Design, synthesis, and biological activities of new thieno[3,2- d]pyrimidines as selective type 4 phosphodiesterase inhibitors

Article abstract of DOI:10.1021/jm981012m

A common pharmacophore for compounds structurally related to nitraquazone has been derived. Using this pharmacophore, new structures have been designed, synthesized, and evaluated for their inhibitory potencies against cyclic adenosine 5'-monophosphate (c

Full text of DOI:10.1021/jm981012m

J . Med. Chem. 1998, 41, 4021-4035
4021
Design , Syn th esis, a n d Biologica l Activities of New Th ien o[3,2-d ]p yr im id in es a s
Selective Typ e 4 P h osp h od iester a se In h ibitor s1
Mar´ıa I. Crespo,* Llu´ıs Page`s, Armando Vega, Victor Segarra, Manel Lo´pez, Teresa Dome´nech,
Montserrat Miralpeix, J ordi Beleta, Hamish Ryder, and J ose´ M. Palacios
Almirall Prodesfarma S.A., Research Center, Cardener 68-74, 08024 Barcelona, Spain
Received March 16, 1998
A common pharmacophore for compounds structurally related to nitraquazone has been derived.
Using this pharmacophore, new structures have been designed, synthesized, and evaluated
for their inhibitory potencies against cyclic adenosine 5′-monophosphate (cAMP) specific
phosphodiesterase (PDE 4). From these compounds, 4-benzylamino-2-butylthieno[3,2-d]-
pyrimidine (4) was selected for optimization. The effects of changes to the lipophilic groups
and the amino linkage on the PDE 4 activity have been investigated. As a result, some potent
PDE 4 inhibitors, selective with respect to PDE 3, have been identified. A selected group of
compounds have been further evaluated for their ability to displace [3H]rolipram from its binding
site and also to potentiate isoprenaline-induced cAMP accumulation in isolated guinea pig
eosinophils. Of these, 2-butyl-4-cyclohexylaminothieno[3,2-d]pyrimidine (33) has an interesting
profile, with an important improvement in PDE 4/[3H]rolipram ratio with respect to reference
drugs, and good activity in cAMP potentiation, consistent with efficient cell penetration.
In tr od u ction
Interest in the potential utility of isoenzyme-selective
phosphodiesterase (PDE) inhibitors has increased in
recent years. The inhibition of PDE activity increases
cellular levels of the key second messengers, cyclic
adenosine 5′-monophosphate (cAMP) and cyclic gua-
nosine 5′-monophosphate (cGMP), thereby activating
specific protein phosphorylation cascades that elicit a
variety of functional responses.2 At the present time
there are seven known PDE isoenzyme families (PDE
1-7) which share the property of hydrolyzing cyclic
nucleotides to their 5-monophosphate counterparts.
For a number of reasons, interest in the potential
therapeutic utility of selective PDE inhibitors has
largely been focused on drugs capable of inhibiting PDE
4, one of the cAMP specific phosphodiesterases. First,
the tissue distribution of PDE 4 strongly suggests that
F igu r e 1. Compounds representative of the three chemical
pathologies related to the central nervous3 and immune4
classes of PDE 4 inhibitors: rolipram, nitraquazone, and
xanthine derivatives (denbufylline).
systems could be treated through the selective inhibition
of PDE 4. On the other hand, the increase in intracel-
lular cAMP concentration, the obvious biochemical
consequence of PDE 4 inhibition, has been well char-
acterized in immuno-competent cells, where it acts as
a deactivating signal.5,6 Furthermore the pathologies
associated with these biological systems represent some
of the more important therapeutic targets for the next
century such as asthma,5,7 atopy,8,9 multiple sclerosis,3,10
arthritis,11-13 type-II diabetes,14-16 and AIDS.17,18
properties in the carrageenan-induced rat paw oede-
ma.23 Additionally, RS-1759724a and RS-14491,24b re-
lated to the potent PDE 4 inhibitor RS-25344,25 have
recently been described as PDE 4 inhibitors with ex-
ceptional biological activity in arachidonic acid-induced
ear mouse oedema. Finally, the new PDE 4 inhibitor
KF-1951426 is related to the bronchodilator but weak
PDE 4 inhibitor KF-1762527 and KF-18280,28 which
exhibits extremely potent activity in the carrageenan-
induced rat paw oedema, zymosan-induced rat paw
oedema, and the reverse passive Arthus reaction-in-
duced paw oedema in the rat, and has a glucocorticoid-
like antiinflammatory profile.
From a structural point of view, selective PDE 4
inhibitors in the public domain can be divided into three
classes:2,19-21 structural analogues of rolipram, struc-
tural analogues of nitraquazone, and structures related
to xanthines (Figure 1). The compound nitraquazone22
is notable for its antiinflammatory and analgesic phar-
macological profile. Figure 2 shows several compounds
structurally related to nitraquazone. The analogue CP-
77,059 has also shown interesting antiinflammatory
However, most PDE 4 inhibitors produce nausea and
emetic effects which limit their therapeutic potential.29
[3H]Rolipram binding activity has been correlated with
these side effects in a study in dogs.30 Hence, reducing
S0022-2623(98)01012-7 CCC: $15.00 © 1998 American Chemical Society
Published on Web 09/12/1998

4022 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
ment of ligands was based on their aromatic and
hydrophobic moieties. Figure 3 shows the superposition
of selected structures on the template molecule. The
program GRID42-46 was selected to calculate putative
interaction sites of different ligands. The energy con-
tours obtained from the GRID computation indicate
favorable positions for interaction with other molecules.
Figure 4 shows the interaction pattern of the five
molecules studied, derived from the GRID computation
using the NH neutral flat amide probe which identifies
the areas of the molecule able to act as hydrogen bond
acceptors.
The common interaction map of nitraquazone and its
related compounds using the NH amide probe is de-
picted in Figure 5a. Three main areas of interaction
are clearly identified. The distances between the points
of maximal energy of interaction (HACC, Figure 5b) are
9.43, 8.06, and 10.86 Å.
In summary, in nitraquazone-related compounds,
three common areas of interaction as hydrogen bond
acceptors have been identified by using the program
GRID. These areas of interaction, taken together with
the aromatic and hydrophobic moieties discussed above,
represent the 3D pharmacophore model for PDE 4
inhibitors related to the structure of nitraquazone
(Figure 6).47
F igu r e 2. Compounds selected for pharmacophore evaluation
and related analogues referenced in the text.
Design of New Com p ou n d s. Once the pharmaco-
phore configuration was defined, several condensed het-
erocyclic systems were evaluated. The model described
above requires the existence of a central planar template
that comprises a heterocyclic condensed moiety, three
hydrogen bond acceptor groups, and two out-of-plane
regions which can be occupied by bulky aromatic or
lipophilic substituents. Therefore new compounds with
structural and electronic profiles compatible with the
proposed pharmacophore were initially explored.
These compounds were computationally evaluated.
First of all, they were built using fragments and stand-
ard geometries in Chem-X. After full conformational
analysis, compounds were fitted with the aromatic and
lipophilic components in the pharmacophore by using
a rigid fitting procedure followed by a flexible fitting
routine (FLEXIFIT). In the flexible fit step, the internal
nonbonded energy of the molecules under study is
minimized with respect to user-defined exocyclic bonds
and the penalty functions (restraints to obtain a maxi-
mum quality fit). The geometries of resulting con-
formations were optimized by the AM1 method as im-
plemented in the program MOPAC. These new struc-
tures were then studied with the program GRID (NH
flat amide probe). On the basis of these considerations
and taking into account synthetic feasibility, the com-
pounds 1-8 (depicted in Figure 7) were selected as
potential leads. All these compounds possess at least
two areas of interaction representative of hydrogen bond
acceptor groups (data not shown), whose distances
correspond to those depicted in the pharmacophore
model, as well as the required three aromatic lipophilic
groups.
the affinity of a compound for the [3H]rolipram binding
site may also reduce emetic potency.
Nitraquazone, despite its marked structural dissim-
ilarity to rolipram, also binds with high affinity to the
[3H]rolipram binding site.31 Other series structurally
related to nitraquazone such as nicotinamide ethers,32
which may be considered as ring-opened derivatives,
also had high affinity for the [3H]rolipram binding site.
In series related to rolipram, several attempts have been
made to reduce the [3H]rolipram binding site affinity
while retaining PDE 4 inhibitory potency.29,33-35
A
recent report on a nitraquazone-related chemical series
shows a great improvement in the PDE 4 inhibition/
[3H]rolipram binding site ratio.36
In the present work, several compounds related to the
nitraquazone structure were initially studied with the
aim to determine a common pharmacophore. This
pharmacophore has been used as starting point to
design new structures which may be potent and selec-
tive PDE 4 inhibitors but with attenuated affinity for
the [3H]rolipram binding site.
Molecu la r Mod elin g Stu d ies a n d Design of New
Com p ou n d s
3D P h a r m a cop h or e Mod el. Nitraquazone (Figure
1) and its structural analogues CP-77,059, RS-17597,
RS-25344, and KF-19154 (Figure 2) were selected for
the pharmacophore evaluation. A rigorous systematic
conformational search on the selected compounds was
performed to determine their lowest energy conforma-
tions. Calculations were performed using the full
molecular mechanics force field implemented in Chem-
X.37,38 Resulting geometries of the low-energy conform-
ers were optimized by means of semiempirical quantum
mechanics calculations using the MOPAC package39,40
(method AM1).41 Superposition of optimized structures
was performed by manual geometric fitting, selecting
the nitraquazone structure as a template. The align-
Compounds 1-8 were, therefore, synthesized and
evaluated as PDE 4 inhibitors.
Ch em istr y
The synthetic strategies used for the preparation of
the majority of the heterocyclic-fused 4-aminopyrim-

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4023
F igu r e 3. Superimposition of nitraquazone (purple) and related analogues: CP-77,059 (cyan), KF-19154 (red), RS-17597 (blue),
and RS-25344 (green).
F igu r e 4. Interaction maps obtained with the N1 probe (NH flat amide) of GRID software. All interaction energy maps were
contoured at -3 kcal/mol.
idines are shown in Scheme 1. The pyrimidinones 41-
54 were synthesized starting from the corresponding
2-aminocarboxylic acid, ester, or carboxamide, following
literature procedures.48 Reaction of the pyrimidinones
with phosphorus oxychloride or thionyl chloride gave
the 4-chloropyrimidines 55-68. The 4-aminopyrim-
idines 4-6, 11-21, and 23-39 (Tables 1, 3, and 4) were
obtained by reaction of the 4-chloropyrimidines 55-68
and different amines, more forcing conditions being
required for the less nucleophilic amines. The synthesis
of compound 22 is shown in Scheme 2. Acylation of
3-amino-2-thiophenecarboxamide with 2-choroacetyl chlo-
ride gave the diamide 70. Nucleophilic substitution,
followed by basic cyclization, afforded the intermediate
72, which was carried through a synthetic sequence
similar to that shown in Scheme 1 to give the thieno-
[3,2-d]pyrimidine 22. Compound 40, in which the
benzylamino group of the thieno[3,2-d]pyrimidine 4 has
been replaced by a benzamido group, was synthesized
as shown in Scheme 3.
The thioether and ether replacements for the amino
linkage in 4 (compounds 9 and 10, Table 2) were syn-
thesized directly from benzyl bromide and the thieno-
[3,2-d]pyrimidin-4-thione 69 (R1 ) butyl) and the 4-chlo-
rothieno[3,2-d]pyrimidine 55 (R1 ) butyl), respectively
(Scheme 1).
The preparation of 3-benzylpyrimidin-4-ones 1 and 2
is also illustrated in Scheme 1. Alkylation with benzyl
bromide of the thieno[3,2-d]pyrimidinone 41 and the
quinazolinone 43 (R1 ) butyl) gave 1 and 2, respectively.

4024 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
F igu r e 5. (a) Common interaction map using the N1 GRID probe of nitraquazone (purple), CP-77,059 (cyan), KF-19154 (red),
RS-17597 (blue), and RS-25344 (green). (b) Distances between the three main interaction minima which represent the areas of
the ligand for accepting hydrogen bonds (HACC) are 9.43, 8.06, and 10.86 Å.
F igu r e 6. 3D pharmacophore model for PDE 4 inhibitors related to the nitraquazone structure.
cies against isolated guinea pig ventricular PDE 4.
Isoenzyme selectivity was obtained by comparing the
IC50 values of compounds against PDE 4 with their
inhibitory activity against cGMP-inhibited PDE (PDE
3) from the same source. A representative group of
compounds were evaluated for their ability to displace
[3H]rolipram from its binding site and also to potentiate
isoprenaline-induced cAMP accumulation in isolated
guinea pig eosinophils (cAMP potentiation). Rolipram,
milrinone,51 and RS-1449124b were used as reference
substances.
The analogous pyridopyrimidinone 3 was synthesized
in two steps starting from 3-amino-2-pyridincarboxylic
acid (Scheme 4).
The syntheses49 of pyrazinones 7 and 8 are sum-
marized in Schemes 5 and 6, respectively. Condensa-
tion of 3-amino-2-benzylaminopyridine50 with 2-oxopen-
tanoic acid gave the pyrido[2,3-b]pyrazinone 7. Com-
pound 8 was prepared starting from 1-fluoro-2-nitroben-
zene and D,L-norvaline. Reduction of the nitro group
of the amino ester 75, followed by oxidation with H2O2,
yielded the intermediate quinoxaline 76, which was
converted to 8 by alkylation with benzyl bromide.
Resu lts a n d Discu ssion
Biologica l Eva lu a tion
Lea d F in d in g. Table 1 shows the biological results
for the set of compounds 1-8 suggested as possible PDE
4 inhibitors on the basis of the pharmacophore. As
To assess structure-activity relationships, all the
final compounds were tested for their inhibitory poten-

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4025
with respect to the reference drugs rolipram and RS-
14491. Although PDE 4 potency and PDE 3/PDE 4
selectivity were maintained by the ether analogue 10,
for reasons of chemical stability we chose as a basis for
continued structure-activity relationship exploration
the amino compound 4.
Modification of the alkyl group at C2 of the thienopy-
rimidine was then examined (Table 3). In agreement
with the proposed pharmacophore, the absence of a
lipophilic group at C2 yielded a less active compound
(11). For alkyl groups at this position, the optimal
volume was found to be between a butyl and a cyclo-
pentylmethyl group, with compounds 4 and 14 being the
most active and selective ones. The introduction of
larger groups, such as a phenoxybutyl group (compound
18) or even a cyclohexylmethyl group (compound 15),
reduced PDE 4 potency 4-fold. Compound 16, with a
benzyl group at C2, had reasonable PDE 4 activity but
was not selective against PDE 3. The introduction of a
pyridyl group at the C2 position led to interesting
activities. PDE 4 activity was enhanced in compound
20 (R1 ) 3-pyridyl), although PDE 3 selectivity was
reduced compared to compound 4. Introduction of a
piperidine at the C2 position (compound 22) resulted in
a complete loss of activity, indicating a lack of tolerance
at the receptor for a positive charge in this region of
the molecule. With respect to the [3H]rolipram binding
site, several compounds (14, 17, 20, and 21) were
identified with a much better PDE 4/[3H]rolipram ratio
than the reference compounds.
In summary, within the synthesized compounds, the
best substitution at the C2 position was the 3-pyridyl
group. Compound 20 was the most active with 25-fold
selectivity over PDE 3 and a good PDE 4/[3H]rolipram
ratio.
F igu r e 7. Chemical structures of new compounds which were
designed based on PDE 4 pharmacophore configuration.
pointed out previously, these compounds fit well in the
pharmacophore model, sharing at least two areas of
interaction, related to hydrogen bond acceptor groups,
and the three aromatic lipophilic centers. All of them
were selective for PDE 4 with respect to the PDE 3
enzyme. Pyrimidinones 1-3, in which the relative
position of one of the hydrogen bond acceptor groups
and one of the lipophilic groups was interchanged,
showed poor PDE 4 activities. The most interesting
compounds were the thienopyrimidine 4, the pyridopy-
razine 7, and the quinoxaline 8, with PDE 4 inhibitory
activities (IC50) of around 1 µM and more than 100-fold
selectivity versus PDE 3. These results highlight the
importance of a hydrogen bond acceptor in position 3 of
the heterocycle ring (a nitrogen for the pirimidine and
an oxo group for the pyrazinones). For compound 4,
substitution of the fused thiophene by either a pyrazine
(compound 5) or an imidazole (compound 6) led to a
reduction in PDE 4 inhibition. These data suggest that
the increase of polarity in the fused aromatic ring could
be detrimental for the activity. Among the more active
structures, compound 4 was initially selected for further
optimization.
Table 4 shows the biological activity of analogues of
4-amino-2-butylthieno[3,2-d]pyrimidine. Again, in agree-
ment with the proposed pharmacophore, a lipophilic
group at C4 is needed in order to obtain active com-
pounds. As was observed previously at C2, there exists
an optimal volume that is exceeded by the bulky
(biphenyl-4-yl)methyl group (compound 30). The intro-
duction of electron-donating and electron-withdrawing
substituents on the benzyl group of compound 4 was
studied. Compounds 23-25, with a methoxy group at
ortho, meta, and para positions showed no significant
changes in PDE 4 activity with respect to compound 4.
The ortho substitution (compound 23) maintained good
PDE 3/PDE 4 selectivity while the meta and para
methoxy groups (compounds 24 and 25) exhibited better
PDE 4/[3H]rolipram ratios. The introduction of a nitro
group at ortho and meta positions led to compounds 26
and 27, respectively, as active against PDE 4 as
compound 4 but considerably less PDE 3 selective.
Compound 28, with a para nitro group, was less active
and selective than 4. PDE 4 potency was enhanced by
a pyridylmethyl group (compound 31) and a phenyl
group (compound 32) at the C4 position, but with some
reduction in selectivity. Other interesting analogues
were obtained by introducing nonaromatic groups at C4
(compounds 33-35). Of particular note is compound 33,
which exhibited PDE 4 activity and PDE 3 selectivity
comparable to rolipram, with the advantage of a 20-fold
better PDE 4/[3H]rolipram ratio. By way of contrast,
Lea d Op tim iza tion . Changes in PDE activity on
modifying the linkage between the benzyl group and the
thienopyrimidine were analyzed first (Table 2). The
substitution of the amino group in compound 4 by an
ether led to compound 10, which is as active in PDE 4
inhibitory potency as 4 but less selective against PDE
3. The thioether 9 was less potent. In all these
compounds, the PDE 4/[3H]rolipram ratio was improved

4026 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
Sch em e 1a
a
Reagents: (a) (R1CO)2O or R1COOH, DCC, DMAP, py, DMF; (b) 2 N NaOH; (c) R1COCl, py, CHCl3 or R1COOH, DCC, DMAP, CH2Cl2;
(d) 30% NH3; (e) (R1CO)2O; (f) POCl3 or SOCl2 under standard conditions; (g) R2NHR3, Et3N, THF or R2NHR3, NaOAc, HOAc, KI or
R2NHR3, K2CO3, DMF; (h) PhCH2OH, NaH, DMSO; (i) Lawesson’s reagent, toluene; (j) PhCH2Br, K2CO3, DMF.
Sch em e 2a
Sch em e 3a
a
Reagents: (a) 30% NH3; (b) BuLi, benzoyl chloride, THF.
rolipram ratios. The introduction of an amide led to a
large loss of activity (compound 40).
cAMP P oten tia tion . A selected group of thieno[3,2-
d]pyrimidines were evaluated for their ability to poten-
tiate isoprenaline-induced cAMP accumulation in iso-
lated guinea pig eosinophils. This cell type was chosen
on account of its prominent pathological role in allergic
asthma52 and because PDE 4 appears to be the only
cAMP-hydrolizing activity present in these cells.53 The
selected compounds and their activities are summarized
in Table 5. In general, there is good correlation between
PDE 4 inhibition and cAMP potentiation. In fact,
a
Reagents: (a) Et3N, THF; (b) piperidine, K2CO3, acetonitrile;
(c) 2 N NaOH.
piperidine 35 exhibited a high [3H]rolipram affinity, in
comparison with the other thienopyrimidines. The PDE
3/PDE 4 selectivity was enhanced by disubstituted
amines at C4, as was shown by compound 37 (R2
)
benzyl; R3 ) methyl). Furthermore, compound 37, being
as active as 4, had one of the most favorable PDE 4/[3H]-

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4027
Sch em e 5a
Ta ble 1. Biological Activity (PDE 4 and PDE 3 Inhibition) of
New Pharmacophore-Based Compounds
compd
PDE 4a,b
PDE 3a,b
1
2
3
4
5
6
7
8
5 ( 1
43 ( 3
16 ( 3
>200
>200
97 ( 5
45 ( 7
10 ( 5
0.9 ( 0.4
6.7 ( 0.3
29 ( 6
>200
>200
>200
0.73 ( 0.03
242 ( 11
a
1.0 ( 0.3
0.9 ( 0.3
Reagents: (a) EtOH, 2 N HCl.
Sch em e 6a
milrinone
rolipram
0.32 ( 0.09
a
b
Data are indicated as IC50 (µM) ( SEM (n ) 3). PDE 3 and
PDE 4 were obtained from guinea pig ventricular tissue,59 and
assays were performed following the procedure of Thomson et al.60
Ta ble 2. Changes to the Linkage of Thieno[3,2-d]pyrimidines
PDE
a
compd
X
PDE 4a,b
PDE 3a,b
3H-ROLa,c
4/3H-ROL
Reagents: (a) NaHCO3, EtOH, H2O; (b) SOCl2, EtOH; (c) Fe,
EtOH, HCl, and then 2 N NaOH, H2O2; (d) benzyl bromide, K2CO3,
DMF.
4
9
10
NH 0.9 ( 0.4
97 ( 5
0.23 ( 0.15
2.8 ( 0.8
3.9
2.1
0.9
53
S
6 ( 1
>200
O
1.1 ( 0.2
0.32 ( 0.09
0.056 ( 0.01
58 ( 8
242 ( 11
5.1 ( 2.0 0.0048 ( 0.001
1.2 ( 0.4
rolipram
RS-14491
0.006 ( 0.004
selected as a lead compound for further optimization.
The biological results reported in Tables 3 and 4 confirm
the importance of the presence of lipophilic and aromatic
groups in both the C2 and C4 positions, as suggested by
the pharmacophore model. In general, the compounds
show a good balance of PDE 4 activity and displacement
of [3H]rolipram from its binding site. For the com-
pounds evaluated, the enzyme activity correlates well
with the potentiation of isoprenaline-induced cAMP
accumulation in isolated guinea pig eosinophils, con-
sistent with good cell penetration. Compounds such as
32, 33, and 37, which exhibit good activity in both PDE
4 inhibition and cAMP potentiation and display an
improved ratio with respect to the [3H]rolipram specific
binding site, are being evaluated in vivo as potential
antiasthmatic agents.
12
a
Data are indicated as IC50 (µM) ( SEM or inhibition percent-
b
age ( SEM at indicated concentration (µM) (n ) 3). PDE 3 and
PDE 4 were obtained from guinea pig ventricular tissue,59 and
assays were performed following the procedure of Thomson et al.60
c [3H]Rolipram tests were performed using rat brain membranes.61
Sch em e 4a
Exp er im en ta l Section
Molecu la r Mod elin g. All molecular modeling calculations
were performed on a Digital Alpha Station 3000. Structures
were built with standard bond lengths and angles by using
the Chem-X molecular modeling package. All structures were
initially optimized using steepest descents and conjugate
gradient methods. Charge distributions were calculated after
semiempirical optimization using the MOPAC program (pro-
gram no. 506, version 6.0, Quantum Chemistry Program
Exchange, QCPE, Bloomington, IN). The PULAY keyword
and the eigenvector following (EF) routine for minimun search
with GNORM ) 0.01 were used in the optimization step.
Electrostatic potential maps were calculated from the semiem-
pirical charges by using the program VSS (program no. 249,
QCPE) and displayed in Chem-X. The energies of interaction,
determined by different chemical probes, were analyzed by
GRID computations (version 14.0, Molecular Discovery, Ox-
ford, U.K.).
a
Reagents: (a) (C4H9CO)2O; (b) PhCH2NH2.
compounds 32, 33, and 35 exhibited submicromolar
activities in both assays, and compounds such as 11 and
22 showed no potency in either. By contrast, pyridines
20 and 21 are less able to potentiate cAMP accumula-
tion in isolated guinea pig eosinophils than their potency
as PDE 4 inhibitors might suggest.
Con clu sion s
From the common pharmacophore for nitraquazone-
related compounds, a series of novel heteroaromatic
compounds have been designed, synthesized, and evalu-
ated as PDE 4 inhibitors. Thienopyrimidine 4 was
Ch em istr y: Gen er a l. Reagents, starting materials, and
solvents were purchased from commercial suppliers and used
as received. All organic solutions are dried over sodium
sulfate. Concentration refers to evaporation under aspirator

4028 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
Ta ble 3. Biological Activity of Analogues of 4-Benzylaminothieno[3,2-d]pyrimidine
compd
R1
PDE 4a,b
PDE 3a,b
3H-ROLa,c
PDE 4/3H-ROL
4
11
12
13
14
15
16
17
18
19
20
21
butyl
H
ethyl
isobutyl
cyclopentylmethyl
cyclohexylmethyl
benzyl
phenethyl
4-phenoxybutyl
phenyl
3-pyridyl
4-pyridyl
0.9 ( 0.4
14 ( 4
2.3 ( 0.6
1.5 ( 0.6
1.1 ( 0.3
4 ( 1
1.5 ( 0.7
2.3 ( 0.6
4.7 ( 0.5
3.0 ( 0.1
0.6 ( 0.1
1.3 ( 0.5
65 ( 7
97 ( 5
68 ( 1
30 ( 5
58 ( 16
0.23 ( 0.15
nt
3.9
41% ( 1 at 10
0.4 ( 0.2
5.8 ( 2.6
1.3 ( 0.1
nt
3.9 ( 0.8
nt
8% ( 3 at 10
1.4 ( 0.4
1.7 ( 0.5
nt
3.7
0.2
3.1
>200
>200
5 ( 1
>200
>200
>200
14 ( 1
>200
>200
242 ( 11
5.1 ( 2.0
0.6
0.4
0.8
22
piperidin-1-yl
rolipram
RS-14491
0.32 ( 0.09
0.056 ( 0.01
0.006 ( 0.004
0.0048 ( 0.001
53
12
a
b
Data are indicated as IC50 (µM) ( SEM or inhibition percentage ( SEM at indicated concentration (µM) (n ) 3). PDE 3 and PDE
4 were obtained from guinea pig ventricular tissue,59 and assays were performed following the procedure of Thomson et al.60 c [3H]Rolipram
tests were performed using rat brain membranes;61 nt ) not tested.
Ta ble 4. Biological Activity of Analogues of 4-Amino-2-butylthieno[3,2-d]pyrimidine
compd
R2
R3
PDE 4a,b
PDE 3a,b
3H-ROLa,c
PDE 4/3H-ROL
4
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
benzyl
H
H
H
H
H
H
H
H
H
H
H
H
H
0.9 ( 0.4
0.7 ( 0.2
1.4 ( 0.1
1.5 ( 0.2
1.0 ( 0.2
0.84 ( 0.08
6.3 ( 0.9
1.00 ( 0.05
97 ( 5
39 ( 18
20 ( 4
6 ( 1
18 ( 8
3.5 ( 0.5
8.5 ( 0.5
64 ( 4
200
29 ( 17
42 ( 1
>200
>200
68 ( 20
44 ( 16
>200
0.23 ( 0.15
0.2 ( 0.02
2.2 ( 0.2
1.7 ( 0.1
0.08 ( 0.02
1.5 ( 0.2
nt
0.6 ( 0.3
nt
nt
0.20 ( 0.01
0.20 ( 0.03
1.1 ( 0.5
0.03 ( 0.01
2.3 ( 1
2.5 ( 0.5
37% ( 14 at 10
37% ( 14 at 10
nt
3.9
3.5
0.6
0.9
12.5
0.6
2-methoxybenzyl
3-methoxybenzyl
4-methoxybenzyl
2-nitrobenzyl
3-nitrobenzyl
4-nitrobenzyl
2,6-difluorobenzyl
(biphenyl-4-yl)methyl
(4-pyridyl)methyl
phenyl
cyclohexyl
cyclohexylmethyl
-(CH2)4-
-CH2-C6H4-CH2CH2-
benzyl
benzyl
1.7
>200
0.6 ( 0.2
0.6 ( 0.2
0.4 ( 0.2
1.0 ( 0.5
0.6 ( 0.3
2.7 ( 1.3
0.8 ( 0.1
2.5 ( 0.5
4.0 ( 1.8
23.0 ( 0.2
0.32 ( 0.09
0.056 ( 0.01
3
2
0.9
20
1.2
0.3
methyl
propyl
benzyl
H
38
39
40
>200
>200
>200
242 ( 11
5.1 ( 2.0
benzyl
benzoyl
rolipram
RS-14491
0.006 ( 0.004
0.0048 ( 0.001
53
12
a
b
Data are indicated as IC50 (µM) ( SEM or inhibition percentage ( SEM at indicated concentration (µM) (n ) 3). PDE 3 and PDE
4 were obtained from guinea pig ventricular tissue,59 and assays were performed following the procedure of Thomson et al.60 c [3H]Rolipram
tests were performed using rat brain membranes;61 nt ) not tested.
vacuum using a Bu¨chi rotatory evaporator. Reaction products
were purified, when necessary, by flash chromatography on
silica gel (40-63 µm) with the solvent system indicated.
Spectroscopic data were recorded on a Varian Gemini 300
spectrometer. Melting points were recorded on a Bu¨chi 535
apparatus. Where analyses are indicated only by symbols of
the elements, results obtained were within 0.4% of the
theoretical values.
Gen er a l P r oced u r es for th e Syn th esis of P yr im id in o-
n es 41-54 (Meth od s A, B, C, a n d D). Meth od A. 2-Bu tyl-
3H-th ien o[3,2-d ]p yr im id in -4-on e (41). A mixture of 50 g
(0.35 mol) of 3-amino-2-thiophenecarboxamide and 80 mL (0.40
mol) of valeric anhydride was heated at 90 °C for 20 min. After
cooling, 600 mL of 2 N NaOH were added. The mixture was
stirred under reflux for 1 h, cooled, and neutralized with 2 N
HCl. The resulting white solid was filtered, washed with
water, and dried to give 67.5 g (91%) of the title compound:
1H NMR (CDCl3) δ 12.77 (br, 1H), 7.82 (d, J ) 5 Hz, 1H), 7.34
(d, J ) 5 Hz, 1H), 2.86 (t, J ) 8 Hz, 2H), 1.87 (m, 2H), 1.48
(m, 2H), 0.98 (t, J ) 8 Hz, 3H).
2-Eth yl-3H-th ien o[3,2-d ]p yr im id in -4-on e (42) was pre-
pared according to method A in 71% yield starting from
3-amino-2-thiophenecarboxamide (2.5 g) and propionic anhy-
dride: 1H NMR (DMSO) δ 12.20 (br, 1H), 8.11 (d, J ) 5 Hz,
1H), 7.32 (d, J ) 5 Hz, 1H), 2.63 (q, J ) 8 Hz, 2H), 1.22 (t, J
) 8 Hz, 3H).

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4029
chloride: 1H NMR (CDCl3) δ 11.66 (br, 1H), 7.83 (d, J ) 5 Hz,
1H), 7.45 (d, J ) 7 Hz, 1H), 7.50-7.25 (m, 5H), 4.10 (s, 2H).
2-P h en eth yl-3H-th ien o[3,2-d ]p yr im id in -4-on e (49) was
prepared according to method C in 70% yield starting from
methyl 3-amino-2-thiophenecarboxylate (4.71 g) and 3-
phenylpropionyl chloride: 1H NMR (CDCl3) δ 11.90 (br, 1H),
8.14 (d, J ) 5 Hz, 1H), 7.37 (d, J ) 5 Hz, 1H), 7.30-7.15 (m,
5H), 3.03 (m, 2H), 2.94 (m, 2H).
2-(4-P h en oxyb u t yl)-3H -t h ien o[3,2-d ]p yr im id in -4-on e
(50) was prepared according to method C in 70% yield starting
from methyl 3-amino-2-thiophenecarboxylate (2.84 g) and
5-phenoxyvaleryl chloride:55 1H NMR (CDCl3) δ 12.46 (br, 1H),
7.83 (d, J ) 5 Hz, 1H), 7.34 (d, J ) 5 Hz, 1H), 7.25 (m, 2H),
6.97-6.84 (m, 3H), 4.03 (t, J ) 6 Hz, 2H), 2.94 (t, J ) 6 Hz,
2H), 2.08 (m, 2H), 1.95 (m, 2H).
2-P h en yl-3H-th ien o[3,2-d ]p yr im id in -4-on e (51) was pre-
pared according to method C in 52% yield starting from methyl
3-amino-2-thiophenecarboxylate (4.71 g) and benzoyl chlo-
ride: 1H NMR (CDCl3) δ 12.66 (br, 1H), 8.15 (m, 2H), 8.11 (d,
J ) 5 Hz, 1H), 7.60-7.48 (m, 3H), 7.41 (d, J ) 5 Hz, 1H).
Meth od D. 2-(3-P yr id yl)-3H-th ien o[3,2-d ]p yr im id in -4-
on e (52). To a solution of 4.50 g (0.0288 mol) of methyl
3-amino-2-thiophenecarboxylate and 3.50 g of nicotinic acid
in 75 mL of CH2Cl2 were added 5.93 g (0.0288 mol) of DCC
and 3.50 g (0.0288 mol) of DMAP. The mixture was stirred
under reflux overnight. The resulting suspension was cooled
and concentrated. The residue was suspended in 150 mL of
30% NH3 and then heated to 120 °C for 2 h in a pressure
reactor. The mixture was cooled, and the insoluble residue
was filtered. The basic solution was neutralized with 2 N HCl.
The resulting white solid was filtered, washed with water, and
dried to give 3.60 g (55%) of 52: 1H NMR (DMSO) δ 12.93 (br,
1H), 9.26 (s, 1H), 8.77 (s, 1H), 8.46 (d, J ) 5 Hz, 1H), 8.22 (d,
J ) 7 Hz, 1H), 7.70-7.45 (m, 2H).
Ta ble 5. cAMP Potentiation of Selected
Thieno[3,2-d]pyrimidines
compd
PDE 4a
cAMP pot.b
4
11
14
15
20
21
22
24
27
32
33
34
35
37
0.9 ( 0.4
14 ( 4
1.8 ( 0.4
18 ( 5
3.4 ( 1.7
2 ( 1
3.5 ( 0.2
7.5 ( 3.8
21 ( 8
1.7 ( 0.2
1.05 ( 0.05
0.7 ( 0.4
0.6 ( 0.2
2.1 ( 1.3
0.2 ( 0.1
1.8 ( 0.2
1.1 ( 0.3
4 ( 1
0.6 ( 0.1
1.3 ( 0.5
65 ( 7
1.5 ( 0.2
0.84 ( 0.08
0.6 ( 0.2
0.4 ( 0.2
1.0 ( 0.5
0.6 ( 0.3
0.8 ( 0.1
a
Data are indicated as IC50 (µM) ( SEM; PDE 4 was obtained
from guinea pig ventricular tissue,59 and assays were performed
following the procedure of Thomson et al.60 b Isoprenaline-induced
cAMP accumulation in isolated guinea pig eosinophils; data are
indicated as EC50 (µM) ( SEM.
2-Bu tyl-3H-qu in a zolin -4-on e (43) was prepared according
to method A in 89% yield starting from anthranilamide (13.6
g) and valeric anhydride: 1H NMR (CDCl3) δ 12.22 (br, 1H),
8.28 (d, J ) 7 Hz, 1H), 7.80-7.70 (m, 2H), 7.46 (m, 1H), 2.83
(t, J ) 7 Hz, 2H), 1.90 (m, 2H), 1.50 (m, 2H), 1.00 (t, J ) 7 Hz,
3H).
Meth od B. 2-Ben zyl-1,9-d ih yd r op u r in -6-on e (44). To
a solution of 1.62 g (0.01 mol) of 4-amino-5-imidazolecarbox-
amide hydrochloride in 25 mL of anhydrous pyridine and 50
mL of anhydrous DMF were added 1.36 g (0.01 mol) of
phenylacetic acid, 2.06 g (0.01 mol) of DCC, and 1.22 g (0.01
mol) of DMAP. The mixture was stirred at 70 °C overnight.
The resulting solution was concentrated, and the residue was
suspended in 40 mL of 2 N NaOH. The mixture was stirred
under reflux for 30 min. The suspension was cooled, the
insoluble residue was filtered, and the basic solution was
neutralized with 2 N HCl. The resulting white solid was
filtered, washed with water, and dried to give 1.24 g (55%) of
44: 1H NMR (CDCl3) δ 12.32 (br, 1H), 8.04 (br, 1H), 7.40-
7.20 (m, 6H), 3.94 (s, 2H).
Met h od C. 2-Isob u t yl-3H -t h ien o[3,2-d ]p yr im id in -4-
on e (45). To a cooled solution of 4.71 g (0.03 mol) of methyl
3-amino-2-thiophenecarboxylate and 4.83 mL (0.06 mol) of
pyridine in 75 mL of CHCl3 was added 5.48 g (0.045 mol) of
isovaleryl chloride. The mixture was stirred at room temper-
ature overnight. The reaction mixture was washed with 2 N
HCl and brine, dried, and concentrated. The resulting solid
was suspended in 75 mL of 30% NH3 and then was heated to
120 °C for 2 h in a pressure reactor. The mixture was cooled
and neutralized. The resulting white solid was filtered,
washed with water, and dried to give 2.49 g (40%) of the
desired product: 1H NMR (CDCl3) δ 12.27 (br, 1H), 7.83 (d, J
) 5 Hz, 1H), 7.34 (d, J ) 5 Hz, 1H), 2.69 (d, J ) 8 Hz, 2H),
2.30 (m, 1H), 1.05 (d, J ) 8 Hz, 6H).
2-Cyclop e n t ylm e t h yl-3H -t h ie n o[3,2-d ]p yr im id in -4-
on e (46) was prepared according to method C in 33% yield
starting from methyl 3-amino-2-thiophenecarboxylate (3.50 g)
and cyclopentylacetyl chloride: 1H NMR (CDCl3) δ 12.10 (br,
1H), 7.82 (d, J ) 5 Hz, 1H), 7.33 (d, J ) 5 Hz, 1H), 2.80 (d, J
) 8 Hz, 2H), 2.44 (m, 1H), 1.90-1.50 (m, 6H), 1.43-1.25 (m,
2H).
2-(4-P yr id yl)-3H-th ien o[3,2-d ]p yr im id in -4-on e (53) was
prepared according to method D in 53% yield starting from
methyl 3-amino-2-thiophenecarboxylate (4.50 g) and isonico-
tinic acid: 1H NMR (DMSO) δ 12.96 (br, 1H), 8.79 (d, J ) 5
Hz, 2H), 8.26 (d, J ) 5 Hz, 1H), 8.06 (d, J ) 5 Hz, 2H), 7.52
(d, J ) 5 Hz, 1H).
2-Bu tyl-3H-p ter id in -4-on e (54). A mixture of 5.00 g
(0.036 mol) of 3-amino-2-pyrazinecarboxylic acid and 40 mL
of valeric anhydride was heated at 140 °C for 1 h. The excess
of valeric anhydride was distilled. The resulting red solid was
suspended in 60 mL of 30% NH3 and then heated to 120 °C
for 2 h in a pressure reactor. The resulting solution was
cooled, neutralized with 2 N HCl, and extracted with EtOAc.
The combined extracts were washed with brine, dried, and
concentrated. The resulting solid was washed with pentane
to give 3.06 g (42%) of the title compound: 1H NMR (CDCl3)
δ 12.60 (br, 1H), 9.04 (s, 1H), 8.86 (s, 1H), 3.00 (t, J ) 7 Hz,
2H), 1.96 (m, 2H), 1.50 (m, 2H), 0.97 (t, J ) 7 Hz, 3H).
Gen er a l P r oced u r es for th e Syn th esis of Ch lor op yr i-
m id in es 55-68 (Meth od s E, F , a n d G). Meth od E. 2-Bu -
tyl-4-ch lor oth ien o[3,2-d ]p yr im id in e (55). Thieno[3,2-d]-
pyrimidinone 41 (8.15 g, 0.039 mol) was dissolved in 38 mL of
POCl3. The mixture was stirred under reflux for 30 min,
cooled, and concentrated. The residue was poured into ice and
extracted with EtOAc. The combined extracts were washed
with saturated NaHCO3 and brine, dried, and concentrated
to give 8.56 g (96%) of 55 as a yellow oil: 1H NMR (CDCl3) δ
7.97 (d, J ) 6 Hz, 1H), 7.50 (d, J ) 6 Hz, 1H), 3.03 (t, J ) 7
Hz, 2H), 1.86 (m, 2H), 1.44 (m, 2H), 0.96 (t, J ) 7 Hz, 3H).
4-Ch lor oth ien o[3,2-d ]p yr im id in e (56) was prepared ac-
cording to method E in 89% yield starting from 3H-thieno-
[3,2-d]pyrimidin-4-one48b (2.00 g): 1H NMR (CDCl3) δ 9.00 (s,
1H), 8.06 (d, J ) 6 Hz, 1H), 7.62 (d, J ) 6 Hz, 1H).
2-Cyc loh e xylm e t h yl-3H -t h ie n o[3,2-d ]p yr im id in -4-
on e (47) was prepared according to method C in 37% yield
starting from methyl 3-amino-2-thiophenecarboxylate (3.77 g)
and cyclohexylacetyl chloride:54 1H NMR (CDCl3) δ 11.93 (br,
1H), 7.83 (d, J ) 5 Hz, 1H), 7.34 (d, J ) 5 Hz, 1H), 2.69 (d, J
) 9 Hz, 2H), 1.94 (m, 1H), 1.82-1.60 (m, 5H), 1.32-1.04 (m,
5H).
2-Ben zyl-3H-th ien o[3,2-d ]p yr im id in -4-on e (48) was pre-
pared according to method C in 48% yield starting from methyl
3-amino-2-thiophenecarboxylate (6.28 g) and phenylacetyl
4-Ch lor o-2-eth ylth ien o[3,2-d ]p yr im id in e (57) was pre-
pared according to method E in 65% yield starting from thieno-
[3,2-d]pyrimidinone 42 (2.33 g): 1H NMR (CDCl3) δ 8.00 (d, J
) 6 Hz, 1H), 7.52 (d, J ) 6 Hz, 1H), 3.10 (q, J ) 8 Hz, 2H),
1.44 (t, J ) 8 Hz, 3H).
4-Ch lor o-2-isobu tylth ien o[3,2-d ]p yr im id in e (58) was
prepared according to method E in 86% yield starting from

4030 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
thieno[3,2-d]pyrimidinone 45 (2.48 g): 1H NMR (CDCl3) δ 8.02
(d, J ) 6 Hz, 1H), 7.53 (d, J ) 6 Hz, 1H), 2.94 (d, J ) 8 Hz,
2H), 2.36 (m, 1H), 1.00 (d, J ) 8 Hz, 6H).
4-Ch lor o-2-cyclop e n t ylm e t h ylt h ie n o[3,2-d ]p yr im i-
d in e (59) was prepared according to method E in 80% yield
starting from thieno[3,2-d]pyrimidinone 46 (1.72 g): 1H NMR
(CDCl3) δ 8.00 (d, J ) 6 Hz, 1H), 7.53 (d, J ) 6 Hz, 1H), 3.04
(d, J ) 8 Hz, 2H), 2.50 (m, 1H), 1.80-1.60 (m, 7H), 1.30 (m,
1H).
4-C h lo r o -2-c y c lo h e x y lm e t h y lt h ie n o [3,2-d ]p y r im i-
d in e (60) was prepared according to method E in 89% yield
starting from thieno[3,2-d]pyrimidinone 47 (2.20 g): 1H NMR
(CDCl3) δ 8.00 (d, J ) 6 Hz, 1H), 7.54 (d, J ) 6 Hz, 1H), 2.92
(d, J ) 8 Hz, 2H), 2.00 (m, 1H), 1.75-1.60 (m, 5H), 1.32-1.02
(m, 5H).
2-Ben zyl-4-ch lor oth ien o[3,2-d ]p yr im id in e (61) was pre-
pared according to method E in 70% yield starting from thieno-
[3,2-d]pyrimidinone 48 (4.64 g): 1H NMR (CDCl3) δ 7.98 (d, J
) 7 Hz, 1H), 7.54 (d, J ) 7 Hz, 1H), 7.43 (m, 2H), 7.38-7.17
(m, 3H), 4.38 (s, 2H).
under reflux for 90 h. The mixture was cooled and partitioned
between EtOAc and water; the organic phase was washed with
brine, dried, and concentrated. To the residue dissolved in
acetone was added HCl in dioxane. The resulting solid was
recrystallized from isopropyl alcohol to give 2.12 g (72%) of
the title compound: mp 264-266 °C; 1H NMR (DMSO) δ 10.50
(t, J ) 6 Hz, 1H), 8.45 (d, J ) 5 Hz, 1H), 7.52 (d, J ) 5 Hz,
1H), 7.45-7.25 (m, 5H), 4.84 (d, J ) 6 Hz, 2H), 2.90 (t, J ) 8
Hz, 2H), 1.75 (m, 2H), 1.30 (m, 2H), 0.86 (t, J ) 8 Hz, 3H).
Anal. (C17H19N3S‚HCl) C, H, N, S.
4-Ben zyla m in ot h ien o[3,2-d ]p yr im id in e, H yd r och lo-
r id e (11) was prepared according to method H starting from
56 (2.00 g). Recrystallization from isopropyl alcohol gave 2.76
1
g (78%) of 11: mp 215-217 °C; H NMR (DMSO) δ 10.50 (br,
1H), 8.89 (br, 1H), 8.46 (d, J ) 5 Hz, 1H), 7.57 (d, J ) 5 Hz,
1H), 7.45-7.25 (m, 5H), 4.84 (d, J ) 7 Hz, 2H). Anal.
(C13H11N3S‚HCl) C, H, N, S.
4-Ben zyla m in o-2-et h ylt h ien o[3,2-d ]p yr im id in e, H y-
d r och lor id e (12) was prepared according to method H
starting from 57 (1.65 g). Recrystallization from isopropyl
alcohol gave 0.87 g (35%) of 12: mp 247-250 °C (D); 1H NMR
(DMSO) δ 15.44 (br, 1H), 10.50 (br, 1H), 8.43 (d, J ) 6 Hz,
1H), 7.54 (d, J ) 6 Hz, 1H), 7.45-7.25 (m, 5H), 4.87 (d, J ) 6
Hz, 2H), 2.93 (q, J ) 8 Hz, 2H), 1.31 (t, J ) 8 Hz, 3H). Anal.
(C15H15N3S‚HCl) C, H, N, S.
4-Ch lor o-2-p h en eth ylth ien o[3,2-d ]p yr im id in e (62) was
prepared according to method E in 75% yield starting from
thieno[3,2-d]pyrimidinone 49 (3.84 g): 1H NMR (CDCl3) δ 7.98
(d, J ) 6 Hz, 1H), 7.52 (d, J ) 6 Hz, 1H), 7.28-7.18 (m, 5H),
3.37 (m, 2H), 3.23 (m, 2H).
4-Ben zyla m in o-2-isobu tylth ien o[3,2-d ]p yr im id in e, Hy-
d r och lor id e (13) was prepared according to method H
starting from 58 (1.79 g). Recrystallization from isopropyl
alcohol gave 1.19 g (46%) of 13: mp 261-262 °C; 1H NMR
(DMSO) δ 15.65 (br, 1H), 10.56 (br, 1H), 8.46 (d, J ) 6 Hz,
1H), 7.54 (d, J ) 6 Hz, 1H), 7.40-7.25 (m, 5H), 4.84 (d, J ) 5
Hz, 2H), 2.80 (d, J ) 6 Hz, 2H), 2.25 (m, 1H), 0.90 (d, J ) 6
Hz, 6H). Anal. (C17H19N3S‚HCl) C, H, N.
4-Ch lor o-2-(4-p h en oxyb u t yl)t h ien o[3,2-d ]p yr im id in e
(63) was prepared according to method E in 66% yield starting
from thieno[3,2-d]pyrimidinone 50 (4.21 g): 1H NMR (CDCl3)
δ 7.97 (d, J ) 6 Hz, 1H), 7.50 (d, J ) 6 Hz, 1H), 7.25 (m, 2H),
6.95-6.84 (m, 3H), 4.00 (t, J ) 7 Hz, 2H), 3.14 (t, J ) 7 Hz,
2H), 2.09 (m, 2H), 1.91 (m, 2H).
4-Ch lor o-2-p h en ylth ien o[3,2-d]p yr im idin e (64) was pre-
pared according to method E in 80% yield starting from thieno-
[3,2-d]pyrimidinone 51 (3.51 g): 1H NMR (CDCl3) δ 8.52 (m,
2H), 8.01 (d, J ) 6 Hz, 1H), 7.61 (d, J ) 6 Hz, 1H), 7.55-7.47
(m, 3H).
4-Ch lor o-2-(3-p yr id yl)th ien o[3,2-d ]p yr im id in e (65) was
prepared according to method E in 54% yield starting from
thieno[3,2-d]pyrimidinone 52 (3.76 g): 1H NMR (CDCl3) δ 9.72
(s, 1H), 8.82-8.70 (m, 2H), 8.08 (d, J ) 6 Hz, 1H), 7.65 (d, J
) 6 Hz, 1H), 7.44 (m, 1H).
4-Ch lor o-2-(4-p yr id yl)th ien o[3,2-d ]p yr im id in e (66) was
prepared according to method E in 95% yield starting from
thieno[3,2-d]pyrimidinone 53 (3.30 g): 1H NMR (CDCl3) δ 8.77
(d, J ) 6 Hz, 2H), 8.35 (d, J ) 6 Hz, 2H), 8.08 (d, J ) 6 Hz,
1H), 7.65 (d, J ) 6 Hz, 1H).
Meth od F . 2-Ben zyl-6-ch lor o-9H-p u r in e (67). To a
suspension of 4.03 g (0.0178 mol) of 44 in 80 mL of CHCl3 was
added under reflux a solution of 6.50 mL (0.0893 mol) of SOCl2
in 7 mL of anhydrous DMF. The mixture was stirred under
reflux for 1.30 h and concentrated. The residue was dissolved
in EtOAc, and the resulting solution was washed with satu-
rated NaHCO3 and brine. The organic extracts were dried and
concentrated to give 2.42 g of a solid that was used in the next
step without further purification (1H NMR purity, 80%): yield
44%; 1H NMR (CDCl3) δ 13.35 (br, 1H), 8.17 (s, 1H), 7.35-
7.15 (m, 5H), 4.34 (s, 2H).
Meth od G. 2-Bu tyl-4-ch lor op ter id in e (68). To a solu-
tion of 4.18 mL (0.0246 mol) of diisopropylethylamine and 1.22
mL (0.0134 mol) of POCl3 in 70 mL of anhydrous toluene was
added 2.74 g (0.0134 mol) of pteridinone 54. The suspension
was stirred under reflux for 2 h. The mixture was cooled and
then diluted with diethyl ether. The resulting solution was
washed with water, 4 N NaOH, and brine, dried, and concen-
trated to give 2.25 g (75%) of 68 as a brown oil: 1H NMR
(CDCl3) δ 9.22 (s, 1H), 9.03 (s, 1H), 3.20 (t, J ) 8 Hz, 2H),
1.97 (m, 2H), 1.48 (m, 2H), 1.00 (t, J ) 8 Hz, 3H).
4-Ben zyla m in o-2-cyclop en tylm eth ylth ien o[3,2-d ]p yr i-
m id in e (14) was prepared according to method H starting
from 59 (1.28 g). Recrystallization from acetonitrile gave 1.28
1
g (78%) of 14: mp 160-161 °C; H NMR (CDCl3) δ 7.65 (d, J
) 6 Hz, 1H), 7.45-7.25 (m, 6H), 5.12 (br, 1H), 4.87 (d, J ) 6
Hz, 2H), 2.87 (d, J ) 8 Hz, 2H), 2.49 (m, 1H), 1.80-1.45 (m,
6H), 1.36-1.21 (m, 2H). Anal. (C19H21N3S) C, H, N.
4-Ben zyla m in o-2-cycloh exylm eth ylth ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (15) was prepared according to
method H starting from 60 (2.11 g). Recrystallization from
isopropyl alcohol gave 0.91 g (31%) of 15: mp 295-297 °C (D);
1H NMR (DMSO) δ 10.40 (br, 1H), 8.43 (d, J ) 6 Hz, 1H), 7.50
(d, J ) 6 Hz, 1H), 7.40-7.25 (m, 5H), 4.88 (d, J ) 4 Hz, 2H),
2.76 (d, J ) 8 Hz, 2H), 1.91 (m, 1H), 1.70-1.50 (m, 5H), 1.17-
0.90 (m, 5H). Anal. (C20H23N3S‚HCl) C, H, N, S.
2-Ben zyl-4-b en zyla m in ot h ien o[3,2-d ]p yr im id in e (16)
was prepared according to method H starting from 61 (2.15
g). Recrystallization from ethyl ether gave 0.93 mg (34%) of
16: mp 125-126 °C; 1H NMR (CDCl3) δ 7.58 (d, J ) 6 Hz,
1H), 7.40 (d, J ) 8 Hz, 2H), 7.34 (d, J ) 6 Hz, 1H), 7.30-7.15
(m, 8H), 5.74 (t, J ) 6 Hz, 1H), 4.80 (d, J ) 6 Hz, 2H), 4.20 (s,
2H). Anal. (C20H17N3S) C, H, N, S.
4-Ben zyla m in o-2-p h en et h ylt h ien o[3,2-d ]p yr im id in e,
Hyd r och lor id e (17) was prepared according to method H
starting from 62 (3.10 g). Recrystallization from isopropyl
alcohol gave 1.98 g (46%) of 17: mp 239-241 °C; 1H NMR
(DMSO) δ 15.67 (br, 1H), 10.56 (br, 1H), 8.37 (d, J ) 6 Hz,
1H), 7.50 (d, J ) 6 Hz, 1H), 7.40-7.10 (m, 10H), 4.84 (d, J )
6 Hz, 2H), 3.23 (m, 2H), 3.15 (m, 2H). Anal. (C21H19N3S‚HCl)
C, H, N, S.
4-Ben zyla m in o-2-(4-p h en oxyb u t yl)t h ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (18) was prepared according to
method H starting from 63 (2.04 g). Recrystallization from
isopropyl alcohol gave 1.02 g (37%) of 18: mp 220-222 °C; 1H
NMR (DMSO) δ 15.56 (br, 1H), 10.52 (br, 1H), 8.42 (d, J ) 5
Hz, 1H), 7.52 (d, J ) 5 Hz, 1H), 7.45-7.20 (m, 7H), 6.90-6.85
(m, 3H), 4.83 (d, J ) 6 Hz, 2H), 3.93 (t, J ) 6 Hz, 2H), 3.00 (t,
J ) 6 Hz, 2H), 1.95 (m, 2H), 1.73 (m, 2H). Anal. (C23H23N3-
OS‚HCl) C, H, N, S.
Gen er a l P r oced u r es for th e Syn th esis of Am in op yr i-
m id in es 4-6, 11-21, a n d 23-39 (Meth od s H, I, a n d J ).
Meth od H. 4-Ben zyla m in o-2-bu tylth ien o[3,2-d ]p yr im i-
d in e, Hyd r och lor id e (4). A solution of 2.00 g (8.83 mmol)
of 55, 0.94 g (9.27 mmol) of triethylamine, and 0.99 g (9.27
mmol) of benzylamine in 50 mL of anhydrous THF was stirred
4-Ben zyla m in o-2-p h en ylth ien o[3,2-d ]p yr im id in e, Hy-
d r och lor id e (19) was prepared according to method H

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4031
starting from 64 (2.56 g). Recrystallization from ethanol gave
1.36 g (37%) of 19: mp 258 °C; H NMR (DMSO) δ 10.44 (br,
2-Bu t yl-4-(4-n it r ob en zyla m in o)t h ien o[3,2-d ]p yr im i-
d in e (28) was prepared according to method H starting from
55 (2.00 g) and 4-nitrobenzylamine.57 Purification by flash
chromatography, eluting with EtOAc/hexane (1:1), gave 1.10
1
1H), 8.55-8.35 (m, 3H), 7.82 (d, J ) 6 Hz, 1H), 7.74-7.55 (m,
3H), 7.47 (d, J ) 8 Hz, 2H), 7.35 (t, J ) 8 Hz, 2H), 7.28 (m,
1H), 4.99 (d, J ) 6 Hz, 2H). Anal. (C19H15N3S‚HCl) C, H, N,
S.
4-Ben zyla m in o-2-(3-p yr id yl)t h ien o[3,2-d ]p yr im id in e,
Hyd r och lor id e (20) was prepared according to method H
starting from 65 (2.20 g). Recrystallization from isopropyl
alcohol gave 2.11 g (68%) of 20: mp 227-229 °C; 1H NMR
(DMSO) δ 10.90 (br, 1H), 9.60 (br, 2H), 9.25 (d, J ) 9 Hz, 1H),
9.00 (d, J ) 5 Hz, 1H), 8.31 (d, J ) 5 Hz, 1H), 8.09 (dd, J ) 9
Hz, J ) 5 Hz, 1H), 7.63 (d, J ) 5 Hz, 1H), 7.48 (d, J ) 7 Hz,
2H), 7.33 (t, J ) 7 Hz, 2H), 7.23 (m, 1H), 4.90 (d, J ) 5 Hz,
2H). Anal. (C18H14N4S‚1.3HCl‚0.5H2O) C, H, N, S.
4-Ben zyla m in o-2-(4-p yr id yl)t h ien o[3,2-d ]p yr im id in e,
Hyd r och lor id e (21) was prepared according to method H
starting from 66 (3.30 g). Purification by flash chromatogra-
phy, eluting with CH2Cl2/MeOH (95:5), gave 1.50 g (34%) of
21: mp 270-272 °C (D); 1H NMR (DMSO) δ 9.15 (br, 1H),
9.01 (d, J ) 6 Hz, 2H), 8.82 (d, J ) 6 Hz, 2H), 8.30 (d, J ) 6
Hz, 1H), 7.60 (d, J ) 6 Hz, 1H), 7.48 (d, J ) 8 Hz, 2H), 7.33 (t,
J ) 8 Hz, 2H), 7.24 (m, 1H), 4.90 (d, J ) 6 Hz, 2H). Anal.
(C18H14N4S‚1.5HCl‚0.4H2O) C, H, N, S.
2-Bu t yl-4-(2-m et h oxyb en zyla m in o)t h ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (23) was prepared according to
method H starting from 55 (2.00 g) and 2-methoxybenzy-
lamine. Recrystallization from acetonitrile gave 1.23 g (40%)
of 23: mp 184-186 °C; 1H NMR (DMSO) δ 15.60 (br, 1H),
10.30 (br, 1H), 8.43 (d, J ) 5 Hz, 1H), 7.53 (d, J ) 5 Hz, 1H),
7.35-7.15 (m, 2H), 7.05 (d, J ) 8 Hz, 1H), 6.90 (m, 1H), 4.80
(d, J ) 5 Hz, 2H), 3.80 (s, 3H), 2.98 (t, J ) 6 Hz, 2H), 1.69 (m,
2H), 1.29 (m, 2H), 0.83 (t, J ) 6 Hz, 3H). Anal. (C18H21N3-
OS‚HCl) C, H, N.
2-Bu t yl-4-(3-m et h oxyb en zyla m in o)t h ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (24) was prepared according to
method H starting from 55 (2.00 g) and 3-methoxybenzy-
lamine. Recrystallization from acetonitrile gave 1.94 g (52%)
of 24: mp 203-204 °C; 1H NMR (DMSO) δ 15.67 (br, 1H),
10.56 (br, 1H), 8.46 (d, J ) 6 Hz, 1H), 7.55 (d, J ) 6 Hz, 1H),
7.24 (t, J ) 9 Hz, 1H), 7.00 (s, 1H), 6.97 (d, J ) 9 Hz, 1H),
6.87 (d, J ) 9 Hz, 1H), 4.82 (d, J ) 6 Hz, 2H), 3.73 (s, 3H),
2.93 (t, J ) 8 Hz, 2H), 1.78 (m, 2H), 1.31 (m, 2H), 0.87 (t, J )
8 Hz, 3H). Anal. (C18H21N3OS‚HCl) C, H, N.
2-Bu t yl-4-(4-m et h oxyb en zyla m in o)t h ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (25) was prepared according to
method H starting from 55 (2.00 g) and 4-methoxybenzy-
lamine. Recrystallization from acetonitrile gave 0.66 g (22%)
of 25: mp 245-246 °C; 1H NMR (DMSO) δ 15.65 (br, 1H),
10.53 (br, 1H), 8.41 (d, J ) 6 Hz, 1H), 7.53 (d, J ) 6 Hz, 1H),
7.35 (d, J ) 6 Hz, 2H), 6.90 (d, J ) 6 Hz, 2H), 4.77 (d, J ) 4
Hz, 2H), 3.74 (s, 3H), 2.93 (t, J ) 8 Hz, 2H), 1.80 (m, 2H), 1.31
(m, 2H), 0.90 (t, J ) 8 Hz, 3H). Anal. (C18H21N3OS‚HCl) C,
H, N.
1
g (36%) of 28: mp 136-138 °C; H NMR (CDCl3) δ 8.16 (d, J
) 8 Hz, 2H), 7.67 (d, J ) 6 Hz, 1H), 7.49 (d, J ) 8 Hz, 2H),
7.33 (d, J ) 6 Hz, 1H), 5.61 (br, 1H), 4.95 (d, J ) 6 Hz, 2H),
2.82 (t, J ) 8 Hz, 2H), 1.72 (m, 2H), 1.34 (m, 2H), 0.87 (t, J )
8 Hz, 3H). Anal. (C17H18N4O2S) C, H, N.
2-Bu tyl-4-(2,6-d iflu or oben zyla m in o)th ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (29) was prepared according to
method H starting from 55 (3.16 g) and 2,6-difluorobenzy-
lamine. Recrystallization from acetonitrile gave 2.77 g (53%)
of 29: mp 225-227 °C; 1H NMR (DMSO) δ 15.75 (br, 1H),
10.31 (br, 1H), 8.43 (d, J ) 6 Hz, 1H), 7.52 (d, J ) 6 Hz, 1H),
7.50 (m, 1H), 7.14 (m, 2H), 4.86 (d, J ) 6 Hz, 2H), 2.92 (t, J )
9 Hz, 2H), 1.76 (m, 2H), 1.30 (m, 2H), 0.91 (t, J ) 9 Hz, 3H).
Anal. (C17H17F2N3S‚HCl) C, H, N, S.
4-[(Bip h en yl-4-yl)m eth yla m in o]-2-bu tylth ien o[3,2-d ]-
p yr im id in e, Hyd r och lor id e (30) was prepared according to
method H starting from 55 (2.27 g) and (biphenyl-4-yl)-
methylamine.58 Recrystallization from ethyl alcohol gave 1.68
g (42%) of 30: mp 274-276 °C; 1H NMR (DMSO) δ 15.30 (br,
1H), 10.46 (br, 1H), 8.44 (d, J ) 6 Hz, 1H), 7.65-7.62 (m, 4H),
7.55-7.40 (m, 5H), 7.35 (m, 1H), 4.87 (d, J ) 5 Hz, 2H), 2.89
(t, J ) 7 Hz, 2H), 1.75 (m, 2H), 1.29 (m, 2H), 0.85 (t, J ) 7 Hz,
3H). Anal. (C23H23N3S‚HCl) C, H, N, S.
2-Bu tyl-4-[(4-p yr id yl)m eth yla m in o]th ien o[3,2-d ]p yr i-
m id in e (31) was prepared according to method H starting
from 55 (2.00 g) and (4-pyridyl)methylamine. Purification by
flash chromatography, eluting with EtOAc/hexane (3:1), gave
0.24 g (9%) of 31: mp 155-157 °C; 1H NMR (CDCl3) δ 8.53 (d,
J ) 4 Hz, 2H), 7.66 (d, J ) 6 Hz, 1H), 7.38 (d, J ) 6 Hz, 1H),
7.27 (d, J ) 4 Hz, 2H), 5.75 (br, 1H), 4.90 (d, J ) 6 Hz, 2H),
2.81 (t, J ) 8 Hz, 2H), 1.73 (m, 2H), 1.35 (m, 2H), 0.91 (t, J )
8 Hz, 3H). Anal. (C16H18N4S) C, H, S, N: calcd, 18.78; found,
18.25.
2-Bu tyl-4-cycloh exylam in oth ien o[3,2-d]pyr im idin e, Hy-
d r och lor id e (33) was prepared according to method H
starting from 55 (1.20 g) and cyclohexylamine. Recrystalli-
zation from acetonitrile gave 0.57 g (33%) of 33: mp 209-211
°C; 1H NMR (DMSO) δ 15.36 (br, 1H), 9.67 (d, J ) 8 Hz, 1H),
8.42 (d, J ) 6 Hz, 1H), 7.50 (d, J ) 6 Hz, 1H), 4.22 (br, 1H),
2.91 (t, J ) 8 Hz, 2H), 1.95-1.65 (m, 7H), 1.50-1.10 (m, 7H),
0.92 (t, J ) 8 Hz, 3H). Anal. (C16H23N3S‚HCl) C, H, N, S.
2-Bu tyl-4-cycloh exylm eth yla m in oth ien o[3,2-d ]p yr im i-
d in e, Hyd r och lor id e (34) was prepared according to method
H starting from 55 (2.27 g) and cyclohexylmethylamine.
Recrystallization from acetonitrile gave 2.11 g (62%) of 34: mp
241-243 °C; 1H NMR (DMSO) δ 15.44 (br, 1H), 9.92 (br, 1H),
8.41 (d, J ) 6 Hz, 1H), 7.50 (d, J ) 6 Hz, 1H), 3.47 (m, 2H),
2.90 (t, J ) 8 Hz, 2H), 1.86-1.57 (m, 8H), 1.38 (m, 2H), 1.16
(m, 2H), 1.08-0.87 (m, 6H). Anal. (C17H25N3S‚HCl) C, H, N,
S.
2-Bu tyl-4-(p ip er id in -1-yl)th ien o[3,2-d ]p yr im id in e, Hy-
d r och lor id e (35) was prepared according to method H
starting from 55 (4.07 g) and piperidine. Purification by flash
chromatography, eluting with EtOAc/hexane (1:2), gave 2.62
g (52%) of 35: mp 218-220 °C; 1H NMR (DMSO) δ 15.77 (br,
1H), 8.55 (d, J ) 6 Hz, 1H), 7.62 (d, J ) 6 Hz, 1H), 4.10 (br,
4H), 2.90 (t, J ) 8 Hz, 2H), 1.85-1.65 (m, 8H), 1.38 (m, 2H),
0.94 (t, J ) 8 Hz, 3H). Anal. (C15H21N3S‚HCl) C, H, N, S.
2-Bu t yl-4-(1,2,3,4-t et r a h yd r oisoq u in olin -2-yl)t h ien o-
[3,2-d ]p yr im id in e, Hyd r och lor id e (36) was prepared ac-
cording to method H starting from 55 (2.26 g) and tetrahy-
droisoquinoline. Recrystallization from acetone gave 1.99 g
2-Bu t yl-4-(2-n it r ob en zyla m in o)t h ien o[3,2-d ]p yr im i-
d in e (26) was prepared according to method H starting from
55 (2.00 g) and 2-nitrobenzylamine.56 Purification by flash
chromatography, eluting with EtOAc/hexane (1:1), gave 0.57
1
g (19%) of 26: mp 107-109 °C; H NMR (CDCl3) δ 8.08 (d, J
) 8 Hz, 1H), 7.83 (d, J ) 8 Hz, 1H), 7.65 (d, J ) 6 Hz, 1H),
7.58 (t, J ) 8 Hz, 1H), 7.44 (t, J ) 8 Hz, 1H), 7.34 (d, J ) 6
Hz, 1H), 5.80 (t, J ) 8 Hz, 1H), 5.14 (d, J ) 8 Hz, 2H), 2.86 (t,
J ) 8 Hz, 2H), 1.79 (m, 2H), 1.39 (m, 2H), 0.93 (t, J ) 8 Hz,
3H). Anal. (C17H18N4O2S) C, H, N.
2-Bu t yl-4-(3-n it r ob en zyla m in o)t h ien o[3,2-d ]p yr im i-
d in e, Hyd r och lor id e (27) was prepared according to method
H starting from 55 (2.00 g) and 3-nitrobenzylamine. Recrys-
tallization from acetonitrile gave 1.29 g (39%) of 27: mp 205-
207 °C; 1H NMR (DMSO) δ 15.56 (br, 1H), 10.62 (br, 1H), 8.46
(d, J ) 6 Hz, 1H), 8.30 (s, 1H), 8.15 (d, J ) 8 Hz, 1H), 7.87 (d,
J ) 8 Hz, 1H), 7.65 (t, J ) 8 Hz, 1H), 7.52 (d, J ) 6 Hz, 1H),
4.95 (d, J ) 4 Hz, 2H), 2.90 (t, J ) 8 Hz, 2H), 1.71 (m, 2H),
1.26 (m, 2H), 0.84 (t, J ) 8 Hz, 3H). Anal. (C17H18N4O2S‚
HCl) C, H, N.
1
(56%) of 36: mp 212-214 °C; H NMR (DMSO) δ 15.74 (br,
1H), 8.60 (d, J ) 6 Hz, 1H), 7.61 (d, J ) 6 Hz, 1H), 7.36-7.24
(m, 4H), 5.25 (s, 2H), 4.30 (t, J ) 6 Hz, 2H), 3.10 (t, J ) 6 Hz,
2H), 2.94 (t, J ) 8 Hz, 2H), 1.84 (m, 2H), 1.40 (m, 2H), 0.93 (t,
J ) 8 Hz, 3H). Anal. (C19H21N3S‚HCl) C, H, N, S.
4-[(N-Ben zyl-N-m eth yl)a m in o]-2-bu tylth ien o[3,2-d ]p y-
r im id in e, Hyd r och lor id e (37) was prepared according to
method H starting from 55 (2.00 g) and N-benzyl-N-methyl-

4032 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
amine. Recrystallization from acetonitrile gave 1.65 g (54%)
of 37: mp 209-211 °C; 1H NMR (DMSO) δ 15.70 (br, 1H), 8.35
(d, J ) 6 Hz, 1H), 7.60 (d, J ) 6 Hz, 1H), 7.46-7.28 (m, 5H),
4.20 (s, 2H), 3.61 (s, 3H), 2.90 (t, J ) 9 Hz, 2H), 1.75 (m, 2H),
1.32 (m, 2H), 0.87 (t, J ) 9 Hz, 3H). Anal. (C18H21N3S‚HCl)
C, H, N, S.
4-[(N-Ben zyl-N-p r op yl)a m in o]-2-bu tylth ien o[3,2-d ]p y-
r im id in e, Hyd r och lor id e (38) was prepared according to
method H starting from 55 (3.00 g) and N-benzyl-N-propyl-
amine. Purification by flash chromatography, eluting with
EtOAc/hexane (2:1), gave 1.85 g (37%) of 38: mp 108-110 °C;
1H NMR (DMSO) δ 15.60 (br, 1H), 8.52 (br, 1H), 7.57 (d, J )
6 Hz, 1H), 7.35-7.25 (m, 6H), 5.21 (s, 2H), 3.83 (t, J ) 8 Hz,
2H), 2.88 (t, J ) 7 Hz, 2H), 1.82-1.65 (m, 4H), 1.29 (m, 2H),
0.94 (t, J ) 8 Hz, 3H), 0.84 (t, J ) 7 Hz, 3H). Anal.
(C20H25N3S‚HCl) C, H, N, S.
4-Ben zyla m in o-2-bu tylp ter id in e, Hyd r och lor id e (5)
was prepared according to method H starting from 68 (2.25 g)
and benzylamine. Recrystallization from ethyl alcohol gave
1.15 g (35%) of 5: mp 249-251 °C (D); 1H NMR (DMSO) δ
11.11 (br, 1H), 9.15 (d, J ) 2 Hz, 1H), 9.03 (d, J ) 2 Hz, 1H),
7.44 (d, J ) 8 Hz, 2H), 7.35-7.25 (m, 3H), 4.89 (d, J ) 6 Hz,
2H), 2.91 (t, J ) 7 Hz, 2H), 1.76 (m, 2H), 1.34 (m, 2H), 0.88 (t,
J ) 7 Hz, 3H). Anal. (C17H19N5‚HCl) C, H, N.
a solution of 4.84 g (0.023 mol) of 41 and 5.50 mL (0.023 mol)
of benzyl bromide in 50 mL of DMF was added 6.35 g (0.046
mol) of K2CO3. The mixture was stirred at room temperature
overnight. The resulting suspension was poured into water
and extracted with EtOAc. The combined extracts were
washed, dried, and concentrated. Recrystallization from n-
heptane gave 3.22 g (47%) of the title compound; mp 82-84
1
°C; H NMR (CDCl3) δ 7.75 (d, J ) 6 Hz, 1H), 7.35-7.15 (m,
6H), 5.61 (s, 2H), 2.73 (t, J ) 8 Hz, 2H), 1.70 (m, 2H), 1.36 (m,
2H), 0.88 (t, J ) 8 Hz, 3H). Anal. (C17H18N2OS) C, H, N.
3-Ben zyl-2-bu tyl-3H-qu in a zolin -4-on e (2) was prepared
according to method K starting from 43 (3.70 g) and benzyl
bromide. Purification by flash chromatography, eluting with
EtOAc/hexane (1:3), gave 2.32 g (43%) of the title compound:
mp 80-81 °C; 1H NMR (CDCl3) δ 8.30 (d, J ) 8 Hz, 1H), 7.75-
7.65 (m, 2H), 7.45 (t, J ) 8 Hz, 1H), 7.35-7.22 (m, 3H), 7.18
(d, J ) 8 Hz, 2H), 5.40 (s, 2H), 2.74 (t, J ) 8 Hz, 2H), 1.75 (m,
2H), 1.39 (m, 2H), 0.90 (t, J ) 8 Hz, 3H). Anal. (C19H20N2O)
C, N, H: calcd, 6.90; found, 7.34.
4-Ben zylth io-2-bu tylth ien o[3,2-d ]p yr im id in e (9) was
prepared according to method K starting from 69 (3.15 g) and
benzyl bromide. Crystallization from pentane at -50 °C gave
1
2.90 g (66%) of the title compound: mp 53-55 °C; H NMR
(CDCl3) δ 7.77 (d, J ) 6 Hz, 1H), 7.55-7.15 (m, 6H), 4.74 (s,
2H), 3.09 (t, J ) 8 Hz, 2H), 1.91 (m, 2H), 1.47 (m, 2H), 0.99 (t,
J ) 8 Hz, 3H). Anal. (C17H18N2S2) C, H, N.
2-Ben zyl-4-ben zyla m in o-9H-p u r in e, Hyd r och lor id e (6)
was prepared according to method H starting from 67 (1.69 g)
and benzylamine. Recrystallization from isopropyl alcohol
4-Ben zyloxy-2-bu tylth ien o[3,2-d ]p yr im id in e (10).
A
1
gave 0.52 g (22%) of 6: mp 255-256 °C; H NMR (DMSO) δ
suspension of 0.45 g (0.011 mol) of 60% NaH in 40 mL of
anhydrous DMSO was stirred under nitrogen at 60 °C for 1
h. The mixture was cooled to room temperature prior to the
addition of 1.05 mL (0.01 mol) of benzyl alcohol. After the
mixture was stirred for 15 min, a solution of 2.30 g (0.01 mol)
of 55 in 10 mL of DMSO was added, and the mixture was
stirred overnight. The reaction mixture was partitioned
between EtOAc and water; the organic phase was washed,
dried, and concentrated. The residue was purified by flash
chromatography, eluting with EtOAc/hexane (1:3). Crystal-
lization of the resulting oil from pentane at -30 °C gave 1.57
g (52%) of the title compound: mp 34-36 °C; 1H NMR (CDCl3)
δ 7.78 (d, J ) 6 Hz, 1H), 7.48-7.28 (m, 6H), 5.64 (s, 2H), 2.96
(t, J ) 8 Hz, 2H), 1.86 (m, 2H), 1.42 (m, 2H), 0.96 (t, J ) 8 Hz,
3H). Anal. (C17H18N2SO) C, H, N.
3-(2-Ch lor oa ceta m id o)-2-th iop h en eca r boxa m id e (70).
To a cooled solution of 1.00 g (7 mmol) of 3-amino-2-thiophen-
ecarboxamide and 1.01 g (10 mmol) of triethylamine in 25 mL
of anhydrous THF was added a solution of 1.17 g (10 mmol)
of 2-chloroacetyl chloride in 5 mL of anhydrous THF. The
mixture was warmed to room temperature and stirred for 2
h. The resulting triethylamine hydrochloride was filtered, and
the organic solution was concentrated. The residue was
partitioned between EtOAc and water; the organic phase was
washed with 2 N HCl, 2 N NaOH, and brine, dried, and
concentrated. The resulting white solid was washed with
diisopropyl ether to give 1.38 g (90%) of the title compound:
1H NMR (CDCl3) δ 8.26 (br, 1H), 8.30 (d, J ) 5 Hz, 1H), 7.40
(d, J ) 5 Hz, 1H), 6.66 (br, 2H), 4.10 (s, 2H).
3-[2-(P ip er id in -1-yl)a ceta m id o]-2-th iop h en eca r boxa m -
id e (71). To a solution of 1.38 g (6.32 mmol) of 70 and 0.54 g
(6.32 mmol) of piperidine in 30 mL of anhydrous acetonitrile
was added 0.87 g (6.32 mmol) of K2CO3. The mixture was
stirred under reflux for 7 h. The resulting suspension was
partitioned between EtOAc and water; the organic phase was
dried and concentrated. The resulting solid was washed with
diisopropyl ether to give 1.11 g (66%) of 71: 1H NMR (CDCl3)
δ 11.91 (br, 1H), 8.27 (d, J ) 6 Hz, 1H), 7.35 (d, J ) 6 Hz,
1H), 5.60 (br, 2H), 3.12 (s, 2H), 2.50 (t, J ) 6 Hz, 4H), 1.72
(m, 4H), 1.50 (m, 2H).
2-(P ip er id in -1-yl)m eth yl-3H-th ien o[3,2-d ]p yr im id in -4-
on e (72). A suspension of 1.1 g (4.1 mmol) of 71 in 20 mL of
2 N NaOH was stirred under reflux for 20 min. The resulting
solution was cooled, neutralized with 2 N HCl, and extracted
with EtOAc. The combined extracts were washed with brine,
dried, and concentrated to give 0.85 g (83%) of 72 as a white
solid: 1H NMR (CDCl3) δ 7.80 (d, J ) 6 Hz, 1H), 7.30 (d, J )
10.24 (br, 1H), 8.55 (br, 1H), 7.40-7.20 (m, 11H), 5.60 (d, J )
4 Hz, 2H), 4.20 (s, 2H). Anal. (C19H17N5‚HCl) C, H, N.
Meth od I. 2-Bu tyl-4-p h en yla m in oth ien o[3,2-d ]p yr i-
m id in e, Hyd r och lor id e (32). To a solution of 2.00 g (8.82
mmol) of 55 and 0.80 mL (8.82 mmol) of aniline in 20 mL of
acetic acid were added 0.72 g (8.82 mmol) of sodium acetate
and 50 mg of potassium iodide. The mixture was stirred under
reflux for 20 h, cooled, and concentrated. The residue was
partitioned between EtOAc and water; the organic phase was
washed with brine, dried, and concentrated. To the residue
dissolved in acetone was added HCl in dioxane. The resulting
solid was recrystallized from isopropyl alcohol to give 2.38 g
(84%) of the title compound: mp 235-237 °C; 1H NMR
(DMSO) δ 11.52 (br, 1H), 8.49 (d, J ) 4 Hz, 1H), 7.72 (d, J )
6 Hz, 2H), 7.60 (d, J ) 4 Hz, 1H), 7.47 (t, J ) 6 Hz, 2H), 7.32
(m, 1H), 2.94 (t, J ) 8 Hz, 2H), 1.80 (m, 2H), 1.37 (m, 2H),
0.90 (t, J ) 8 Hz, 3H). Anal. (C16H17N3S‚HCl) C, H, N.
Meth od J . 2-Bu tyl-4-[(N,N-d iben zyl)a m in o]th ien o[3,2-
d ]p yr im id in e, Hyd r och lor id e (39). To a solution of 2.00 g
(8.82 mmol) of 55 and 1.84 mL (9.30 mmol) of dibenzylamine
in 50 mL of DMF was added 2.43 g (17.6 mmol) of K2CO3. The
mixture was stirred at 140 °C for 24 h. The resulting
suspension was poured into water and extracted with EtOAc.
The combined extracts were washed, dried, and concentrated.
The residue was purified by flash chromatography, eluting
with EtOAc/hexane (1:2). The resulting solid was dissolved
in acetone, and HCl in dioxane was added. The crystallized
hydrochloride was washed and dried to give 0.84 g (23%) of
the title compound: mp 116-118 °C; 1H NMR (DMSO) δ 8.46
(d, J ) 5 Hz, 1H), 7.60 (d, J ) 5 Hz, 1H), 7.45-7.25 (m, 11H),
5.20 (s, 4H), 2.95 (t, J ) 8 Hz, 2H), 1.71 (m, 2H), 1.30 (m, 2H),
0.83 (t, J ) 8 Hz, 3H). Anal. (C24H25N3S‚HCl‚0.75H2O) H, S,
C: calcd, 65.82; found, 65.34; N: calcd, 9.60; found, 9.15.
2-Bu tyl-3H-th ien o[3,2-d ]p yr im id in -4-th ion e (69). To a
solution of 3.08 g (0.0127 mol) of 41 in 75 mL of toluene was
added 3.84 g (9.50 mmol) of Lawesson’s reagent. The mixture
was stirred under reflux for 1.5 h. The solution was washed
twice with 2 N NaOH, and the combined basic extracts were
neutralized with 2 N HCl. The resulting yellow solid was
filtered, washed, and dried to give 3.15 g (95%) of 69: 1H NMR
(CDCl3) δ 12.27 (br, 1H), 7.96 (d, J ) 5 Hz, 1H), 7.38 (d, J )
5 Hz, 1H), 2.89 (t, J ) 8 Hz, 2H), 1.86 (m, 2H), 1.48 (m, 2H),
0.97 (t, J ) 8 Hz, 3H).
Gen er a l P r oced u r e for th e Alk yla tion of P yr im id in o-
n es 41 a n d 43 a n d P yr im id in th ion e 69. Meth od K.
3-Ben zyl-2-bu tyl-3H-th ien o[3,2-d ]p yr im id in -4-on e (1). To

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4033
6 Hz, 1H), 3.54 (s, 2H), 2.51 (t, J ) 5 Hz, 4H), 1.64 (m, 4H),
1.50 (m, 2H).
mL of H2O was added 14.1 g (0.168 mol) of NaHCO3. The
mixture was stirred under reflux for 66 h. The resulting red
suspension was concentrated, acidified with 2 N HCl, and
extracted with EtOAc. The combined extracts were washed
twice with brine, dried, and concentrated. The resulting yellow
solid was dissolved in 100 mL of EtOH, and the solution cooled
at 0 °C. To the mixture was added 6 mL (0.0825 mol) of
thionyl chloride, and the solution was stirred at room tem-
perature overnight. The reaction mixture was concentrated
and diluted with EtOAc. The resulting solution was washed
twice with brine, dried, and concentrated to give 8.30 g (86%)
of 75 as an orange oil: 1H NMR (CDCl3) δ 8.30 (d, J ) 8 Hz,
1H), 8.17 (d, J ) 8 Hz, 1H), 7.44 (m, 1H), 6.77-6.67 (m, 2H),
4.22 (m, 3H), 1.92 (m, 2H), 1.50 (m, 2H), 1.27 (t, J ) 8 Hz,
3H), 0.98 (t, J ) 8 Hz, 3H).
4-Ch lor o-2-[(p ip er id in -1-yl)m et h yl]t h ien o[3,2-d ]p yr i-
m id in e (73) was prepared according to method E in 79% yield
starting from thieno[3,2-d]pyrimidinone 72 (2.97 g): 1H NMR
(CDCl3) δ 8.01 (d, J ) 6 Hz, 1H), 7.61 (d, J ) 6 Hz, 1H), 3.90
(s, 2H), 2.56 (t, J ) 5 Hz, 4H), 1.65 (m, 4H), 1.46 (m, 2H).
4-Ben zyla m in o-2-[(p ip er id in -1-yl)m eth yl]th ien o[3,2-d ]-
p yr im id in e, h yd r och lor id e (22) was prepared according to
method H starting from 73 (2.51 g). Recrystallization from
isopropyl alcohol gave 1.96 g (52%) of 22: mp 253-257 °C; 1H
NMR (DMSO) δ 10.72 (br, 1H), 9.97 (br, 1H), 8.37 (d, J ) 6
Hz, 1H), 7.50 (d, J ) 6 Hz, 1H), 7.40-7.20 (m, 5H), 4.91 (d, J
) 6 Hz, 2H), 4.47 (s, 2H), 3.26 (br, 4H), 1.73 (br, 4H), 1.47 (br,
2H). Anal. (C19H22N4S‚1.7HCl‚0.6H2O) C, H, N, S.
3-P r op yl-1H-qu in oxa lin -2-on e (76). To a solution of 8.30
g (0.031 mol) of ethyl pentanoate 75 in 50 mL of EtOH were
added 4.0 g (0.072 mol) of iron and 30 mL of 6 N HCl in EtOH.
The mixture was stirred under reflux overnight, cooled, and
concentrated. The resulting residue was diluted with 100 mL
of water, and the pH was adjusted to 5 with NaHCO3. The
suspension was extracted with EtOAc, and the combined
extracts were washed with brine, dried, and concentrated. The
resulting solid was suspended in 70 mL of 2 N NaOH, and
the suspension warmed to 80 °C prior to the addition of 4.70
mL (0.054 mol) of 35% of H2O2. The mixture was stirred at
80 °C for 3 h, cooled, and neutralized with 2 N HCl. The
resulting white solid was filtered, washed with water, and
dried to give 4.20 g (72%) of the desired product 76: 1H NMR
(CDCl3) δ 12.00 (br, 1H), 7.74 (d, J ) 8 Hz, 1H), 7.20-7.15
(m, 3H), 2.85 (t, J ) 7 Hz, 2H), 1.92 (m, 2H), 1.04 (t, 3H).
1-Ben zyl-3-p r op yl-1H-qu in oxa lin -2-on e (8). To a solu-
tion of 4.2 g (0.022 mol) of quinoxaline 76 in 75 mL of DMF
were added 3.81 g (0.022 mol) of benzyl bromide and 6.15 g
(0.045 mol) of K2CO3. The mixture was stirred at room
temperature overnight. The suspension was poured into 200
mL of water, and extracted with EtOAc. The combined
extracts were washed with brine, dried, and concentrated. The
solid residue was washed with pentane, filtered, and recrystal-
lized from EtOH to give 1.2 g (20%) of the title compound as
a colorless solid: mp 101-102 °C; 1H NMR (CDCl3) δ 7.83 (d,
J ) 8 Hz, 1H), 7.40-7.20 (m, 8H), 5.49 (s, 2H), 2.98 (t, J ) 8
Hz, 2H), 1.87 (m, 2H), 1.07 (t, J ) 8 Hz, 3H). Anal.
(C18H18N2O) C, H, N.
4-Am in o-2-bu tylth ien o[3,2-d ]p yr im id in e (74). A solu-
tion of 3.00 g (0.0132 mol) of 55 in 50 mL of ethyl alcohol and
50 mL of 30% NH3 was stirred at 120 °C for 4 h in a pressure
reactor. The mixture was cooled and partitioned between
EtOAc and water. The organic phase was washed with brine,
dried, and concentrated to give 2.05 g (75%) of 74 as a brown
solid: 1H NMR (CDCl3) δ 7.74 (d, J ) 6 Hz, 1H), 7.35 (d, J )
6 Hz, 1H), 6.28 (br, 2H), 2.83 (t, J ) 8 Hz, 2H), 1.80 (m, 2H),
1.43 (m, 2H), 0.95 (t, J ) 8 Hz, 3H).
4-Ben za m id o-2-bu tylth ien o[3,2-d ]p yr im id in e, Hyd r o-
ch lor id e (40). To a solution of 2.00 g (9.70 mmol) of 74 in 20
mL of anhydrous THF was added 6.64 mL (0.011 mmol) of
1.6 M butyllithium at -78 °C. The mixture was stirred for
0.5 h at the same temperature, prior to the addition of a
solution of 1.3 mL (0.011 mol) of benzoyl chloride in 10 mL of
anhydrous THF. The resulting solution was stirred at -78
°C for 0.5 h and at room temperature overnight. The mixture
was concentrated and partitioned between EtOAc and water.
The organic phase was washed with brine, dried, and concen-
trated. The residue was purified by flash chromatography,
eluting with EtOAc/hexane (1:3). To the resulting solid
dissolved in ethyl ether and EtOAc was added 6 N HCl to
provide 0.46 g (15%) of the title compound: mp 103-105 °C;
1H NMR (DMSO) δ 11.48 (br, 1H), 8.35 (d, J ) 5 Hz, 1H), 8.10
(d, J ) 8 Hz, 2H), 7.66 (m, 1H), 7.55 (t, J ) 8 Hz, 2H), 7.50
(d, J ) 5 Hz, 1H), 2.93 (t, J ) 8 Hz, 2H), 1.82 (m, 2H), 1.38
(m, 2H), 0.93 (t, J ) 8 Hz, 3H). Anal. (C17H17N3OS‚HCl) C,
H, N.
3-Ben zyl-2-bu tyl-3H-p yr id o[3,2-d ]p yr im id in -4-on e, Hy-
d r och lor id e (3). A suspension of 2.50 g (0.018 mol) of
3-amino-2-pyridincarboxylic acid in 15 mL of valeric anhydride
was stirred at 140 °C for 1 h. The mixture was cooled, and
the excess of the valeric anhydride was distilled. The resulting
solid was suspended in 1.25 mL (0.018 mol) of benzylamine
and stirred at 200 °C for 1 h. The residue was purified by
flash chromatography, eluting with EtOAc/hexane (2:3). To
the resulting oil dissolved in acetone was added HCl in dioxane
Biology. Milrinone was obtained from IMPEX Qu´ımica
(Barcelona, Spain). Rolipram and RS-14491 were synthesized
in the Medicinal Chemistry Department of Almirall-Prodes-
farma. Reagents were purchased from commercial suppliers
and used as received.
Purification and characterization of cyclic nucleotide phos-
phodiesterases 3 and 4 and PDE 3 and PDE 4 enzyme assays
have been described previously.36,59,60 The [3H]Rolipram dis-
placement was performed according to described protocols.36,61
1
to give 0.49 g (8%) of 3: mp 204-206 °C; H NMR (DMSO) δ
9.07 (d, J ) 5 Hz, 1H), 8.77 (d, J ) 8 Hz, 1H), 7.72 (dd, J ) 5
Hz, J ) 8 Hz, 1H), 7.45-7.10 (m, 5H), 5.44 (s, 2H), 2.88 (t, J
) 8 Hz, 2H), 1.67 (m, 2H), 1.31 (m, 2H), 0.81 (t, J ) 8 Hz,
3H). Anal. (C18H19N3O‚HCl) C, H, N.
Cyclic AMP Accu m u la tion . P r ep a r a tion of Gu in ea
P ig Eosin op h ils. Male Dunkin-Hartley guinea pigs (300-
400 g) were injected intraperitoneally with 0.5 mL of donor
horse serum once per week for 6-7 weeks. Four days after
the last injection, the animals were anaesthetized with ether
and killed by jugular vein bleeding. A ventral incision was
made and 80 mL of HBSS (Hank’s balanced salt serum)
without Ca2+ and Mg2+ was poured into the abdominal cavity.
The abdomen was gently massaged for 5 min and then the
peritoneal exudate was aspirated and centrifuged at 250 G
for 10 min at 4 °C. The supernatant was discarded, and the
pellet was resuspended in 2 mL of RPMI-1640 medium
containing 10% fetal bovine serum (FBS). The enriched
eosinophil fraction was separated using discontinuous gradi-
ents of Percoll. For the preparation of gradients, 9 parts of
Percoll were mixed with 1 part of 1.5 M NaCl, and the mixture
was diluted with RPMI-1640 containing 10% FBS to obtain
densities of 1.075, 1.085, and 1.105 g/mL. Starting with the
most dense layer, 2 mL of decreasing densities of Percoll were
carefully layered into a 15 mL conical centrifuge tube, on top
of which the cells were placed in a 2 mL volume. The gradients
4-Ben zyl-2-p r op yl-4H -p yr id o[2,3-b]p yr a zin -3-on e (7).
To a solution of 2.23 g (0.016 mol) of 2-oxopentanoic acid,
sodium salt, in 10 mL of EtOH and 8 mL of 2 N HCl was added
a solution of 3.22 g (0.016 mol) of 3-amino-2-benzylaminopy-
ridine50 in 35 mL of EtOH. The mixture was stirred under
reflux overnight, cooled, and concentrated. The residue was
extracted with EtOAc. The combined extracts were washed,
dried, and concentrated. The resulting solid was purified by
flash chromatography eluting with EtOAc/hexane (1:3) to
provide 7 as a white solid: yield 0.86 g (19%); mp 90-91 °C;
1H NMR (DMSO) δ 8.56 (d, J ) 6 Hz, 1H), 8.43 (d, J ) 8 Hz,
1H), 7.42 (dd, J ) 8 Hz, J ) 6 Hz, 1H), 7.40-7.15 (m, 5H),
5.55 (s, 2H), 2.86 (t, J ) 8 Hz, 2H), 1.76 (m, 2H), 0.98 (t, J )
8 Hz, 3H). Anal. (C17H17N3O) C, H, N.
Eth yl 2-(2-n itr op h en yla m in o)p en ta n oa te (75). To a
solution of 4.94 g (0.035 mol) of 1-fluoro-2-nitrobenzene and
8.30 g (0.070 mol) of D,L-norvaline in 125 mL of EtOH and 30

4034 J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21
Crespo et al.
(13) Elliott, M. J .; Maini, R. N.; Feldmann, M.; Kalden, J . R.; Antoni,
C.; Smolen, J . S.; Leeb, B.; Breedveld, F. C.; Macfarlane, J . D.;
Bijl, H.; Woody, J . N. Randomised Double-bind Comparison of
Chimeric Monoclonal Antibody to Tumour Necrosis Factor R
(cA2) Versus Placebo in Rheumatoid Arthritis. Lancet 1994, 344,
1105-1110.
(14) Malaisse, W. J .; Malaisse-Lagae, F.; Mayhew, D. A Possible Role
for the Adenylcyclase System in Insulin Secretion. J . Clin. Invest.
1967, 46, 1724-1734.
were centrifuged in a swing-out rotor at 425 G for 20 min at
18 °C. The eosinophil-rich cell band was collected and washed
twice with HBSS containing Ca2+ and Mg2+
. Cell viability,
as determined by Trypan Blue exclusion, was greater than
98%, and eosinophil purity, as determined by May-Gru¨nwald-
Giemsa staining, was greater than 95%.
Deter m in a tion of Eosin op h il Cyclic AMP Accu m u la -
tion . Freshly prepared eosinophils ((1-2) × 106), resuspended
in HBSS containing Ca2+ and Mg2+, were incubated with the
compounds at the indicated concentrations for 10 min at 37
°C. Incubations were continued for a another 2 min in the
presence of isoprenaline (10 µM) and terminated by addition
of two volumes of ice-cold ethanol. The samples were centri-
fuged (2000 G) for 15 min at 4 °C, and the supernatant was
transferred to a clean tube. The samples were dried by gassing
with nitrogen at 60 °C, and the pellet was resuspended in
water. Cyclic AMP was quantified using an enzyme-immu-
noassay kit from Amersham Life Sciences (U.K.) following the
protocol without acetylation. For the calculations, the cAMP
levels obtained in the presence of isoprenaline were considered
as 100%, and the values obtained in the presence of compounds
were corrected accordingly. EC50 values were calculated by
adjusting the data to a sigmoid curve using the program Inplot.
Four independent measurements made in two different days
were obtained for each drug concentration.
(15) Zawalich, W. S.; Rasmussen, H. Control of Insulin Secretion: a
Model Involving Ca2+, cAMP and Diacylglycerol. Mol. Cell.
Endocrinol. 1990, 70, 119-137.
(16) Leibowitz, M. D.; Biswas, C.; Brady, E. J .; Conti, M.; Cullinan,
C. A.; Hayes, N. S.; Manganiello, V. C.; Saperstein, R.; Wang,
L. H.; Zafian P. T.; Berger, J . A. Novel Insulin Secretagogue Is
a Phosphodiesterase Inhibitor. Diabetes 1995, 44, 67-74.
(17) La¨hdevirta, J .; Maury, C. P.; Teppo A. M.; Repo, H. Elevated
Levels of Circulating Cachectin/Tumour Necrosis Factor in
Patients with Acquired Immunodeficiency Syndrome. Am. J .
Med. 1988, 85, 289-291.
(18) Kobayashi, S.; Hamamoto, Y.; Kobayuashi, N.; Yamamoto, N.
Serum Level of TNFR in HIV-Infected Individuals. AIDS 1990,
4, 169-170.
(19) Lowe III, J . A.; Cheng, J . B. The PDE 4 Family of Calcium-
Independent Phosphodiesterases Enzymes. Drugs Future 1992,
17, 799-807.
(20) Torphy, T. J .; Livi, G. P.; Christensen, S. B. Novel Phosphodi-
esterase Inhibitors for the Therapy of Asthma. Drugs News
Perspect. 1993, 6, 203-214.
(21) Palfreyman, M. N. Phosphodiesterase Type 4 Inhibitors as
Antiinflammatory Agents. Drugs Future 1995, 20, 793-804.
(22) Glaser, T.; Traber, J . TVX 2706. A New Phosphodiesterase
Inhibitor with Antiinflammatory Action. Biochemical Charac-
terization. Agents Actions, 1984, 15, 341-348.
(23) Lowe III, J . A.; Archer, R. L.; Chapin, D. S.; Cheng, J . B.; Helweg,
D.; J ohnson, J . L.; Koe, B. K.; Lebel, L. A.; Moore, P. F.; Nielsen,
J . A.; Russo, L. L.; Shirley, J . T.; Structure-Activity Relationship
of Quinazolinedione Inhibitors of Calcium-Independent Phos-
phodiesterase. J . Med. Chem. 1991, 34, 624-628.
(24) (a) Wilhem, R.; Loe, B.; Alvarez, R.; Devens, B.; Fong, A. Pyrido-
[2,3-d]pyridazinones as Potent and Selective Type 4 Phosphodi-
esterase Inhibitors. 8th RSC-SCI Medicinal Chemistry Sympo-
sium, Cambridge, 1995, P32. (b) Alvarez, R.; Daniels, D. V.;
Shelton, E. R.; Baecker, P. S.; Fong, T. A. T.; Devens, B.;
Wilhelm, R.; Eglen, R. M.; Cont, M. An Isoform-Selective
Inhibitor of Cyclic AMP-Specific Phosphodiesterase (PDE 4) with
Antiinflammatory Properties. In Phosphodiesterase Inhibitors;
Schudt, C., Dent, G., Rabe, K. F., Eds.; Academic Press: London,
1996; p 161-171.
(25) Alvarez, R.;. Wilhelm, R.; Shelton, E. R.; Daniels, D. V.; Yang,
D.; Kelly, K.; Eglen, R. M. Potent and Selective Inhibition of
Cyclic AMP Specific Phosphodiesterase (PDE 4) by Structural
Analogues of Nitraquazone. Can. J . Physiol. Pharmacol. 1994,
72 (Suppl. 1), Abst. P18.2.11, p 510.
Ack n ow led gm en t. We thank Mr. Ramo´n Roy and
Mr. Pere Vilaseca for their technical assistance and Mr.
J osep M. Huerta and Mr. Amadeu Gavalda` for their
kind support in analytical and binding evaluations.
Refer en ces
(1) A preliminary report was presented at XIVth International
Symposium on Medicinal Chemistry, Maastricht, September
1996, P5.14 and P5.36.
(2) Palacios, J . M.; Beleta, J .; Segarra, V. Second Messengers
Systems as Targets for New Therapeutic Agents: Focus on
Selective Phosphodiesterase Inhibitors. Il Farmaco, 1995, 50,
819-827.
(3) Sommer, N.; Lo¨schmann, P. A.; Northoff, G. H.; Weller, M.;
Steinbrecher, A.; Steinbach, J . P.; Lichtenfels, R.; Meyermann,
R.; Riethmu¨ller, A.; Fontana, A.; Dichgans, J .; Martin, R. The
Antidepressant Rolipram Suppresses Cytokine Production and
Prevents Autoimmune Encephalomyelitis. Nat. Med. 1995, 1,
244-248.
(4) Essayan, D. M.; Kagey-Sobotka, A.; Lichtenstein, L. M.; Huang,
S. K. Differential Regulation of Human Antigen-specific Th1 and
Th2 Lymphocyte Responses by Isoenzyme Selective Cyclic
Nucleotide Phosphodiesterase Inhibitors. J . Pharmacol. Exp.
Ther. 1997, 282, 505-512.
(5) Bourne, H. R.; Lichtenstein, L. M.; Melmon, K. L.; Henney, C.
S.; Weinstein, Y.; Shearer, G. M. Modulation of Inflammation
and Immunity by Cyclic AMP. Science 1974, 184, 19-28.
(6) Kammer, G. M. The Adenylate Cyclase-cAMP-Protein Kinase
A Pathway and Regulation of the Immune Response. Immunol.
Today. 1988, 9, 222-229.
(7) Epstein, P. M.; Hachisu, R. Cyclic Nucleotide Phosphodiesterase
in Normal and Leukemic Human Lymphocytes and Lympho-
blasts. Adv. Cyclic Nucleotide Protein Phosphorylation Res. 1984,
16, 303-24.
(8) Klein-Tebbe, J .; Wicht, H.; Gagne´, L.; Friese, A.; Schunack, W.;
Schudt, C.; Kunkel, G. Inhibition of IgE-Mediated Histamine
Release from Human Peripheral Leukocytes by Selective Phos-
phodiesterase Inhibitors. Agent Actions 1992, 36, 200-206.
(9) Chan, S. C.; Reifsnyder, D.; Beavo, J . A.; Hanifin, J . M.
Immunochemical Characterisation of the Distinct Monocyte
Cyclic AMP-Phosphodiesterase from Patients with Atopic Der-
matitis. J . Allergy Clin. Immunol. 1993, 91, 1179-1188.
(10) Genain, C. P.; Roberts, T.; Davis, R. L.; Nguyen, M. N.; Uccelli,
A.; Faulds, D.; Li, Y.; Hedgpeth, J .; Hauser, S. L. Prevention of
(26) Manabe, H.; Ohnuma, K.; Ichimura, M.; Kawakita, T.; Suzuki,
F.; Ohmori, K. Pharmacological Properties of a New Bronchodi-
lator, KF19514. 68th Annual Meeting of the J apanese Pharma-
cological Society, Nagoya, J apan, 1995, S24-4.
(27) Suzuki, F.; Kuroda, T.; Kawakita, T.; Manabe, H.; Kitamura,
S.; Ohmori, K.; Ichimura, M.; Kase, H.; Ichikawa, S. New
Bronchodilators. 3. Imidazo[4,5-c][1,8]naphthyridin-4(5H)-ones.
J . Med. Chem. 1992, 35, 4866-4874.
(28) Suzuki, F.; Kuroda, T.; Tamura, T.; Sato, S.; Ohmori, K.;
Ichikawa, S. New Antiinflammatory Agents. 2. 5-Phenyl-3H-
imidazo[4,5-c][1,8]naphthyridin-4(5H)-ones: A New Class of
Nonsteroidal Antiinflammatory Agents with Potent Activity Like
Glucocorticoids. J . Med. Chem. 1992, 35, 2863-2870.
(29) Duplantier, A. J .; Biggers, M. S.; Chambers, R. J .; Cheng, J . B.;
Cooper, K.; Damon, D. B.; Eggler, J . F.; Kraus, K. G.; Marfat,
A.; Masamune, H.; Pillar, J . S.; Shirley, J . T.; Umland, J . P.;
Watson, J . W. Biarylcarboxylic Acids and Amides: Inhibition
of Phosphodiesterase Type
4
versus [3H]Rolipram Binding
Activity and their Relationship to Emetic Behaviour in the
Ferret. J . Med. Chem. 1996, 39, 120-125.
(30) Barnette. M. S. Challenges for Drug Discovery. New Drugs for
Asthma-III, Montebello, Quebec, J uly, 1994.
Autoimmune Demyelination in Non-Human Primates by
a
(31) Russo, L. L.; Lebel, L. A.; Koe, B. K. Effects of Selected
Phosphodiesterase PDE Inhibitors on Calcium-Independent PDE
Activity and Rolipram Binding Sites of Cerebral Cortex. Soc.
Neurosci. Abstr. 1987, 13, 903.
(32) Vinick, F. J .; Saccomano, N. A.; Koe, B. K.; Nielsen, J . A.;
Williams, I. H.; Thadeio, P. F.; J ung, S.; Meltz, M.; J ohnson, J .,
J r.; Lebel, L. A.; Russo, L. L.; Helweg, D. Nicotinamide Ethers:
Novel Inhibitors of Calcium-Independent Phosphodiesterase and
[3H]Rolipram Binding. J . Med. Chem., 1991, 34, 86-89.
cAMP-Specific Phosphodiesterase Inhibitor. Proc. Natl. Acad.
Sci. U.S.A. 1995, 92, 3601-3605.
(11) Hopkins, S. J .; Meager, A. Cytokines in Synovial Fluid: II. The
Presence of Tumour Necrosis Factor and Interferon. Clin. Exp.
Immunol. 1988, 73, 88-92.
(12) Saxne, T.; Palladino, M. A. J r.; Heinegard, D.; Talal, N.;
Wollheim, F. A . Detection of Tumour Necrosis Factor R but not
Tumour Necrosis Factor â in Rheumatoid Arthritis Synovial
Fluid and Serum. Arthritis Rheum. 1988, 31, 1041-1045.

Design of New Thieno[3,2-d]pyrimidines
J ournal of Medicinal Chemistry, 1998, Vol. 41, No. 21 4035
(33) Masamune, H.; Cheng, J . B.; Cooper, K.; Eggler, J . F.; Marfat,
A.; Marshall, S. C.; Shirley, J . T.; Tickner, J . E.; Umlamd, J . P.;
Vazquez, E.; Discovery of Micromolar PDE 4 Inhibitors that
Exhibit Much Reduced Affinity for the [3H]Rolipram Binding
Site: 3-Norbornyloxy-4-methoxyphenylmethylene Oxindoles.
Bioorg. Med. Chem. Lett. 1995, 5, 1965-1968.
rimidines. I. 4-Oxo-3,4-dihydrothieno[3,2-d]pyrimidine. Bull.
Soc. Chim. Fr. 1970, 10, 3630-3636. (c) Robba, M.; Lecomte, J .
M.; Cugnon de Se´vricourt, M. Thienopyrimidines. II. Thieno-
[3,2-d]pyrimidines. Tetrahedron 1971, 27 (2), 487-499. (d)
Elderfield, R. C. Heterocyclic Compounds; J . Wiley & Sons: New
York, 1957; Vol. 6, pp 324-376.
(34) Cheng, J . B.; Cooper, K.; Duplantier, A. J .; Eggler, J . F.; Kraus,
K. G.; Marshall, S. C.; Marfat, A.; Masamune, H.; Shirley, J . T.;
Tickner, J . E.; Umland, J . P. Synthesis and In Vitro Profile of a
Novel Series of Catechol Benzimidazoles. The Discovery of
Potent, Selective Phosphodiesterase Type 4 Inhibitors with
Greatly Attenuated Affinity for the [3H]Rolipram Binding Site.
Biorg. Med. Chem. Lett. 1995, 5, 1969-1972.
(49) Suzuki, Y.; Yasso, M.; Nishimura, K.; Sacki, K.; Takayanagi, N.;
Saito, T.; Hayashi, E. 6-Substituted Alkoxy-2-oxo-1,2-dihydro-
quinoxaline Derivatives. Eur. Pat. Applic. 311378 A2, 1989 (to
Toyo J ozo KK).
(50) J anssens, F.; Torremans, J .; J anssen, M.; Stokbroekx, R. A.;
Luyckx, M. New Antihistaminic N-Heterocyclic 4-Piperidi-
namines. 3. Synthesis and Antihistaminic Activity of N-(4-
Piperidinyl)-3H-imidazo[4,5-b]pyridin-2-amines. J . Med. Chem.
1985, 28, 1943-1947.
(51) Alousi, A. A.; Carter, J . M.; Montenaro, M. J .; Fort, D. J .; Ferrari,
R. A. Cardiotonic Activity of Milrinone, a New and Potent
Cardiac Bipyridine, on the Normal and Failing Heart of Experi-
mental Animals. J . Cardiovasc. Pharmacol. 1983, 5, 792-803.
(52) Barnes, P. J .; Chung, K. F.; Page, C. P. Inflammatory Mediators
and Asthma. Pharmacol. Rev. 1988, 40, 49-84.
(53) Souness, J . E.; Carter, C. M.; Diocee, B. K.; Hassall, G. A.; Wood,
L. J .; Turner, N. C. Characterization of Guinea Pig Eosinophil
Phosphodiesterase Activity. Assessment of its Involvement in
Regulating Superoxide Generation. Biochem. Pharmacol. 1991,
42, 937-945.
(54) Wamser, C. C.; Wagner, W. R. Type II Photoelimination from
R-Cycloalkylacetophenones and a Polystyrene-Bound Analog. J .
Am. Chem. Soc., 1981, 103 (24), 7232-7234.
(55) Barger, G.; Robinson, R.; Smith, L. H. Synthetical Experiments
Related to Carpaine. III. Some Derivatives of Tetrahydrofuran
and Intermediates of the Aliphatic Series. J . Chem. Soc. 1937,
718-725.
(56) Ong-Lee, A.; Sylvester, L.; Wasley, J . W. F. Synthesis of 1,3,4,-
14b-Tetra hydro-2H,10H-pyra zino[1,2-a ]pyrrolo[2,1-c][1,4]-
dibenzoazepines. J . Heterocycl. Chem. 1983, 20, 1565-1569.
(57) Andrews, A. F.; Smith, D. M.; Hodson, H. F.; Thorogood, P. B.
Synthetic Approaches to Some Aza Analogs of Benzimidazole
N-Oxides. Part 1. The Imidazo[4,5-b]pyridine Series. J . Chem.
Soc., Perkin Trans. 1 1982, (12), 2995-3006.
(58) Shoppee, C. W. Symmetrical Triad Prototropic Systems. IX. The
Influence of Polynuclear Aryl Groups upon Mobility and Equi-
librium in the R,γ-Diarylmethyleneazomethine System. J . Chem.
Soc. 1933, 37-45.
(35) Pinto, I. L.; Buckle, D. R.; Readshaw, S. A.; Smith, D. G. The
Selective Inhibition of Phosphodiesterase
4 by Benzopyran
Derivatives of Rolipram. Bioorg. Med. Chem. Lett. 1993, 3,
1743-1746.
(36) Dal Piaz, V.; Giovannoni, M. P.; Castellana, C.; Palacios, J . M.;
Beleta, J .; Dome´nech, T.; Segarra, V. Novel Heterocyclic-Fused
Pyridazinones as Potent and Selective Phosphodiesterase 4
Inhibitors. J . Med. Chem. 1997, 40, 1417-1421.
(37) Chem-X, version J an96, Chemical Design Limited, Oxfordshire
OX7 5SR, England.
(38) Davies, E. K.; Murrall, N. W. How Accurate Does a Force Field
Need to Be? Comput. Chem. 1989, 13, 149-156.
(39) MOPAC version 6.00 (program no. 506 QCPE, Bloomington, IN).
(40) Stewart, J . J . P. MOPAC: A Semiempirical Molecular Orbital
Program. J . Comput. Aided Mol. Des. 1990, 4, 1-105.
(41) Dewar, M. J . S.; Zoebish, E. G.; Healy, E. F.; Stewart, J . J . P.
AM1: A New General Purpose Quantum Mechanical Molecular
Model. J . Am. Chem. Soc. 1985, 107, 3902-3909.
(42) GRID, version 14, Molecular Discovery Limited, Oxford OX2
9LL, England.
(43) Goodford, P. J . A Computational Procedure for Determining
Energetically Favorable Binding Sites on Biologically Important
Macromolecules. J . Med. Chem. 1985, 28, 849-857.
(44) Wade, R. C.; Clark, K. J .; Goodford, P. J . Further Development
of Hydrogen Bond Functions for Use in Determining Energeti-
cally Favorable Binding Sites on Molecules of Known Structure.
1. Ligand Probe Groups with the Ability To Form Two Hydrogen
Bonds. J . Med. Chem. 1993, 36, 140-147.
(45) Wade, R. C.; Goodford, P. J . Further Development of Hydrogen
Bond Functions for Use in Determining Energetically Favorable
Binding Sites on Molecules of Known Structure. 2. Ligand Probe
Groups with the Ability To Form More Than Two Hydrogen
Bonds. J . Med. Chem. 1993, 36, 148-156.
(46) Boobbyer, D. N. A.; Goodford, P. J .; McWhinnie, P. M.; Wade,
R. C. New Hydrogen Bond Potentials for Use in Determining
Energetically Favorable Binding Sites on Molecules of Known
Structure. J . Med. Chem. 1989, 32, 1083-1094.
(47) A similar pharmacophore model was simultaneously presented
at the XIVth International Symposium on Medicinal Chemistry,
Maastrich, September 1996. Polymeropoulos, E. E.; Ho¨fgen, N.
A Pharmacophore Model for PDE 4 Inhibitors. P5.44.
(48) (a) Mueller, R.; Gebert, L.; Pech, R.; Bohm, R. On Thieno
Compounds. Part 17. Preparation of Some New Derivatives of
Thieno[2,3-d]pyrimidine. Pharmazie 1993, 48 (8), 585-587. (b)
Robba, M.; Lecomte, J . M.; Cugnon de Se´vricourt, M. Thienopy-
(59) Gristwood, R. W.; Beleta, J .; Bou, J .; Cardelus, I.; Ferna´ndez,
A. G.; Llenas, J .; Berga, P. Studies on the Cardiac Actions of
Flosequinan in Vitro. Br. J . Pharmacol. 1992, 105, 985-991.
(60) Thompson, W. J .; Strada, S. J .; Cyclic Nucleotide Phosphodi-
esterase (PDE). In Method of Enzymatic Analysis, 3th ed.;
Bergmayer, H. U., Ed.; Verlag-Chemie: Weinheim, 1984; Vol.
IV, pp 127-134.
(61) Schneider, H. H.; Schmiechen, R.; Brezinski, M.; Seidler, J .
Stereospecific Binding of Antidepressant Rolipram to Brain
Protein Structures. Eur. J . Pharmacol. 1986, 127, 105-115.
J M981012M