Synthesis and antituberculosis activity of novel 5-styryl-4-(hetero)aryl- pyrimidines via combination of the Pd-catalyzed Suzuki cross-coupling and S NH reactions

Article abstract of DOI:10.1016/j.bmcl.2014.05.006

Combination of the Suzuki cross-coupling and nucleophilic aromatic substitution of hydrogen (S_N^H) reactions proved to be a convenient method for the synthesis of 5-styryl-4-(hetero)aryl substituted pyrimidines from commercially avail

Full text of DOI:10.1016/j.bmcl.2014.05.006

Accepted Manuscript
Synthesis and antituberculosis activity of novel 5-styryl-4-(hetero)aryl-pyrimi-
H
dines via combination of the Pd-catalyzed Suzuki cross-coupling and SN
reactions
Marionella A. Kravchenko, Egor V. Verbitskiy, Igor D. Medvinskiy, Gennady
L. Rusinov, Valery N. Charushin
PII:
S0960-894X(14)00493-4
DOI:
http://dx.doi.org/10.1016/j.bmcl.2014.05.006
BMCL 21619
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Revised Date:
Accepted Date:
20 December 2013
2 May 2014
5 May 2014
Please cite this article as: Kravchenko, M.A., Verbitskiy, E.V., Medvinskiy, I.D., Rusinov, G.L., Charushin, V.N.,
Synthesis and antituberculosis activity of novel 5-styryl-4-(hetero)aryl-pyrimidines via combination of the Pd-
catalyzed Suzuki cross-coupling and SN H reactions, Bioorganic & Medicinal Chemistry Letters (2014), doi: http://
dx.doi.org/10.1016/j.bmcl.2014.05.006
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Bioorganic & Medicinal Chemistry Letters
jo u rn al h ome p ag e : w ww .e lse vie r .c om
Synthesis and antituberculosis activity of novel 5-styryl-4-(hetero)aryl-
pyrimidines via combination of the Pd-catalyzed Suzuki cross-coupling and SN
reactions
H
Marionella A. Kravchenkoa, Egor V. Verbitskiyb,c, Igor D. Medvinskiya, Gennady L. Rusinovb,c,
Valery N. Charushinb,c
aUral Research Institute for Phthisiopulmonology, 22 Parts’ezda, 50, Ekaterinburg, 620039, Russia
bI. Postovsky Institute of Organic Synthesis, Ural Branch of the Russian Academy of Sciences, S. Kovalevskoy Str., 22, Ekaterinburg, 620990, Russia
cUral Federal University, Mira St. 19, Ekaterinburg, 620002, Russia
ARTICLE INFO
ABSTRACT
Article history:
Received
Combination of the Suzuki cross-coupling and nucleophilic aromatic substitution of hydrogen
(SNH) reactions proved to be a convenient method for the synthesis of 5-styryl-4-(hetero)aryl
substituted pyrimidines from commercially available 5-bromopyrimidine. All intermediate 5-
Revised
Accepted
bromo-4-(hetero)aryl substituted
pyrimidines and also the targeted 5-styryl-4-
Available online
(hetero)arylpyrimidines were found to be active in micromolar concentrations in vitro against
Mycobacterium tuberculosis H37Rv, avium, terrae, and multi-drug-resistant strain isolated from
tuberculosis patients in Ural region (Russia). It has been found that some of these compounds
possess a low toxicity and have a bacteriostatic effect, comparable and even higher with that of
first-line antituberculosis drugs.
Keywords:
Pyrimidine
Antimicobacterial
Tuberculosis
Cross-coupling
2014 Elsevier Ltd. All rights reserved.
Nucleophilic aromatic substitution of hydrogen
Tuberculosis∗ (TB) is still a very serious disease, which
claims approximately two millions of deaths per year world
wide.1 The wide spread of TB is due to the following major
factors: the susceptibility of people infected with the acquired
immune deficiency syndrome (AIDS), which enhances the risk
of developing TB in 100 times, and increasing resistance to the
existing drugs.2
trans-styryl or heteroaryl substituents in the 6-position; the
most promising compound containing the furyl substituent was
active at the minimal inhibitory concentration (MIC)
comparable with that for rifampicin.12
R'
X
N
Although a long term solution of the problem is associated
with the development of a better vaccine, in the nearest future
the major role will certainly belong to the chemotherapy,3
requiring novel, effective, non-toxic antituberculosis agents.1,3,4
N
N
N
N
R
N
N
R
N
Ar
Ar
I (X= S or O)
II
It has been established that some pyrimidines,5-9 and purine
Figure 1. General structures of antimycobacterial 9-substituted purines
derivatives1,10-12 possess
a high antituberculosis activity.
Our research studies were focused on the synthesis and
biological testing of novel 4,5-disubstituted pyrimidines
containing both 2-thienyl- (2-furyl-) and trans-styryl
fragments. The target 5-styryl-4-(hetero)arylpyrimidines 7a-d
were synthesized from commercially available 5-bromo-
pyrimidine (1), as shown in Scheme 1. At the first stage, we
obtained 5-bromo-4-(hetero)arylpyrimidines 3a-d by using the
reaction of nucleophilic aromatic substitution of hydrogen
(SNH). It has previously been shown that 5-bromopyrimidine
(1) reacts with thiopene (2a), bithiophene (2b) and 2-
phenylthiophene (2c) in CF3COOH to afford the corresponding
Screening of the series of 6-substituted and 2,6-disubstituted 9-
benzylpurines revealed several compounds with a promising
level of activity against M. tuberculosis, with a general trend
that 6-substituted derivatives to be more effective than 2,6-
disubstituted compounds. Indeed, a high inhibitory activity was
found for purines of general formula I and II (Fig. 1), bearing
———
∗
Corresponding author. Tel./fax: +7 343 3693058;
E-mail address: Verbitsky@ios.uran.ru.
1

N
X
N
B(OH)2
R
Br
Br
N
N
i, ii
iii
6a-d
+
+
SHN- reaction
R
X
R'
X
N
N
1
R
4,
R'= H;
5,R'= Ph
2a-d
3a-d
N
X
N
R
a: X= S, R= H; b: X= S, R= Ph; c: X= S, R= thiophen-2-yl; d: X= O, R= furan-2-yl
7a-d
Scheme 1. Reagents and conditions: (i) CF3COOH, r.t., 24 hours; (ii) K3Fe(CN)6, KOH, H2O, r.t., 6 hours;
(iii) Pd(PPh3)4, K2CO3, H2O, THF, 155 ºC, MW, 20 min.
σH-adducts
–
5-bromo-4-(hetero)aryl-3,4-dihydropyrimidinium
Bromo derivatives 3a-d have been involved in the Suzuki
reaction with trans-2-styrylboronic acids and under
microwave irradiation (155 °C, 20 min). The corresponding
cross-coupling products, (E)-5-arylethenyl-4-(hetero)aryl-
pyrimidines 6a-d and 7a-d, have been obtained in high yields
(Scheme 1, Table 1).
salts.13 Oxidation of these compounds with K3Fe(CN)6 in an
aqueous solution of KOH gave 5-bromo-4-(hetero)
arylpyrimidines 3a-c. The reaction 5-bromopyrimidine (1) with
furan (2d) under the same conditions leads to the corresponding
5-bromo-4-(furyl-2’l)pyrimidine (3d) in a good yield (64%).
4
5
5-Bromo- (3a-d) and 5-arylethenyl- (6a-d and 7a-d)
substituted 4-(hetero)arylpyrimidines were screened for their
activity in vitro against Mycobacterium tuberculosis H37Rv,
avium, terrae, and multi-drug-resistant strains. The antimy-
cobacterial activities for 5-bromo- (3a-c) and 5-arylethenyl- (6a-c
and 7a-c) substituted 4-(hetero)arylpyrimidines were correlated
with activities of recently obtained 5-unsubsituted 4-
(hetero)arylpyrimidines (Scheme 2).13
Table 1. The microwave-assisted Suzuki cross-coupling
reaction of 5-bromo-4-(hetero)arylpyrimidines (3a-d) with
trans-2-styrilboronic acids (4 and 5)
Isolated
yield (%)
Entry
Reaction
Products
N
1
3a + 4
6a – 88
H
H
N
N
i, ii
S
N
+
SHN- reaction
R
S
S
N
N
8
R
2a-c
9a-c
N
a: R= H; b: R= Ph; c: R= thiophen-2-yl
2
3
4
3b + 4
3c + 4
3d + 4
6b – 71
6c – 80
6d – 95
S
N
Scheme 2. Reagents and conditions: (i) CF3COOH, r.t., 24 hours; (ii)
K3Fe(CN)6, KOH, H2O, r.t., 6 hours.
The synthesized pyrimidine derivatives were compared to the
commercially available drugs Isoniazid and Pyrazinamide under
the same experimental conditions (Table 2).
N
S
N
S
All 5-unsubsituted 4-thiehylpyrimidines 9a-c demonstrated a
low level of tuberculostatic activity (MIC = 12.5 µg/mL).
However, 5-bromo substituted pyrimidines 3a-d proved to
exhibit significant inhibitory properties against Mycobacterium
tuberculosis H37Rv (MIC from 6.25 to 0.7 µg/mL). As a rule,
substitution of bromo at C-5 with an arylethenyl fragment results
in a decrease of the tuberculostatic activity to 12.5 µg/mL.
Introduction of [2,2′]bithiophenyl substituent into C-4 position is
accompanied with a considerable increase in anti-TB activities of
pyrimidines 6c and 7c (MIC= 1.5 µg/mL). For the most active
compound 3a, 3b and 3d the acute toxicity on white mice was
determined (Table 2). These compounds were less toxic for white
mice than Isoniazid. For compounds 6c and 7c acute oral toxicity
was three or four times higher than that for compounds 3a,b,d.
Since the ability of some synthesized 5-substituted pyrimidines to
inhibit the growth of mycobacteria is comparable with
Pyrazinamide or even higher, these structures can be regarded as
potential prototypes for design of new antituberculosis drugs.
N
O
N
5
6
7
3a + 5
3b + 5
3c + 5
7a – 54
7b – 52
7c – 62
N
S
N
N
S
N
N
In summary, we have obtained a novel series of 5-arylethenyl-
4-(hetero)arylpyrimidines by using a convenient two step
procedure. All compounds have exhibited a reasonable level of
activity against tuberculosis and a low toxicity. Being not active
enough to be therapeutics, they certainly can be considered as
promising structures for further studies aimed at the development
of novel effective agents to combat resistant forms of TB.
S
N
S
8
3d + 5
7d – 52
N
O
N
2

Table 2. In vitro antituberculosis activity of 5-bromo-4-(hetero)arylpyrimidines (3a-d), (E)-5-arylethenyl-4-
(hetero)arylpyrimidines (6a-d and 7a-d), 4-(hetero)arylpyrimidines (9a-c) and their acute in vivo toxicity in mice
Antimycobacterial activity against different strains
of Mycobacterium tuberculosis (MIC in µg/mL)
LD50 mouse,
mg/kg
Entry
Compound
H37Rv
M. avium
M. terrae
MDRa
1
3a
3b
3c
3d
6a
6b
6c
6d
7a
7b
7c
7d
9a
9b
9c
0.7
0.3
0.3
0.7
315
2
1.5
0.7
0.7
1.5
225
n.d.
405
n.d.
n.d.
100
n.d.
n.d.
n.d.
78
3
6.25
1.5
n.d.
0.7
n.d.
0.7
n.d.
1.5
4
5
12.5
12.5
1.5
n.d.
n.d.
0.7
n.d.
n.d.
0.7
n.d.
n.d.
1.5
6
7
8
6.25
12.5
12.5
1.5
n.d.
n.d.
n.d.
0.7
n.d.
n.d.
n.d.
0.7
n.d.
n.d.
n.d.
1.5
9
10
11
12
13
14
15
12.5
12.5
12.5
12.5
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.ab
1680c
133b
151c
PZA
INH
12.5
0.1
n.d.
0.1
n.d.
0.1
-
-
n.d. - not determined; n.a. - data not available; INH – Isoniazid; PZA – Pyrazinamide;
aMDR (multi-drug-resistant tuberculosis strain) – Rifampin and Isoniazid resistant Mycobacterium tuberculosis strain having Beijing genotype with a
combination of mutations Ser 531 - Leu 315 and Ser-Thr in rpoB and katG genes, respectively.
bLD50 Oral mouse14
cLD50 Intraperitoneal mouse14
9.
Shmalenyuk, E. R.; Chernousova, L. N.; Karpenko, I. L.;
Kochetkov, S. N.; Smirnova, T. G.; Andreevskaya, S. N.;
Chizhov, A. O.; Efremenkova, O. V.; Alexandrova L. A. Bioorg.
Med. Chem. 2013, 21, 4874.
Acknowledgments
This work was supported by the Ural Branch of the Russian
Academy of Sciences (Grants № 12-P-3-1014, 12-P-3-1030, 12-
T-3-1025, 12-T-3-1031 and 13-3-019-UMA), the Russian
Foundation for Basic Research (research projects No. 13-03-
96049-r_ural_a, 13-03-90606-Arm_a and 14-03-31040-mol_а),
the Council on Grants at the President of the Russian Federation
(Program of State Support for Leading Scientific Schools of the
Russian Federation and Young Scientists, Grant MK-
3939.2014.3).
10. Janin Y. L. Bioorg. Med. Chem. 2007, 15, 2479.
11. Pathak, A. K.; Pathak, V.; Seitz, L. E.; Suling, W. J.; Reynolds
R. C. Bioorg. Med. Chem. 2013, 21, 1685.
12. (a) Bakkestuen, A. K.; Gundersen, L.-L.; Langli, G.; Liu, F.;
Nolsøe, J. M. J. Bioorg. Med. Chem. Lett. 2000, 10, 1207; (b)
Gundersen, L.-L.; Nissen-Meyer, J.; Spilsberg B. J. Med. Chem.
2002, 45, 1383; (c) Brændvang, M.; Charnock, C.; Gundersen L.-
L. Bioorg. Med. Chem. Lett. 2009, 19, 3297; (d) Read, M. L.;
Brændvang, M.; Miranda, P. O.; Gundersen L.-L. Bioorg. Med.
Chem. 2010, 18, 3885; (e) Khoje, A. D.; Kulendrn, A.; Charnock,
C.; Wan, B.; Franzblau, S.; Gundersen L.-L. Bioorg. Med. Chem.
2010, 18, 7274.
References and notes
13. (a) Verbitskiy, E. V.; Cheprakova, E. M.; Slepukhin, P. A.;
Kodess, M. I.; Ezhikova, M. A.; Pervova, M. G.; Rusinov, G.
L.; Chupakhin, O. N.; Charushin V. N. Tetrahedron 2012, 68,
5445; (b) Verbitskiy, E. V.; Cheprakova, E. M.; Zhilina, E. F.;
Kodess, M. I.; Ezhikova, M. A.; Pervova, M. G.; Slepukhin, P.
A.; Subbotina, J. O.; Schepochkin, A. V.; Rusinov, G. L.;
Chupakhin, O. N.; Charushin V. N. Tetrahedron 2013, 69,
5164.
1.
Bhowruth, V.; Dover, L. G.; Besra, G. S. Prog. Med. Chem.
2007, 45, 169. and references therein.
2. (a) Bloom, B. R.; Murray, C. J. L. Science 1992, 257, 1055; (b)
Snider, D. E. J.; Roper, W. L. N. Engl. J. Med. 1992, 326, 703;
(c) Bass, J. B. L.; Hopewell, P. C.; O’Brein, R.; Jacobs, R. F.;
Ruben, F.; Dixie, E.; Snider, J.; Thornton, G. Am. J. Respir. Crit.
Care. Med. 1994, 149, 1359; (d) Nakajima, H. World Health
1993, 46, 3; (e) Ma, Z.; Lienhardt, C.; McIlleron, H.; Nunn, A.
J.; Wang, X. Lancet 2010, 375, 2100.
14. Handbook of Anti-Tuberculosis Agents. Tuberculosis. 2008, 88,
85.
3.
Orme, I.; Secrist, J.; Anathan, S.; Kwong, C.;Maddry, J.;
Reynolds, R.; Poffenberger, A.; Michael, M.; Miller, L.;
Krahenbuh, J.; Adams, L.; Biswas, A.; Franzblau, S.;Rouse, D.;
Winfield, D.; Brooks, J. Antimicrob. Agents Chemother. 2001,
45, 1943.
Supplementary Material
4.
5.
Reddy, V. M.; Nadadhur, G.; Daneluzzi, D.; Osullivan, J. F.;
Gangadharam, P. R. J. Antimicrob. Agents Chemother. 1996, 40,
633.
Supplementary
material
(experimental
procedures,
characterization of final compounds and biological assays
protocols) associated with this article can be found in the online
version.
Rewcastle, G. W. In Comprehensive Heterocyclic Chemistry III;
Katritzky, A. R.; Ramsden C. A.; Scriven E. F. V.; Taylor R. J.
K., Eds.; Elsevier Science Ltd.: New York, 2008; Vol. 8, Chapter
8.02., 117-272.
6.
7.
8.
Morgan, J.; Haritakul, R.; Keller, P. A. Bioorg. Med. Chem. Lett.
2003, 13, 1755.
El-Hamamsy, M. H. R. I.; Smith, A. W.; Thompson, A. S.;
Threadgill, M. D. Bioorg. Med. Chem. 2007, 15, 4552.
Matyugina, E.; Khandazhinskaya, A.; Chernousova, L.;
Andreevskaya, S.; Smirnova, T.; Chizhov, A.; Karpenko, I.;
Kochetkov, S.; Alexandrova L. Bioorg. Med. Chem. 2012, 20,
6680.
3

R'
X
N
N
N
N
N
N
R
N
R
N
Ar
Ar
I (X= S or O)
II
Figure 1. General structures of antimycobacterial 9-substituted purines
4

N
X
N
B(OH)2
R
Br
Br
N
i, ii
N
iii
+
6a-d
+
SHN- reaction
R
X
R'
X
N
N
1
R
4,
R'= H;
5,R'= Ph
2a-d
3a-d
N
X
N
R
a: X= S, R= H; b: X= S, R= Ph; c: X= S, R= thiophen-2-yl; d: X= O, R= furan-2-yl
7a-d
Scheme 1. Reagents and conditions: (i) CF3COOH, r.t., 24 hours; (ii)
K3Fe(CN)6, KOH, H2O, r.t., 6 hours;
(iii) Pd(PPh3)4, K2CO3, H2O, THF, 155 ºC, MW, 20 min.
5

H
H
N
i, ii
N
+
SNH- reaction
R
S
S
N
N
8
R
2a-c
9a-c
a: R= H; b: R= Ph; c: R= thiophen-2-yl
Scheme 2. Reagents and conditions: (i) CF3COOH, r.t., 24 hours; (ii)
K3Fe(CN)6, KOH, H2O, r.t., 6 hours.
6

Table 1. The microwave-assisted Suzuki cross-coupling
reaction of 5-bromo-4-(hetero)arylpyrimidines (3a-d) with
trans-2-styrilboronic acids (4 and 5)
Isolated
yield (%)
Entry
Reaction
Products
N
1
3a + 4
6a – 88
S
N
N
2
3
4
3b + 4
3c + 4
3d + 4
6b – 71
6c – 80
6d – 95
S
N
N
S
N
S
N
O
N
5
6
7
3a + 5
3b + 5
3c + 5
7a – 54
7b – 52
7c – 62
N
S
N
N
S
N
N
S
N
S
8
3d + 5
7d – 52
N
O
N
7

Table 2. In vitro antituberculosis activity of 5-bromo-4-
(hetero)arylpyrimidines
(hetero)arylpyrimidines
(hetero)arylpyrimidines (9a-c) and their acute in vivo toxicity
(3a-d),
(6a-d
(E)-5-arylethenyl-4-
and 7a-d), 4-
in mice
Antimycobacterial activity against different strains
of Mycobacterium tuberculosis (MIC in µg/mL)
LD50 mouse,
mg/kg
Entry
Compound
H37Rv
M. avium
M. terrae
MDRa
1
3a
3b
3c
3d
6a
6b
6c
6d
7a
7b
7c
7d
9a
9b
9c
0.7
0.3
0.3
0.7
315
2
1.5
0.7
0.7
1.5
225
n.d.
405
n.d.
n.d.
100
n.d.
n.d.
n.d.
78
3
6.25
1.5
n.d.
0.7
n.d.
0.7
n.d.
1.5
4
5
12.5
12.5
1.5
n.d.
n.d.
0.7
n.d.
n.d.
0.7
n.d.
n.d.
1.5
6
7
8
6.25
12.5
12.5
1.5
n.d.
n.d.
n.d.
0.7
n.d.
n.d.
n.d.
0.7
n.d.
n.d.
n.d.
1.5
9
10
11
12
13
14
15
12.5
12.5
12.5
12.5
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.ab
1680c
133b
151c
PZA
INH
12.5
0.1
n.d.
0.1
n.d.
0.1
-
-
n.d. - not determined; n.a. - data not available; INH – Isoniazid; PZA –
Pyrazinamide;
aMDR (multi-drug-resistant tuberculosis strain) – Rifampin and Isoniazid
resistant Mycobacterium tuberculosis strain having Beijing genotype with a
combination of mutations Ser 531 - Leu 315 and Ser-Thr in rpoB and katG
genes, respectively.
bLD50 Oral mouse14
cLD50 Intraperitoneal mouse14
8

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Synthesis and antituberculosis activity of
novel 5-styryl-4-(hetero)arylpyrimidines via
combination of the Pd-catalyzed Suzuki
cross-coupling and SNH reactions
M. A. Kravchenko, E. V. Verbitskiy, I. D. Medvinskiy, G. L. Rusinov, V. N. Charushin
I. Postovsky Institute of Organic Synthesis, S. Kovalevskoy Str., 22, 620990 Ekaterinburg, Russia
R'
B(OH)2
, Pd0
Suzuki cross-coupling
Br
, [o]
R
R'
X
N
N
SNH- reaction
X
N
N
R
X= S or O; R= H, Ph, 2-thienyl; R'= H, Ph
MIC = 1.5 - 12.5 µg/mL
Mycobacterium tuberculosis
9