)2
CH2CH3
i-C3H7
C4H9
CH3
CH3
CH3
CH3
phenoxy
4-fluorophenoxy
CH2OH
CH2OCH2CH2OCH3
H
H
H
H
CH3
H
CH3
H
CH3
H
CH3
H
CH3
H
H
H
H
H
H
H
CH3
H
CH3
H
5.8
13
6.8
6.9
6.8
1.5
19
6.3
2.7
2.6
NSe
3.8
20
24
59
phenyl
(42%)
(43%)
(60%)
(40%)
(54%)
NSe
1; 8
3-methylphenyl
3-fluorophenyl
3-chlorophenyl
3-bromophenyl
phenyl
phenyl
phenyl
phenyl
pyrid-2-yl
2.2; 44
7.2; 70.2
4; 31
1.8
2.7
5.2
2.6
2.6
7.2; 26
NSe
40% (32 µM)
14
24
8.6
26
8
1; 184
pyrid-3-yl
pyrid-4-yl
NS (32 µM)e
NS (32 µM)e
6-methylpyrid-2-yl
6-methylpyrid-2-yl
6-fluoropyrid-2-yl
6-fluoropyrid-2-yl
6-chloropyrid-2-yl
6-chloropyrid-2-yl
6-bromopyrid-2-yl
6-bromopyrid-2-yl
6-methoxypyrid-2-yl
6-methoxypyrid-2-yl
7-chloroquinol-4-yl
quinol-3-yl
benzothiazol-2-yl
benzoxazol-2-yl
3-pyrid-2-ylphenyl
3-pyrid-4-ylphenyl
3-thien-2-ylphenyl
3-thien-2-ylphenyl
3-thien-3-ylphenyl
3-fur-2-ylphenyl
3-thiazol-2-ylphenyl
6-thien-2-ylpyrid-2-yl
6-pyrid-2-ylpyrid-2-yl
5.5
3.8
9.1
6.6
5.6
3.5
3.5
2.4
7.9
1.7
3
6.0
4.8
1.6
4.6
3.1
3.5
7.1
17
1; 99
1; 125
16
2; 122
2; 48
1; 79
2; 21
1; 101
1; 85
4; 85
1; 58
2; 123
NSe
41
(57%)
(26%)
9.5
5.5
NS (32 µM)e
NS (32 µM)e
NS (32 µM)e
NS (32 µM)e
30
30
2.1
2.8
4.9
4.8
8.8
5.5
5.3
8.4
1; 3
4; 1
1; 2
1; 7
NSe
35% (32 µM)
8.2
6
H
H
37
>50
a
A broken cell 20000g supernatant from rat basophilic leukemia (RBL) cells with 5-LO activity. The 95% confidence limits were
b
<(50% of the mean value, and assays were done in duplicate. Human whole blood stimulated with calcium ionophore (A23187) and
LTB4 was measured by enzyme immunoassay. The 95% confidence limits were <(50% of the mean value and assays were done in
duplicate. c Rat anaphylaxis leukotriene formation with a 1 h inhibitor pretreatment time, an oral dose of 200 µmol/kg was used to screen
compounds for in vivo activity, and data are reported as the mean of eight rats (percent inhibition of LTE4 from peritoneal fluids). Dose
response studies were conducted for the more promising compounds, the ED50 values in mg/kg are the mean of eight rats with 90%
d
confidence limits <(50% of the mean value. Plasma drug levels evaluated in rats dosed by oral gavage with 200 µmol/kg of test compound
and blood samples were drawn via the tail vein at approximately 1 h time intervals and analyzed by HPLC. Values are the mean of data
from two animals. e NS indicates that the activity observed was not significantly different from that of the control group.
terest from this initial screen of activity were examined
further in a human whole blood assay. Stimulation of
human blood with calcium ionophore A23187 resulted
in leukotriene formation. This assay provided a second

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3941
Sch em e 3. Method B: Synthesis of Triazinones 10, 13,
19, 27, 29-31, and 33-50 from 2-Chloroethyl
Isocyanatesa
Sch em e 7. Method F: Synthesis of 4-N-Substituted
Triazinones 60-62a
a
Reagents: (a) Et3N, H2NNHR1, CH2Cl2, room temperature for
2 h; (b) NaI, DMF, 75 °C, 48 h.
a
Reagents: (a) RCOCl, Et3N, CH2Cl2, 0 °C for 1 h then room
Sch em e 4. Method C: Synthesis of Triazinones 21-26
and 28 From 1-Amino-2-hydroxyethanesa
temperature for 16 h; (b) LiAlH4, THF, room temperature for 1 h
then reflux for 16 h; (c) ClCO2CH3, Et3N, THF, -5 °C, 2 h then
room temperature for 16 h; (d) NaNO2, 12 N HCl, H2O, 5 °C, 2 h;
(e) Zn dust, H2O, CH3CO2H, 5-15 °C for 1 h then room temper-
ature for 1 h; (f) EtMgBr, CH2Cl2, -40 °C for 0.5 h then 40 °C for
48 h.
Sch em e 8. Method G: Synthesis of
4-N-Hydroxy-Substituted Triazinones 63-65a
a
Reagents: (a) ClCO2CH3, K2CO3, CH3CN, -20 °C for 1 h then
room temperature for 16 h; (b) (COCl)2, Et3N, DMSO, CH2Cl2, -78
°C for 0.5 h then room temperature for 1 h; (c) NaBH3CN, CH3OH,
pH 5-6, room temperature for 16 h; (d) NaNO2, 12 N HCl, H2O,
5 °C, 2 h; (e) Zn dust, H2O, CH3CO2H, 5-15 °C for 1 h then room
temperature for 1 h; (f) EtMgBr, CH2Cl2, -40 °C for 0.5 h then 40
°C for 48 h.
a
Reagents: (a) Na2CO3, chloroacetone oxime, CH2Cl2, reflux
for 18 h; (b) BH3/pyridine, ethanol, room temperature for 1 h then
0 °C, 6 N ethanolic HCl; (c) KOtC4H9, THF, room temperature
for, 18 h.
Sch em e 5. Method D: Synthesis of Triazinone 32 from
Sch em e 9. Synthesis of
an Amino Acida
1-Phenyl-3H-trihydro-1,3,4-oxadiazin-2-one (67)a
a
Reagents: (a) NH2R1, EtMgBr, THF, CH2Cl2, 0 °C for 1 h, then
a
Reagents: (a) EtN(iPr)2, HOCH2CH2Br, toluene, 80 °C, 48 h;
reflux for 72 h; (b) LiAlH4, THF, room temperature for 1 h then
reflux for 16 h; (c) ClCO2CH3, Et3N, THF, -5 °C for 2 h then room
temperature for 1 h; (d) NaNO2, 12 N HCl, H2O, 5 °C, 2 h; (e) Zn
dust, H2O, CH3CO2H, 5-15 °C for 1 h then room temperature for
1 h; (f) EtMgBr, CH2Cl2, -40 °C for 0.5 h then 40 °C for 48 h.
(b) 6 N HCl, reflux, 2 h; (c) carbonyldiimidazole, CH2Cl2, room
temperature, 3 h.
at 200 µmol/kg in rats was conducted with measurement
of compound plasma concentrations by HPLC as a
preliminary evaluation of pharmacokinetic properties.
A rat anaphylaxis assay was used to evaluate the in
vivo leukotriene inhibitory activity of orally adminis-
tered compounds. In this in vivo assay leukotriene
biosynthesis was induced in the peritoneal cavity of rats
in response to an antibody-antigen reaction.20 An oral
dose of 200 µmol/kg was selected with a 1 h pretreat-
ment time for the initial screen of 5-LO inhibitors. A
dose-response study was conducted on the more potent
analogs identified. The most promising inhibitors were
evaluated in dog and monkey for ex vivo leukotriene
inhibition of calcium ionophore stimulated blood samples
taken over a 24 h period after oral dosing. The drug
plasma levels were also measured by HPLC. Finally,
due to the toxicity problems seen with earlier progeni-
tors of this series, a preliminary evaluation of blood
Sch em e 6. Method E: Synthesis of Triazinones 51-59
by Pd-Catalyzed Coupling with Trimethylstannyl
Heterocyclesa
a
Reagents: (a) [(C6H5)3P]4Pd0, toluene, reflux, 4 h.
measure of inhibitory activity in the complex medium
of whole blood containing intact leukotriene producing
cells. In general the triazinone analogs showed similar
or better inhibitory activity in human blood than in the
broken RBL cell assay. Routine oral dosing of inhibitors

3942 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
Bhatia et al.
abnormalities was conducted by examining blood samples
for Heinz bodies after multiple daily oral doses in rats
and dogs.
n-butyl (25) (IC50 ) 1.5 µM) analogs. However the
opposing trend of reduced activity in vivo continued as
the plasma levels in rats of these analogs (oral dose of
200 µmol/kg) decreased accordingly: 21 (Cmax 141 µM),
22 (Cmax 20 µM), 23 (Cmax 59 µM), 24 (Cmax 48 µM), and
25 (Cmax 8 µM). Several other analogs were examined
which did not provide any clues to overcome this
dichotomy.
The design of the parent triazinone 4 evolved as a
means to reduce the hypothetical potential for metabolic
cleavage of the pyridazinone 3 to form reactive and toxic
phenylhydrazine intermediates such as 3b. Replacing
the 4-CH2 for a 4-NH group in the tranformation of 3
to 4 resulted in a 5-fold loss of inhibitory activity in the
broken cell (IC50 ) 4.3 versus 21 µM) and an 8-fold loss
of potency in the human blood 5-LO assay (IC50 ) 0.7
versus 5.8 µM). However the in vivo leukotriene inhibi-
tory activity of the two compounds was similar with
ED50 values of 6 mg/kg for 3 and 7 mg/kg for 4. Thus
the triazinone 4 represented a promising new 5-LO
inhibitor for further optimization.
SAR of 1-N-Heter oar yltr iazin on es. The next phase
of SAR evaluation was to replace the 1-N-phenyl sub-
stituent in 4 with heteroaryl substituents to test the
idea that the presence of heteroatoms might change the
SAR profile. In the pyridyl series the position of the
nitrogen atom proved to be critical. The 2-pyridyl
analog 34 (IC50 ) 26 µM) had potency similar to that of
4, whereas the 3-pyridyl (35) and 4-pyridyl (36) analogs
were inactive at the screening dose of 32 µM. Com-
pound 34 had activity similar to that of 4 in the human
whole blood assay (IC50 ) 8.6 versus 5.8 µM) and in vivo
in the rat (ED50 ) 8 versus 7 mg/ kg), and 34 (Cmax 184
µM) also gave higher plasma levels upon oral dosing in
rats compared to 4 (Cmax 130 µM). The promising
properties of 34 stimulated the evaluation of several
analogs substituted in the 6-position of the pyridyl group
and the 5-position of the triazinone (Table 1). The
results were similar to those observed in the phenyl-
substituted series, and no unexpectedly superior analogs
were identified. Other heteroaryl substituents sur-
veyed, 4-quinolyl (47), 3-quinolyl (48), 2-benzothiazolyl
(49), and 2-benzoxazolyl (50), were all inactive against
5-LO at 32 µM.
SAR of (Heter oa r ylp h en yl)tr ia zin on es. The next
phase of the SAR evaluation was to add a heteroaryl
substituent to the 1-N-phenyl group of 4 in the meta
position. The 3-pyrid-2-yl (51) and 3-pyrid-4-yl (52)
analogs had similar in vitro potency. The thienylphenyl
analogs 53-55 and furylphenyl analog 56 were at least
10-fold more potent in vitro but unfortunately gave very
low plasma levels in rats. The 3-thiazol-2-yl analog 57
was a weak inhibitor. Two analogs with heteroaryl
combinations were examined for the 2-pyridyl series.
The thienylpyridyl analog, 58 and the pyridylpyridyl
analog 59 showed much less activity in the human blood
assay than in the broken cell assay, which might be
attributed to increased interference from plasma protein
binding effects. None of these analogs were considered
promising enough to conduct in vivo testing.
SAR of P h en yl Su bstitu tion . The initial plan was
to survey several analogs with the objective of identify-
ing structural modifications which might enhance in-
hibitory activity. Substituents on the phenyl group of
4 were examined (Table 1). The results were analogous
to the previously reported pyridazinone system where
inhibitory potency was significantly attenuated by any
substituent in the 2- or 4-positions.13 For example, in
the toluene series, ortho-substituted 5 and para-
substituted 7 were inactive below 100 µM, whereas
meta-substituted 6 (IC50 ) 15 µM) had similar potency
to the parent 4. The 3,5-dimethyl analog 8 (IC50 ) 12
µM) maintained potency in vitro but had about a 6-fold
lower plasma level compared to that of 4 in orally dosed
rats which correlated with its weak activity in vivo. The
3-ethyl analog 9 (IC50 ) 13 µM) also demonstrated
reduced plasma levels and attenuated in vivo activity
(ED50 ) 24 mg/kg). Halogen-substituted analogs 10-
14 were examined and found to offer no significant
advantages. The 3-methoxy analog 15 (IC50 ) 47 µM)
was less potent than the 3-methyl thiol analog 16 (IC50
) 14 µM), further supporting the conclusion that
inhibitory activity preferred less polar substituents.
Increasing the lipophilicity of the meta substituent as
in the 3-phenylmethoxy analog 17 (IC50 ) 1.1 µM in the
broken cell and 0.2 µM in human blood) provided the
most potent in vitro inhibitor prepared. Unfortunately,
17 had very low plasma levels after oral dosing in the
rat and, as expected from this, poor activity in vivo.
It became clear that the optimization of this series
would prove challenging in view of opposing phenomena
where increased lipophilicity resulted in greater in vitro
inhibitory potency but reduced oral bioavailability which
in turn attenuated in vivo activity. Three analogs with
polar hydrogen bonding substituents, 3-hydroxy (18),
3-nitro (19), and 3-amino (20), were found to have weak
in vitro inhibition. It was concluded that lipophilic
substituents in the meta position of the phenyl group
provided enhanced in vitro 5-LO inhibitory activity
which did not result in improved in vivo activity due to
diminished oral bioavailability.
SAR of 4-N-Su bstitu ted Tr ia zin on es. The out-
come of 4-N substituents was examined (Table 2). In
the series, methyl (60), ethyl (61), and phenylmethyl
(62) only the latter showed 5-LO inhibitory activity
below the screening dose of 32 µM. In the context of
our experience with hydroxamate and N-hydroxyureas4
the corresponding 4-N-OH analogs 63-65 were evalu-
ated and found to have IC50 values in the range of 4-13
µM in both the broken cell and human blood assays.
The N-methoxy analog 66 had in vitro potency similar
to that of the parent compound 4. In the rat anaphyl-
axis assay, 62 and 63 had 14% and 55% leukotriene
inhibition with a single 200 µmol/kg oral dose.
SAR of 5-Su bstitu ted Tr ia zin on es. Alternative
sites for modification that might lead to optimized
potency were investigated. A series of 5-substituted
analogs were evaluated (Table 1). The trend for im-
proved in vitro potency with increased liphophilicity of
the 5-substituent is supported by comparing the methyl
(21) (IC50 ) 11 µM), dimethyl (22) (IC50 ) 6.8 µM), ethyl
(23) (IC50 ) 6.9 µM), isopropyl (24) (IC50 ) 6.8 µM), and
SAR of Miscella n eou s An a logs. Replacing 4-CH2
in 3 for 4-O provided 1-phenyl-3H-trihydro-1,3,4-oxa-
diazin-2-one (67) which had an IC50 ) 21 µM in the
broken cell and 5.8 µM in human blood. Transforming
4 into the corresponding thiocarbonyl analog 68 resulted

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3943
Ta ble 2. Inhibitory Activity for Selected Triazinone Analogs
IC50, µM
compd
(synth meth)
R1
R2
R4
CH3
CH2CH3
CH2C6H5
OH
OH
OH
OCH3
R5
broken cella
HWBLb
60
61
62
63
64
65
66
(F)
(F)
(F)
(G)
(G)
(G)
phenyl
phenyl
phenyl
phenyl
3-chlorophenyl
3-methylphenyl
phenyl
H
H
H
H
H
H
H
H
H
H
CH3
CH3
CH3
CH3
NS (32 µM)c
NS (32 µM)c
8.3
6
3.9
8.4
22
13
3.6
4.8
a
A broken cell 20000g supernatant from rat basophilic leukemia (RBL) cells with 5-LO activity. The 95% confidence limits were
b
<(50% of the mean value, and assays were done in duplicate. HWBL, human whole blood stimulated with calcium ionophore (A23187)
and LTB4 was measured by enzyme immunoassay. The 95% confidence limits were <(50% of the mean value, and assays were done in
duplicate. c NS indicates that the activity observed was not significantly different from that of the control group.
Ta ble 3. Selectivity of in Vitro Inhibition of 5-Lipoxygenase and Related Enzymes
a
IC50
,
µM or (% inhibition)
enzyme
phenidone (1)
3
4
5-LO broken RBL
12-LO human platelet
15-LO soybean
1.9 (1.5-2.8)
0.10 (0.09-0.10)
0.42 (0.34-0.51)
94 (78-115)
4.3 (4.1-4.5)
21 (18-24)
(7% at 100 µM)b
(25% at 100 µM)b
(5% at 300 µM)b
(14% at 100 µM)b
(14% at 100 µM)b
(4% at 100 µM)b
COX sheep seminal vesicle
a
b
95% confidence limits shown in parentheses; assays done in duplicate. Percent inhibition at highest tested concentration.
Ta ble 4. In Vitro Inhibition of 5-LO and Cyclooxygenase (COX) in Intact Cells and Human Blood
a
IC50
,
µM or (% inhibition)
enzyme (cell type)
phenidone (1)
3
4
5-LO rat PMNLc
0.52 (0.33-0.74)
13 (11-16)
1.3 (1.1-1.6)
0.3 (0.25-0.4)
12 (4-20)
1.2 (0.8-1.9)
4.6 (4.5-4.8)
COX rat PMNLd
(3% at 300 µM)b
1.0 (0.1-2.3)
(28% at 100 µM)b
9.2 (4.7-17.0)
5.8 (4.4-7.4)
5-LO human PMNLe
5-LO human bloodf
COX human bloodg
0.7 (0.1-1.3)
(13% at 100 µM)b
(0% at 100 µM)b
95% confidence limits shown in parentheses, assays done in duplicate. Percent inhibition at highest tested concentration. c Rat
a
b
d
polymorphonuclear leukocytes stimulated with calcium ionophore (A23187) and LTB4 was measured by enzyme immunoassay. Rat
polymorphonuclear leukocytes stimulated with calcium ionophore (A23187) and thromboxane was measured by enzyme immunoassay.
e Human polymorphonuclear leukocytes stimulated with calcium ionophore (A23187) and LTB4 was measured by enzyme immunoassay.
g
f Human whole blood stimulated with calcium ionophore (A23187) and LTB4 was measured by enzyme immunoassay. Human whole
blood stimulated with calcium ionophore (A23187) and thromboxane was measured by enzyme immunoassay.
Ta ble 5. Plasma Levels of 4 after Oral Dosinga
in an inhibitor with similar potency in the broken cell
assay (IC50 ) 17 µM). However, 68 was less active in
vivo, and toxicity was observed in the rat at oral doses
of 200 µmol/kg. Further study of the thiocarbonyl series
was discontinued.
oral dose,
species
µmol/kg
Tmax, h
Cmax, µM
T1/2 est, h
rat
dog
monkey
200
113
113
1.0
0.3
2.0
130
132
74
3.3
0.8
1.0
F u r th er Biologica l Eva lu a tion of 4. This survey
of SAR did not reveal any compounds with overall
properties superior to the parent compound 4, which
had the most consistent in vivo inhibition on repeated
oral dose response studies. Compound 4 exhibited
selectivity for 5-LO inhibition with little activity against
related enzymes 12-LO from human platelets, 15-LO
from soybean, and cyclooxygenase (COX) from sheep
seminal vesicle (Table 3). In both intact cell assays and
human blood, compound 4 was a selective inhibitor of
calcium ionophore (A23187) stimulated 5-LO activity
and did not inhibit COX activity (Table 4). Preliminary
pharmacokinetic studies in rat, dog, and monkey showed
satisfactory drug plasma levels after oral administration
(Table 5). In both dog (Figure 1) and monkey (Figure
2), oral administration of 10 mg/kg of 4 provided greater
than 50% ex vivo leukotriene inhibition in blood samples
taken over the first 4 h. The plasma concentration of 4
a
Values are the mean of data from two animals.
was also measured and corresponded to the degree of
ex vivo inhibition observed in the monkey. However,
in the dog the shape of the plasma versus inhibition
plots indicated that an active metabolite was being
formed (not identified).
Two analogs were found to have comparable proper-
ties to 4. The 5-methyl analog (21) had a longer
elimination half-life in dog (4.2 h) compared to that of
4 (0.8 h) but was about 2-fold less active than 4 in the
rat anaphylaxis model. The pyridyl analog (34) had
similar inhibitory activity (Table 1) and pharmacoki-
netics in rat (n ) 2, dose 200 µmol/kg po, Tmax 1 h, Cmax
184 µM, T1/2 2.9 h), dog (n ) 2, dose 112 µmol/kg po,
Tmax 0.7 h, Cmax 109 µM, T1/2 1 h), and monkey (n ) 2,
dose 112 µmol/kg po, Tmax
1.5 h, Cmax 37 µM, T1/2 1.1 h)
as 4 (Table 5). Oral administration of 10 mg/kg of 34

3944 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
Bhatia et al.
tetramethylsilane as an internal standard. Elemental analysis
(C, H, N) were performed by Abbott Laboratories Pharmaceu-
tical Products Division or Robertson Microlit Laboratories, Inc.,
Madison, NJ . Silica gel 60 (E. Merck, 230-400 mesh) was
used for preparative column chromatography. Reactions were
performed under dry nitrogen atmosphere unless stated
otherwise. THF was freshly distilled from sodium benzophe-
none ketyl. Other solvents were HPLC grade. Reagents were
obtained commercially and used without further purification.
Chemical yields reported are unoptimized specific examples
of one prepartion. Analytical TLC using E. Merck F254
commercial plates was used to follow the course of reactions.
Meth od A (Sch em e 2). 1-P h en yl-2H,4H-tetr a h yd r o-
1,2,4-tr ia zin -3-on e (4). To a stirred 0 °C suspension of
bromoethylamine hydrochloride (255 g, 1.25 mol) in dichlo-
romethane (700 mL) was added triethylamine (252 g, 2.5 mol)
followed by dropwise addition of ethyl chloroformate (135.3 g,
1.25 mol) in dichloromethane (200 mL) while the reaction
termperature was maintained between 0 and 4 °C. The
mixture was stirred for 1 h; water (500 mL) was added; and
the organic layer was separated and washed with saturated
aqueous NaCl, dried (MgSO4), filtered, and concentrated in
vacuo to give ethyl N-(2-bromoethyl)carbamate as a colorless
oil (218 g, 88%).
F igu r e 1. Plasma concentrations of 4 and ex vivo inhibition
of LTB4 after a 10 mg/kg oral dose in dog. Plasma concentra-
tions were determined by HPLC. Data presented are the mean
from two dogs.
A solution of aniline (83.7 g, 0.9 mol), ethyl N-(2-bromoeth-
yl)carbamate (190.0 g, 0.92 mol), and diisopropylethylamine
(90.3 g, 0.7 mol) in benzene (700 mL) was heated at reflux for
24 h. The reaction mixture was allowed to cool to room
temperature, water (500 mL) was added, and the organic layer
was separated, washed with saturated aqueous NaCl, dried
(MgSO4), filtered, and concentrated in vacuo to give ethyl N-[2-
(phenylamino)ethyl]carbamate (118 g, 63%).
To ethyl N-[2-(phenylamino)ethyl]carbamate (118 g, 0.57
mol) was added an ice cold solution of concentrated HCl (120
mL) dissolved in water (200 mL). With mechanical stirring
at 0 °C, a solution of NaNO2 (41.4 g, 0.6 mol) in water (100
mL) was added dropwise. The mixture was stirred for 2 h at
0 °C, and a solid precipitate formed. The solid was collected
by filtration, washed with water (2 × 100 mL), and dried in
vacuo to provide ethyl N-[2-(phenylnitrosoamino)ethyl]car-
bamate as a greenish yellow solid (115 g, 85%).
To a mechanically stirred suspension of zinc dust (115 g,
0.48 mol) in water (350 mL) was added dropwise a solution of
N-nitroso intermediate (115 g) dissolved in acetic acid (230
mL). The temperature was controlled between 5-15 °C during
the addition after which the cooling bath was removed, and
the mixture was stirred at 30 °C for 1 h. Dichloromethane
(700 mL) was added, and the mixture was filtered. The
organic layer was separated, washed with 10% Na2CO3 (2 ×
350 mL) and saturated aqueous NaCl, dried (MgSO4), filtered,
and concentrated in vacuo to give N-[2-[(ethoxycarbonyl)-
amino]ethyl]-N-phenylhydrazine (93 g, 87%).
To a stirred solution of the hydrazine intermediate (93 g,
0.42 mol) in dichloromethane (350 mL) cooled to -40 °C was
added dropwise ethylmagnesium bromide (225 mL, 2.0 M in
tetrahydrofuran) while the temperature was maintained below
-30 °C. After the addition, the mixture was allowed to warm
to room temperature and then heated at 40 °C for 48 h. Ice
chips (500 g) were added, and the mixture was acidified with
3 N HCl to pH 3. The phases were separated, and the aqueous
phase was extracted with 5% methanol in dichloromethane
(2 × 300 mL). The combined organic extracts were washed
with saturated aqueous NaCl, dried (MgSO4), filtered, and
concentrated in vacuo. The residue was suspended in 2%
MeOH in dichloromethane (100 mL), and the solids were
collected by filtration to provide 4 (28.7 g, 39%): mp 219 °C;
1H NMR (300 MHz, DMSO-d6) 3.01-3.07 (2 H, m), 3.63 (2 H,
t, J ) 5.5 Hz), 6.64 (1 H, br s), 6.87 (1 H, t, J ) 7.5 Hz), 7.03
(2 H, d, J ) 8 Hz), 7.27 (2 H, t, J ) 8 Hz), 8.42 (1 H, br s).
Anal. (C9H11N3O) C, H, N.
F igu r e 2. Plasma concentrations of 4 and ex vivo inhibition
of LTB4 after a 10 mg/kg oral dose in cynomolgus monkeys.
Plasma concentrations were determined by HPLC. Data
presented are the mean from two monkeys.
to cynomolgus monkeys (n ) 2) provided greater than
50% ex vivo leukotriene inhibition in blood samples
taken over the first 4 h.
In pilot subacute toxicity testing, 4 did not produce
methemoglobinemia in rats after repeated oral daily
doses of 400 mg/kg for 9 days. In a dog toxicity study,
oral daily doses of 200 mg/kg of 4 for 1 month also
showed no methemoglobinemia. Therefore, the phenyl-
triazinone heterocycle represented by 4 provided a 5-LO
inhibitor pharmacophore which was devoid of the meth-
emoglobinemia-associated toxicity problem previously
observed for the pyridazinone series represented by 3.13
The SAR study did not identify triazinone analogs with
in vitro potency below 1 µM and at the same time exhibit
in vivo activity with ED50 values below 10 mg/kg.
Triazinone 4 is an easily synthesized, low molecular
weight, selective, orally bioavailable 5-LO inhibitor
useful for in vivo pharmacologic studies involving bio-
synthetic products derived from 5-lipoxygenase.
1-(2-Met h ylp h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
on e (5). Meth od A: 25% overall yield; mp 230 °C; NMR (300
MHz, DMSO-d6) 2.23 (3 H, s), 2.99-2.93 (2 H, m), 3.22 (2 H,
t, J ) 5 Hz), 6.78 (1 H, br s), 6.99 (1 H, dt, J ) 6, 1 Hz), 7.04
(1 H, d, J ) 7 Hz), 7.16 (2 H, d, J ) 7 Hz), 8.19 (1 H, br s).
Anal. (C10H13N3O) C, H, N.
Exp er im en ta l Section
Ch em istr y. Gen er a l. Melting points were determined in
open glass capillaries and are uncorrected. 1H NMR spectra
were recorded on a GE QE300 spectrometer and chemical
shifts are reported in parts per million (ppm, δ) relative to

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3945
1-(3-Met h ylp h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
on e (6). Meth od A: 18% overall yield; mp 235-236 °C; H
NMR (300 MHz, DMSO-d6) 2.26 (3 H, s), 3.05-2.98 (2 H, m),
3.61 (2 H, t, J ) 5 Hz), 6.63 (1 H, br s), 6.68 (1 H, d, J ) 7.0
Hz), 6.87-6.80 (2 H, m), 7.14 (1 H, t, J ) 7.5 Hz), 8.38 (1 H,
br s). Anal. (C10H13N3O) C, H, N.
Hz), 3.73 (3 H, s), 6.47 (1 H, dd, J ) 9, 2 Hz), 6.58 (1 H, t, J )
2.5 Hz), 6.63 (1 H, dd, J ) 9, 1 Hz), 6.75 (1 H, br s), 7.16 (1 H,
t, J ) 8 Hz), 8.46 (1 H, br s). Anal. (C10H13N3O2) C, H, N.
1-[3-(Meth ylth io)ph en yl]-2H,4H-tetr ah ydr o-1,2,4-tr iazin -
3-on e (16). Meth od A: 28% overall yield; mp 179-180 °C;
1H NMR (300 MHz, DMSO-d6) 2.45 (3 H, s), 3.01-3.07 (2 H,
m), 3.63 (2 H, t, J ) 6 Hz), 6.67 (1 H, br s), 6.76 (1 H, d, J )
7 Hz), 6.82 (1 H, dd, J ) 9, 1.5 Hz), 6.9 (1 H, t, J ) 1.5 Hz),
7.21 (1 H, t, J ) 8 Hz), 8.56 (1 H, br s). Anal. (C10H13N3-
OS‚0.25H2O) C, H, N.
1-[3-(P h en ylm eth oxy)p h en yl]-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (17). Meth od A: 18% overall yield; mp 169 °C;
1H NMR (300 MHz, DMSO-d6) 3.01-3.07 (2 H, m), 3.62 (2 H,
t, J ) 6 Hz), 5.06 (2 H, s), 6.54 (1 H, dd, J ) 9, 1.5 Hz), 6.61-
6.66 (2 H, m), 6.68 (1 H, t, J ) 2 Hz), 7.17 (1 H, t, J ) 8 Hz),
7.28-7.47 (5 H, m), 8.42 (1 H, d, J ) 1 Hz). Anal.
(C16H17N3O‚0.75H2O) C, H, N.
1-(3-Hyd r oxyp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-
on e (18). A mixture of 17 (0.5 g, 1.8 mmol) in methanol (50
mL) was hydrogenated at 4 atm with 20% Pd/C (0.2 g) at room
temperature for 4 h. The mixture was filtered, concentrated
in vacuo, and crystallized from ethyl acetate to provide 18 (0.22
g, 60%): mp 244 °C; 1H NMR (300 MHz, DMSO-d6) 3.03 (2 H,
m), 3.53 (2 H, t, J ) 7 Hz), 6.27 (1 H, m), 6.44 (2 H, m), 6.62
(1 H, br s), 7.03 (1 H, t, J ) 7.5 Hz), 8.73 (1 H, br s), 9.28 (1 H,
br s). Anal. (C9H11N3O2) C, H, N.
1
1-(4-Met h ylp h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
1
on e (7). Meth od A: 23% overall yield; mp 218-220 °C; H
NMR (300 MHz, CDCl3, CD3OD) 2.22 (3 H, s), 3.14 (2 H, t, J
) 5 Hz), 3.52 (2 H, t, J ) 5 Hz), 6.87 (2 H, d, J ) 9 Hz), 7.03
(2 H, d, J ) 9 Hz); MS m/e M+ 191. Anal. (C10H13N3O‚0.5H2O)
C, H, N.
1-(3′,5′-Dim eth ylph en yl)-2H,4H-tetr ah ydr o-1,2,4-tr iazin -
3-on e (8). Meth od A: 27% overall yield; mp 255-257 °C; 1H
NMR (300 MHz, DMSO-d6) 2.20 (6 H, s), 2.99-3.06 (2 H, m),
3.58 (2 H, t, J ) 5 Hz), 6.51 (1 H, s), 6.6 (1 H, br s), 6.65 (2 H,
s), 8.33 (1 H, d, J ) 1.5 Hz). Anal. (C11H15N3O) C, H, N.
1-(3-E t h ylp h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
1
on e (9). Meth od A: 22% overall yield; mp 204-205 °C; H
NMR (300 MHz, DMSO-d6) 1.16 (3 H, t, J ) 7.5 Hz), 2.56 (2
H, q, J ) 7 Hz), 3.01-3.07 (2 H, m), 3.62 (2 H, t, J ) 5 Hz),
6.63 (1 H, br s), 6.73 (1 H, d, J ) 7.5 Hz), 6.82-6.9 (2 H, m),
7.16 (1 H, t, J ) 7.5 Hz), 8.38 (1 H, br s). Anal. (C11H15N3O)
C, H, N.
Meth od B (Sch em e 3). 1-(3-F lu or op h en yl)-2H,4H-tet-
r a h yd r o-1,2,4-tr ia zin -3-on e (10). To a stirred suspension
of (3-fluorophenyl)hydrazine hydrochloride (4.06 g, 25 mmol)
in dichloromethane (150 mL) was added triethylamine (2.5 g,
25 mmol), and the mixture was stirred for 15 min, after which
2-chloroethyl isocyanate (2.6 g, 25 mmol) was added dropwise.
The mixture was stirred for 2 h, water (100 mL) was added,
and the organic layer was separated, washed with brine, dried
(MgSO4), filtered, and concentrated in vacuo to give N-(4-
fluorophenyl)-N ′-[[(2-chloroethyl)amino]carbonyl]hydrazine (5.03
g, 87%).
A mixture of N-(4-fluorophenyl)-N ′-[[(2-chloroethyl)amino]-
carbonyl]hydrazine (2.3 g, 10 mmol) in DMF (20 mL) and NaI
(2.99 g, 20 mmol) was stirred at 75 °C for 48 h and allowed to
cool to room temperature, and water (10 mL) was added
followed by dropwise addition of a saturated solution of
NaHSO3 to consume the iodine color. The pH of the mixture
was adjusted to 8-9 by adding 2 N NaOH and was concen-
trated in vacuo to give an oil which was purified by chroma-
tography (silica gel, 7% methanol in dichloromethane) to
provide 10 (0.8 g, 46%): mp 194-196 °C; 1H NMR (300 MHz,
DMSO-d6) 2.17-2.23 (2 H, m), 2.8 (2 H, t, J ) 6 Hz), 5.79 (1
H, dt, J ) 8, 1.5 Hz), 5.85 (1 H, br s), 5.94-6.04 (2 H, m),
6.36-6.44 (1 H, m), 7.76 (1 H, br s). Anal. (C9H10FN3O) C,
H, N.
1-(3-Nit r op h e n yl)-2H ,4H -t e t r a h yd r o-1,2,4-t r ia zin -3-
1
on e (19). Meth od B: 42% overall yield; mp 205-208 °C; H
NMR (300 MHz, DMSO-d6) 3.06-3.14 (2 H, m), 3.79 (2 H, t, J
) 5 Hz), 6.82 (1 H, br s), 7.49-7.59 (2 H, m), 7.68-7.74 (1 H,
m), 7.83 (1 H, t, J ) 7 Hz), 8.73 (1 H, br s). Anal. (C9H10N4O3)
C, H, N.
1-(3-Am in op h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
on e (20). A solution of 19 in ethyl acetate was hydrogenated
at 4 atm with 20% Pd/C at room temperature for 4 h to provide
20 (0.085 g, 40%): mp 214-216 °C; 1H NMR (300 MHz,
DMSO-d6) 3.02 (2 H, m), 3.5 (2 H, t, J ) 7 Hz), 4.97 (2 H, s),
6.08 (1 H, dd, J ) 9, 1.5 Hz), 6.18 (1 H, dd, J ) 9, 1.5 Hz),
6.55 (1 H, br s), 6.88 (1 H, t, J ) 7.5 Hz), 8.21 (1 H, br s).
Anal. (C9H12N4O) C, H, N.
Meth od C (Sch em e 4). 5-Meth yl-1-p h en yl-2H,4H-tet-
r a h yd r o-1,2,4-tr ia zin -3-on e (21). A solution of D,L-2-amino-
1-propanol (7.51 g, 0.10 mol) in acetonitrile (100 mL) and
K2CO3 (27.6 g, 0.20 mol) was mechanically stirred at -20 °C
while a solution of methyl chloroformate (9.45 g, 0.10 mol) in
acetonitrile (20 mL) was added over 0.5 h. The reaction
mixture was stirred overnight at room temperature, filtered,
and concentrated in vacuo to give 2-[(methoxycarbonyl)amino]-
1-propanol (8.47 g, 64%).
1-(3-Ch lor op h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
on e (11). Meth od A: 18% overall yield; mp 244-245 °C; H
To a stirred solution of oxalyl chloride (4.36 g, 34.0 mmol)
in methylene chloride (30 mL) at -78 °C was added, dropwise,
1
NMR (300 MHz, DMSO-d6) 3.03-3.09 (2 H, m), 3.67 (2 H, t, J
) 5 Hz), 6.72 (1 H, br s), 6.9 (1 H, dd, J ) 9, 1.5 Hz), 7.01 (1
H, dd, J ) 9, 1.5 Hz), 7.07 (1 H, t, J ) 2.5 Hz), 7.27 (1 H, t, J
) 8 Hz), 8.53 (1 H, br s). Anal. (C9H10ClN3O) C, H, N.
1-(3-Br om op h en yl)-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
a
solution of dimethyl sulfoxide (5.28 g, 68.0 mmol) in
methylene chloride (10 mL) over 15 min. The mixture was
stirred for another 15 min at -78 °C followed by the addition
of 2-[(methoxycarbonyl)amino]-1-propanol (3.01 g, 22.6 mmol)
in methylene chloride (30 mL). The mixture was stirred for 1
h at -78 °C, then triethylamine (7.26 g, 72.0 mmol) was added,
and the mixture was allowed to warm to room temperature,
filtered, and concentrated in vacuo at 30 °C to provide
2-[(methoxycarbonyl)amino]-1-propanal (2.40 g, 80%).
To a solution of 2-[(methoxycarbonyl)amino]-1-propanal in
methanol (100 mL) was added aniline (2.05 g, 22.0 mmol) and
the mixture adjusted to pH 5-6 with 10% ethanolic HCl.
Sodium cyanoborohydride (1.57 g, 25 mmol) was added slowly,
and the mixture was stirred at room temperature for 16 h,
maintaining the pH at 5-6 with occasional addition of 10%
ethanolic HCl. The mixture was acidified to pH 1 with 10%
ethanolic HCl and concentrated in vacuo at 30 °C. The residue
was dissolved in methylene chloride (100 mL) and washed with
10% K2CO3 (20 mL), the organic layer was dried over MgSO4
and concentrated in vacuo, and the residue was purified by
chromatography (silica gel, 10% methylene chloride/ether) to
provide methyl N-[1-(phenylamino)prop-2-yl]carbamate (2.09
g, 45%).
1
on e (12). Meth od A: 21% overall yield; mp 260-262 °C; H
NMR (300 MHz, DMSO-d6) 3.03-3.09 (2 H, m), 3.67 (2 H, t,
J ) 6 Hz), 6.77 (1 H, br s), 7.01-7.07 (2 H, m), 7.22 (2 H, t,
J ) 7.5 Hz), 8.54 (1 H, br s). Anal. (C9H10BrN3O‚0.25H2O)
C, H, N.
1-(3,5-Diflu or op h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (13). Meth od B: 39% overall yield; mp 238-240 °C;
1H NMR (300 MHz, DMSO-d6) 3.07-3.13 (2 H, m), 3.69 (2 H,
t, J ) 5 Hz), 6.65 (1 H, tt, J ) 8, 2 Hz), 6.73 (1 H, d, J ) 2 Hz),
6.77 (1 H, d, J ) 2 Hz), 6.82 (b, 1 H), 8.62 (br s). Anal.
(C9H9F2N3O) C, H, N.
1-[3-(Tr iflu or om eth yl)p h en yl]-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (14). Meth od A: 16% overal yield; mp 239-
241 °C; 1H NMR (300 MHz, DMSO-d6) 3.04-3.11 (2 H, m),
3.76 (2 H, t, J ) 5.5 Hz), 6.79 (1 H, br s), 7.20 (1 H, d, J ) 7
Hz), 7.36 (2 H, d, J ) 10 Hz), 7.5 (1 H, t, J ) 7.5 Hz), 8.63 (1
H, d, J ) 1.5 Hz). Anal. (C10H10F3N3O) C, H, N.
1-(3-Meth oxyp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-
on e (15). Meth od A: 23% overall yield; mp 183 °C; 1H NMR
(300 MHz, DMSO-d6) 3.01-3.09 (2 H, m), 3.62 (2 H, t, J ) 6
To a mechanically stirred solution of the carbamate inter-
mediate (4.54 g, 0.021 mol) in water (20 mL) chilled to -10

3946 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
Bhatia et al.
°C was added concentrated HCl (4.5 mL) followed by dropwise
addition of sodium nitrite (1.66 g, 0.02 mol) in water (4 mL).
Dimethoxyethane (15 mL) was added to facilitate the stirring.
After 1 h at 0 °C the mixture was extracted with benzene and
the extracts were dried (MgSO4) and concentrated in vacuo to
yield methyl N-[1-(phenylnitrosoamino)prop-2-yl]carbamate
(5.05 g, 97%).
5-Met h yl-1-(3-ch lor op h en yl)-2H ,4H -t et r a h yd r o-1,2,4-
tr ia zin -3-on e (28). Meth od C: 21% overall yield; mp 246-
247 °C; 1H NMR (300 MHz, DMSO-d6) 1.02 (3 H, d, J ) 7 Hz),
2.92 (1 H, dd, J ) 13, 10 Hz), 3.22-3.33 (2 H, m), 4.08 (1 H,
dd, J ) 13, 4 Hz), 6.77 (1 H, br s), 6.88 (1 H, d, J ) 7.5 Hz),
7.01 (1 H, dd, J ) 7.5, 1.0 Hz), 7.27 (1 H, t, J ) 7.5 Hz), 8.53
(1 H, d, J ) 1 Hz). Anal. (C11H12ClN3O) C, H, N.
A suspension of zinc dust (5.30 g, 0.08 mol) in water (15
mL) was stirred and cooled to 15 °C while a solution of the
N-nitroso intermediate (5.05 g, 0.021 mol) in acetic acid (12
mL) was added dropwise while a reaction temperature of 15-
20 °C was maintained. The mixture was allowed to warm to
room temperature and stirred for 1 h, after which water (100
mL) and methylene chloride (100 mL) were added and the pH
was adjusted to 6 with dilute NaOH. The layers were decanted
from the zinc residue, and the organic layer was separated,
dried (MgSO4), filtered, and concentrated in vacuo to provide
crude N-[2-[(methoxycarbonyl)amino]propyl-N-phenylhydra-
zine (4.28 g, 90%).
1-(3-Br om op h en yl)-5-m et h yl-2H ,4H -t et r a h yd r o-1,2,4-
tr ia zin -3-on e (29). Meth od B: 36% overall yield; mp 230-
232 °C; 1H NMR (300 MHz, DMSO-d6) 1.1 (3 H, d, J ) 7 Hz),
2.91 (1 H, dd, J ) 14, 10 Hz), 3.22-3.34 (2 H, m), 3.9 (1 H, dd,
J ) 14, 3 Hz), 6.79 (1 H, s), 7.02 (2 H, t, J ) 9 Hz), 7.20 (2 H,
t, J ) 8 Hz), 8.02 (1 H, s). Anal. (C10H12BrN3O) C, H, N.
5-P h en oxy-1-p h en yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-
on e (30). Meth od B: 34% overall yield; mp 161 °C; 1H NMR
(300 MHz, DMSO-d6) 3.08 (2 H, m), 3.62 (2 H, t, J ) 5.5 Hz),
6.45 (1 H, dd, J ) 9, 2 Hz), 6.67 (1 H, br s), 6.74 (1 H, t, J )
2 Hz), 6.83 (1 H, dd, J ) 9, 2 Hz), 6.98 (2 H, m), 7.12 (1 H, t,
J ) 7.5 Hz), 7.25 (1 H, t, J ) 7.5 Hz), 7.38 (2 H, m), 8.45 (1 H,
d, J ) 3 Hz). Anal. (C15H15N3O2) C, H, N.
To a stirred solution of the hydrazine intermediate (4.28 g,
0.019 mol) in methylene chloride (25 mL) at -25 °C was added
dropwise EtMgBr (2 M solution in THF, 12.5 mL, 0.025 mol),
and then the mixture was heated at 45 °C for 48 h, treated
5-(4-F lu or op h en oxy)-1-p h en yl-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (31). Meth od B: 37% overall yield; mp 199
°C; 1H NMR (300 MHz, DMSO-d6) 3.03 (2 H, m), 3.70 (2 H, t,
J ) 5.5 Hz), 6.38 (1 H, dd, J ) 9, 2 Hz), 6.62 (1 H, br s), 6.68
(1 H, t, J ) 2 Hz), 6.77 (1 H, dd, J ) 9, 2 Hz), 7.01 (2 H, m),
with ice water, and adjusted to pH 3-4 with 6 N HCl.
A
portion of the precipitated product was isolated by filtration,
while the remainder was isolated by drying and evaporation
of the methylene chloride layer. The combined solids were
crystallized from ethanol to provide 21 (1.15 g, 32%): mp 238-
7.15 (3 H, m), 8.41 (1 H, d, J ) 3 Hz). Anal. (C15H14
FN3O2‚0.25H2O) C, H, N.
-
Meth od D (Sch em e 5). 5-(Hyd r oxym eth yl)-1-p h en yl-
2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (32). To a solution
of aniline (2.79 g, 0.03 mol) in dichloromethane (10 mL) cooled
to 0 °C was added dropwise EtMgBr (2 M in THF, 15.0 mL,
0.03 mol). The mixture was stirred for 30 min, followed by
portionwise addition of DL-serine methyl ester hydrochloride
(1.56 g, 0.01 mol) over a 1 h period. The thick reaction mixture
was diluted with additional dichloromethane (10 mL) and then
heated at reflux for 3 days. The mixture was cooled to room
temperature, saturated aqueous NH4Cl (50 mL) was added,
the mixture was extracted with dichloromethane (2 × 100 mL),
dried (MgSO4), and concentrated in vacuo, and the residue was
triturated in pentane/ether (1:1) to give DL-serine phenylamide
(0.47 g, 26%): mp 116-118 °C.
To a solution of DL-serine phenylamide (1.96 g, 0.01 mol) in
THF (50 mL) was added dropwise a suspension of LiAlH4 (0.84
g, 0.02 mol) in THF (35 mL) with stirring at room temperature.
The mixture was stirred at reflux overnight, cooled, and
quenched by dropwise addition of water (0.9 mL), followed by
15% NaOH solution (0.9 mL) and then more water (2.7 mL).
The mixture was stirred for 3 h and filtered, and the filtrate
was concentrated in vacuo. The residue which began to
solidify within a short time was triturated in ether (10 mL)
and the solid collected to give 2-amino-1-(phenylamino)-3-
propanol (1.2 g, 67%): mp 89-92 °C.
2-Amino-1-(phenylamino)-3-propanol (1.20 g, 0.007 mol) and
triethylamine (0.73 g, 7 mmol) in THF (50 mL) was stirred at
-5 °C while a solution of methyl chloroformate (0.68 g, 7 mmol)
in THF (15 mL) was added dropwise. The mixture was stirred
at -5 °C for 2 h, allowed to warm to room temperature,
filtered, and concentrated in vacuo to give methyl N-[1-
(phenylamino)-3-hydroxyprop-2-yl]carbamate (1.73 g, 99%).
A mixture of methyl N-[1-(phenylamino)-3-hydroxyprop-2-
yl]carbamate (1.73 g, 7.7 mmol), concentrated HCl (2.0 mL),
water (10 mL), and dimethoxyethane (6 mL) was stirred at 0
°C and NaNO2 (0.59 g, 8.5 mmol) in water (1.0 mL) was added
dropwise. The mixture was stirred at 0 °C for 1 h, extracted
with dichloromethane (100 mL), and concentrated in vacuo to
give methyl N-[1-(nitrosophenylamino)-3-hydroxyprop-2-yl]-
carbamate (1.78 g, 91%).
To a stirred suspension of zinc dust (1.80 g, 0.027 mol) in
water (5.3 mL) cooled to 15 °C was added dropwise a solution
of methyl N-[1-(nitrosophenylamino)-3-hydroxyprop-2-yl]car-
bamate (1.78 g, 0.007 mol) in acetic acid (4.5 mL). After the
addition, the cooling bath was removed and the mixture was
stirred at room temperature for 1 h. Dichloromethane (10 mL)
was added, the mixture was adjusted to pH 6 with 15% NaOH,
and the organic and aqueous layers were decanted from the
zinc residue which was extracted with a fresh portion (10 mL)
1
240 °C; H NMR (300 MHz, DMSO-d6) 1.00 (3 H, d, J ) 6.0
Hz), 2.89 (1 H, dd, J ) 15, 10.5 Hz), 3.25 (1 H, m), 4.03 (1 H,
dd, J ) 13.5, 4.5 Hz), 6.68 (1 H, s), 6.86 (1 H, t, J ) 7.5 Hz),
7.03 (2 H, d, J ) 7.5 Hz), 7.25 (2 H, t, J ) 7.5 Hz), 8.42 (1 H,
d, J ) 1.5 Hz). Anal. (C10H13N3O) C, H, N.
5,5-Dim eth yl-1-p h en yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (22). Meth od C: 14% overall yield; mp 273 °C; 1H NMR
(300 MHz, DMSO-d6) 0.95 (6 H, s), 3.58 (2 H, s), 6.72 (1 H, t,
J ) 7.5 Hz), 6.81 (1 H, br s), 7.01 (2 H, d, J ) 7.5 Hz), 7.2 (2
H, t, J ) 7.5 Hz), 8.39 (1 H, br s). Anal. Calcd for
C11H15N3O: C, 64.37; H, 7.37; N, 20.47. Found: C, 64.39; H,
7.34; N, 20.55.
5-E t h yl-1-p h e n yl-2H ,4H -t e t r a h yd r o-1,2,4-t r ia zin -3-
on e (23). Meth od C: 41% overall yield; mp 205 °C; 1H NMR
(300 MHz, DMSO-d6) 0.82 (3 H, t, J ) 7 Hz), 1.3-1.45 (2 H,
m), 2.99 (1 H, dd, J ) 12, 10 Hz), 3.03-3.12 (2 H, m), 4.04 (1
H, dd, J ) 13, 3 Hz), 6.68 (1 H, br s), 6.85 (1 H, t, J ) 7.5 Hz),
7.03 (2 H, d, J ) 7.5 Hz), 7.25 (2 H, t, J ) 7.5 Hz), 8.39 (1 H,
br s). Anal. (C11H15N3O) C, H, N.
5-Isop r op yl-1-p h en yl-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -
3-on e (24). Meth od C: 18% overall yield; mp 181-182 °C;
1H NMR (300 MHz, DMSO-d6) 0.85 (6 H, d, J ) 7.5 Hz), 1.64
(1 H, m), 2.96-3.12 (2 H, m), 4.00 (1 H, d, J ) 7.5 Hz), 6.56 (1
H, br s), 6.86 (1 H, t, J ) 7.5 Hz), 7.03 (2 H, d, J ) 7.5 Hz),
7.26 (2 H, t, J ) 7.5 Hz), 8.44 (1 H, d, J ) 1.5 Hz). Anal.
(C12H17N3O) C, H, N.
5-n -Bu t yl-1-p h en yl-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
1
on e (25). Meth od C: 35% overall yield; mp 153-155 °C; H
NMR (300 MHz, DMSO-d6) 0.83 (3 H, t, J ) 7.5 Hz), 1.10-
1.50 (6 H, m), 2.99 (1 H, dd, J ) 15, 10.5 Hz), 3.09-3.19 (1 H,
m), 4.03 (1 H, dd, J ) 13.5, 4.5 Hz), 6.62 (1 H, s), 6.86 (1 H, t,
J ) 7.5 Hz), 7.04 (2 H, d, J ) 7.5 Hz), 7.26 (2 H, t, J ) 7.5
Hz), 8.42 (1 H, d, J ) 1.5 Hz). Anal. (C13H19N3O‚0.125H2O)
C, H, N.
5-Meth yl-1-(3-m eth ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (26). Meth od C: 14% overall yield; mp 228-
229 °C; 1H NMR (300 MHz, DMSO-d6) 1.0 (3 H, d, J ) 6 Hz),
2.26 (3 H, s), 2.86 (1 H, dd, J ) 13, 10 Hz), 3.21-3.31 (1 H,
m), 4.03 (1 H, dd, J ) 13, 4.5 Hz), 6.57-6.62 (2 H, m), 6.8-
6.87 (2 H, m), 7.13 (1 H, t, J ) 7.5 Hz), 8.39 (1 H, br s). Anal.
(C11H15N3O) C, H, N.
1-(3-F lu or op h en yl)-5-m et h yl-2H ,4H -t et r a h yd r o-1,2,4-
tr ia zin -3-on e (27). Meth od B: 42% overall yield; mp 238-
240 °C; 1H NMR (300 MHZ, DMSO-d6) 1.1 (3 H, d, J ) 6 Hz),
2.91 (1 H, dd, J ) 14 Hz, J ) 10 Hz), 3.24-3.34 (2 H, m), 4.09
(1 H, dd, J ) 14 Hz, J ) 3 Hz), 6.14 (1 H, dt, J ) 8 Hz, J ) 2
Hz), 6.77 (1 H, br s), 6.8-6.89 (2 H, m), 7.23-7.32 (1 H, m),
8.52 (1 H, br s). Anal. (C10H12FN3O) C, H, N, F.

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3947
of dichloromethane. The combined organic layers were dried
(MgSO4) and concentrated in vacuo to yield N-[2-[(methoxy-
carbonyl)amino]-3-hydroxyprop-1-yl]-N-phenylhydrazine (1.53
g, 92%).
°C dec; 1H NMR (300 MHz, DMSO-d6) 1.03 (3 H, d, J ) 6 Hz),
3.02 (1 H, dd, J ) 14, 9 Hz), 3.31-3.42 (1 H, m), 4.37 (1 H, dd,
J ) 14, 4 Hz), 6.85 (1 H, d, J ) 7 Hz), 6.93 (1 H, d, J ) 7 Hz),
6.95 (1 H, br s), 6.69 (1 H, t, J ) 8 Hz), 8.69 (1 H, br s). Anal.
To a solution of the hydrazine intermediate (1.53 g, 0.006
mol) in dichloromethane (10 mL), chilled to -30 °C, was added
dropwise EtMgBr (2 M in THF, 12.0 mL, 0.024 mol), and the
mixture was heated at 50 °C for 4 days, cooled, and treated
with a solution of concentrated HCl (2.0 mL) in methanol (25
mL). The mixture was concentrated in vacuo, the residue was
extracted with 10% methanol in dichloromethane (4 × 25 mL),
dried (MgSO4), and concentrated in vacuo, and the residue
was purified by chromatography (silica gel, 1% methanol/
dichloromethane) to give 32 (9 mg, 0.7%): mp 223-224 °C dec;
1H NMR (300 MHz, DMSO-d6) 3.08-3.30 (4 H, m), 4.01 (1 H,
d, J ) 10.5 Hz), 4.77 (1 H, t, J ) 6.0 Hz), 6.46 (1 H, br s), 6.86
(1 H, t, J ) 7.5 Hz), 7.02 (2 H, d, J ) 7.5 Hz), 7.26 (2 H, t, J
) 7.5 Hz), 8.47 (1 H, d, J ) 1.5 Hz). Anal. (C10H13N3O2) C,
H, N.
(C9H11ClN4O‚0.125H2O) C, H, N.
1-(6-Br om op yr id -2-yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (43). Meth od B: 13% overall yield; mp 237-239.5 °C;
1H NMR (300 MHz, DMSO-d6) 3.10 (2 H, m), 3.86 (2 H, t,
J ) 5.25 Hz), 6.84 (1 H, br S), 6.99 (1 H, d, J ) 9.0 Hz), 7.86
(1 H, dd, J ) 3.0, 9.0 Hz), 8.26 (1 H, d, J ) 3.0 Hz), 8.67 (1 H,
d, J ) 1.5 Hz); MS m/e 257 (M + H)+. Anal. (C8H9BrN4O) C,
H, N.
1-(6-Br om opyr id-2-yl)-5-m eth yl-2H,4H-tetr ah ydr o-1,2,4-
tr ia zin -3-on e (44). Meth od B: 12% overall yield; mp 252-
253 °C; 1H NMR (300 MHz, DMSO-d6) 1.04 (3 H, d, J ) 6 Hz),
3.03 (1 H, dd, J ) 14, 9 Hz), 3.31-3.43 (1 H, m), 4.35 (1 H, dd,
J ) 14, 4 Hz), 6.94-7.0 (3 H, m), 7.58 (1 H, t, J ) 8 Hz), 8.69
(1 H, br s). Anal. (C9H11BrN4O‚0.25H2O) C, H, N.
1-(6-Meth oxypyr id-2-yl)-2H,4H-tetr ah ydr o-1,2,4-tr iazin -
3-on e (45). Meth od B: 18% overall yield; mp 235-236 °C;
1H NMR (300 MHz, DMSO-d6) 3.07-3.13 (2 H, m), 3.79 (3 H,
s), 3.86 (2 H, t, J ) 5 Hz), 6.22 (1 H, d, J ) 7 Hz), 6.53 (1 H,
d, J ) 7 Hz), 6.74 (1 H, br s), 7.57 (1 H, t, J ) 8 Hz), 8.61 (s).
Anal. (C9H12N4O2‚0.5H2O) C, H, N.
5-[(Meth oxyeth oxy)m eth yl]-1-ph en yl-2H,4H-tetr ah ydr o-
1,2,4-tr ia zin -3-on e (33). Meth od B: 2% overall yield; mp
118-120 °C; 1H NMR (300 MHz, DMSO-d6) 3.35 (3 H, s), 3.44-
3.96 (9 H, m), 5.42 (1 H, m), 6.85 (2 H, d, J ) 7.5 Hz), 6.92 (1
H, t, J ) 7.5 Hz), 7.28 (2 H, t, J ) 7.5 Hz), 8.50 (1 H, s). Anal.
(C13H19N3O3) C, H, N.
5-Met h yl-1-(6-m et h oxyp yr id -2-yl)-2H ,4H -t et r a h yd r o-
1,2,4-tr ia zin -3-on e (46). Meth od B: 9% overall yield; mp
196-197 °C; 1H NMR (300 MHz, DMSO-d6) 1.02 (3 H, d, J )
6 Hz), 3.01 (1 H, dd, J ) 14, 9 Hz), 3.27-3.41 (4 H, m, s), 4.43
(1 H, dd, J ) 14, 3 Hz), 6.2 (1 H, d, J ) 7 Hz), 6.51 (1 h, d, J
) 7 Hz), 6.82 (1 H, br s), 7.55 (1 H, t, J ) 8 Hz), 8.61 (1 H, br
s). Anal. (C10H14N4O2‚0.25H2O) C, H, N.
1-P yr id -2-yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (34).
Meth od B: 35% overall yield; mp 174-176 °C; 1H NMR (300
MHz, DMSO-d6) 3.06-3.13 (2 H, m), 3.87 (2 H, t, J ) 6 Hz),
6.77 (1 H, br s), 6.80-6.85 (1 H, m), 7.03 (1 H, d, J ) 7.5 Hz),
7.64-7.7 (1 H, m), 8.15-8.2 (1 H, m), 8.63 (1 H, br s). Anal.
(C8H10N4O‚0.125H2O) C, H, N.
1-P yr id -3-yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (35).
1-(7-Ch lor oqu in ol-4-yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
1
1
Meth od B: 5% overall yield; H NMR (300 MHz, DMSO-d6)
3-on e (47). Meth od B: 26% overall yield; mp >250 °C; H
3.08 (2 H, m), 3.69 (2 H, t, J ) 5.5 Hz), 6.75 (1 H, br s), 7.29
(1 H, m), 7.41 (1 H, m), 8.08 (1 H, dd, J ) 6, 1.5 Hz), 8.38 (1
H, d, J ) 3 Hz), 8.55 (1 H, br s). Anal. (C8H10N4O) C, H, N.
1-P yr id -4-yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (36).
Meth od B: 21% overall yield; mp 179-181 °C; 1H NMR (300
MHz, DMSO-d6) 3.07 (2 H, q, J ) 6 Hz), 3.22-3.29 (2 H, m),
5.72 (1 H, t, J ) 7 Hz), 7.27 (1 H, br s), 7.72 (2 H, d, J ) 6 Hz),
8.31 (2 H, d, J ) 6 Hz). Anal. (C8H10N4O) C, H, N.
1-(6-Meth ylp yr id -2-yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (37). Meth od B: 36% overall yield; mp 191-193 °C;
1H NMR (300 MHz, DMSO-d6) 2.35 (1 H, s), 3.04-3.11 (2 H,
m), 3.87 (2 H, t, J ) 5 Hz), 6.68 (1 H, d, J ) 7 Hz), 6.73 (1 H,
bs), 6.8 (1 H, d, J ) 8 Hz), 7.55 (1 H, t, J ) 8 Hz) 8.58 (1 H, s);
MS m/e 193 (M + H)+. Anal. Calcd for C9H12N4O: C, 56.23;
H, 6.29; N, 29.14. Found: C, 55.80; H, 5.97; N, 29.05.
5-Meth yl-1-(6-m eth ylpyr id-2-yl)-2H,4H-tetr ah ydr o-1,2,4-
tr ia zin -3-on e (38). Meth od B: 12% overall yield; mp 253-
254 °C; 1H NMR (300 MHz, DMSO-d6) 1.02 (3 H, d, J ) 7 Hz),
2.33 (3 H, s), 2.9 (1 H, dd, J ) 14, 10 Hz), 3.29-3.34 (1 H, m),
4.53 (1 H, dd, J ) 14, 4 Hz), 6.65 (1 H, d, J ) 7 Hz), 6.76 (1 H,
br s), 6.78 (1 H, d, J ) 7 Hz), 7.53 (1 H, t, J ) 8 Hz), 8.56 (1
H, br s). Anal. (C10H14N4O) C, H, N.
1-(6-F lu or o-2-p yr id yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (39). Meth od B: 32% overall yield; mp 181-183 °C;
1H NMR (300 MHz, DMSO-d6) 3.07-3.15 (2 H, m), 3.82 (2 H,
t, J ) 6 Hz), 6.51 (1 H, dd, J ) 9, 1.5 Hz), 6.85-6.90 (2 H, br
m), 7.83 (1 H, q), 8.73 (1 H, br s); MS m/e 197 (M + H)+. Anal.
(C8H9FN4O‚0.25H2O) C, H, N.
1-(6-Flu or opyr id-2-yl)-5-m eth yl-2H,4H-tetr ah ydr o-1,2,4-
tr ia zin -3-on e (40). Meth od B: 15% overall yield; mp 240
°C dec; 1H NMR (300 MHz, DMSO-d6) 1.03 (3 H, d, J ) 7 Hz),
3.0 (1 H, dd, J ) 14, 9 Hz), 3.31-3.41 (1 H, m), 4.33 (1 H, dd,
J ) 14, 4 Hz), 6.47 (1 H, dd, J ) 8, 3 Hz), 6.85 (1 H, dd, J )
8, 3 Hz), 6.81 (1 H, q), 8.71 (1 H, br s). Anal. (C9H11FN4O) C,
H, N.
NMR (300 MHz, DMSO-d6) 3.41 (2 H, t, J ) 7.5 Hz), 3.51 (2
H, m), 6.75 (1 H, d, J ) 6 Hz), 7.01 (1 H, br s), 7.51 (1 H, dd,
J ) 9, 3 Hz), 7.84 (1 H, d, J ) 1.5 Hz), 8.15 (1 H, d, J ) 9 Hz),
8.45 (1 H, d, J ) 9 Hz), 9.31 (1 H, br s). Anal. (C12H11
ClN4O‚0.5H2O) C, H, N.
-
1-Qu in ol-3-yl-2H,4H-tetr ah ydr o-1,2,4-tr iazin -3-on e (48).
Meth od B: 12% overall yield; mp 184-186 °C; 1H NMR (300
MHz, DMSO-d6) 3.1-3.17 (2 H, m), 3.83 (2 H, t, J ) 5 Hz),
6.82 (1 H, br s), 7.5-7.6 (2 H, m), 7.73 (1 H, d, J ) 3.0 Hz),
7.84-7.87 (1 H, m), 7.91-7.96 (1 H, m), 8.67 (1 H, d, J ) 1.5
Hz), 9.87 (1 H, d, J ) 3 Hz). Anal. (C12H12N4O) C, H, N.
1-Ben zoth ia zol-2-yl-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-
1
on e (49). Meth od B: 13% overall yield; mp 249-252 °C; H
NMR (300 MHz, DMSO-d6) 3.31-3.36 (2 H, m), 3.53-3.6
(2 H, t, J ) 7 Hz), 7.09 (1 H, dt, J ) 7.5, 1 Hz), 7.15 (1 H, br
s), 7.27 (1 H, dt, J ) 7.5, 1 Hz), 7.45 (1 H, d, J ) 7.5 Hz), 7.75
(1 H, d, J ) 7.5 Hz), 9.75 (1 H, br s). Anal. (C10H10N4OS) C,
H, N.
1-Ben zoxa zol-2-yl-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
1
on e (50). Meth od B: 8% overall yield; mp 240-241 °C; H
NMR (300 MHz, DMSO-d6) 3.28-3.34 (2 H, m), 3.83 (2 H, t, J
) 5 Hz), 7.08 (1 H, br s), 7.16 (1 H, dt, J ) 7.5, 1.5 Hz), 7.25
(1 H, dt, J ) 7.5, 1.5 Hz), 7.45 (1 H, d, J ) 7.5 Hz), 7.55 (1 H,
d, J ) 7.5 Hz), 9.05 (1 H, br s). Anal. (C10H10N4O‚0.25H2O)
C, H, N.
1-(3-P yr id -2-ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (51). Meth od E: 17% overall yield; mp 261 °C dec; 1H
NMR (300 MHz, DMSO-d6) 3.08 (2 H, m), 3.71 (2 H, t, J )
5.25), 6.67 (1 H, d, J ) 1.5 Hz), 7.12 (1 H, dd, J ) 1.5, 7.5 Hz),
7.32-7.42 (2 H, m), 7.56 (1 H, d, J ) 7.5 Hz), 7.79 (1 H, t, J )
1.5 Hz), 7.83-7.96 (2 H, m), 8.51 (1 H, d, J ) 1.5 Hz), 8.66 (1
H, d, J ) 4.5 Hz). Anal. (C14H14N4O) C, H, N.
1-(3-P yr id -4-ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
1
3-on e (52). Meth od E: 13%; mp 250 °C dec; H NMR (300
MHz, DMSO-d6) 3.04 (2 H, m), 3.75 (2 H, t, J ) 5.25 Hz), 6.69
(1 H, d, J ) 1.5 Hz), 7.16 (1 H, dd, J ) 1.5, 7.5 Hz), 7.29 (1 H,
d, J ) 7.5 Hz), 7.38-7.47 (2 H, m), 7.67 (2 H, d, J ) 6.0 Hz),
1-(6-Ch lor op yr id -2-yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (41). Meth od B: 29% overall yield; mp 194-196 °C;
1H NMR (300 MHz, DMSO-d6) 3.08-3.16 (2 H, m), 3.84 (2 H,
t, J ) 5 Hz), 6.85 (1 H, d, J ) 7 Hz), 6.89 (1 H, br s), 6.95 (1
H, d, J ) 7 Hz), 7.70 (1 H, t, J ) 8 Hz), 8.71 (1 H, s). Anal.
(C8H9ClN4O) C, H, N.
8.54 (1 H, s), 8.64 (2 H, d,
(C14H14N4O‚0.5H2O) C, H, N.
J
) 6.0 Hz). Anal.
1-(3-Th ien -2-ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (53). Meth od E: 15% overall yield; mp 212-215 °C;
1H NMR (300 MHz, DMSO-d6) 3.08 (2 H, dt, J ) 2, 5 Hz),
3.70 (2 H, t, J ) 5 Hz), 6.68 (1 H, d, J ) 2 Hz), 6.99 (1 H, dd,
1-(6-Ch lor opyr id-2-yl)-5-m eth yl-2H,4H-tetr ah ydr o-1,2,4-
tr ia zin -3-on e (42). Meth od B: 12% overall yield; mp 255

3948 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
Bhatia et al.
J ) 2, 7 Hz), 7.15 (2 H, m), 7.30 (2 H, m), 7.47 (1 H, dd, J )
1, 3 Hz), 7.54 (1 H, dd, J ) 1, 4 Hz), 8.50 (1 H, d, J ) 2 Hz).
Anal. (C13H13N3OS) C, H, N.
) 6.0 Hz), 6.87 (1 H, t, J ) 7.5 Hz), 7.03 (2 H, d, J ) 7.5 Hz),
7.26 (2 H, t, J ) 7.5 Hz), 8.43 (1 H, s). Anal. (C11H15N3O) C,
H, N.
Meth od E (Sch em e 6). 5-Meth yl-1-(3-th ien -2-ylp h en -
yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (54). To a stirred
solution of thiophene (4.88 g, 58 mmol) in ether (60 mL) at 0
°C was added dropwise n-butyllithium (2.5 M in hexanes, 23.0
mL, 58 mmol). After 3 h of stirring at 0 °C, the mixture was
cooled to -78 °C and a solution of trimethyltin chloride (11.5
g, 58 mmol) in ether (60 mL) was added dropwise. The
mixture was allowed to warm to room temperature, filtered,
and concentrated in vacuo. The residue was distilled at
reduced pressure to give 2-(trimethylstannyl)thiophene (9.61
g, 67%), bp 79-81 °C at 12 mmHg.
A mixture of 2-(trimethylstannyl)thiophene (1.92 g, 7.8
mmol), 3-bromophenyl-2H,4H-tetrahydro-1,2,4-triazin-3-one
(29, 1.05 g, 3.9 mmol), and tetrakis(triphenylphosphine)-
palladium(0) (0.23 g, 0.2 mmol) in toluene (15 mL) was stirred
under nitrogen at reflux 4 h, cooled to room temperature, and
poured into saturated aqueous NH4Cl (50 mL). The mixture
was extracted with 15-20% methanol/dichloromethane (2 ×
85 mL), dried (MgSO4), filtered, and concentrated in vacuo.
The crude produce was purified by chromatography (silica gel,
2-3% methanol/dichloromethane) followed by two crystalliza-
tions from ethanol to provide 54 (0.27 g, 26%): mp 198.5-
200.5 °C; 1H NMR (300 MHz, DMSO-d6) 1.02 (3 H, d, J ) 6.0
Hz), 2.93 (1 H, dd, J ) 10.5, 13.5 Hz), 3.32 (1 H, m), 4.13 (1 H,
dd, J ) 4.5, 13.5 Hz), 6.73 (1 H, s), 6.99 (1 H, dd, J ) 2.5, 7.5
Hz), 7.14 (2 H, m), 7.29 (2 H, m), 7.47 (1 H, m), 7.54 (1 H, m),
8.51 (1 H, d, J ) 1.5 Hz). Anal. (C14H15N3OS) C, H, N.
1-(3-Th ien -2-ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-tr ia zin -
3-on e (55). Meth od E: 32% overall yield; mp 227 °C dec; 1H
NMR (300 MHz, DMSO-d6) 3.08 (2 H, m), 3.72 (2 H, t, J )
5.25 Hz), 6.66 (1 H, d, J ) 1.5 Hz), 6.97 (1 H, m), 7.16-7.39 (3
H, m), 7.52 (1 H, dd, J ) 1.5, 6.0 Hz), 7.64 (1 H, dd, J ) 3.0,
4.5 Hz), 7.81 (1 H, dd, J ) 1.5, 3.0 Hz), 8.47 (1 H, d, J ) 1.5
Hz). Anal. (C13H13N3OS) C, H, N.
Meth od F (Sch em e 7). 4-(P h en ym eth yl)-1-p h en yl-
2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (62). To a stirred
mixture of N-phenylethylenediamine (24.4 g, 0.179 mol) and
triethylamine (18.1 g, 0.179 mol) in dichloromethane (100 mL)
cooled to 0 °C was added dropwise a solution of benzoyl
chloride (25.2 g, 0.179 mol) in dichloromethane (50 mL) while
the reaction temperature was maintained below 10 °C. The
mixture was then stirred at room temperature overnight and
washed with 5% NH4Cl (2 × 75 mL), and the organic layer
was dried over MgSO4 and concentrated in vacuo to yield
benzoic acid 2-(phenylamino)ethylamide (37.8 g, 87%).
To a stirred suspension of LiAlH4 (3.80 g, 0.10 mol) in THF
(100 mL) was added dropwise a solution of the benzoic acid
amide intermediate (12.02 g, 0.05 mol) in THF (50 mL). The
mixture was stirred at reflux overnight, cooled in an ice bath,
followed by dropwise addition of water (3.8 mL), followed by
15% NaOH (3.8 mL), and then water (11.3 mL), stirred at room
temperature for 1.5 h, filtered, and concentrated in vacuo to
provide 2-(phenylamino)-1-[(phenymethyl)amino]ethane (11.29,
99%).
To a stirred solution of 2-(phenylamino)-1-[(phenylmethyl)-
amino]ethane (11.29 g, 0.049 mol) and triethylamine (5.00 g,
0.049 mol) in dichloromethane (100 mL) cooled to 0 °C was
added dropwise a solution of methyl chloroformate (4.68 g,
0.049 mol). The mixture was then stirred at room temperature
overnight and washed with 5% NH4Cl (2 × 75 mL), and the
organic layer was dried (MgSO4) and concentrated in vacuo
to yield methyl N-[2-(phenylamino)ethyl]-N-(phenylmethyl)-
carbamate (13.8 g, 99%).
To a stirred solution of methyl N-[2-(phenylamino)ethyl]-
N-(phenylmethyl)carbamate (13.8 g, 0.048 mol) in dimethoxy-
ethane (20 mL) at 5 °C was added concentrated HCl (9.1 mL)
in water (45 mL) followed by dropwise addition of a solution
of NaNO2 (3.33 g, 0.048 mol) in water (6 mL). The product
which separated as a heavy oil was extracted into dichlo-
romethane, dried over MgSO4, and concentrated in vacuo to
give methyl N-[2-(nitrosophenylamino)ethyl]-N-(phenylmeth-
yl)carbamate (15 g, 99%).
A solution of the N-nitroso intermediate (17.0 g, 0.054 mol)
in glacial acetic acid (30 mL) was introduced dropwise into a
mechanically stirred suspension of zinc dust (14.2 g) in water
(35 mL), which was cooled in ice during the addition to
maintain the reaction temperature between 15-20 °C. The
cooling bath was removed and the stirring continued for
another hour, during which time the reaction temperature was
maintained below 40 °C. The reaction mixture was treated
with dichloromethane (200 mL) and water (100 mL) and
filtered through a Celite pad to remove the solid zinc residue.
The filtrate was treated with 6 N NaOH to pH 6, and the
layers were separated, the organic extract was dried over
MgSO4 and concentrated in vacuo to give N-[2-[N ′-(phenyl-
methyl)-N ′-(methoxycarbonyl)amino]ethyl]-N-phenylhydra-
zine (16.0 g, 99%).
5-Meth yl-1-(3-fu r -2-ylp h en yl)-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (56). Meth od E: 64% overall yield; mp 212-
1
214 °C; H NMR (300 MHz, DMSO-d6) 1.02 (3 H, d, J ) 6.0
Hz), 2.93 (1 H, dd, J ) 10.5, 13.5 Hz), 3.30 (1 H, m), 4.11 (1 H,
dd, J ) 4.5, 13.5 Hz), 6.59 (1 H, m), 6.72 (1 H, s), 6.91 (1 H, d,
J ) 3.0 Hz), 6.97 (1 H, dd, J ) 1.5, 7.5 Hz), 7.17-7.39 (3 H,
m), 7.74 (1 H, d, J ) 1.5 Hz), 8.48 (1 H, d, J ) 1.5 Hz). Anal.
(C14H15N3O2) C, H, N.
1-(3-Th iazol-2-ylph en yl)-2H,4H-tetr ah ydr o-1,2,4-tr iazin -
3-on e (57). Meth od E: 15% overall yield; mp 239 °C dec; 1H
NMR (300 MHz, DMSO-d6) 3.09 (2 H, m), 3.72 (2 H, t, J )
5.25 Hz), 6.72 (1 H, s), 7.17 (1 H, d, J ) 7.5 Hz), 7.34-7.48 (2
H, m), 7.66 (1 H, s), 7.78 (1 H, d, J ) 3.0 Hz), 7.92 (1 H, d, J
) 3.0 Hz), 8.56 (1 H, d, J ) 1.5 Hz). Anal. (C12H12N4OS) C,
H, N.
1-(6-Th ien -2-ylp yr id -2-yl)-2H ,4H -t et r a h yd r o-1,2,4-t r i-
a zin -3-on e (58). Meth od E: 35% overall yield; mp 253 °C
dec; 1H NMR (300 MHz, DMSO-d6) 3.15 (2 H, m), 3.95 (2 H, t,
J ) 5.25 Hz), 6.80 (1 H, d, J ) 1.5 Hz), 6.90 (1 H, d, J ) 7.5
Hz), 7.14 (1 H, dd, J ) 3.0, 4.5 Hz), 7.33 (1 H, d, J ) 7.5 Hz),
7.59 (1 H, dd, J ) 1.5, 4.5 Hz), 7.67-7.78 (2 H, m), 8.67 (1 H,
d, J ) 1.5 Hz). Anal. (C12H12N4OS) C, H, N.
To a stirred solution of the hydrazine intermediate (15.0 g,
0.05 mol) in dichloromethane (50 mL) at -30 °C was added
dropwise EtMgBr (2 M in THF, 31.0 mL, 0.062 mol) under
nitrogen. The mixture was gently warmed at 45 °C for 3 days,
cooled, and treated with ice and 3 N HCl to pH 4-5, and the
1-(6-P yr id-2-ylpyr id-2-yl)-2H,4H-tetr ah ydr o-1,2,4-tr iazin -
3-on e (59). Meth od E: 47% overall yield; mp 243 °C dec; 1H
NMR (300 MHz, DMSO-d6) 3.18 (2 H, m), 4.05 (2 H, t, J )
5.25 Hz), 6.80 (1 H, d, J ) 1.5 Hz), 7.09 (1 H, dd, J ) 1.5, 7.5
Hz), 7.38-7.46 (1 H, m), 7.77-7.88 (2 H, m), 7.92 (1 H, dt, J
) 1.5, 7.5 Hz), 8.33 (1 H, d, J ) 7.5 Hz), 8.62-8.68 (1 H, m),
8.70 (1 H, d, J ) 1.5 Hz). Anal. (C13H13N5O) C, H, N.
4-Met h yl-1-p h en yl-2H ,4H -t et r a h yd r o-1,2,4-t r ia zin -3-
layers were separated. The organic layer was dried (MgSO4
)
and concentrated in vacuo and the residue triturated in ether.
The crude product was crystallized from benzene to give 62
1
(3.79 g, 28%): mp 164-166 °C; H NMR (300 MHz, DMSO-
d6) 3.02 (2 H, t, J ) 6.0 Hz), 3.75 (2 H, t, J ) 6.0 Hz), 4.42 (2
H, s), 6.89 (1 H, t, J ) 7.5 Hz), 7.02-7.15 (4 H, m), 7.17-7.34
(5 H, m), 8.71 (1 H, s). Anal. (C16H17N3O) C, H, N.
1
on e (60). Meth od F : 30% overall yield; mp 174-178 °C; H
NMR (300 MHz, DMSO-d6) 2.73 (3 H, s), 3.11 (2 H, t, J ) 6.0
Hz), 3.74 (2 H, t, J ) 6.0 Hz), 6.87 (1 H, t, J ) 7.5 Hz), 7.02 (2
H, d, J ) 7.5 Hz), 7.26 (2 H, t, J ) 7.5 Hz), 8.49 (1 H, s). Anal.
(C10H13N3O) C, H, N.
4-E t h yl-1-p h e n yl-2H ,4H -t e t r a h yd r o-1,2,4-t r ia zin -3-
on e (61). Meth od F : 22% overall yield; mp 152.5-154.5 °C;
1H NMR (300 MHz, DMSO-d6) 0.94 (3 H, t, J ) 7.5 Hz), 3.11
(2 H, t, J ) 6.0 Hz), 3.18 (2 H, q, J ) 7.5 Hz), 3.73 (2 H, t, J
Meth od G (Sch em e 8). 4-Hyd r oxy-5-m eth yl-1-p h en yl-
2H,4H-tetr a h yd r o-1,2,4-tr ia zin -3-on e (63) To a stirred ice
cold solution of phenylhydrazine (27.0 g, 0.25 mol) in meth-
ylene chloride (350 mL) was added pyridine (19.7 g, 0.25 mol),
and the mixture was stirred for 15 min, after which methyl
chloroformate (24.5 g, 0.26 mol) in methylene chloride (150
mL) was added dropwise over 0.5 h while the reaction
temperature was maintained below 15 °C. The mixture was

5-Lipoxygenase Inhibitors
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20 3949
stirred for 1 h at room temperature, water (200 mL) was
added, the organic layer was separated, washed with 1 N HCl
and brine, dried (MgSO4), filtered, and concentrated in vacuo.
The crude product was slurried in pentane (150 mL) and
filtered to provide 1-(methoxycarbonyl)-2-phenylhydrazine
(37.6 g, 90%), mp 110-111 °C.
To a stirred solution of 1-(methoxycarbonyl)-2-phenylhy-
drazine (25 g, 0.15 mol) and Na2CO3 (47.7 g, 0.45 mol) in
dichloromethane (350 mL) was added chloroacetone oxime
(21.9 g, 0.2 mol), and the mixture was heated at reflux for 18
h. The mixture was cooled to room temperature, filtered,
concentrated in vacuo, and chromatographed (silica gel, 3%
methanol/dichloromethane) to provide 1-(methoxycarbonyl)-
2-(phenylhydrazino)acetone oxime (31.25 g, 88%).
To a stirred solution of 1-(methoxycarbonyl)-2-(phenylhy-
drazino)acetone oxime (31.0 g, 0.13 mol) in ethanol (300 mL)
was added dropwise borane/pyridine (37.5 g, 0.4 mol), and the
mixture was stirred for 1 h at room temperature and then
cooled in an ice bath as 6 N ethanolic HCl (180 mL) was added
dropwise. The ethanol was removed in vacuo, water (200 mL)
was added, and the mixture was extracted with ether (2 ×
150 mL). The aqueous layer was brought to pH 11 by addition
of 6 N NaOH, extracted with dichloromethane (3 × 150 mL),
washed with brine, dried (MgSO4), filtered, and concentrated
in vacuo. Recrystallization from benzene/ether gave [1-[1-
(methoxycarbonyl)-2-phenylhydrazino]-2-propyl]hydroxy-
lamine (16.4 g, 50%).
To a solution of [1-[1-(methoxycarbonyl)-2-phenylhydrazino]-
2-propyl]hydroxylamine (0.5 g, 2 mmol) in THF (15 mL) was
added potassium tert-butoxide (0.24 g, 2 mmol), the mixture
was stirred at room temperature for 18 h, ice cold water was
added followed by 6 N HCl to pH 5, and the mixture was
extracted with dichloromethane (2 × 50 mL), dried (MgSO4),
filtered, concentrated in vacuo, and recrystallized from ether/
pentane to provide 63 (0.22 g, 54%): mp 174-176 °C; 1H
NMR (300 MHz, DMSO-d6) 1.06 (3 H, d, J ) 6 Hz), 3.41 (1 H,
dd, J ) 15, 3 Hz), 3.55 (1 H, m), 4.08 (1 H, dd, J ) 15, 3 Hz),
6.86 (1 H, t, J ) 7 Hz), 7.05 (2 H, d, J ) 7.0 Hz), 7.26 (2 H, t,
J ) 7 Hz), 8.61 (1 H, s), 9.01 (1 H, s). Anal. (C10H13N3O2) C,
H, N.
purified by chromatography (silica gel, 10% methanol in
dichloromethane) to provide acetyl-2-(2-hydroxyethyl)-2-phen-
ylhydrazine (30.4 g, 32%).
Acetyl-2-(2-hydroxyethyl)-2-phenylhydrazine (13.5 g, 0.07
mol) in 6 N HCl (50 mL) was refluxed under nitrogen for 2 h.
The mixture was cooled in an ice bath, and 6 N KOH was
added to pH 7. The mixture was extracted with ethyl acetate
(5 × 100 mL), dried (MgSO4), filtered, and concentrated in
vacuo to give 1-(2-hydroxyethyl)-1-phenylhydrazine (10.2 g,
95%).
To a solution of 1-(2-hydroxyethyl)-1-phenylhydrazine (10.2
g, 0.067 mol) in THF (50 mL) was added carbonyldiimidazole
(10.9 g, 0.067 mol), and the mixture was stirred at room
temperature for 3 h. Dichloromethane (100 mL) was added,
and the organic layer was separated, washed with water (2 ×
50 mL) and brine (50 mL), dried (MgSO4), filtered, concen-
trated in vacuo, and purified by chromatography (silica gel,
7% methanol in dichloromethane) followed by recrystallization
from ether to provide 67 (2.28 g, 20%): mp 99-100 °C; 1H
NMR (300 MHz, CDCl3) 3.74 (2 H, t, J ) 10 Hz), 4.29 (2H t,
J ) 10 Hz), 7.08 (3 H, m), 7.23 (1 H, br), 7.33 (2 H, m). Anal.
(C9H10N2O2) C, H, N.
1-P h en yl-2H,4H-tetr ah ydr o-1,2,4-tr iazin e-3-th ion e (68).
To a suspension of 1-phenyl-2H,4H-triazin-3-one (4.0 g, 23
mmol) in toluene (150 mL) was added Lawesson’s reagent (2,4-
bis(4-methoxyphenyl)-1,3-dithia-2,4-diphosphetane 2,4-disul-
fide, 10.0 g). The mixture was heated at 80 °C for 15 h and
then cooled, and a mixture of methanol/dichloromethane (1:1,
50 mL) and water (100 mL) was added. The organic layer was
collected and concentrated to provide a residue which was
purified by chromatography (silica gel, 5% methanol in dichlo-
1
romethane) to provide 68 (1.8 g, 42%): mp 180 °C; H NMR
(300 MHz, DMSO-d6) 2.95-3.02 (2 H, m), 3.63 (2 H, t, J ) 5
Hz), 6.91-7.01 (3 H, m), 7.30 (2 H, t, J ) 7.5 Hz), 8.39 (1 H,
br s), 9.97 (1 H, br s). Anal. (C9H11N3S) C, H, N.
Biologica l Meth od s. The biological methods for the
inhibition assays in vitro and in vivo have been previously
reported.2 Percent inhibition was computed by comparing
individual values in treatment groups to the mean value of
the control group. Statistical significance was determined
using one-way analysis of variance and Tukey’s multiple
comparison procedure. Linear regression was used to estimate
IC50 and ED50 values.
Deter m in a tion of Dr u g P la sm a Con cen tr a tion s. Com-
pounds for oral administration were suspended in 0.2% HPMC
with a Potter-Elvehjem homogenizer equipped with a Teflon-
coated pestle and administered orally to cynomolgus monkeys.
Blood samples were collected at various times following
compound administration to dogs or cynomolgus monkeys.
Blood samples were centrifuged, and the plasma was removed
and stored frozen until assayed. Plasma samples were thawed,
2 volumes of methanol was added, and precipitated plasma
proteins were removed by centrifugation. Supernatants were
injected directly onto a C18 reversed phase column (Adsorbo-
sphere HL 7 micron column) and chromatographed using a
mobile phase composed of 55% acetonitrile containing 10 mM
acetohydroxamic acid and 8 mM triethylamine acetate, pH 6.5,
at a flow rate of 1 mL/min. Compound peaks were quantitated
by UV absorbance at 260 nm using an external calibration
curve. Data presented are means from two animals.
4-Hydr oxy-5-m eth yl-1-(3-ch lor oph en yl)-2H,4H-tetr ah y-
d r o-1,2,4-tr ia zin -3-on e (64). Meth od G: 26% yield; mp
180-182 °C; 1H NMR (300 MHz, DMSO-d6) 1.08 (3 H, d, J )
7 Hz), 3.43 (1 H, m), 3.60 (1 H, m), 4.11 (1 H, dd, J ) 15, 4.5
Hz), 6.88 (1 H, dd, J ) 9, 3 Hz), 7.06 (2 H, m), 7.27 (2 H, t, J
) 9 Hz), 8.71 (1 H, s), 9.13 (1 H, s). Anal. Calcd for C10H12
-
ClN3O2: C, 49.69; H, 5.01; N, 17.38. Found: C, 49.74; H, 5.13;
N, 16.82.
4-Hydr oxy-5-m eth yl-1-(3-m eth ylph en yl)-2H,4H-tetr ah y-
d r o-1,2,4-tr ia zin -3-on e (65). Meth od G: 17% yield; mp
177-179 °C; 1H NMR (300 MHz, DMSO-d6) 1.06 (3 H, d, J )
7 Hz), 2.27 (3 H, s), 3.46 (1 H, m), 3.55 (1 H, m), 4.06 (1 H, dd,
J ) 15, 4.5 Hz), 6.88 (1 H, d, J ) 9 Hz), 6.85 (2 H, m), 7.13 (2
H, t, J ) 9 Hz), 8.56 (1 H, s), 9.08 (1 H, s). Anal.
(C11H15N3O2‚0.125H2O) C, H, N.
4-Meth oxy-5-m eth yl-1-p h en yl-2H,4H-tetr a h yd r o-1,2,4-
tr ia zin -3-on e (66). To a solution of 4-hydroxy-5-methyl-1-
phenyl-2H,4H-tetrahydro-1,2,4-triazin-3-one (63) (0.2 g, 1
mmol) in acetone (15 mL) was added K2CO3 (1.0 g, 7 mmol)
and methyl iodide (0.21 g, 1.5 mmol), and the mixture was
stirred for 18 h at room temperature, filtered, concentrated
in vacuo, chromatographed (silica gel, 1:1 ether/dichlo-
romethane), and recrystallized from ether to provide 72 (0.08
Ex Vivo Leu k otr ien e B4 Biosyn th esis Assa y. Com-
pounds were suspended in 0.2% HPMC with
a Potter-
Elvehjem homogenizer equipped with a Teflon-coated pestle
and administered orally to dogs or cynomolgus monkeys.
Heparinized blood samples were obtained before and at various
times after compound administration. Aliquots of blood were
incubated at 37 °C with 50 µM calcium ionophore, A23187.
After 30 min, the blood was placed in an ice bath and
centrifuged at 400g for 20 min, and the plasma samples were
analyzed for LTB4. The level of LTB4 in aliquots of the extracts
was analyzed by EIA.
1
g, 36%): mp 138-140 °C; H NMR (300 MHz, DMSO-d6) 1.1
(3 H, d, J ) 6 Hz), 3.45 (1 H, dd, J ) 15, 4.5 Hz), 3.56 (3 H, s),
3.65 (1 H, m), 4.03 (1 H, dd, J ) 15, 4.5 Hz), 6.88 (1 H, t, J )
7 Hz), 7.04 (2 H, d, J ) 7.0 Hz), 7.28 (2 H, m), 8.82 (1 H, s).
Anal. (C11H15N3O2) C, H, N.
4-P h en yl-3H-tr ih yd r o-1,3,4-oxa d ia zin -2-on e (67). A so-
lution of acetyl-2-phenylhydrazine (75.3 g, 0.5 mol), 2-bromo-
ethanol (64.0 g, 0.55 mol), and diisopropylethylamine (68.0 g,
0.55 mol) in toluene (300 mL) was refluxed with stirring for
48 h. The mixture was allowed to cool to room temperature,
and dichloromethane (500 mL) and water (250 mL) were
added. The organic layer was washed with water (2 × 250
mL), dried (MgSO4), filtered, concentrated in vacuo, and
Eva lu a tion of Meth em oglobin em ia in Vivo. When
oxidative denaturation of hemoglobin to methemoglobin occurs
in circulating erythrocytes, intracellular precipitates known
as Heinz bodies are formed that can be detected histologically.
Blood taken from rats or dogs dosed with test compound or

3950 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 20
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drug-induced methemoglobinemia was assessed by determin-
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Heinz bodies.
Ack n ow led gm en t. We thank the members of the
Leukotriene Biosynthesis Regulators Project and sup-
porting departments at Abbott Laboratories for techni-
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