Design, synthesis, and pharmacological evaluation of monocyclic pyrimidinones as novel inhibitors of PDE5

Article abstract of DOI:10.1021/jm301159y

Cyclic nucleotide phosphodiesterase type 5 (PDE5) is a prime drug target for treating the diseases associated with a lower level of the cyclic guanosine monophosphate (cGMP), which is a specific substrate for PDE5 hydrolysis. Here we report a series of novel PDE5 inhibitors with the new scaffold of the monocyclic pyrimidin-4(3H)-one ring developed using the structure-based discovery strategy. In total, 37 derivatives of the pyrimidin-4(3H)-ones, were designed, synthesized, and evaluated for their inhibitory activities to PDE5, resulting in 25 compounds with IC₅₀ ranging from 1 to 100 nM and 11 compounds with IC₅₀ ranging from 1 to 10 nM. Compound 5, 5,6-diethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl] pyrimid-4(3H)-one, the most potent compound, has an excellent IC₅₀ (1.6 nM) in vitro and a good efficacy in a rat model of erection. It thus provides a potential candidate for the further development into a new drug targeting PDE5.

Full text of DOI:10.1021/jm301159y

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Article
Design, Synthesis, and Pharmacological Evaluation of
Monocyclic Pyrimidinones as Novel Inhibitors of PDE5
Guan Wang, Zheng Liu, Tiantian Chen, Zhen Wang, Huaiyu Yang, Mingyue Zheng, Jing
Ren, Guanghui Tian, Xiaojun Yang, Li Li, Jianfeng Li, Jin Suo, Rongxia Zhang, Xiang-
Rui Jiang, Nicholas Kenneth Terrett, Jing-Shan Shen, Yechun Xu, and Hualiang Jiang
J. Med. Chem., Just Accepted Manuscript • Publication Date (Web): 08 Nov 2012
Downloaded from http://pubs.acs.org on November 17, 2012
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Design, Synthesis, and Pharmacological
Evaluation of Monocyclic Pyrimidinones as
Novel Inhibitors of PDE5
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†
,#
§,#
†,#
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Guan Wang, Zheng Liu, Tiantian Chen, Zhen Wang, Huaiyu Yang,
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Mingyue, Zheng, Jing Ren, Guanghui Tian, Xiaojun Yang, Li Li,
†
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Jianfeng Li, Jin Suo, Rongxia Zhang, Xiangrui Jiang, Nicholas
§
*,†
*,†
*,†
Kenneth Terrett, Jingshan Shen, Yechun Xu, and Hualiang Jiang
†
CAS Key Laboratory for Receptor Research, Shanghai Institute of
Materia Medica, Chinese Academy of Sciences, 555 Zuchongzhi Road,
Shanghai 201203, China
§
Vitargeta Therapeutics Inc. Plainsboro, NJ 08536, USA.
ABSTRACT
Cyclic nucleotide phosphodiesterase type 5 (PDE5) is a prime drug
target for treating the diseases associated with a lower level of the cyclic
guanosine monophosphate (cGMP), which is a specific substrate for
PDE5 hydrolysis. Here we report a series of novel PDE5 inhibitors with
the new scaffold of monocyclic pyrimidinꢀ4(3H)ꢀone ring developed
using the structureꢀbased discovery strategy. In total, 37 derivatives of the
pyrimidinꢀ4(3H)ꢀones, were designed, synthesized, and evaluated for
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their inhibitory activities to PDE5, resulting in 25 compounds with IC50
ranging from 1 to 100 nM and 11 compounds with IC ranging from 1
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to10 nM. Compound 5,
,6ꢀdiethylꢀ2ꢀ[2ꢀnꢀpropoxyꢀ5ꢀ(4ꢀmethylꢀ1ꢀpiperazinylsulfonyl)phenyl]pyr
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imidꢀ4(3H)ꢀone, the most potent compound, has an excellent IC (1.6
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nM) in vitro and a good efficacy in a rat model of erection. It thus
provides a potential candidate for the further development into a new
drug targeting PDE5.
INTRODUCTION
Cyclic nucleotide phosphodiesterase type 5 (PDE5), mostly distributed
in smooth muscle and found in corpus cavernosum, heart, lung, platelets,
1ꢀ4
prostate, urethra, bladder, liver, brain, and stomach, is responsible for
specifically cleaving the second messenger, cyclic guanosine
5
monophosphate (cGMP). Inhibitors of PDE5 prevent the hydrolysis of
cGMP, leading to their potential to treat the diseases associated with low
6
cGMP levels. PDE5 is thus a prime target for the development of
inhibitors which have served as drugs in the treatment of male erectile
dysfunction (ED) and cardiovascular diseases. For example, sildenafil,
the prototypical member of the PDE5 inhibitors, was approved by FDA in
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1998 as the first oral medicine for the treatment of ED, and was also
launched for the treatment of pulmonary arterial hypertension (PAH) in
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005.
Two other agents, vardenafil and tadalafil, are also used to
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treat ED and PAH.
Moreover, tadalafil has received the FDA approval
for the treatment of lower urinary tract symptoms (LUTS) in men with
benign prostatic hyperplasia (BPH). Another inhibitor, avanafil, as a
second generation PDE5 inhibitor, was also approved for the treatment of
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ED. Despite deficiencies with these inhibitors,
such as the lack of
sufficient selectivity against other PDE isozymes, most notably PDE6 or
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PDE11, it has been proved that the inhibition of PDE5 is an effective
therapy for the diseases mentioned above and there is continuing interest
in discovering novel PDE5 inhibitors. Additionally, the medical
application of PDE5 inhibitors for other diseases such as stroke,
Raynaud’s disease, overactive bladder, and premature ejaculation has also
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been indicated.
The solved crystal structures of the PDE5 catalytic domain in complex
with different inhibitors have revealed the characteristic binding
interactions of inhibitors with the enzyme. These include mono or
bidentate hydrogen bonds (Hꢀbonds) between inhibitors and the residue
Q817, hydrophobic interactions, in particular the πꢀπ stacking
interactions between the aromatic rings of inhibitors and the hydrophobic
clamp mainly composed of residues V782 and F820, and occasionally
2+
interactions also formed between inhibitors and two metal ions (Zn and
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Mg in most cases) coordinated at the active site of the catalytic
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domain
. For example, in the crystal structure of PDE5/sildenafil
(
pdb codes 1UDT and 2H42), bidentate Hꢀbonds are formed between the
amide moiety of the pyrazolopyrimidinone of sildenafil and the
sideꢀchain of Q817. A third Hꢀbond is formed between the N2 atom of
the pyrazol ring and a water molecule that simultaneously forms Hꢀbonds
with the sideꢀchain of Y612, the mainꢀchain of D764, and a water
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molecule coordinating to Zn (Figure 1A). The pyrazole ring as well as
the ethoxyphenyl group of sildenafil also interact with residues V782,
L785, Y612, F820, F786, A783, and L804, in particular, including a
19, 21
faceꢀtoꢀface πꢀπ stacking interaction with the phenyl ring of F820
.
All these available crystal structures associated with the pharmacological
data enable us to carry out structureꢀbased discovery or design of more
potent and pharmacologically favorable inhibitors of PDE5.
We previously designed and synthesized a series of
quinazolinꢀ4(3H)ꢀones by introducing the phenyl ring influenced by the
structure of sophoflavescenol instead of the pyrazol ring in sildenafil so
as to improve the hydrophobic interactions with the residues of PDE5 and
reduce or cancel the interactions of the inhibitor with the metal ions. This
finally resulted in inhibitors of PDE5 such as compound 1 (Scheme 1)
with similar potency to sildenafil but improved selectivity of PDE5 over
2
2
PDE6. In order to reduce the size of the compound but still keep its the
hydrophobic interactions with the enzyme, we designed and synthesized
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compound 2, bearing a fused cyclopentene ring instead of the
sophoflavescenolꢀderived substituted phenyl ring in compound 1.
However, it turns out that compound 2 displayed a dramatic loss in PDE5
affinity: only 24% inhibition at 1 ꢁM compound concentration. We
determined the crystal structure of PDE5 in complex with compound 2
and superimposed it to the structure of PDE5/sildenafil. As we can see
from the superimposition of compound 2 and sildenafil in Figure 1B, the
scaffold of compound 2 is similar to sildenafil but it is much simplified
around the bicyclic rings. Because of its hydrophobic character, the
cyclopentene ring in compound 2 binds to the subꢀpocket (SP1) created
primarily by residues L765, I768, and F820, resulting a shift of the whole
compound, thus breaking its perfect πꢀπ stacking interactions with F820
compared to the binding of sildenafil (Figures 1B, 1C and 2B). However,
it is surprising to see that the water molecule which forms Hꢀbonds with
both residues (Y612 and D764) and the water molecule coordinating to
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2+
Zn also appear in the complex structure of PDE5/2, although there is no
Hꢀbond formed between the compound 2 and the water molecule (Figure
1
B). We also find this water molecule in the apo structure of PDE5 (pdb
code 2H40) and five other crystal structures of PDE5 in complex with the
inhibitors which have a similar scaffold structure and binding position as
the pyrazolopyrimidinone of sildenafil. These structures are PDE5 in
complex with sildenafil (pdb code 1UDT and 2H42), IBMX (pdb code
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RKP ),
ꢀethoxyꢀ4ꢀ(1ꢀmethylꢀ7ꢀoxoꢀ3ꢀpropylꢀ6,7ꢀdihydroꢀ1Hꢀpyrazolo[4,3ꢀd]pyr
imidinꢀ5ꢀyl)thiopheneꢀ2ꢀsulfonamide (pdb code 3BJC), and vardenafil
pdb code 3B2R) (Figure 1C). The resolution of these five selected
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complex structures is all above 2.5 angstroms in order to see the key
water molecules that Hꢀbond with the inhibitors as well as Y612 and
D764. It is notable that a water molecule is always present in the
structures, and that the hydrophobic substituents on the pyrazol, pyrrole
or other heterocyclic ring occupy the same subꢀpockets SP1 and SP2
(
Figure 1C). This indicates that the binding of the water molecule to the
enzyme is strong and acts essentially as a residue of the enzyme.
Hꢀbonding of the polar atom in the inhibitor with this water molecule
increases its binding affinity to the enzyme. Meanwhile, the hydrophobic
substituents on the heterocyclic ring interact with the two subꢀpockets
nearby. Accordingly, the hydrophobic cyclopentene ring of compound 2
has to bind into the subꢀpocket SP1 so as to avoid a close contact with
this water molecule.
These structural insights inspired us to design and synthesize
compound 3, opening the cyclopentene ring and retaining only a planar
monocyclic structure in the compound so as to enable the two
hydrophobic substituent groups on the pyrimidinꢀ4(3H)ꢀone ring to fit
into the two subꢀpockets SP1 and SP2. Fortunately, compound 3
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exhibited a significant increase in the potency to PDE5 compared to that
of compound 2. After that, we designed and synthesized a series of
substituted pyrimidones represented by a general structure of 4 to
systematically study the monocyclic pyrimidones as inhibitors of PDE5.
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Among these compounds, compound 5 has an excellent IC (1.6 nM),
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good selectivity (29ꢀfold) over PDE6 in vitro and good efficacy in a rat
model of erection. The binding position of compound 5 in its crystal
structure with PDE5 is superimposed well on that of sildenafil as shown
in Figure 1D, which proved that the ring opening could fully overcome
the disadvantages of the cyclopentene ring in compound 2, shift the
whole compound back to the same binding location as sildenafil, and thus
recover or even improve the potency of the compound against PDE5.
Accordingly, our study not only obtained more potent inhibitors of PDE5
but also provided a novel scaffold for further selectivity and metabolism
optimization of PDE5 inhibitors. This study therefore presents a good
example of the structureꢀbased discovery of potent PDE5 inhibitor.
1
. CHEMISTRY
We describe herein the synthesis of novel inhibitors of PDE5 bearing a
monocyclic pyrimidinone core. To gain a rapid exploration of SAR, two
different synthetic routes for bulk preparation of two series of designed
derivatives were applied. The synthetic approach to obtain
2
ꢀphenylpyrimidinꢀ4(3H)ꢀones 2, 3, 5, and 9ꢀ31 is outlined in Scheme 2.
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Compound 7, 2ꢀPropoxyꢀ5ꢀ(4ꢀmethylpiperazinꢀ1ꢀyl)
sulphonylbenzamidine, is the key intermediate, and the synthesis of this
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compound was reported preciously. The reaction of 7 with diethyl
malonate, ethyl cyanoacetate, and the appropriate βꢀketoesters 8, which
were either commercially available or prepared by the procedure
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described in the literature, in DMF with the presence of anhydrous
potassium carbonate at 100℃ to obtain substituted pyrimidinꢀ4(3H)ꢀones.
In addition, compound 9 was acetylated using acetic anhydride to yield
compound 11, and halogenation of compounds 13 and 15 resulted in
compounds 21ꢀ26.
The effect of the different substituent at the 2’ꢀposition of the phenyl
ring of pyrimidinꢀ4(3H)ꢀones was investigated by synthesizing
compounds 36ꢀ39 using an alternative route outlined in scheme 3. The
intermediates, compounds 34, prepared in the manner described in
Scheme 2, was halogenated, then treated with chlorosulfonic acid, and
finally followed by Nꢀmethyl piperazinethe, resulting in the compounds
36ꢀ39.
2
. RESULTS AND DISCUSSION
3.1 Structure determination of compounds 2 and 5 in complex with
the PDE5 catalytic domain
The crystal structures of compounds 2 and 5 in complex with the
catalytic domain of PDE5 have been determined by soaking the
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compounds into the apo PDE5 crystals (space group, P3 21), which
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diffracted to 2.4, and 2.28 angstroms, respectively (Table S1). The (F ꢀF )
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difference electronꢀdensity maps contoured at 3.0 σ for compounds 2 and
in two crystal structures were shown in Figure S1. Similar to the
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binding of sildenafil with PDE5, both compounds fitted into the substrate
binding pocket located in the centre of the Cꢀterminal helical bundle
domain of PDE5. However, as mentioned above, the orientation of the
two compounds within the binding pocket is different (Figures 1B and D)
and their interactions are different too. The program LIGPLOT was used
to plot the interactions between the compounds and PDE5 (Figures 2A
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and B). Compound 2 only formed the key bidentate Hꢀbonds with
residue Q817, while besides these Hꢀbonds compound 5 also interacts
with residue Q775 through a water molecule. Compound 5 formed
hydrophobic interactions with multiple residues, Y612, L765, V782,
A783, F786, M816, and F820 (Figure 2B). The pyrimidinone ring of 5 is
located just between the hydrophobic clamp composed of V782 and F820,
and has perfect πꢀπ stacking interactions with the phenyl ring of F820
(
Figure S2). Two ethyl groups on the pyrimidinone ring simultaneously
occupy two subꢀpockets SP1 (created primarily by residues L765, I768,
and F820) and SP2 (created primarily by residues Y612, H613, V782,
and F786) (Figure 2D). However, in the complex of PDE5/2, as discussed
above, the hydrophobic cyclopentene ring of the compound has to bind to
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the subꢀpocket SP1 in order to avoid close contact with the crucial water
molecule which forms Hꢀbonds with two residues and another water
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molecule coordinating to Zn at the active site of the enzyme (Figures
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B, 1C, 1D, and 2D). The interaction at SP1 results in a shift of the whole
compound and thereby breaks the perfect πꢀπ stacking interactions
between the pyrimidinone ring and F820 seen in both complexes of
PDE5/5 and PDE5/sildenafil. Consequently, hydrophobic interactions
were only formed between compound 2 and four residues L725, V782,
F786, and F820 (Figure 2A). Compound 2 formed less hydrophobic and
Hꢀbond interactions with PDE5, which leads to its much lower potency
(only 24% inhibition of PDE5 at 1 ꢁM concentration ) than compound 5
(
IC , 1.6 nM). Therefore, the structural analysis of these two compounds
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together with other inhibitors including sildenafil binding with PDE5
provide new insights into the substrate binding pocket of the catalytic
domain. Consideration of the critical water molecule and the two distinct
subꢀpockets provides a rationale for designing more potent PDE5
inhibitors with new scaffold such as the monocyclic pyrimidinone.
3
.2 SAR of substitution at the 6-position of the 4(3H)-pyrimidinone
ring.
The initial investigation on the structureꢀactivity relationship (SAR) of
the pyrimidinꢀ4(3H)ꢀone derivatives were carried out by substituting the
6ꢀposition of the pyrimidinone ring and fixing the
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ꢀ[(4ꢀmethylꢀ1ꢀpiperazinyl)sulfonyl]ꢀ2ꢀpropoxyphenyl ring. The
inhibition activities of these substituted compounds to PDE5 were
presented in Table 1. Compounds 9 and 10 with an amino and a hydroxyl
group, respectively, at the 6ꢀposition of the 4(3H)ꢀpyrimidinone ring
showed little inhibition of PDE5, while compound 11 bearing an
acetamido group had a great increase in potency, with an IC of 450 nM,
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5
0
suggesting that a hydrophilic substitution at this position is unfavorable to
the interaction of inhibitors with the residues in the substrate binding
pocket. After the hydrophilic group was replaced by other hydrophobic
groups, the resulted compounds 12-16 displayed more potent inhibition
activity to PDE5 than those of 2, 9, and 10, which is consistent with our
structural understanding mentioned above that the substitute on the
6
ꢀposition of the pyrimidinone ring tends to form hydrophobic
interactions with the enzyme and it indeed binds into the SP2 subꢀpocket
which has been seen in the PDE5/5 complex structure. Compounds 13
and 15 showed higher PDE5 inhibitory activity than 12, 14, and 16,
indicating that the length of alkyl maybe influence the interaction with
the subꢀpocket and the 2ꢀcarbon substituent such as an ethyl group or
iꢀpropyl group may be suitable for the pocket. Furthermore, replacing the
alkyl group with a phenyl group resulted in a decrease in potency
(compound 17), which also implies that the subꢀpocket is more favorable
in holding an alkyl substituent than an aromatic ring. In the case of
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compound 18, a significant loss of potency in comparison with that of 12
might be ascribed to the partial loss of the stacking interaction of the
phenyl ring of F820 with the pyrimidinone moiety because of the lack of
electron in the pyrimidinone ring caused by the substitution of the
strongly electronꢀwithdrawing trifluoromethyl group in this compound.
0
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3
.3 SAR of substitution at 5-position of the 4(3H)-pyrimidinone ring
On the basis of the above results, we decided to retain the 2ꢀcarbon
chain, the ethyl or iꢀpropyl group, at the 6ꢀposition while surveying
various 5ꢀsubstituted pyrimidinꢀ4(3H)ꢀones, and the results are
summarized in Table 2. The introduction of an acetylamino group at the
5ꢀposition of pyrimidinꢀ4(3H)ꢀone drastically decreased the potency in
comparison with that of 13. It was quickly discovered that introducing a
halogen atom except for fluorin at the 5ꢀposition led to a remarkable
increase in potency comparable to or even higher than that of 13. Among
them, the iodo derivative, compound 23, has the highest IC of 7.2 nM.
5
0
Most significantly, we find that compound 5 with an ethyl group at both
the 6ꢀ and 5ꢀposition of the pyrimidinꢀ4(3H)ꢀone ring exhibited better
PDE5 inhibitory activity than sildenafil and showed the highest IC (1.6
50
nM) among all the compounds reported in the present study. Compounds
and 27 with a shorter and longer alkyl group than 2ꢀcarbon one,
3
respectively, both had less inhibitory activity to PDE5 than that of 5,
indicating that the subꢀpocket of the enzyme to accommodate the
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substituted group at the 5ꢀposition of the pyrimidinꢀ4(3H)ꢀone ring is also
favorable in interaction with an 2ꢀcarbon alkyl group, which is verified by
the crystal structure of PDE5/5 complex. Although the introduction of an
iꢀpropyl group at the 6ꢀposition of the pyrimidinone ring in compound 15
resulted in better inhibitor activity than that of 13 with an ethyl group at
the same position, compound 29 with an iꢀpropyl group at the 6ꢀposition
of the pyrimidinone ring showed a potency 2ꢀfold less than 5.
10
11
12
13
14
15
16
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18
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Subsequently, the ethyl group was selected for the 5ꢀposition substituent
and the methyl and nꢀpropyl groups were individually chosen for the
substituent at the 6ꢀposition. The compounds 29, 30, and 31 had ~2ꢀfold,
8ꢀfold, and 3ꢀfold, respectively, loss in potency in comparison with that of
5
. Therefore, the SAR results presented above indicate that the substituent
at both the 5ꢀ and the 6ꢀposition of the pyrimidinꢀ4(3H)ꢀone moiety are
pivotal for determining the inhibition potency of the compounds to PDE5
and consequently, the 5, 6ꢀdiethylꢀpyrimidinꢀ4(3H)ꢀone showed the
strongest inhibition potency among the series of pyrimidinꢀ4(3H)ꢀone
analogues.
Using compounds 13 (H), 20 (F), 21(Cl), 22 (Br), and 23 (I), we have
previously reported that the halogen bond formed between the halogen
atom of the compound and the hydroxyl oxygen atom of Y612 in the
active site of PDE5 plays important role in binding of the inhibitors with
23
PDE5. The order of halogenꢀbinding contribution to IC of these
5
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compounds to PDE5 is 23 (I)> 22 (Br)> 21 (Cl)> 20 (F). Here, the
chemical structure of compound 5 is similar to those of the four
compounds except the substitution at 5ꢀposition of the
0
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4
(3H)ꢀpyrimidinone ring is an ethyl group, instead of the halogen atoms.
The IC of compound 5 is even higher than that of compound 23. It is
50
suggested that the hydrophobic interactions of the ethyl group of 5 with
the subꢀpocket SP2, mainly composed of the residues F786, V782, Y612,
and H613, compensate the missed halogen bonding interactions in
PDE5/5 complex, contributing greatly to the binding affinity of
compound 5 with PDE5. It is thereby concluded that the optimization of
the substitutions at the 5ꢀposition of the 4(3H)ꢀpyrimidinone is important
for the potency of the pyrimidinꢀ4(3H)ꢀones.
3.4 SAR of substitution at the 2’-position of the phenyl ring
In parallel to establishing SAR at the 6ꢀ and 5ꢀposition substitutions of
the pyrimidinone moiety, we devoted a part of effort to evaluating the
effect of the oꢀalkyl sideꢀchain at the 2’ꢀposition of the phenyl ring on its
hydrophobic interaction with the enzyme. As shown in Table 3, it is clear
that a 3ꢀcarbon chain in compounds 24 and 25 resulted in high inhibition
potency to PDE5. Therefore, the nꢀpropoxyl group is selected for the
substitution at the 2’ꢀposition of the phenyl ring of the inhibitors, which
contributes the proper hydrophobic interaction with PDE5.
3
.5 Selectivity and in vivo study of compound 5
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The inhibitions of compound 5 versus 11 PDEs were measured by
using sildenafil as a control (Table 4). The inhibition of the compound to
PDE2, PDE3, PDE4, PDE7A1, PDE8A1, PDE9A2, PDE10A2 is very
weak (IC > 10000 nM). The IC of 5 to PDE11A4, PDE1, and PDE6C
10
11
12
13
14
15
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17
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22
23
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5
0
50
are 2127ꢀ, 469ꢀ, and 29ꢀfold higher than the IC of this compound to
50
PDE5A, respectively. The selectivity profile of sildenafil to the 11 PDEs
is similar to that of compound 5.
With excellent enzyme inhibition of PDE5 and also good selectivity
over other PDEs in vitro, compound 5 was selected for further testing in
vivo. The pharmacokinetic data of compound 5 is summarized in Table 5
and the profile generated based on these data is shown in Figure 3. After
oral administration of a 10 mg/kg dose of 5 to male rats, a Cmax of 468
ng/ml was obtained at 20 min, and the oral bioavailability of the
26
compound was 10.0%, lower than that of sildenafil (23%) .
In vivo efficacy of compound 5 was evaluated in the rat model of
erection (Figure 4). Intracavernous pressure (ICP) and arterial blood
pressure (BP) were simultaneously monitored during electric stimulation
before and after the oral administration of compound 5. At a dose of 10
mg/kg, compound 5 significantly increased the ICP/BP value at 30 min,
compared to vehicle. It showed a good efficacy as sildenafil did.
3. CONCLUSIONS
The structural analysis based on crystal structures of the PDE5
1
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catalytic domain in complex with our compounds as well as other
wellꢀknown PDE5 inhibitors revealed new characteristics of the inhibitor
binding pocket. This includes the two separated subꢀpockets and a crucial
water molecule, which enable us to design the monocyclic
0
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0
pyrimidinꢀ4(3H)ꢀone as a new scaffold to synthesize the novel, potent,
and selective PDE5 inhibitors. Systematical exploration of SAR of the
substitutions on the pyrimidineꢀ4(3H)ꢀone moiety resulted in the
identification of 25 compounds with IC ranging from 1 to100 nM and
5
0
11 compounds with IC ranging from 1 to10 nM. In particular, the
5
0
compound 5 has the best IC , with a value of 1.6 nM, among all these
5
0
compounds. The crystal structures of PDE5/5, PDE5/2, and
PDE5/sildenafil complexes confirmed our rationale to design simpler
monocyclic pyrimidinꢀ4(3H)ꢀones working as potent inhibitors of PDE5.
The structures also revealed that simultaneously occupying two
subꢀpockets by two ethyl groups of monocyclic pyrimidinꢀ4(3H)ꢀones
compensated the Hꢀbond formed with the crucial water molecule which
has been seen in the PDE5/sildenafil complex but missed in
pyrimidinꢀ4(3H)ꢀones in complex with PDE5. Moreover, in vivo efficacy
studies have revealed that compound 5 exhibited a significant effect on
intracavernosal pressure in rats. Further investigation to improve the
druggability of this compound through structural modification to seek a
potential candidate for clinical study is ongoing.
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4
. EXPERIMENTAL SECTION
5
.1 Chemistry
1
HꢀNMR spectra was determined using a Mercury 300 MHz, FT NMR
spectrometer. HRMS were performed on a Finnigan MAT95 mass
spectrometer. Reaction solvents were purchased and used without further
purification. HPLC conditions were as follows: column, YMCꢀPack CN 5
ꢁM, 4.6 mm × 250 mm; solvent system, (A) MeCN; (B) 0.02 M
KH PO (pH 6.0); step gradient, time 0, 30% A; time 20 min, 70% A;
10
11
12
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2
4
stop time, 25 min; flow rate 1.0 mL/min; UV detection, 220 nm; injection
volume, 5 ꢁL; temperature, 30℃. The purity of all target compounds
was >95% as confirmed by HPLC.
General Procedure for the Synthesis of 2, 3, 5, 9, 10, 12-20 and 27-31
2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]cyclopenta
pyrimid-4(3H)-one (2) A mixture of
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide (340 mg, 1
mmol), ethyl 2ꢀoxoꢀcyclopentaneꢀ1ꢀcarboxylate (312 mg, 2 mmol), and
potassium carbonate (276 mg, 2 mmol) in N,NꢀDimethylformamide (5
mL) was stirred at 100 ℃ for 4 h. After cooled to room temperature, the
reaction mixture was poured into ice water, and extracted with
dichloromethane (3×10 ml). The organic layer was washed with saturated
brine, dried over anhydrous sodium sulfate, and concentrated. The residue
was purified by flash column chromatography. After crystallization from
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ethyl acetate, compound 2 was obtained as a white solid (84 mg, yield:
1
9%).
1
HꢀNMR (CDCl ) δ: 10.96 (1H, br), 8.84 (1H, d), 7.82 (1H, dd), 7.15 (1H,
3
0
1
2
3
4
5
6
7
8
9
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2
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5
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d), 4.25 (2H, t), 3.09 (4H, br), 2.93 (2H, t), 2.85 (2H, t), 2.51 (4H, br),
+
2
.28 (3H, s), 2.16ꢀ1.98 (4H, m), 1.15 (3H,t). HRMS (ESI) calcd [M + H]
for C H N O S 433.1910, found 433.1916.
21
29
4
4
5-methyl-6-ethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimid-4(3H)-one (3) Compound 3 was prepared in 63% yield
from ethyl 2ꢀmethylpropionylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
11.02 (1H, br), 8.78 (1H, d), 7.80 (1H, dd), 7.14 (1H, d), 4.22 (2H, t),
3.10 (4H, br), 2.65 (2H, q), 2.54 (4H, br), 2.30 (3H, s), 2.09 (3H, s), 1.99
+
(
2H, m), 1.23 (3H, t), 1.12 (3H, t). HRMS (ESI) calcd [M + H] for
C H N O S 435.2066, found 435.2066.
2
1
31
4
4
5
,6-diethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]
pyrimid-4(3H)-one (5) Compound 5 was prepared in 65% yield from
ethyl 2ꢀethylpropionylacetate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR
(DMSOꢀd ) δ: 12.06 (1H, br), 7.88 (1H, d), 7.82 (1H, dd), 7.38 (1H, d),
6
4
.12 (2H, t), 2.88 (4H, br), 2.57 (2H, q), 2.46 (2H, q), 2.36 (4H, br), 2.14
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(
3H, s), 1.75 (2H, m), 1.17 (3H, t), 1.04 (3H, t), 0.96 (3H, t). HRMS (ESI)
+
calcd [M + H] for C H N O S 449.2223, found 449.2227.
22
33
4
4
6
-amino-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]py
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
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35
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rimid-4(3H)-one (9) Compound 9 was prepared in 38% yield from ethyl
cyanoacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR
(
DMSOꢀd ) δ: 11.25 (1H, br), 7.90 (1H, d), 7.81 (1H, dd), 7.38 (1H, d),
6
6.56 (2H, br), 5.00 (1H, s), 4.12 (2H, t), 2.87 (4H, br), 2.36 (4H, br), 2.14
+
(
3H, s), 1.75 (2H, m), 0.97 (3H, t). HRMS (ESI) calcd [M + H] for
C H N O S 408.1706, found 408.1709.
1
8
26
5
4
6
-hydroxy-2-[2-n-propoxy-5-(4-methyl-1-piperazinytsulfonyl)phenyl]
pyrimid-4(3H)-one (10) Compound 10 was prepared in 41% yield from
diethyl malonate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR
(DMSOꢀd ) δ: 8.09 (1H, d), 7.76 (1H, dd), 7.36 (1H, d), 4.52 (1H, s), 4.15
6
(
2H, t), 2.87 (4H, br), 2.36 (4H, br), 2.13 (3H, s), 1.79 (2H, m), 1.01 (3H,
+
t). HRMS (ESI) calcd [M + H] for C H N O S 409.1546, found
1
8
25
4
5
4
09.1549.
6-methyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]p
yrimid-4(3H)-one (12) Compound 12 was prepared in 62% yield from
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2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
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4
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5
5
5
5
5
5
6
ethyl acetoacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
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0
11.03 (1H, br), 8.83 (1H, d), 7.85(1H, dd), 7.16 (1H, d), 6.23 (1H, s), 4.25
(
2H, t), 3.07 (4H, br), 2.48 (4H, br), 2.35 (3H, s), 2.26 (3H, s), 2.03 (2H,
+
m), 1.14 (3H, t). HRMS (ESI) calcd [M + H] for C H N O S 407.1753,
1
9
27
4
4
found 407.1744.
-ethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]pyr
6
imid-4(3H)-one (13) Compound 13 was prepared in 60% yield from
ethyl propionylacetate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR
(DMSOꢀd ) δ: 7.89 (1H, d), 7.84 (1H, dd), 7.39 (1H, d), 4.15 (2H, t), 2.89
6
(
4H, br), 2.45 (2H, q), 2.36 (4H, br), 2.16 (3H, s), 1.75 (2H, m), 1.13 (3H,
+
t), 0.96 (3H, t). HRMS (ESI) calcd [M + Na] for C H N O NaS
20
29
4
4
4
43.1729, found 443.1736.
6-n-propyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]
pyrimid-4(3H)-one (14) Compound 14 was prepared in 58% yield from
ethyl butyrylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
11.01 (1H, br), 8.85 (1H, d), 7.85 (1H, dd), 7.15 (1H, d), 6.22 (1H, d),
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.25 (2H, t), 3.07 (4H, br), 2.56 (2H, t), 2.48 (4H, br), 2.26 (3H, s), 2.03
(
2H, m), 1.74 (2H, m), 1.14 (3H, t), 0.98 (3H, t). HRMS (ESI) calcd [M +
+
Na] for C H N O SNa 457.1885, found 457.1898.
21 30
4
4
10
11
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58
59
60
6
-isopropyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl
]
pyrimid-4(3H)-one (15) Compound 15 was prepared in 55% yield from
ethyl isobutyrylacetate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
10.99 (1H, br), 8.87 (1H, d), 7.86 (1H, dd), 7.15 (1H, d), 6.24 (1H, s),
4
.25 (2H, t), 3.08 (4H, br), 2.83 (1H, m), 2.49 (4H, br), 2.26 (3H, s), 2.03
+
(2H, m), 1.26 (6H, d), 1.15 (3H, t). HRMS (ESI) calcd [M + H] for
C H N O S 435.2066, found 435.2069.
2
1
31
4
4
6-isobutyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]
pyrimid-4(3H)-one (16) Compound 16 was prepared in 65% yield from
ethyl isovalerylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
11.02 (1H, br), 8.86 (1H, d), 7.86 (1H, dd), 7.15 (1H, d), 6.19 (1H, s),
4.25 (2H, t), 3.07 (4H, br), 2.48 (4H, br), 2.44 (2H, d), 2.26 (3H, s), 2.13
(
1H, m), 2.01 (2H, m), 1.15 (3H, t), 0.96 (6H, d). HRMS (ESI) calcd [M
+
+ H] for C H N O S 449.2223, found 449.2210.
2
2
33
4
4
6
-phenyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phenyl]p
2
1
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9
1
1
1
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2
2
2
2
2
2
2
2
2
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5
6
yrimid-4(3H)-one (17) Compound 17 was prepared in 67% yield from
ethyl benzoylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
similar procedure to that described for synthesis of 2. HꢀNMR
(
DMSOꢀd ) δ: 12.44 (1H, br), 8.10ꢀ8.05 (2H, m), 8.01 (1H, d), 7.87 (1H,
6
dd), 7.53ꢀ7.47 (3H, m), 7.43 (1H, d), 6.91 (1H, s), 4.15 (2H, t), 2.92 (4H,
br), 2.37 (4H, br), 2.14 (3H, s), 1.76 (2H, m), 0.95 (3H, t). HRMS (ESI)
+
calcd [M + H] for C H N O S 469.1910, found 469.1916.
24
28
4
4
6-trifluoromethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)
phenyl]pyrimid-4(3H)-one (18) Compound 18 was prepared in 37%
yield from ethyl 4,4,4ꢀtrifluoroacetoacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
8
.84 (1H, d), 7.92 (1H, dd), 7.19 (1H, d), 6.75 (1H, s), 4.29 (2H, t), 3.08
(4H, br), 2.49 (4H, br), 2.27 (3H, s), 2.04 (2H, m), 1.17 (3H, t). HRMS
+
(
ESI) calcd [M + H] for C H F N O S 461.1470, found 461.1470.
19
24
3
4
4
5-acetamido-6-ethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfony
l)phenyl]pyrimid-4(3H)-one (19) Compound 19 was prepared in 42%
yield from ethyl 2ꢀacetamidopropionylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR
(
DMSOꢀd ) δ: 12.48 (1H, br), 9.22 (1H, br), 7.88 (1H, d), 7.84 (1H, dd),
6
2
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.39 (1H, d), 4.13 (2H, t), 2.91 (4H, br), 2.43 (2H, q), 2.40 (4H, br), 2.16
(
3H, s), 2.02 (3H, s), 1.76 (2H, m), 1.13 (3H, t), 0.96 (3H, t). HRMS (ESI)
+
calcd [M + H] for C H N O S 478.2124, found 478.2126.
22 32
5
5
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
60
5
-fluoro-6-ethyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phe
nyl]pyrimidin-4(3H)-one (20) Compound 20 was prepared in 72% yield
from methyl 2ꢀfluoroꢀ3ꢀoxopentanoate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following
following a similar procedure to that described for synthesis of 2.
1
HꢀNMR (CDCl ) δ: 8.80 (1H, d), 7.86 (1H, dd), 7.16 (1H, d), 4.26 (2H,
3
t), 3.09 (4H, br), 2.74 (2H, q), 2.51 (4H, br), 2.28 (3H, s), 2.03 (2H, m),
+
1.28 (3H, t), 1.15 (3H, t). HRMS (ESI) calcd [M + Na] for
C H N O NaSF 461.1635, found 461.1630.
2
0
27
4
4
5-propyl-6-ethyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimid-4(3H)-one (27) Compound 27 was prepared in 39% yield
from ethyl 2ꢀpropylpropionylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
1
0.97 (1H, br), 8.84 (1H, d), 7.82 (1H, dd), 7.13 (1H, d), 4.23 (2H, t),
3.07 (4H, br), 2.65 (2H, q), 2.52 (2H, t), 2.49 (4H, br), 2.26 (3H, s), 2.02
(
2H, m), 1.55 (2H, m), 1.26 (3H, t), 1.15 (3H, t), 0.99 (3H, t). HRMS
+
(ESI) calcd [M + H] for C H N O S 463.2379, found 463.2379.
23
35
4
4
5
-methyl-6-isopropyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfon
2
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9
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2
2
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5
5
5
5
5
5
6
yl)phenyl]pyrimid-4(3H)-one (28) Compound 28 was prepared in 54%
yield from ethyl 2ꢀmethylisobutyrylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
1
3
0.93 (1H, br), 8.86 (1H, d), 7.83 (1H, dd), 7.13 (1H, d), 4.24 (2H, t),
.16 (1H, m), 3.08 (4H, br), 2.48 (4H, br), 2.26 (3H, s), 2.12 (3H, s), 2.02
+
(2H, m), 1.22 (6H, d), 1.15 (3H, t). HRMS (ESI) calcd [M + H] for
C H N O S 449.2223, found 449.2227.
2
2
33
4
4
5-ethyl-6-isopropyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl
phenyl]pyrimid-4(3H)-one (29) Compound 29 was prepared in 41%
yield from ethyl 2ꢀethylisobutyrylacetate and
)
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
1
0.95 (1H, br), 8.85 (1H, d), 7.82 (1H, dd), 7.13 (1H, d), 4.23 (2H, t),
3.15 (1H, m), 3.07 (4H, br), 2.58 (2H, q), 2.47 (4H, br), 2.25 (3H, s), 2.02
(
2H, m), 1.23 (6H, d), 1.14 (3H, t), 1.12 (3H, t). HRMS (ESI) calcd [M +
+
H] for C H N O S 463.2379, found 463.2379.
2
3
35
4
4
5
-ethyl-6-methyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimid-4(3H)-one (30) Compound 30 was prepared in 42% yield
from ethyl 2ꢀethylacetoacetate and
5ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
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1
1.00 (1H, br), 8.77 (1H, d), 7.80 (1H, dd), 7.12 (1H, d), 4.22 (2H, t),
3
.05 (4H, br), 2.55 (2H, q), 2.48(4H, br), 2.37 (3H, s), 2.25 (3H, s), 1.98
+
(
2H, m), 1.13 (3H, t), 1.11 (3H, t). HRMS (ESI) calcd [M + H] for
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
60
C H N O S 435.2066, found 435.2068.
2
1
31
4
4
5-ethyl-6-propyl-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimid-4(3H)-one (31) Compound 31 was prepared in 38% yield
from ethyl 2ꢀethylbutyrylacetate and
5
ꢀ(4ꢀmethylpiperazinꢀ1ꢀylsulfonyl)ꢀ2ꢀpropoxybenzimidamide following a
1
similar procedure to that described for synthesis of 2. HꢀNMR (CDCl ) δ:
3
1
0.95 (1H, br), 8.82 (1H, d), 7.82 (1H, dd), 7.13 (1H, d), 4.24 (2H, t),
3.07 (4H, br), 2.63ꢀ2.55 (4H, m), 2.49 (4H, br), 2.27 (3H, s), 2.03 (2H, m),
1
.74 (2H, m), 1.15 (3H, t), 1.14 (3H, t), 1.00 (3H, t). HRMS (ESI) calcd
+
[M + H] for C H N O S 463.2379, found 463.2379.
23
35
4
4
6
-Acetamido-2-[2-n-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phen
yl]pyrimid-4(3H)-one (11) The compound 9 (0.20 g, 0.5 mmol) was
suspended in acetic anhydride (5 ml), and the mixture was stirred at 100
˚C for 1 h. The cooled reaction mixture was poured into ice water to
generate a white solid. The solid was collected by filtration, washed with
clear water (3×10 ml), and dried at 60˚C to give compound 11 (0.12 g,
1
yield: 54%). HꢀNMR (DMSOꢀd ) δ: 12.16 (1H, br), 10.54 (1H, br), 7.92
6
(1H, d), 7.84 (1H, dd), 7.40 (1H, d), 6.89 (1H, s), 4.12 (2H, t), 2.88 (4H,
br), 2.36 (4H, br), 2.14 (3H, s), 2.08 (3H, s), 1.74 (2H, m), 0.95 (3H, t).
2
5
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Page 26 of 63
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9
1
1
1
1
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2
2
2
2
2
2
3
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3
3
3
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4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
HRMS (ESI) calcd [M + H] for C H N O S 450.1811, found 450.1811.
20 28
5
5
General Procedure for the Synthesis of 21-26
-Chloro-6-ethyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phe
5
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
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7
8
9
0
1
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4
5
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7
8
9
0
nyl]pyrimidin-4(3H)-one (21) Chlorine gas was bubbled into an iceꢀcold
solution of 13 (100 mg, 0.24 mmol) and pyridine (40 ꢁL, 0.5 mmol) in
dichloromethane (10 mL) for 1 min. The resulting mixture was washed
with 1N Na S O (aq, 5 mL) and water (5 mL). The organic layer was
2
2
3
dried over anhydrous sodium sulfate and concentrated in vacuo. After
crystallization from ethyl acetate, compound 21 was obtained as white
1
crystals (93 mg, yield 85%). HꢀNMR (CDCl ) δ: 11.25 (1H, br), 8.85
3
(1H, d), 7.87 (1H, dd), 7.16 (1H, d), 4.27 (2H, t), 3.08 (4H, br), 2.85 (2H,
q), 2.49 (4H, br), 2.27 (3H, s), 2.03 (2H, m), 1.29 (3H, t), 1.15 (3H, t).
HRMS (ESI) calcd [M + Na]+ for C H N O NaSCl 477.1339, found
2
0
27
4
4
4
77.1339.
5-Bromo-6-ethyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl)phe
nyl]pyrimidin-4(3H)-one (22) Bromine (40 mg, 0.27 mmol) was added
to an iceꢀcold solution of 13 (100 mg, 0.24 mmol) and pyridine (22 ꢁL,
0
.27 mmol) in dichloromethane (10 mL). The mixture was stirred for 15
min at 0˚C and washed with 1 N Na S O (aq, 5 mL) and water (5 mL).
2
2
3
The organic layer was dried over anhydrous sodium sulfate and
concentrated. After crystallization from ethyl acetate, compound 22 was
1
obtained as a white solid (107 mg, yield 90%). HꢀNMR (CDCl ) δ: 11.12
3
2
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9
(
1H, br), 8.86 (1H, d), 7.88 (1H, dd), 7.16 (1H, d), 4.27 (2H, t), 3.09 (4H,
br), 2.88 (2H, q), 2.50 (4H, br), 2.28 (3H, s), 2.03 (2H, m), 1.29 (3H, t),
+
1
5
.15 (3H, t). HRMS (ESI) calcd [M + Na] for C H N O NaSBr
20 27
4
4
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
60
21.0834, found 521.0834.
5-Iodo-6-ethyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl)pheny
l]pyrimidin-4(3H)-one (23) Iodine (254 mg, 1 mmol) was added to a
solution of 13 (420 mg, 1 mmol) and silver nitrate (170 mg, 1 mmol) in
methanol (10 mL) at 0˚C. The mixture was stirred at room temperature
for 30 min. After filtration, the filtrate was poured into water (25 mL) and
extracted with dichloromethane (20 mL). The organic layer was washed
with 1N Na S O (aq, 5 mL) and water (5 mL), dried, and concentrated.
2
2
3
After crystallization from ethyl acetate, 23 was obtained as a white solid
1
(410 mg, 75%). HꢀNMR (CDCl ) δ: 11.12 (1H, br), 8.88 (1H, d), 7.88
3
(
1H, dd), 7.16 (1H, d), 4.27 (2H, t), 3.10 (4H, br), 2.92 (2H, q), 2.52 (4H,
br), 2.30 (3H, s), 2.03 (2H, m), 1.28 (3H, t), 1.16 (3H, t). HRMS (ESI)
+
calcd [M + Na] for C H N O NaSI 569.0695, found 569.0704.
2
0
27
4
4
5-Chloro-6-isopropyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl
phenyl]pyrimidin-4(3H)-one (24) Compound 24 was prepared in 72%
)
yield from 15 following a similar procedure to that described for
1
synthesis of 21. HꢀNMR (CDCl ) δ: 11.20 (1H, br), 8.87 (1H, d), 7.89
3
(1H, dd), 7.17 (1H, d), 4.28 (2H, t), 3.50 (1H, m), 3.10 (4H, br), 2.51 (4H,
br), 2.28 (3H, s), 2.05 (2H, m), 1.27 (6H, d), 1.16 (3H, t). HRMS (ESI)
2
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9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
calcd [M + H] for C H N O SCl 469.1676, found 469.1676.
21 30
4
4
5
-Bromo-6-isopropyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl
)
phenyl]pyrimidin-4(3H)-one (25) Compound 25 was prepared in 68%
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
yield from 15 following a similar procedure to that described for
1
synthesis of 22. HꢀNMR (CDCl ) δ: 11.16 (1H, br), 8.86 (1H, d), 7.89
3
(
1H, dd), 7.16 (1H, d), 4.28 (2H, t), 3.50 (1H, m), 3.09 (4H, br), 2.49 (4H,
br), 2.27 (3H, s), 2.03 (2H, m), 1.26 (6H, d), 1.15 (3H, t). HRMS (ESI)
+
calcd [M+H] for C H N O SBr 513.1711, found 513.1715.
21
30
4
4
5-Iodo-6-isopropyl-2-[2-propoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimidin-4(3H)-one (26) Compound 26 was prepared in 62%
yield from 15 following a similar procedure to that described for synthsis
1
of 23. HꢀNMR (CDCl ) δ: 8.90 (1H, d), 7.89 (1H, dd), 7.16 (1H, d), 4.27
3
(2H, t), 3.46 (1H, m), 3.09 (4H, br), 2.49 (4H, br), 2.27 (3H, s), 2.04 (2H,
+
m), 1.25 (6H, d), 1.16 (3H, t).. HRMS (ESI) calcd [M + H] for
C H N O SI 561.1032, found 561.1035.
2
1
30
4
4
5
-Bromo-6-isopropyl-2-[2-ethoxy-5-(4-methyl-1-piperazinylsulfonyl)p
henyl]pyrimidin-4(3H)-one (36) A mixture of 2ꢀethoxybenzonitrile
20.5 g, 0.14 mol), potassium carbonate (38.5 g, 0.28 mol), and
(
hydroxylamine hydrochloride (19.2 g, 0.28 mol) in a mixed solution of
ethanol (100 mL) and water (100 mL) was refluxed for 10 h and the
ethanol was removed by vacuum distillation. After the residue was cooled,
white solid 32a (18.1 g, 72%) was separated out and collected by
2
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filtration.
The solid 32a (18 g, 0.5 mol) was solved in acetic acid (200 mL) and
degassed. A catalytic amount of 10% Pd/C was added, and the solution
was placed on the shaker under 3 MPa hydrogen at 65℃ for 8h. After the
solvent was removed by vacuum distillation, the residue was 33a acetate
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
60
(
13.2 g, 80%) which was not purified.
A mixture of 33a acetate (9.4g, 42mmol), ethyl isobutyrylacetate (7.3 g,
6 mmol), and potassium carbonate (11.6 g, 84 mmol) in
4
N,NꢀDimethylformamide (80 mL) was stirred at 100℃ for 4 h. The
mixture was then cooled to room temperature and taken up in water (400
mL), and the solid was collected by filtration and purified by
recrystallization from ethyl acetic to yield 34a (7.7 g, 71%) as white solid.
1
HꢀNMR (CDCl ) δ: 11.22 (1H, br), 8.52 (1H, dd), 7.48 (1H, t), 7.12 (1H,
3
t), 7.03 (1H, d), 6.20 (1H, s), 4.29 (2H, q), 2.82 (1H, m), 1.59 (3H, t),
1.27 (6H, d).
Bromine (1.9g, 12mmol) was added to an iceꢀcold solution of 34a (3.2 g,
12 mmol) and pyridine (1 mL) in dichloromethane (150 mL). The
mixture was stirred for 10min at 0℃ and washed with 1 N Na S O (aq,
2
2
3
45 mL) and water (50 mL). The organic layer was dried over anhydrous
sodium sulfate and concentrated. After crystallization from
1
etherꢀacetonitrile, 35a (3.3 g, yield 82%) was obtained. HꢀNMR (CDCl )
1
3
δ: 8.53 (1H, d), 7.51 (1H, t), 7.13 (1H, t), 7.04 (1H, d), 4.32 (2H, q), 3.51
2
9
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