Design, synthesis and biological evaluation of N-arylphenyl-2,2- dichloroacetamide analogues as anti-cancer agents

Article abstract of DOI:10.1016/j.bmcl.2012.07.057

Our earlier research has shown that N-phenyl-2,2-dichloroacetamide analogues had much higher anti-cancer activity than the lead compound sodium dichloroacetate (DCA). In this current study, a variety of N-arylphenyl-2,2- dichloroacetamide analogues were synthesized via Suzuki coupling reaction and their anti-cancer activity was evaluated. The results showed that N-terphenyl-2,2-dichloroacetamide analogues had satisfactory anti-cancer activity. Among them, N-(3,5-bis(benzo[d][1,3]dioxol-5-yl)phenyl)-2,2- dichloroacetamide (6 k) had an IC₅₀ of 2.40 μM against KB-3-1 cells, 1.04 μM against H460 cells and 1.73 μM against A549 cells.

Full text of DOI:10.1016/j.bmcl.2012.07.057

Bioorganic & Medicinal Chemistry Letters 22 (2012) 7268–7271
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, synthesis and biological evaluation
of N-arylphenyl-2,2-dichloroacetamide analogues as anti-cancer agents
Tianwen Li a, Yongchong Yang a, Changmei Cheng a, , Amit K. Tiwari b, Kamlesh Sodani b, Yufen Zhao a,
⇑
Ioana Abraham b, Zhe-Sheng Chen b,
⇑
a Key laboratory of Bioorganic Phosphorus and Chemical Biology, The Ministry of Education, Department of Chemistry, Tsinghua University, Beijing 100084, PR China
b Department of Pharmaceutical Sciences, College of Pharmacy and Allied Health Professions, St. John’s University, Queen, NY 11439, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Our earlier research has shown that N-phenyl-2,2-dichloroacetamide analogues had much higher anti-
cancer activity than the lead compound sodium dichloroacetate (DCA). In this current study, a variety
of N-arylphenyl-2,2-dichloroacetamide analogues were synthesized via Suzuki coupling reaction and
their anti-cancer activity was evaluated. The results showed that N-terphenyl-2,2-dichloroacetamide
analogues had satisfactory anti-cancer activity. Among them, N-(3,5-bis(benzo[d][1,3]dioxol-5-yl)phe-
Received 18 April 2012
Revised 29 June 2012
Accepted 13 July 2012
Available online 23 July 2012
nyl)-2,2-dichloroacetamide (6 k) had an IC50 of 2.40
and 1.73 M against A549 cells.
l
M against KB-3-1 cells, 1.04
lM against H460 cells
Keywords:
Sodium dichloroacetate (DCA)
Anti-cancer activity
l
Ó 2012 Elsevier Ltd. All rights reserved.
N-Arylphenyl dichloroacetamide
Dichloroacetate (DCA) is a highly bioavailable, small molecule
that inhibits pyruvate dehydrogenase kinase.1–7 It has been used
for decades in the symptomatic treatment of inherited defects in
mitochondrial metabolism2,8–14 and lactic acidosis.15–20 Recently,
Bonnet et al.21,22 found that DCA could reverse the suppressed
mitochondrial apoptosis in cancer cells but exerted no apparent
toxicities on the normal cells. It has also been reported that DCA
could inhibit cell growth of a wide range of tumor cells such as
endometrial cancer cells,23 metastatic breast cancer cells24 and
prostate cancer cells25 by similar mechanism. Combination ther-
apy with DCA has also been suggested. For example, DCA was
found to enhance tumor cell death in combination with oncolytic
adenovirus armed with MDA-7/IL-2426 and increase the cytotoxic-
ity of some platinum compounds.27
evaluation showed that optimized compounds 2,2-dichloro-N-
(3-iodophenyl) acetamide (Fig. 1, 1a, IC50 = 4.76 M for A549 cells)
and 2,2-dichloro-N-(3,5-diiodophenyl)acetamide (Fig. 1, 1b, IC50
2.84 M for A549 cells) had much higher potency than DCA. SAR
study showed that the cytotoxicity was completely dependent on
the nature and positions of the substituents on the benzene ring.
In particular, hydrophobic and moderately electron-withdrawing
substituents at the meta position were beneficial for the
activities.30,31 As a continuing effort to develop and screen more
active compounds, in this work, a series of N-arylphenyl-2,2-
dichloroacetamide analogues were designed and synthesized,
and their anti-cancer activity and SAR were evaluated.
The synthesis of N-phenyl-2,2-dichloroacetamide heterocyclic
derivatives is outlined in Scheme 1. Suzuki coupling reaction of
heterocyclic bromide and 3-aminophenylboronic acid affords the
3-hetero aniline intermediates, and then the intermediates reacted
with 1.3 mol equiv dichloroacetyl chloride in toluene to give the
desired compounds (Scheme 1). N-biphenyl-2,2-dichloroacetamide
derivatives were synthesized via Suzuki coupling reaction
l
=
l
Because of its simple structure, low price, and especially more
than 30 years of clinical history,16,18,19 DCA is a very promising
drug and rapidly enter clinical phase II testing only three years
after report of its anti-cancer activity.28,29 However, The therapeu-
tic dosage of DCA is very high.23,25–27,30 In addition, DCA’s anti-
cancer activity is indeed not ideal, for example its IC50 against
A549 cancer cell is more than 1000 l
M.30 Modification of DCA to
enhance its anti-cancer activity is imperative.
NHCOCHCl2
NHCOCHCl2
We have previously designed and synthesized a series of
N-phenyl-2,2-dichloroacetamide analogues.30,31 The biological
I
I
I
⇑
1b
Corresponding authors. Tel./fax: +86 10 62784642 (C.C.); tel.: +1 718 990 1432;
fax: +1 718 990 1877 (Z.-S.C.).
1a
E-mail addresses: chengcm@mail.tsinghua.edu.cn (C. Cheng), chenz@stjohns.edu
(Z.-S. Chen).
Figure 1. Structure of 2,2-dichloro-N-(3-iodophenyl)acetamide (1a) and 2,2-
dichloro-N-(3,5-diiodophenyl)acetamide (1b).
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.07.057

T. Li et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7268–7271
7269
O
NH2
NH2
Cl
K2CO3
HN
Toluene
reflux, 2h
PPh3/Pd(OAc)2
CHCl2COCl
Cl
R
R-Br
Toluene/EtOH
60 °C, 12 h
OH
R
B
OH
R= furan-3-y,thiophen-2-yl, thiophen-3-yl,pyridin-2-yl, thiazol-2-yl
Scheme 1. Synthesis of N-phenyl-2,2-dichloroacetamide heterocyclic derivatives.
Table 1
O
O
Structure and in vitro activity data of mono-substituted N-biphenyl-2,2-
dichloroacetamide analogues and N-phenyl-2,2-dichloroacetamide heterocyclic
derivatives
K2CO3
Cl
Cl
HN
HN
OH
OH
PPh3/Pd(OAc)2
Cl
Ar
Cl
I
Ar
B
Toluene/EtOH
60 °C, 12 h
NHCOCHCl2
NHCOCHCl2
R
NHCOCHCl2
NHCOCHCl2
Scheme 2. Synthesis of N-([1,10-biphenyl]-3-yl)-2,2-dichloroacetamide derivatives.
R
R
R
2a-f
O
O
5a-g
4a-g
3a-e
K2CO3
PPh3/Pd(OAc)2
Cl
Cl
HN
HN
OH
OH
Cl
Ar
Cl
I
Ar
B
2
Toluene/EtOH
60 oC, 12 h
Ar
I
Entry
R
IC50 error (lM)
KB-3-1
H460
A549
Scheme 3. Synthesis of N-([1,10:30,100-terphenyl]-50-yl)-2,2-dichloroacetamide
DCA
2a
2b
2c
2d
2e
2f
3a
3b
3c
3d
3e
4a
4b
4c
4d
4e
4f
5883 359
46.77 9.18
>50
>50
>50
3892 127
20.19 6.18
37.6 3.17
>50
>50
>50
4286 375
8.90 3.42
8.48 0.79
47.59 3.24
>50
derivatives.
Ph
Furan-3-yl
Thiophen-2-yl
Thiophen-3-yl
Pyridin-2-yl
Thiazol-2-yl
Me
F
Cl
OMe
CF3
Me
F
Cl
OMe
CF3
NH2
NHCOCHCl2
Me
F
Cl
OMe
CF3
Ph
of 2,2-dichloro-N-(3-iodophenyl)acetamide and substituted
phenylboronic acid (Scheme 2). A similar method was used to
synthesize N-terphenyl-2,2-dichloroacetamide derivatives as out-
lined in Scheme 3. 2,2-dichloro-N-(3-iodophenyl)acetamide and
2,2-dichloro-N-(3,5-diiodophenyl)-acetamide were synthesized
according to the procedures we have previously reported.30,31
The cytotoxicity of all synthesized compounds was analyzed
in vitro by MTT assay using Human epidermoid carcinoma cell line
(KB-3-1), non-small cell lung cancer (A549) and large cell lung
cancer cell line NCI-H460 (H460).32,33 Since N-arylphenyl-2,2-
dicholoroactetamide derivatives showed higher activities against
the A549 cell line, the IC50 against A549 was selected as the activity
index for the SAR discussion. The cytotoxicity of these compounds
against the other two cancer cell lines is discussed later.
In our earlier study on N-phenyl-2,2-dichloroacetamide deriva-
tives, a systematic modification on the benzene ring led to the dis-
covery of much higher potent compounds 1a and 1b.30,31 In order
to further explore and enhance their cytotoxic activity, we further
modified of 1a and 1b and obtained SAR of resultant potent
compounds. SAR efforts were initiated with substituting the
phenyl or hetero group with the iodine group of 1a. As Table 1
shows, a number of N-phenyl-2,2-dichloroacetamide heterocyclic
derivatives were synthesized (2a–f). Most of the compounds
did not show good potency, while N-([1,10-biphenyl]-3-yl)-2,2-
dichloroacetamide (2a) was found to be most potent against all
cancer cell lines used in this study. The potency of furan derivative
2b was comparable to 2a. However it is much easier to introduce
substituents to benzene ring and obtain SAR of resultant com-
pounds, and this compound 2b was regrettably abandoned in the
further modifications. Hence, we chose phenyl group to substitute
the iodine group of 1a and 1b. Thereafter, a number of N-arylphe-
nyl-2,2-dichloroacetamide analogues were synthesized and their
anti-cancer activities were evaluated (Table 1 and Table 2).
Unexpectedly all N-([1,10-biphenyl]-3-yl)-2,2-dichloroacetamide
analogues did not show better activities than the lead compound
1a (Table 1). Specifically, the compounds possessing substituents
such as Ph, COOMe, NHCOCHCl2, which could conjugate with
>50
>50
>50
>50
>50
41.95 3.8
48.34 4.31
>50
13.07 2.15
35.90 8.09
>50
>50
>50
43.33 10.41
>50
26.14 5.54
>50
31.58 3.79
26.80 4.38
>50
40.39 4.87
7.55 2.84
24.25 0.49
>50
9.86 3.45
13.21 2.98
22.56 4.19
13.13 4.06
7.50 0.80
8.70 3.25
9.53 3.85
21.43 4.61
4.31 1.25
18.50 4.03
>50
47.85 2.05
8.30 6.13
7.84 2.07
36.24 5.83
8.53 1.94
23.54 4.49
>50
>50
33.76 6.31
30.15 4.79
>50
>50
8.00 2.83
>50
4g
5a
5b
5c
5d
5e
5f
>50
26.17 6.58
38.16 4.57
>50
>50
>50
>50
43.27 6.24
33.55 4.74
10.94 1.23
>50
5g
COOMe
>50
>50
IC50 is 50% inhibitory concentration.
benzene ring to form a big
seen from the IC50 values: 4g (3-NHCOCHCl2), >50
>50 M; 5g (4-COOMe), >50 M.
p-system, had lower potencies, as can be
l
M; 5f (4-Ph),
l
l
Table 2 shows the IC50 values of N-([1,10:30,100-terphenyl]-50-yl)-
2,2-dichloroacetamide derivatives against cancer cell line and
some of them displayed improvement in the activity compared
to the lead compound 1b. Specifically, the 2, 3, 4-OMe derivatives
(6a, 6b, 6c) were less potent than the 2, 3, 4-OEt derivatives (6d,
6e, 6f) respectively. These data suggest that bulky substituents at
the benzene ring are acceptable, as also can be seen from these
IC50 values: 6c (4-OMe), 20
lM; 6f (4-OEt), 8.05 lM; 6j (4-OiPr),
3.82 M. The potency of 4-OiPr derivative (6j) was comparable to
l
1b, while the 4-iPr derivative (6m) led to a sixfold loss of potency.
These data suggest that the hydrogen bond acceptor is tolerated.
Hence some disubstituted derivatives with hydrogen bond accep-

7270
T. Li et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7268–7271
Table 2
Structure and in vitro activity data of N-terphenyl-2, 2-dichloroacetamide analogues
NHCOCHCl2
R
R
6a-o
Entry
R
IC50 error (lM)
KB-3-1
H460
A549
DCA
6a
6b
6c
6d
6e
6f
6g
6h
6i
6j
6k
6l
6m
6n
6o
5883 359
>30
>30
3892 127
>30
>30
4286 375
25.50 4.82
9.40 2.15
20.78 2.87
7.28 0.95
8.83 0.91
8.05 0.69
6.51 1.25
15.90 2.13
22.30 1.89
3.82 0.47
1.73 0.18
2.75 0.49
17.41 0.94
1.95 0.23
8.99 1.24
2-OMe
3-OMe
4-OMe
2-OEt
3-OEt
4-OEt
2-SMe
3-SMe
>30
>30
8.73 1.02
14.83 1.78
23.95 2.98
>30
>30
>30
8.85 0.79
2.40 0.47
7.22 0.86
>30
2.94 0.39
5.99 0.86
5.09 0.82
4.95 0.59
6.08 0.52
>30
>30
>30
2.86 0.29
1.04 0.32
2.42 0.68
24.80 1.93
1.76 0.26
2.90 0.68
4-SMe
4-OiPr
3,4-diO-CH2
4-(Tetrahydro-2H-pyran-2-yl)oxy
4-iPr
3-F-4-OMe
3-F-5-OMe
IC50 is 50% inhibitory concentration.
tor (e.g., 6k, 6n, 6o) exhibited the best potencies, as can be seen
References and notes
from the IC50 values: 6k (3,4-diO-CH2), 1.73
1.95 M; 6o(3-F-5-OMe), 8.99 M.
lM; 6n (3-F-4-OMe),
1. Whitehouse, S.; Cooper, R. H.; Randle, P. J. Biochem. J. 1974, 141, 761.
2. Stacpoole, P. W. Metabolism 1989, 38, 1124.
l
l
SAR gleaned from MTT assay results using cancer cell lines KB-
3-1 and H460 cells (Table 1 and 2) were in line with that seen in
the A549 cells. The general trends were the same. For example,
6k, 6l, 6n were identified as the most potent compounds against
all of the cell lines, while 6a, 6b, 6c and 6m were identified as
the least potent compounds. Moreover, it was also found that
bulky substituents were acceptable. For example, the 2, 3, 4-OMe
derivatives (6a, 6b, 6c) were less potent than the 2, 3, 4-OEt deriv-
atives (6d, 6e, 6f) respectively. 4-OiPr derivative (6j) was much
more potent than 4-iPr derivative (6m), as can be seen from the
3. Kato, M.; Li, J.; Chuang, J. L.; Chuang, D. T. Structure 2007, 15, 992.
4. Aicher, T. D.; Anderson, R. C.; Gao, J.; Shetty, S. S.; Copola, G. M.; Stanton, J. L.;
Knorr, D. C.; Sperbeck, D. M.; Brand, L. J.; Vinluan, C. C.; Kaplan, E. L.; Dragland,
C. J.; Tomaselli, H. C.; Islam, A.; Lozito, R. J.; Liu, X.; Maniara, W. M.; Fillers, W.
S.; Delgrande, D.; Walter, R. E.; Mann, W. R. J. Med. Chem. 2000, 43, 236.
5. Kolobova, E.; Tuganova, A.; Boulatnikov, I.; Popov, K. M. FEBS Lett. 1997, 419,
197.
6. Aicher, T. D.; Anderson, R. C.; Bebernitz, G. R.; Coppola, G. M.; Jewell, C. F.;
Knorr, D. C.; Liu, C.; Sperbeck, D. M.; Brand, L. J.; Strohschein, R. J.; Gao, J.;
Vinluan, C. C.; Shetty, S. S.; Dragland, C.; Kaplan, E. L.; DelGrande, D.; Islam, A.;
Liu, X.; Lozito, R. J.; Maniara, W. M.; Walter, R. E.; Mann, W. R. J. Med. Chem.
1999, 42, 2741.
7. Rebernitz, G. R.; Aicher, T. D.; Stanton, J. L.; Gao, J.; Shetty, S. S.; Knorr, D. C.;
Strohschein, R. J.; Tan, J.; Brand, L. J.; Liu, C.; Wang, W. H.; Vinluan, C. C.; Kaplan,
E. L.; Dragland, C. J.; Delgrande, D.; Islam, A.; Lozito, R. J.; Liu, X.; Maniara, W.
M.; Mann, W. R. J. Med. Chem. 2000, 43, 2248.
8. Blackshear, P. J.; Holloway, P. A. H.; Alberti, K. G. M. M. Biochem. J. 1974, 142, 279.
9. Stacpoole, P. W.; Moore, G. W.; Kornhauser, D. M. N. Engl. J. Med. 1978, 298, 526.
10. Wells, P. G.; Moore, G. W.; Rabin, D.; Wilkinson, G. R.; Oates, J. A. Diabetologia
1980, 19, 109.
IC50 values: 6j (4-OiPr), 8.85
2.86 M against H460 cells; 6m (4-iPr), >30
cells and 24.80 M against H460 cells. These data suggest that
lM
against KB-3-1 cells and
l
lM against KB-3-1
l
the hydrogen bond acceptor is tolerated, which also can be found
from the IC50 values against the A549 cells.
In conclusion,
a variety of substituted N-arylphenyl-2,2-
11. Bersin, R. M.; Stacpoole, P. W.; Bersin, R. M.; Stacpoole, P. W. Am. Heart J. 1997,
134, 841.
dichloroacetamide analogues were designed and synthesized, and
their cytotoxic activity and SAR were evaluated. Among them
substituted N-([1,10:30,100-terphenyl]-50-yl)-2,2-dichloroacetamide
derivatives showed potent cytotoxic activity against not only
A549 cells but also H460 and KB-3-1 cells. Specifically, N-(3,5-
bis(benzo[d][1,3]dioxol-5-yl)phenyl)-2,2-dichloro-acetamide (6k)
12. Preiser, J. C.; Moulart, D.; Vincent, J. L. Circ. Shock. 1990, 30, 221.
13. Johnson, G. A. H.; Alberti, K. G. M. M. Biochem. Soc. Trans. 1977, 5, 1387.
14. Stacpoole, P. W.; Nagaraja, N. V.; Hutson, A. D. J. Clin. Pharmacol. 2003, 43, 683.
15. Henderson, G. N.; Curry, S. H.; Derendorf, H.; Wright, E. C.; Stacpoole, P. W. J.
Clin. Pharmacol. 1997, 37, 416.
16. Stacpoole, P. W.; Kerr, D. S.; Barnes, C.; Bunch, S. T.; Carney, P. R.; Fennell, E. M.;
Felitsyn, N. M.; Gilmore, R. L.; Greer, M.; Henderson, G. N.; Hutson, A. D.;
Neiberger, R. E.; O’Brien, R. G.; Perkins, L. A.; Quisling, R. G.; Shroads, A. L.;
Shuster, J. J.; Silverstein, J. H.; Theriaque, D. W.; Valenstein, E. Pediatrics 2006,
117, 1519.
had an IC50 of 1.04
3-1 cells and 1.73
l
M against H460 cells, 2.40
lM against KB-
l
M against A549 cells. Therefore, compound
6k is potentially valuable for drug development.
17. Stacpoole, P. W.; Nagaraja, N. V.; Hutson, A. D. J. Clin. Pharmacol. 2003, 43, 683.
18. Stacpoole, P. W.; Henderson, G. N.; Yan, Z. M.; Cornett, R.; James, M. O. Drug
Metab. Rev. 1998, 3, 499.
Acknowledgments
19. Stacpoole, P. W.; Gilbert, L. R.; Neiberger, R. E.; Carney, P. R.; Valenstein, E.;
Theriaque, D. W.; Shuster, J. J. Pediatrics 2008, 5, 1223.
We would like to thank the National Natural Science Founda-
tion of China for financial support (No. 20872078).
20. Stacpoole, P. W.; Nagaraja, N. V.; Hutson, A. D. J. Clin. Pharmacol. 2003, 43, 683.
21. Bonnet, S.; Archer, S. L.; Allalunis-Turner, J.; Haromy, A.; Beaulieu, C.;
Thompson, R.; Lee, C. T.; Lopaschuk, G. D.; Puttagunta, L.; Bonnet, S.; Harry,
G.; Hashimoto, K.; Porter, C. J.; Andrade, M. A.; Thebaud, B.; Michelakis, E. D.
Cancer Cell 2007, 11, 37.
Supplementary data
22. Michelakis, E. D.; Webster, L.; Mackey, J. R. Br. J. Cancer 2008, 99, 989.
23. Wong, J. Y. Y.; Huggins, G. S.; Debidda, M.; Munshi, N. C.; Vivo, I. D. Gynecol.
Oncol. 2008, 109, 394.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.07.057.

T. Li et al. / Bioorg. Med. Chem. Lett. 22 (2012) 7268–7271
7271
24. Sun, R. C.; Fadia, M.; Dahlstrom, J. E.; Parish, C. R.; Board, P. G.; Blackburn, A. C.
Breast Cancer Res. Treat. 2010, 120, 253.
25. Cao, W. G.; Yacoub, S.; Shiverick, K. T.; Namiki, K.; Sakai, Y.; Porvasnik, S.;
Urbanek, C.; Rosser, C. J. Prostate 2008, 11, 1223.
30. Yang, Y. C.; Shang, P. H.; Cheng, C. M.; Wang, D. C.; Yang, P.; Zhang, F.; Li, T. W.;
Lu, A. J.; Zhao, Y. F. Eur. J. Med. Chem. 2010, 45, 4300.
31. Li, T. W.; Yang, Y. C.; Cheng, C. M.; Wang, D. C.; Lu, A. J.; Zhao, Y. F. Acta Pharm.
Sin. 2012, 47, 354.
26. Xiao, L.; Li, X.; Niu, N.; Qian, J.; Xie, G.; Wang, Y. Mol. Cell. Biochem. 2010, 340, 31.
27. Olszewski, U.; Poulsen, T. T.; Ulsperger, E.; Poulsen, H. S.; Geissler, K.; Hamilton,
G. Clin. Pharmacol. 2010, 2, 177.
32. Huang, M.; Gao, H. Y.; Chen, Y.; Zhu, H.; Cai, Y. J.; Zhang, X. W.; Miao, Z. H.;
Jiang, H. L.; Shen, H. W.; Lin, L. P.; Lu, W.; Ding, J. Clin. Cancer Res. 2007, 13,
1298.
28. Tennant, D. A.; Durán, R. V.; Gottlieb, E. Nat. Rev. Cancer 2010, 10, 267.
29. Wang, F.; Ogasawara, M. A.; Huang, P. Mol. Aspects Med. 2010, 31, 75.
33. Yang, Y.; Lv, Q. J.; Du, Q. Y.; Yang, B. H.; Lin, R. X.; Wang, S. Q.; World, J.
Gastroenterology 2005, 16, 2491.