Synthesis and biological evaluation of 5-carbamoyl-2-phenylpyrimidine derivatives as novel and potent PDE4 inhibitors

Article abstract of DOI:10.1016/j.bmc.2013.09.013

5-Carbamoyl-2-phenylpyrimidine derivative 2 has been identified as a phosphodiesterase 4 (PDE4) inhibitor with moderate PDE4B inhibitory activity (IC₅₀ = 200 nM). Modification of the carboxylic acid moiety of 2 gave N-neopentylacetamide derivative 10f, which had high in vitro PDE4B inhibitory activity (IC₅₀ = 8.3 nM) and in vivo efficacy against lipopolysaccharide (LPS)-induced pulmonary neutrophilia in mice (ID₅₀ = 16 mg/kg, ip). Furthermore, based on the X-ray crystallography of 10f bound to the human PDE4B catalytic domain, we designed 7,8-dihydro-6H-pyrido[4,3-d] pyrimidin-5-one derivative 39 which has a fused bicyclic lactam scaffold. Compound 39 exhibited excellent inhibitory activity against LPS-induced tumor necrosis factor alpha (TNF-α) production in mouse splenocytes (IC ₅₀ = 0.21 nM) and in vivo anti-inflammatory activity against LPS-induced pulmonary neutrophilia in mice (41% inhibition at a dose of 1.0 mg/kg, i.t.).

Full text of DOI:10.1016/j.bmc.2013.09.013

Accepted Manuscript
Synthesis and biological evaluation of 5-carbamoyl-2-phenylpyrimidine deriv‐
atives as novel and potent PDE4 inhibitors
Taiji Goto, Akiko Shiina, Toshiharu Yoshino, Kiyoshi Mizukami, Kazuki
Hirahara, Osamu Suzuki, Yoshitaka Sogawa, Tomoko Takahashi, Tsuyoshi
Mikkaichi, Naoki Nakao, Mizuki Takahashi, Masashi Hasegawa, Shigeki
Sasaki
PII:
S0968-0896(13)00793-1
DOI:
http://dx.doi.org/10.1016/j.bmc.2013.09.013
BMC 11097
Reference:
To appear in:
Bioorganic & Medicinal Chemistry
Received Date:
Revised Date:
Accepted Date:
19 June 2013
31 July 2013
6 September 2013
Please cite this article as: Goto, T., Shiina, A., Yoshino, T., Mizukami, K., Hirahara, K., Suzuki, O., Sogawa, Y.,
Takahashi, T., Mikkaichi, T., Nakao, N., Takahashi, M., Hasegawa, M., Sasaki, S., Synthesis and biological
evaluation of 5-carbamoyl-2-phenylpyrimidine derivatives as novel and potent PDE4 inhibitors, Bioorganic &
Medicinal Chemistry (2013), doi: http://dx.doi.org/10.1016/j.bmc.2013.09.013
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Synthesis and biological evaluation of 5-carbamoyl-2-phenylpyrimidine derivatives
as novel and potent PDE4 inhibitors
a,d,
b
a
c
Taiji Goto *, Akiko Shiina , Toshiharu Yoshino , Kiyoshi Mizukami , Kazuki
a
a
a
a
Hirahara , Osamu Suzuki , Yoshitaka Sogawa , Tomoko Takahashi , Tsuyoshi
a
a
a
a
d
Mikkaichi , Naoki Nakao , Mizuki Takahashi , Masashi Hasegawa , Shigeki Sasaki
a
R&D Division, Daiichi Sankyo Co.,Ltd, 1-2-58, Hiromachi, Shinagawa-ku, Tokyo
1
40-8710, Japan
b
Quality & Safety Management Division, Daiichi Sankyo Co.,Ltd, 3-5-1,
Nihonbashihoncho, Chuo-ku, Tokyo 103-8426, Japan
c
Corporate Strategy Division, Daiichi Sankyo Co.,Ltd, 3-5-1, Nihonbashihoncho,
Chuo-ku, Tokyo 103-8426, Japan
d
Graduate School of Pharmaceutical Sciences, Kyushu University, 3-1-1 Maidashi,
Higashi-ku, Fukuoka 812-8582, Japan
*
Tel.: +81 334923131 E-mail address: goto.taiji.kt@daiichisankyo.co.jp
Abstract
-Carbamoyl-2-phenylpyrimidine derivative
phosphodiesterase 4 (PDE4) inhibitor with moderate PDE4B inhibitory activity (IC50
5
2
has been identified as
a
=
200 nM). Modification of the carboxylic acid moiety of 2 gave N-neopentylacetamide
derivative 10f, which had high in vitro PDE4B inhibitory activity (IC50 = 8.3 nM) and in
vivo efficacy against lipopolysaccharide (LPS) -induced pulmonary neutrophilia in mice

(
ID50 = 16 mg/kg, i.p.). Furthermore, based on the X-ray crystallography of 10f bound
to the human PDE4B catalytic domain, we designed
,8-dihydro-6H-pyrido[4,3-d]pyrimidin-5-one derivative 39 which has a fused bicyclic
7
lactam scaffold. Compound 39 exhibited excellent inhibitory activity against
LPS-induced tumor necrosis factor alpha (TNF-α) production in mouse splenocytes
(
IC50 = 0.21 nM) and in vivo anti-inflammatory activity against LPS-induced pulmonary
neutrophilia in mice (41% inhibition at a dose of 1.0 mg/kg, i.t.).
1. Introduction
1
Phosphodiesterase 4 (PDE4) is an enzyme responsible for the degradation of cyclic
adenosine monophosphate (cAMP) via hydrolysis to 5’-AMP. PDE4 is predominantly
expressed in most inflammatory and immune cells such as lymphocytes, monocytes,
1b,2
macrophages, neutrophils, eosinophils, and mast cells.
In the inflammatory cells,
PDE4 inhibition causes an elevation of the intracellular level of cAMP which plays a
3
critical role as a negative regulator of the inflammatory responses. Therefore, PDE4
has been established as a promising target for the treatment of various inflammatory
4
diseases such as asthma, chronic obstructive pulmonary disease (COPD), and so forth.
We have previously reported that the 2-phenylpyrimidine derivative 1 had high in
vitro PDE4B inhibitory activity (IC50 = 25 nM) and moderate inhibitory activity against
lipopolysaccharide (LPS) -induced tumor necrosis factor alpha (TNF-α) production in
5
mouse splenocytes (IC50 = 390 nM). (Figure 1) In the course of further exploration of
this series, we designed and synthesized a 5-carbamoyl-2-phenylpyrimidine derivative 2
as a new lead compound, because the oxygen atom of the primary amide group was

expected to form hydrogen bonds with the surface of the enzyme in a similar manner to
the sulfone oxygen atoms of compound 1. As predicted, compound 2 had PDE4B
inhibitory activity with an IC50 value of 200 nM and inhibited LPS-induced TNF-α
production in mouse splenocytes (IC50 = 690 nM), leading to further derivatization of 2.
Herein, we report the syntheses, structure-activity relationships (SAR), and X-ray
crystal structure analysis of the series of 5-carbamoyl-2-phenylpyrimidine derivatives.
2. Chemistry
The 5-carbamoyl-2-phenylpyrimidine derivatives were synthesized as shown in
Schemes 1-5.
In Scheme 1, the syntheses of compounds 2 and 9b-d are depicted. Starting material 3
and benzamidine were condensed to form pyrimidine 4. The 4-hydroxyl group of 4 was
converted to a chlorine atom by treatment with phosphoryl chloride to produce 5.
Hydrolysis of the ethyl ester, followed by condensation with several amines afforded
amide compounds 7a-d. Substitution of the chlorine atom with ethyl
2
-(4-aminophenyl)acetate gave 8a-d, which were hydrolyzed to afford the target
compounds, 2 and 9b-d. Amide compounds (10a-h) were synthesized from 2 by a
condensation reaction with corresponding amines using
N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide (WSC), 1-hydroxybenzotriazole
hydrate (HOBt) and N,N-diisopropylethylamine (DIPEA).
The preparation of 6-methyl derivative 19 is outlined in Scheme 2. Pyrimidine ring
formation with diethyl ethylidenemalonate (11) and benzamidine was achieved in basic
condition to give 12. Bromination on 5-position of 12 with N-bromosuccinimide (NBS)

6
gave bromide 13. An elimination reaction of hydrogen bromide and a successive
chlorination reaction were performed by treatment with phosphoryl chloride to give 14.
Hydrolysis of 14 gave a 1:1 mixture of 15 and alkoxylated compound 16. As the
separation of 15 and 16 was difficult, byproducts derived from 16 were separated after
the synthesis of 19. The mixture of 15 and 16 was subjected to a series of reactions for
amide formation, a substitution reaction of the chlorine atom with ethyl
2-(4-aminophenyl)acetate, and hydrolysis of the ester to produce target compound 19
together with byproducts derived from 16. Finally, 19 was isolated by recrystallization.
Compound 24 was synthesized as shown in Scheme 3. Commercially available 20
7
reacted with benzamidine to give 21 , which was treated with phosphoryl chloride to
give chloride 22. Substitution of 4-chlorine atom with ethyl 2-(4-aminophenyl)acetate
gave 23. Hydrolysis of terminal ester and 5-cyano group afforded compound 24.
Compounds 29a-f and 30a-c were synthesized via intermediate 28 as outlined in
Scheme 4. Initial compound 25 was treated with phosphoryl pentachloride and
8
phosphoryl chloride to give chloride 26. The reaction of the acid chloride with
ammonia and selective substitution of the chlorine atom on the 4-position with
2
-(4-aminophenyl)-N-(2,2-dimethylpropyl)acetamide gave intermediate 28. Various
substituents were introduced at the 2-position by the Suzuki-Miyaura coupling reaction
or substitution reaction to afford the target compounds (29a-f, 30a-c).
The syntheses of the fused bicyclic lactam compounds (38, 39) are shown in Scheme
5
. Ethyl 3-aminopropanoate (31) reacted with ethyl malonyl chloride (32) to afford 33.
9
A Dieckmann reaction of 33 gave lactam 34 , which was condensed with benzamidine
using DBU to give pyrimidine 35. Chlorination of the hydroxyl group with phosphoryl

chloride, and the subsequent introduction of a side chain yielded ester 37. Hydrolysis of
the ester gave compound 38, which was converted to N-isobutylacetamide derivative 39
by a condensation reaction.
3. Results and discussion
The PDE4 inhibitory activities of synthesized compounds were evaluated using
1
0
human PDE4B enzyme prepared according to the reported protocols. For selected
compounds, an inhibition of LPS-induced TNF-α production in isolated mouse
splenocytes was also assessed. Furthermore, in vivo anti-inflammatory activity against
LPS-induced pulmonary neutrophilia in mice was evaluated for representative
compounds (10f, 39). These results are summarized in Tables 1-5.
3.1. Modification of pyrimidine core
Initially, the influence of substituents on the pyrimidine ring or primary amide moiety
of 2 was evaluated (Table 1). Introduction of methyl and methylthio groups at the
-position of the pyrimidine ring decreased the PDE4B inhibitory activity (19, 24).
6
Substitution on the amide moiety also decreased the potency (9b-d). Among these
compounds, only 2 showed moderate PDE4B inhibition.
3
.2. Amidation of carboxylic acid
Next, in vitro cell-free and cell-based evaluations were performed using the amide
derivatives of the carboxylic acid of the side chain (Table 2). In our previous study for 1,

we found that a conversion of the carboxylic acid of the side chain to an alkyl
5
substituted amide was effective at improving the in vitro profiles. It was similarly
found in this study that the primary and alkyl substituted amide compounds (10a-f)
exhibited moderate to good PDE4B inhibitory activity. Hydroxamic acid 10g also
showed strong PDE4B inhibition, although sulfonyl amide 10h decreased the potency.
Considering the result that n-propyl amide and t-butyl amide compounds (10b, 10e)
exhibited weaker PDE4B inhibitory activity than other alkyl amide compounds(10c,
1
0d, 10f), the elongated alkyl groups that have methylene linked terminal bulky moiety
were found to be an appropriate form for potent inhibition.
As expected, advanced inhibitory activities against LPS-induced TNF-α production in
mouse splenocytes were observed in all compounds except 10h because of their
improved membrane permeability (data not shown). In particular, the neopentyl amide
compound (10f) exhibited strong inhibitory activity against LPS-induced TNF-α
production with an IC50 value of 3.0 nM.
3.3. Modification on 2-position of the pyrimidine core
We further evaluated PDE4B inhibitory activity to search for suitable substituents on
the 2-position of the pyrimidine core (Table 3). Introduction of the 3-pyridinyl group
retained the PDE4B inhibitory potency (29a), whereas the 5-pyrimidinyl group
exhibited decreased potency (29b). Although the 3-thienyl group led to the loss of
potency, 3-furyl and 5-indolyl groups were shown to be positive for PDE4B inhibitory
activity (29c-e). However, 29d and 29e showed only weak inhibitory activity against
LPS-induced TNF-α production in mouse splenocytes. Besides these aromatic

substituents, we also evaluated the influence of alkenyl and amino groups, which
include 2-propenyl, dimethylamino, pyrrolidino, and piperidylamino groups (29f,
3
0a-c). Although the piperidylamino compound (30c) showed a decrease in PDE4B
inhibitory activity, the other three compounds strongly inhibited both PDE4B activity
and TNF-α production in mouse splenocytes. From these results, it was revealed that
phenyl, pyridinyl, alkenyl, and amino groups were preferred at the 2-position of the
pyrimidine ring.
3
.4. In vivo evaluation of 10f
The in vitro physicochemical properties and in vivo pharmacokinetic properties of 10f
are shown in Table 4. Compound 10f had acceptable in vitro metabolic stability and in
vivo exposure by intraperitoneal administration (i.p.). As an in vivo study, inhibitory
activity against LPS-induced pulmonary neutrophilia in mice was assessed. In this study,
1
0f inhibited the increase of neutrophils in the bronchoalveolar lavage fluid with an ID50
of 16 mg/kg (i.p.).
3.5. X-ray crystallography
In order to rationalize the high inhibitory activity of 10f for PDE4B, 10f was
co-crystallized with a human PDE4B catalytic domain and the structure of the complex
was solved by X-ray crystallography (Figure 2). The 5-carbamoyl moiety forms
water-bridged hydrogen bonding networks with Asn395 and Gln443. The potency loss
of 9b-d in Table 1 would be led from obstruction of these hydrogen bonding networks
by substitution on 5-carbamoyl moiety. The amino group at 4-position and an oxygen

atom of the 5-carbamoyl group are thought to form an intramolecular hydrogen bond,
which presumably contributes to fixing the conformation. Since this fixed conformation
of the 5-carbamoyl group is thought to be obstructed by the substitution on 6-position,
19 and 24 showed weaker inhibitory activity than 2 (Table 1). The pyrimidine ring
forms offset π-π stacking interaction with Phe446, while one of the nitrogen atoms on
the pyrimidine ring interacts with Gln443. The 2-phenyl group on the pyrimidine ring
has no clear interaction with the enzyme surface, which allows a variety of substrates in
this position such as pyridinyl, indolyl, furyl, propenyl, and amino groups in Table 3.
The neopentyl amide moiety, a methylene linked bulky alkyl group, occupies a
lipophilic pocket. To fill this lipophilic pocket, the alkyl group of amide seems
necessary to have an appropriate length that can reach into the pocket, as well as the
volume. In this point of view, the methylene moiety of the neopentyl group could be a
good linker between the amide bond and terminal t-butyl moiety. This might illustrate
why 10f had superior PDE4B inhibitory activity to that of t-butyl compound 10e (Table
2
).
Thus, the co-crystal structure suggests an excellent fit of 10f with the surface of the
enzyme. On the other hand, it is also shown that there is an empty pocket near the
6-position of pyrimidine moiety, which is supposed to be a hydrophobic pocket for an
5
interaction with the aliphatic moiety of the sulfone ring of the derivative of 1.
Accordingly, we next tested 38 and 39 as to whether the fused lactam ring, a mimic of
the sulfone ring moiety, can occupy a hydrophobic pocket to increase the inhibitory
activity.

3
.6. In vitro and in vivo evaluation of lactam compounds
The in vitro profile of the lactam compounds is shown in Table 5. The carboxylic
acid 38 showed a superior in vitro profile to 2. Moreover, N-isobutylacetamide
derivative 39 showed higher inhibitory activity against LPS-induced TNF-α production
1
1
than Roflumilast , demonstrating the highest potency among all 5-carbamoyl
derivatives. These results have shown the validity of the design based on the complex
structure as shown in Figure 2. Finally, 39 was subjected to in vivo evaluation and
intratracheal administration (1.0 mg/kg) of 39 inhibited LPS-induced pulmonary
neutrophilia in mice by 41%, which is nearly equal efficacy to that of Roflumilast.
4. Conclusions
In summary, we identified a novel class of PDE4 inhibitors that have
5-carbamoyl-2-phenylpyrimidine and 7,8-dihydro-6H-pyrido[4,3-d]pyrimidin-5-one
scaffolds. The optimization of substituents on the pyrimidine core and the side chain of
led to compound 10f, which had potent in vitro PDE4B inhibitory activity (IC50 = 8.3
nM) and high in vivo efficacy against LPS-induced pulmonary neutrophilia in mice
ID50 = 16 mg/kg, i.p.). Furthermore, from the result of the X-ray crystallography
2
(
analysis, we designed and synthesized 7,8-dihydro-6H-pyrido[4,3-d]pyrimidin-5-one
derivative 39, which had excellent inhibitory potency against LPS-induced TNF-α
production in mouse splenocytes (IC50 = 0.21 nM) and exhibited a high in vivo
inhibitory effect against LPS-induced pulmonary neutrophilia in mice (41% inhibition
at a dose of 1.0 mg/kg, i.t.). Further pharmacological exploration of this series of potent
PDE4 inhibitors is currently in progress.

5. Experimental
5.1. Chemistry
5.1.1. General
Unless otherwise noted, materials were obtained from commercial suppliers and used
without further purification. Column chromatography was performed with Biotage
FLASH Si columns. Thin-layer chromatography (TLC) was performed on Merck
1
pre-coated TLC glass sheets with Silica Gel 60 F254. The H NMR spectra were
recorded on a JEOL JNM-EX-400 spectrometer, and chemical shifts are given in ppm
(δ) from tetramethylsilane as an internal standard. The special splitting patterns are
designated as follows: s, singlet; d, doublet; dd, double of doublet; t, triplet; q, quartet;
and m, multiple. ESI mass spectra were recorded on a SCIEX API-150EX spectrometer.
The high-resolution mass (HRMS) spectra were recorded on a JEOL JMS-100LP
spectrometer or Waters Xevo Q-Tof spectrometer. IR spectra were recorded on a
HORIBA FT-720 spectrometer. Elemental analyses were performed by a Perkin-Elmer
CHNS/O 2400II, Leco CHNS-932 and a YOKOKAWA analysis IC7000RS.
5
.1.2. Ethyl 4-hydroxy-2-phenyl-pyrimidine-5-carboxylate (4)
To a solution of diethyl (ethoxymethylene)malonate (1.52 g, 7.02 mmol) and
benzamidine hydrochloride (1.00 g, 6.39 mmol) in EtOH (10 mL) was added NaOH
0.281 g, 7.02 mmol) and the mixture was warmed to 100 °C. After stirring at 100 °C
(

for 4 h, the reaction mixture was cooled to room temperature and the resulting
precipitate was collected by filtration to give the title compound (1.30 g, 5.32 mmol,
1
8
8%) as a colorless solid. H NMR (CDCl
3
) δ: 1.35 (3H, t, J = 6.9 Hz), 4.32 (2H, q, J =
6.9 Hz), 7.42-7.52 (3H, m), 8.30 (2H, d, J = 7.5 Hz), 8.92 (1H, s); MS (ESI) m/z: 245
+
(
M+H) .
5
.1.3. Ethyl 4-chloro-2-phenyl-pyrimidine-5-carboxylate (5)
A solution of 4 (1.20 g, 4.91 mmol) in phosphoryl chloride (8 mL) was stirred at 100
°C for 3 h. After cooling, the reaction mixture was concentrated in vacuo and the
residue was dissolved in EtOH (5 mL). To the resulting mixture, H O was added
2
dropwise over 5 min and the precipitate was collected by filtration and washed with
1
water to give the title compound (0.765 g, 2.91 mmol, 59%) as a colorless solid. H
NMR (CDCl
3
) δ: 1.46 (3H, t, J = 7.1 Hz), 4.48 (2H, q, J = 7.1 Hz), 7.52-7.59 (3H, m),
+
8
.51-8.53 (2H, m), 9.21 (1H, s); MS (ESI) m/z: 263 (M+H) .
5.1.4. 4-Chloro-2-phenyl-pyrimidine-5-carboxylic acid (6)
To a solution of 5 (0.700 g, 2.66 mmol) in EtOH (5 mL) and THF (5 mL) was added
1N NaOH aqueous solution (5.3 mL) and the reaction mixture was stirred at room
temperature. After 2 h, the organic solvent was evaporated and 1 N HCl aqueous
solution (10 mL) was added to the residue. The resulting mixture was extracted 3 times
3
with i-PrOH/CHCl (1:3, v/v) and the combined extract was washed with brine, dried

over Na
2
SO
4
and concentrated in vacuo to give the title compound (0.622 g, 2.65 mmol,
1
9
9%) as a colorless solid. H NMR (CDCl
3
) δ: 7.55-7.63 (3H, m), 8.38-8.43 (2H, m),
+
9
.25 (1H, s); MS (ESI) m/z: 235 (M+H) .
5.1.5. 4-Chloro-2-phenyl-pyrimidine-5-carboxamide (7a)
2 2
To a solution of 6 (3.73 g, 15.9 mmol) in CH Cl (50 mL) were added oxalyl chloride
(
2.73 mL, 31.8 mmol) and DMF (0.1 mL) at 0 °C. The reaction mixture was allowed to
warm to room temperature and stirred for 3 h. The reaction mixture was concentrated in
vacuo and the residue was dissolved in THF (50 mL). The resulting mixture was cooled
to -10 °C and 28% NH
min, 1 N HCl aqueous solution (40 mL) was added and the reaction mixture was
concentrated in vacuo. The resulting residue was triturated in THF (3 mL), H O (15
mL), and sat. NaHCO aqueous solution (10 mL). The resulting precipitate was
3
(2.14 mL, 31.8 mmol) was added. After stirring at -10 °C for 30
2
3
collected by filtration to give the title compound (2.52 g, 10.8 mmol, 68%) as a
1
colorless solid. H NMR (DMSO-d
6
) δ: 7.56-7.63 (3H, m), 7.97 (1H, s), 8.19 (1H, s),
+
8
.36-8.39 (2H, m), 8.97 (1H, s); MS (ESI) m/z: 234 (M+H) .
5
.1.6. 4-Chloro-N-methyl-2-phenyl-pyrimidine-5-carboxamide (7b)
2 2
To a solution of 6 (150 mg, 0.639 mmol) in CH Cl (5 mL) were added oxalyl
chloride (0.110 mL, 1.28 mmol) and DMF (0.01 mL) at 0 °C. The reaction mixture was
allowed to warm to room temperature and stirred for 1 h. The reaction mixture was

concentrated in vacuo and the residue was dissolved in CH
mixture, MeNH (0.196 mL, 1.92 mmol, 9.8 mol/L solution in MeOH) was added at
room temperature. After stirring for 40 min, H O was added and the reaction mixture
was extracted twice with CH Cl . The combined extract was washed with brine, dried
2 2
Cl (5 mL). To the resulting
2
2
2
2
over Na
2
SO
4
, and concentrated in vacuo to give the title compound (108 mg, 0.436
1
mmol, 68%) as a pale yellow solid. H NMR (CDCl
3
) δ: 3.09 (3H, d, J = 5.2 Hz), 6.66
+
(
1H, s), 7.49-7.55 (3H, m), 8.44-8.48 (2H, m), 9.18 (1H, s); MS (ESI) m/z: 248 (M+H) .
5
.1.7. 4-Chloro-N,N-dimethyl-2-phenyl-pyrimidine-5-carboxamide (7c)
Compound 7c was synthesized from 6 according to the procedure used to prepare 7b
1
(
colorless amorphous solid, Yield 24%) H NMR (CDCl
3
) δ: 2.99 (3H, s), 3.18 (3H, s),
7
.48-7.56 (3H, m), 8.44-8.46 (2H, m), 8.69 (1H, s).
5
.1.8.
N-[tert-Butyl(dimethyl)silyl]oxy-4-chloro-2-phenyl-pyrimidine-5-carboxamide
(
7d)
Compound 7d was synthesized from 6 according to the procedure used to prepare 7b
1
(
pale brown amorphous solid, Yield 25%) H NMR (CDCl
3
) δ: 0.27 (6H, s), 1.03 (9H,
s), 7.47-7.54 (3H, m), 8.45-8.48 (2H, m), 8.64 (1H, s), 9.08 (1H, s); MS (ESI) m/z: 364
+
(
M+H) .

5
.1.9. Ethyl 2-[4-[(5-carbamoyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetate (8a)
To a solution of 7a (600 mg, 2.57 mmol) in DMF (5 mL) was added ethyl
-(4-aminophenyl)acetate (690 mg, 3.85 mmol) and the reaction mixture was stirred at
0 °C for 4 h. After cooling, the reaction mixture was poured into 0.2 N HCl aqueous
2
8
solution (20 mL). The resulting precipitate was collected by filtration to give the title
1
compound (961 mg, 2.55 mmol, 99%) as a pale yellow solid. H NMR (CDCl
3
) δ: 1.20
(
3H, t, J = 6.9 Hz), 3.68 (2H, s), 4.10 (2H, q, J = 6.9 Hz), 7.34 (2H, d, J = 8.6 Hz),
7.53-7.60 (3H, m), 7.74 (2H, d, J = 8.6 Hz), 7.89 (1H, s), 8.35-8.37 (2H, m), 8.46 (1H,
+
s), 9.01 (1H, s), 11.49 (1H, s); MS (ESI) m/z: 377 (M+H) .
5
2
.1.10.
Ethyl
-[4-[[5-(methylcarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetate (8b)
To a solution of 7b (108 mg, 0.436 mmol) in DMF (2 mL) was added ethyl
2-(4-aminophenyl)acetate (117 mg, 0.654 mmol) and the reaction mixture was stirred at
6
2
0 °C for 4 h. After cooling, H O was added, and the reaction mixture was extracted
twice with EtOAc/hexane (1:1, v/v). The combined organic layer was washed with
brine, dried over Na SO , and concentrated in vacuo. The resulting residue was
2
4
chromatographed (NH silica gel, EtOAc/hexane = 0% to 60%) to give the title
1
compound (78.0 mg, 0.199 mmol, 46%) as a colorless amorphous solid. H NMR
(
3
CDCl ) δ: 1.28 (3H, t, J = 6.9 Hz), 3.06 (3H, d, J = 5.2 Hz), 3.63 (2H, s), 4.18 (2H, q, J

=
=
6.9 Hz), 6.23-6.25 (1H, m), 7.33 (2H, d, J = 8.6 Hz), 7.47-7.53 (3H, m), 7.78 (2H, d, J
+
8.6 Hz), 8.40-8.43 (2H, m), 8.61 (1H, s), 10.88 (1H, s); MS (ESI) m/z: 391 (M+H) .
5
2
.1.11.
Ethyl
-[4-[[5-(dimethylcarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetate (8c)
To a solution of 7c (19.0 mg, 0.0726 mmol) in DMF (0.5 mL) was added ethyl
2-(4-aminophenyl)acetate (26.0 mg, 0.145 mmol) and the reaction mixture was stirred at
8
2
0 °C for 16 h. After cooling, H O was added and the reaction mixture was extracted
twice with EtOAc. The combined organic layer was washed with brine, dried over
Na SO , and concentrated in vacuo. The resulting residue was chromatographed (NH
2
4
silica gel, EtOAc/hexane = 0% to 50%) to give the title compound (23.0 mg, 0.0569
1
mmol, 78%) as a colorless amorphous solid. H NMR (CDCl
3
) δ: 1.27 (3H, t, J = 7.2
Hz), 3.20 (6H, s), 3.63 (2H, s), 4.17 (2H, q, J = 7.2 Hz), 7.33 (2H, d, J = 8.6 Hz),
7.47-7.50 (3H, m), 7.73 (2H, d, J = 8.6 Hz), 8.40-8.42 (2H, m), 8.46 (1H, s), 9.22 (1H,
+
s); MS (ESI) m/z: 405 (M+H) .
5
2
.1.12.
Ethyl
-[4-[[5-(hydroxycarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetate (8d)
To a solution of 7d (38.0 mg, 0.104 mmol) in DMF (2 mL) was added ethyl
-(4-aminophenyl)acetate (114 mg, 0.639 mmol) and the reaction mixture was stirred at
0 °C for 13 h. The tert-butyl(dimethyl)silyl group was deprotected during under this
2
6

condition. After cooling, H
with THF/CHCl (1:3, v/v). The combined organic layer was washed with brine, dried
over Na SO , and concentrated in vacuo. The resulting residue was purified by reverse
phase preparative HPLC (0.1% HCO
2
O was added and the reaction mixture was extracted twice
3
2
4
2
H MeCN / 0.1% HCO
2
H aq. = 0% to 100%) to
1
give the title compound (19.0 mg, 0.0484 mmol, 46%) as a colorless solid. H NMR
(
3
CDCl ) δ: 1.25 (3H, t, J = 6.9 Hz), 3.65 (2H, s), 4.15 (2H, q, J = 6.9 Hz), 7.33 (2H, d, J
=
8.6 Hz), 7.47-7.53 (3H, m), 7.76 (2H, d, J = 8.6 Hz), 8.35 (2H, dd, J = 8.0, 1.7 Hz),
+
8
.66 (1H, s); MS (ESI) m/z: 393 (M+H) .
5.1.13. 2-[4-[(5-Carbamoyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetic acid (2)
2
To a solution of 8a (960 mg, 2.55 mmol) in THF (15 mL), EtOH (15 mL), and H O
(
2.5 mL) was added 1 N NaOH aqueous solution (5.10 mL) and the mixture was stirred
at room temperature. After 14 h, 1 N HCl aqueous solution (6.0 mL) was added to the
reaction mixture and the organic solvent was evaporated. H O was added to the residue
2
and the mixture was triturated. The resulting precipitate was collected by filtration to
1
give the title compound (854 mg, 2.45 mmol, 96%) as a pale yellow solid. H NMR
(
6
DMSO-d ) δ: 3.58 (2H, s), 7.33 (2H, d, J = 8.6 Hz), 7.54-7.57 (3H, m), 7.73-7.75 (2H,
m), 7.87 (1H, s), 8.36-8.38 (2H, m), 8.44 (1H, s), 9.00 (1H, s), 11.45 (1H, s); MS (ESI)
+
m/z: 349 (M+H) ; HRMS (ESI) Calcd for C19
H
16
N
4
O
3
-H: 347.1144. Found: 347.1140.
5.1.14. 2-[4-[[5-(Methylcarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetic
acid (9b)

Compound 9b was synthesized from 8b according to the procedure used to prepare 2.
1
(
yellow solid, Yield 97%) H NMR (DMSO-d
6
) δ: 2.83 (3H, d, J = 4.6 Hz), 3.56 (2H,
s), 7.30 (2H, d, J = 8.6 Hz), 7.51-7.55 (3H, m), 7.71-7.73 (2H, m), 8.33-8.35 (2H, m),
+
8
.91-8.93 (2H, m), 11.29 (1H, s); MS (ESI) m/z: 363 (M+H) ; HRMS (ESI) Calcd for
20 18 4 3
C H N O +H: 363.1457. Found: 363.1458.
5.1.15. 2-[4-[[5-(Dimethylcarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetic
acid (9c)
Compound 9c was synthesized from 8c according to the procedure used to prepare 2.
1
(
(
(
6
colorless solid, Yield 86%) H NMR (DMSO-d ) δ: 3.00 (6H, s), 3.55 (2H, s), 7.27
2H, d, J = 8.6 Hz), 7.48-7.51 (3H, m), 7.64 (2H, d, J = 8.6 Hz), 8.28-8.30 (2H, m), 8.43
+
1H, s), 9.12 (1H, s); MS (ESI) m/z: 377 (M+H) ; HRMS (ESI) Calcd for
21 20 4 3
C H N O -H: 375.1457. Found: 375.1458.
5.1.16. 2-[4-[[5-(Hydroxycarbamoyl)-2-phenyl-pyrimidin-4-yl]amino]phenyl]acetic
acid (9d)
Compound 9d was synthesized from 8d according to the procedure used to prepare 2.
1
(
6
yellow solid, Yield 71%) H NMR (DMSO-d ) δ: 3.56 (2H, s), 7.31 (2H, d, J = 8.6
Hz), 7.52-7.54 (3H, m), 7.71 (2H, d, J = 8.6 Hz), 8.32-8.33 (2H, m), 8.76 (1H, s), 10.88
+
(
1H, s), 11.65 (1H, s); MS (ESI) m/z: 365 (M+H) ; HRMS (ESI) Calcd for
19 16 4 4
C H N O -H: 363.1093. Found: 363.1094.

5.1.17.
4-[4-(2-Amino-2-oxo-ethyl)anilino]-2-phenyl-pyrimidine-5-carboxamide
(10a)
2 2
To a solution of 2 (250 mg, 0.718 mmol) in CH Cl (5 mL) were added oxalyl
chloride (0.246 mL, 2.87 mmol) and DMF (three drops), and the reaction mixture was
stirred at room temperature. After stirring for 1 h, the reaction mixture was concentrated
in
vacuo
to
give
crude
2-[4-[(5-carbamoyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetyl chloride (270 mg) as
a pale yellow solid. This compound was used for next reaction without further
purification.
To a solution of 2-[4-[(5-carbamoyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetyl
chloride (30.0 mg, 0.0818 mmol) in THF (0.5 mL) and DMF (0.5 mL) was added 28%
3
NH aqueous solution (0.0276 mL, 0.409 mmol) at 0 °C, and the reaction mixture was
stirred at same temperature. After stirring for 1.5 h, the reaction mixture was
concentrated in vacuo. The resulting residue was purified by reverse phase preparative
HPLC (0.1% HCO
2
H MeCN / 0.1% HCO
2
H aq. = 0% to 100%) to give the title
1
compound (8.00 mg, 0.0230 mmol, 28%) as a colorless solid. H NMR (DMSO-d
6
) δ:
3.38 (2H, s), 3.40 (2H, s), 6.91 (1H, s), 7.32 (2H, d, J = 7.9 Hz), 7.49 (1H, s), 7.54-7.57
(
(
3H, m), 7.72 (2H, d, J = 7.9 Hz), 7.87 (1H, s), 8.36-8.44 (2H, m), 9.00 (1H, s), 11.41
+
1H, s); MS (ESI) m/z: 348 (M+H) ; HRMS (ESI) Calcd for C19
17
H N
5
2
O +H: 348.1460.
Found: 348.1465.

5
4
.1.18.
-[4-[2-Oxo-2-(propylamino)ethyl]anilino]-2-phenyl-pyrimidine-5-carboxamide
(
10b)
To a solution of 2 (200 mg, 0.574 mmol) in DMF (4 mL) were added
N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide (165 mg, 0.861 mmol),
-hydroxybenzotriazole hydrate (116 mg, 0.861 mmol), N,N-diisopropylethylamine
0.300 mL, 1.72 mmol), and n-propylamine (0.0710 mL, 0.861 mmol), and the reaction
mixture was stirred at room temperature. After stirring for 17 h, the reaction mixture
was concentrated in vacuo. The resulting residue was chromatographed (THF/CH Cl
% to 50%) to give the title compound (112 mg, 0.288 mmol, 50%) as a colorless solid.
1
(
2
2
=
0
1
6
H NMR (DMSO-d ) δ: 0.84 (3H, t, J = 7.4 Hz), 1.38-1.45 (2H, m), 3.02 (2H, q, J = 6.3
Hz), 3.40 (2H, s), 7.31 (2H, d, J = 8.6 Hz), 7.52-7.58 (3H, m), 7.71 (2H, d, J = 8.6 Hz),
7
.84 (1H, s), 8.01 (1H, t, J = 5.4 Hz), 8.36-8.42 (3H, m), 9.00 (1H, s), 11.38 (1H, s); MS
+
(
ESI) m/z: 390 (M+H) ; HRMS (ESI) Calcd for C22
H
23
N
5
O
2
+H: 390.1930. Found:
-
1
3
1
6
90.1930; IR (KBr) cm : 3297, 3196, 2962, 2931, 1674, 1646, 1602, 1558, 1515, 1488,
418, 790, 697; Anal. Calcd for C22
7.47; H, 5.93; N, 17.70.
23 5 2
H N O : C, 67.85; H, 5.95; N, 17.98. Found: C,
5
4
.1.19.
-[4-[2-(Cyclopropylmethylamino)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-car
boxamide (10c)

Compound 10c was synthesized from 2 according to the procedure used to prepare
1
1
0b. (colorless solid, Yield 43%) H NMR (DMSO-d
6
) δ: 0.14-0.17 (2H, m), 0.38-0.42
(2H, m), 0.87-0.95 (1H, m), 2.96 (2H, t, J = 6.3 Hz), 3.42 (2H, s), 7.32 (2H, d, J = 8.6
Hz), 7.52-7.57 (3H, m), 7.71 (2H, d, J = 8.6 Hz), 7.84 (1H, s), 8.11 (1H, t, J = 5.4 Hz),
+
8
.36-8.42 (3H, m), 9.00 (1H, s), 11.38 (1H, s); MS (ESI) m/z: 402 (M+H) ; HRMS
-
1
(
23 5 2
ESI) Calcd for C23H N O +H: 402.1930. Found: 402.1922; IR (KBr) cm : 3304,
3196, 1672, 1643, 1603, 1558, 1515, 1489, 1419, 1385, 790, 697.
5
4
.1.20.
-[4-[2-(Isobutylamino)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-carboxamide
(
10d)
Compound 10d was synthesized from 2 according to the procedure used to prepare
1
1
1
7
8
6
0b. (colorless solid, Yield 55%) H NMR (DMSO-d ) δ: 0.83 (6H, d, J = 6.9 Hz),
.64-1.72 (1H, m), 2.89 (2H, t, J = 6.3 Hz), 3.42 (2H, s), 7.32 (2H, d, J = 8.6 Hz),
.52-7.58 (3H, m), 7.71 (2H, d, J = 8.6 Hz), 7.84 (1H, s), 8.02 (1H, t, J = 5.7 Hz),
+
.36-8.41 (3H, m), 9.00 (1H, s), 11.37 (1H, s). MS (ESI) m/z: 404 (M+H) ; HRMS
-
1
(
ESI) Calcd for C23
H
25
N
5
O
2
+H: 404.2087. Found: 404.2041; IR (KBr) cm : 3300,
197, 2958, 1675, 1644, 1602, 1558, 1515, 1489, 1418, 1320, 1211, 790, 696; Anal.
: C, 68.47; H, 6.25; N, 17.36. Found: C, 68.31; H, 6.31; N, 17.08.
3
25 5 2
Calcd for C23H N O

5
4
.1.21.
-[4-[2-(tert-Butylamino)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-carboxamide
(10e)
Compound 10e was synthesized from 2 according to the procedure used to prepare
1
1
6
0b. (colorless solid, Yield 87%) H NMR (DMSO-d ) δ: 1.26 (9H, m), 3.36 (2H, s),
7
.30 (2H, d, J = 8.6 Hz), 7.52-7.58 (3H, m), 7.67 (1H, s), 7.70 (2H, d, J = 8.6 Hz), 7.84
+
(
1H, s), 8.36-8.44 (3H, m), 9.00 (1H, s), 11.37 (1H, s); MS (ESI) m/z: 404 (M+H) ;
-1
25 5 2
HRMS (ESI) Calcd for C23H N O +H: 404.2087. Found: 404.2086; IR (KBr) cm :
3335, 3196, 2867, 1677, 1651, 1601, 1558, 1515, 1486, 1448, 1419, 1212, 788, 697.
5
4
.1.22.
-[4-[2-(2,2-Dimethylpropylamino)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-car
boxamide (10f)
Compound 10f was synthesized from 2 according to the procedure used to prepare
1
1
0b. (colorless solid, Yield 76%) H NMR (DMSO-d
6
) δ: 0.83 (9H, s), 2.90 (2H, d, J =
6.3 Hz), 3.46 (2H, s), 7.34 (2H, d, J = 8.6 Hz), 7.52-7.56 (3H, m), 7.70 (2H, d, J = 8.6
Hz), 7.84 (1H, s), 7.93 (1H, t, J = 6.3 Hz), 8.36-8.41 (3H, m), 9.00 (1H, s), 11.36 (1H,
+
s); MS (ESI) m/z: 418 (M+H) ; HRMS (ESI) Calcd for C24
27
H N
5
O
2
+H: 418.2243.
-
1
Found: 418.2242; IR (KBr) cm : 3321, 3198, 2959, 1677, 1649, 1622, 1602, 1559,
1
516, 1487, 1418, 1209, 791, 697.

5
4
.1.23.
-[4-[2-(Hydroxyamino)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-carboxamide
(10g)
To a solution of 2 (20.0 mg, 0.0574 mmol) in DMF (1 mL) were added
N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide (16.5 mg, 0.0861 mmol),
-hydroxybenzotriazole hydrate (11.6 mg, 0.0861 mmol), N,N-diisopropylethylamine
0.0300 mL, 0.172 mmol), and hydroxylamine hydrochloride (5.98 mg, 0.0861 mmol),
1
(
and the reaction mixture was stirred at room temperature. After stirring for 15 h, the
reaction mixture was concentrated in vacuo. The resulting residue was purified by
revers phase preparative HPLC (0.1% HCO
2
2
H MeCN / 0.1% HCO H aq. = 0% to
1
1
00%) to give the title compound (14.0 mg, 0.0385 mmol, 67%) as a colorless solid. H
6
NMR (DMSO-d ) δ: 3.29 (2H, s), 7.32 (2H, d, J = 8.5 Hz), 7.52-7.58 (3H, m), 7.73 (2H,
d, J = 8.5 Hz), 7.87 (1H, s), 8.36-8.39 (2H, m), 8.42 (1H, s), 8.86 (1H, s), 9.00 (1H, s),
+
1
0.67 (1H, s), 11.40 (1H, s); MS (ESI) m/z: 364 (M+H) ; HRMS (ESI) Calcd for
19 17 5 3
C H N O +H: 364.1410. Found: 364.1406.
5
.1.24.
-[4-[2-(Methanesulfonamido)-2-oxo-ethyl]anilino]-2-phenyl-pyrimidine-5-carboxa
4
mide (10h)
Compound 10h was synthesized from 2 according to the procedure used to prepare
1
1
0g. (pale brown solid, Yield 29%) H NMR (DMSO-d
6
) δ: 3.23 (3H, s), 3.62 (2H, s),
7
.33 (2H, d, J = 8.5 Hz), 7.54-7.58 (3H, m), 7.76 (2H, d, J = 8.5 Hz), 7.87 (1H, s),

+
8
.34-8.44 (3H, m), 9.01 (1H, s), 11.44 (1H, s); MS (ESI) m/z: 426 (M+H) ; HRMS
(
ESI) Calcd for C20 S+H: 4426.1236. Found: 426.1247.
19 5 4
H N O
5.1.25. Ethyl 4-methyl-6-oxo-2-phenyl-4,5-dihydro-1H-pyrimidine-5-carboxylate
(
12)
A solution of diethyl 2-ethylidenemalonate (1.10 g, 5.91 mmol), benzamidine
hydrochloride (925 mg, 5.91 mmol), and N,N-diisopropylethylamine (1.03 mL, 5.91
mmol) in MeOH (10 mL) was heated in a microwave reactor at 110 °C for 45 min.
After cooling, the reaction mixture was concentrated in vacuo. The resulting residue
was chromatographed (EtOAc/hexane = 0% to 50%) to give the title compound (918
1
mg, 3.53 mmol, 60%) as a colorless solid. H NMR (CDCl
3
) δ: 1.30 (3H, t, J = 7.2 Hz),
1.43 (3H, d, J = 6.9 Hz), 3.27-3.32 (1H, m), 3.81 (1H, s), 4.25-4.29 (3H, m), 7.44-7.47
+
(
3H, m), 7.74-7.77 (2H, m), 8.42 (1H, s); MS (ESI) m/z: 261 (M+H) .
5
5
.1.26.
Ethyl
-bromo-4-methyl-6-oxo-2-phenyl-1,4-dihydropyrimidine-5-carboxylate (13)
To a solution of 12 (910 mg, 3.50 mmol) in CCl
N-bromosuccinimide (622 mg, 3.50 mmol), K CO (4.84 g, 35.0 mmol), and benzoyl
peroxide (42.4 mg, 0.175 mmol). The reaction mixture was refluxed for 1 h. After
cooling, H O was added and the mixture was extracted 3 times with CH Cl . The
combined organic layer was washed with brine, dried over Na SO , and concentrated in
4
(40 mL) were added
2
3
2
2
2
2
4
vacuo. The resulting residue was chromatographed (EtOAc/hexane = 0% to 50%) to
1
give the title compound (214 mg, 0.631 mmol, 18%) as a colorless amorphous solid. H

3
NMR (CDCl ) δ: 1.25 (3H, t, J = 7.2 Hz), 1.64 (3H, d, J = 6.9 Hz), 4.23-4.31 (3H, m),
7.46-7.50 (2H, m), 7.53-7.56 (1H, m), 7.76 (2H, d, J = 7.4 Hz), 8.21 (1H, s); MS (ESI)
+
m/z: 339 (M+H) .
5.1.27. Ethyl 4-chloro-6-methyl-2-phenyl-pyrimidine-5-carboxylate (14)
3
A solution of 13 (114 mg, 0.336 mmol) in POCl (2 mL) was stirred at 100 °C for 1
h. After cooling, the reaction mixture was concentrated in vacuo. The resulting residue
was chromatographed (EtOAc/hexane = 0% to 30%) to give the title compound (68.0
1
mg, 0.246 mmol, 73%) as a colorless solid. H NMR (CDCl
3
) δ: 1.44 (3H, t, J = 7.4
Hz), 2.63 (3H, s), 4.48 (2H, q, J = 7.4 Hz), 7.47-7.53 (3H, m), 8.44-8.47 (2H, m); MS
+
(
ESI) m/z: 277 (M+H) .
5
.1.28. 2-[4-[(5-Carbamoyl-6-methyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetic
acid (19)
To a solution of 14 (125 mg, 0.452 mmol) in THF (2 mL) and EtOH (2 mL) were
added 1 N NaOH aqueous solution (0.903 mL) and H O (0.100 mL). After stirring at
2
room temperature for 5 h, the reaction mixture was warmed to 50 °C and stirred for 3.5
h. After cooling, 1 N HCl aqueous solution (1.00 mL) was added and the organic
solvent was evaporated. H
2
O was added to the residue and the mixture was extracted
. The combined organic layer was washed with brine, dried over Na SO
concentrated in vacuo to give 1:1 mixture of
with CH
and
2
Cl
2
2
4
,
a

4
4
-chloro-6-methyl-2-phenyl-pyrimidine-5-carboxylic
acid
(15)
and
-ethoxy-6-methyl-2-phenyl-pyrimidine-5-carboxylic acid (16). The resulting mixture
(130 mg) was used for the next reaction without further purification. 15; MS (ESI) m/z:
+
+
2
49 (M+H) . 16; MS (ESI) m/z: 259 (M+H) .
2 2
To a solution of the mixture (15 and 16) (130 mg) in CH Cl (2 mL) were added
oxalyl chloride (0.125 mL, 1.36 mmol) and DMF (0.01 mL) at 0 °C. The reaction
mixture was allowed to warm to room temperature and stirred for 20 min. The reaction
mixture was concentrated in vacuo and the residue was dissolved in THF (5 mL). 28%
NH
stirring for 1 h, H
CHCl . The combined extract was washed with brine, dried over Na
concentrated in vacuo to give crude mixture (132 mg)
3
aqueous solution (1.00 mL) was added to the resulting mixture, at 0 °C. After
O was added and the reaction mixture was extracted twice with
SO , and
including
2
3
2
4
a
4-chloro-6-methyl-2-phenyl-pyrimidine-5-carboxamide (17). This mixture was used for
+
the next reaction without further purification. MS (ESI) m/z: 248 (M+H) .
To a solution of the mixture including 17 (132 mg) in DMF (3 mL) was added ethyl
2
-(4-aminophenyl)acetate (191 mg, 1.07 mmol) and the reaction mixture was stirred at
0 °C for 5.5 h. After cooling, H O was added and the reaction mixture was extracted
SO
and concentrated in vacuo. The resulting residue was chromatographed (EtOAc/hexane
0% to 80%) to give crude mixture (106 mg) including ethyl
-[4-[(5-carbamoyl-6-methyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetate (18). This
8
2
twice with EtOAc. The combined extract was washed with brine, dried over Na
2
4
,
=
a
2
mixture was used for the next reaction without further purification. MS (ESI) m/z: 391
+
(
M+H) .

To a solution of the mixture including 18 (106 mg) in THF (2 mL), EtOH (2 mL),
and H O (1 mL) was added 1 N NaOH aqueous solution (0.407 mL) and the mixture
was stirred for 4 h. After 1 N HCl aqueous solution (0.5 mL) was added, the solvent
was evaporated. To the residue, H O was added and the precipitate was collected by
2
2
filtration to give the title compound (47.0 mg, 0.130 mmol, 29% from 14) as a colorless
1
solid. H NMR (DMSO-d
6
) δ: 2.52 (3H, s), 3.57 (2H, s), 7.26 (2H, d, J = 8.6 Hz),
7.49-7.54 (3H, m), 7.63 (2H, d, J = 8.6 Hz), 7.94 (1H, s), 8.15 (1H, s), 8.23-8.25 (2H,
+
18 4 3
m); MS (ESI) m/z: 363 (M+H) ; HRMS (ESI) Calcd for C20H N O +H: 363.1457.
Found: 363.1408.
5.1.29. 4-Chloro-6-methylsulfanyl-2-phenyl-pyrimidine-5-carbonitrile (22)
To a solution of ethyl 3,3-bis(methylthio)-2-cyanoacrylate (20) (1.74 g, 8.00 mmol)
and benzamidine hydrochloride (1.25 g, 8.00 mmol) in EtOH (100 mL) was added
triethylamine (5.56 mL, 40.0 mmol) and the reaction mixture was refluxed for 5 h. After
cooling, the reaction mixture was concentrated in vacuo. 1N HCl aqueous solution (100
mL) and H
2
O (100 mL) were added to the residue and the resulting precipitate was
by filtration to give
collected
4-methylsulfanyl-6-oxo-2-phenyl-1H-pyrimidine-5-carbonitrile (21) (1.81 g, 7.42
mmol, 93%) as a colorless solid.
3
To a solution of 21 (900 mg, 3.70 mmol) in POCl (10 mL) was added
N,N-diethylaniline (1.18 mL, 7.40 mmol) and the reaction mixture was stirred at 100 °C
for 2.5 h. After cooling, the reaction mixture was concentrated in vacuo. The resulting

residue was chromatographed (CH
2 2
Cl = 100%) to give the title compound (829 mg,
1
3
.17 mmol, 86%) as a colorless solid. H NMR (CDCl
3
) δ: 2.81 (3H, s), 7.51-7.54 (2H,
+
m), 7.58-7.61 (1H, m), 8.47-8.49 (2H, m); MS (ESI) m/z: 262 (M+H) .
5
2
.1.30.
Ethyl
-[4-[(5-cyano-6-methylsulfanyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetate
(
23)
To a solution of 22 (480 mg, 1.83 mmol) in DMF (6 mL) was added ethyl
-(4-aminophenyl)acetate (658 mg, 3.67 mmol) and the reaction mixture was stirred at
0 °C for 1.5 h. After cooling, H O was added and the reaction mixture was extracted
SO
and concentrated in vacuo. The resulting residue was chromatographed (EtOAc/CH Cl
0% to 10%) to give the title compound (620 mg, 1.53 mmol, 83%) as a colorless
2
8
2
twice with EtOAc. The combined extract was washed with brine, dried over Na
2
4
,
2
2
=
1
3
solid. H NMR (CDCl ) δ: 1.28 (3H, t, J = 7.2 Hz), 2.77 (3H, s), 3.65 (2H, s), 4.19 (2H,
q, J = 7.2 Hz), 7.12 (1H, s), 7.36 (2H, d, J = 8.6 Hz), 7.47-7.55 (3H, m), 7.64 (2H, d, J =
+
8
.6 Hz), 8.41-8.44 (2H, m); MS (ESI) m/z: 405 (M+H) .
5
2
.1.31.
-[4-[(5-Cyano-6-methylsulfanyl-2-phenyl-pyrimidin-4-yl)amino]phenyl]acetic acid
(
24)

Compound 23 (60.0 mg, 0.148 mmol) was dissolved in conc. H
2
SO
4
(1.5 mL), and
O were added
the reaction mixture was stirred at 70 °C for 2 h. After cooling, ice and H
2
to the reaction mixture and the resulting precipitate was collected by filtration to give a
yellow solid. The resulting solid was purified by reverse phase preparative HPLC (0.1%
HCO
2
H MeCN / 0.1% HCO
2
H aq. = 0% to 100%) to give the title compound (3.00 mg,
1
0
.00761 mmol, 5%) as a colorless solid. H NMR (DMSO-d
6
) δ: 2.63 (3H, s), 3.56 (2H,
s), 7.26 (2H, d, J = 8.5 Hz), 7.52-7.56 (3H, m), 7.65 (2H, d, J = 8.5 Hz), 8.29 (1H, s),
+
8
.33-8.36 (2H, m), 8.55 (1H, s); MS (ESI) m/z: 395 (M+H) ; HRMS (ESI) Calcd for
20 18 4 3
C H N O S+H: 395.1178. Found: 395.1223.
5.1.32. 2,4-Dichloropyrimidine-5-carboxamide (27)
To a solution of 2,4-dihydroxypyrimidine-5-carboxylic acid (25) (2.98 g, 19.1 mmol)
3 5
in POCl (7.10 mL, 76.4 mmol) was added PCl (13.1 g, 63.0 mmol) and the reaction
mixture was refluxed for 4.5 h. After cooling, the reaction mixture was concentrated in
vacuo. To the resulting residue, toluene (20 mL) was added and the mixture was
2 2
concentrated in vacuo again. The residue was dissolved in CH Cl (20 mL) and filtered.
The filtrate was concentrated in vacuo to give 2,4-dichloropyrimidine-5-carbonyl
chloride (26) as a yellow oil (4.38 g). This compound was used for next reaction
without further purification.
To a solution of 26 (4.38 g) in CH
3 mL) and H O (3 mL) at -10 °C, and the reaction mixture was stirred at -10 °C for 5
min. The organic solvent was removed under reduced pressure and the resulting
2 2 3
Cl (5 mL) were added 28% NH aqueous solution
(
2

precipitate was collected by filtration to give the title compound (3.13 g, 16.3 mmol,
1
8
8
5% from 25) as a pale yellow solid. H NMR (DMSO-d
6
) δ: 8.05 (1H, s), 8.17 (1H, s),
+
.90 (1H, s); MS (ESI) m/z: 192 (M+H) .
5
2
.1.33.
-Chloro-4-[4-[2-(2,2-dimethylpropylamino)-2-oxo-ethyl]anilino]pyrimidine-5-carb
oxamide (28)
To a solution of 27 (706 mg, 3.68 mmol) in MeCN (15 mL) were added
2
-(4-aminophenyl)-N-(2,2-dimethylpropyl)acetamide (810 mg, 3.68 mmol) and
N,N-diisopropylethylamine (0.640 mL, 3.68 mmol), and the reaction mixture was stirred
at 80 °C for 3 h. After cooling to 0 °C, H O (50 mL) was added to the reaction mixture.
2
The resulting precipitate was triturated and collected by filtration to give the title
1
compound (1.29 g, 3.44 mmol, 93%) as a colorless solid. H NMR (CDCl
3
) δ: 0.82 (9H,
s), 2.89 (2H, d, J = 6.3 Hz), 3.44 (2H, s), 7.29 (2H, d, J = 8.6 Hz), 7.53 (2H, d, J = 8.6
Hz), 7.91-7.95 (2H, m), 8.42 (1H, s), 8.76 (1H, s), 11.45 (1H, s); MS (ESI) m/z: 376
+
(
M+H) .
5
4
.1.34.
-[4-[2-(2,2-Dimethylpropylamino)-2-oxo-ethyl]anilino]-2-(3-pyridyl)pyrimidine-5-
carboxamide (29a)