Synthesis of novel substituted pyrimidine derivatives bearing a sulfamide group and their in vitro cancer growth inhibition activity

Article abstract of DOI:10.1016/j.bmcl.2016.11.052

The synthesis of two series of novel substituted pyrimidine derivatives bearing a sulfamide group have been described and their in vitro cancer growth inhibition activities have been evaluated against three human tumour cell lines (HT-29, M21, and MCF7). In general, growth inhibition activity has been enhanced by the introduction of a bulky substituent on the aromatic ring with the best compound having GI₅₀?

Full text of DOI:10.1016/j.bmcl.2016.11.052

Accepted Manuscript
Synthesis of novel substituted pyrimidine derivatives bearing a sulfamide group
and their in vitro cancer growth inhibition activity
Carole-Anne Lefebvre, Elsa Forcellini, Sophie Boutin, Marie-France Côté,
René C.-Gaudreault, Patrick Mathieu, Patrick Lagüe, Jean-François Paquin
PII:
S0960-894X(16)31205-7
DOI:
http://dx.doi.org/10.1016/j.bmcl.2016.11.052
BMCL 24447
Reference:
Bioorganic & Medicinal Chemistry Letters
To appear in:
Received Date:
Accepted Date:
15 November 2016
18 November 2016
Please cite this article as: Lefebvre, C-A., Forcellini, E., Boutin, S., Côté, M-F., C.-Gaudreault, R., Mathieu, P.,
Lagüe, P., Paquin, J-F., Synthesis of novel substituted pyrimidine derivatives bearing a sulfamide group and their
in vitro cancer growth inhibition activity, Bioorganic & Medicinal Chemistry Letters (2016), doi: http://dx.doi.org/
10.1016/j.bmcl.2016.11.052
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Carole-Anne Lefebvre, Elsa Forcellini, Sophie Boutin, Marie-France Côté, René C.-Gaudreault, Patrick
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Bioorganic & Medicinal Chemistry Letters
Synthesis of novel substituted pyrimidine derivatives bearing a sulfamide group
and their in vitro cancer growth inhibition activity
Carole-Anne Lefebvrea,†, Elsa Forcellinia,†, Sophie Boutina, Marie-France Côtéb, René C.-Gaudreaultb,c,
Patrick Mathieud, Patrick Lagüee and Jean-François Paquina,
a PROTEO, Département de chimie, Université Laval, Québec, QC, Canada G1V 0A6
b CHU de Québec Research Centre, Oncology Division, Hôpital Saint-François d’Assise, Québec, QC, Canada G1L 3L5
c Department of Molecular Medicine, Faculty of Medicine, Université Laval, Québec, QC, Canada G1V 0A6
d Laboratory of Cardiovascular Pathobiology, Quebec Heart and Lung Institute/Research Center, Department of Surgery, Université Laval, QC, Canada G1V-
4G5
e PROTEO, IBIS, Département de biochimie, de microbiologie et de bio-informatique, Université Laval, Québec, QC, Canada G1V 0A6
ARTICLE INFO
ABSTRACT
Article history:
Received
Revised
Accepted
Available online
The synthesis of two series of novel substituted pyrimidine derivatives bearing a sulfamide
group have been described and their in vitro cancer growth inhibition activities have been
evaluated against three human tumour cell lines (HT-29, M21, and MCF7). In general, growth
inhibition activity has been enhanced by the introduction of a bulky substituent on the aromatic
ring with the best compound having GI50 < 6 μM for all the human tumour cell lines. The
MCF7 selective compounds were evaluated on four additional human invasive breast ductal
carcinoma cell lines (MDA-MB-231, MDA-MB-468, SKBR3, and T47D) and were selective
against T47D cell line in all cases except one, suggesting a potential antiestrogen activity.
Keywords:
Pyrimidine
Sulfamide
Anticancer activities
Structure-activity relationships (SARs)
Growth inhibition
2009 Elsevier Ltd. All rights reserved.
Cancer is one of the most devastating diseases in the world
with millions of deaths every year.1 Despite the development of
new treatments,2 deleterious effects and drug-resistance
sometimes result in therapy failure.3 Thus, the continuous
development of new chemotherapeutic scaffolds that could
potentially circumvent these shortcomings is still of primary
importance.
On the one hand, pyrimidine and its analogues are naturally
present in nucleobases which composed deoxyribonucleic acid
(DNA) and ribonucleic acid (RNA). Importantly, pyrimidine
derivatives are found in many bioactive compounds with a broad
spectrum of biological effects including antibiotic and anticancer
activities.4,5 On the other hand, the sulfamide group is a valuable
and versatile substituent in medicinal chemistry as it can be
considered as a bioisostere of sulfamate, sulfonamide, urea,
carbamate and amide functionalities. Hence, it is not surprising to
find that bioactive molecules bearing a sulfamide group display a
wide range of activity including antiepileptic, antibacterial and
antiviral activities.6,7
Considering the above-mentioned elements, these compounds
were also tested for their potential antitumour activities and
demonstrated promising results against MCF-7 cancer cell line
(Figure 1). Hence, we decided to further investigate the potential
antitumour activities of this class of compounds. We report
herein the synthesis of novel substituted pyrimidines and their in
vitro growth inhibition activity against various cancer cell lines.
The targeted compounds were built around 4,6-
dichloropyrimidine (3), and prepared by a Suzuki-Miyaura cross-
coupling using various arylboronic acids to functionalize one
chloro-position while a SNAr reaction using sulfamide-containing
piperidine 4 or 5 allowed us to obtain the final compounds 1 or 2
(Figure 1).
In the context of a screening campaign for inhibition of
ectonucleotide pyrophosphatase/phosphodiesterase 1 (NPP1),8
we had access to some compounds bearing both functionalities.
———
 Corresponding author. Tel.: +1-418-656-2131 ext. 11430; fax: +1-418-656-7916; e-mail: jean-francois.paquin@chm.ulaval.ca
† These authors contributed equally to this work.

Figure 1. Initial results, targeted substituted pyrimidine
derivatives bearing a sulfamide group, and retrosynthetic
analysis.
We first investigated the monoarylation of 4,6-
dichloropyrimidine (3) as shown in Scheme 1. Suzuki-Miyaura
conditions inspired by ones developed for the arylation of 6-aryl-
2,4-dichloropyrimidine were used.9 To avoid diarylation and thus
facilitate purification, the stoichiometry was adjusted by
increasing the amount of 4,6-dichloropyrimidine. Under those
conditions, a wide range of monoarylated pyridimine (6a-n) were
obtained in moderate to excellent yields (39-90%) using 4-, 3- or
2-substituted arylboronic acids (Scheme 1).
Scheme 2. Synthesis of piperidine 4 (n = 1). Reagents and
conditions: (a) BnBr, K2CO3, EtOH, reflux, 16 h; (b) LiAlH4,
THF, reflux, 5 h, 81% from 7 (2 steps); (c) 11, i-Pr2EtN, CH2Cl2,
rt, 16 h, 77%; (d) H2, Pd/C, PdCl2, CH3OH, 16 h, 71%; (e) 4 M
HCl/dioxane, 2.5 h, 100%.
For the synthesis of 5, 4-piperidone monohydrate
hydrochloride (12) was first Boc-protected under standard
conditions to afford 1313 in 93% yield. A Horner–Wadsworth–
Emmons
reaction
between
13
and
diethyl(cyanomethyl)phosphonate provided the acrylonitrile
derivative 14 in 93% yield.14 Reduction of the alkene and the
nitrile12 followed by sulfamoylation using reagent 1112 gave
product 15 in 61% for the 2 steps. Finally, removal of the Boc
was accomplished using 4M HCl/dioxane to give amine 5 as the
hydrochloride salt in a quantitative yield.
Scheme 1. Synthesis of 4-chloro-6-arylpyridimine 6a-n.
Inspired by a partial synthetic sequence reported by Patel,10
the synthesis of the sulfamide-containing piperidines 4 and 5 are
shown in Schemes 2 and 3, respectively.
Scheme 3. Synthesis of piperidine 5 (n = 2). Reagents and
conditions: (a) Boc2O, NaOH, THF/H2O, rt, 2 h, 93%; (b) diethyl
cyanomethylphosphonate, Et3N, LiBr, THF, rt, 16 h. 96%; (c) H2,
Ni Raney, Pd/C, LiOH·H2O, dioxane/H2O, rt, 16 h; (d) 11, i-
Pr2EtN, CH2Cl2, rt, 16 h, 61% from 14 (2 steps); (e) 4 M
HCl/dioxane, 2.5 h, 100%.
For the synthesis of 4, the primary amine 8 was first obtained
from 4-piperidinecarboxamide (7) through a two-step sequence.11
Introduction of a benzyl group on the amine followed by
reduction of the amide using LiAlH4 provided 8 in 81% over two
steps. Sulfamoylation using reagent 1112 gave 9 in 77% yield.
Removal of the benzyl group was achieved under palladium
catalysis to provide amine 10 in 71% yield. Finally, Boc-
deprotection was performed using 4 M HCl/dioxane to give
amine 4 as the hydrochloride salt in a quantitative yield.
With both partners in hands, the SNAr reaction could be
realized using K2CO3 as a base and DMF as the solvent at 90 °C
for 16 h (Scheme 4). Moderate yields (28-66%) in the case of 4,
and moderate to good yields (26-80%) in the case of 5 were
obtained. The use of a different reaction conditions (K2CO3,

CH3CN, reflux, 16 h) was also evaluated, but generally provided
lower yields.
2g
1h
2h
1i
22.6
22.9
19.6
5.7
34.8
14.1
19.6
5.0
15.2
17.7
17.6
4.1
2i
6.8
9.0
6.2
1j
47.5
31.6
>100
74.3
>100
52.9
80.4
51.8
6.8
62.5
50.7
>100
96.9
>100
93.9
>100
81.8
6.6
47.1
31.2
10.1
69.8
62.4
44.8
48.5
>100
12.1
5.9
2j
1k
2k
1l
2l
1m
2m
1n
2n
3.5
1.8
a
Values are means of three experiments and the deviation from the mean is
<10% of the mean value.
The in vitro growth inhibitory data revealed that substituted
pyrimidine derivatives 2c, 1-2d, 2h, 1-2i, and 1-2n had
significant GI50 values below 20 μM for all the cell lines. For
those compounds, the length of the linker chain between the
piperidine and the sulfamide group (n = 1 for 1, n = 2 for 2) had
no significant effect on the activity except for 1-2c. Comparison
of the nature of the aryl substituent showed that bulky
substituents were well tolerated, in particular naphthalene-
substituted compound 2n was the most active against HT-29,
M21, and MCF7 cell lines with GI50 = 3.5, 1.8, and 5.9 μM. As a
point of comparison, tamoxifen showed a GI50 value of 11.2 μM
against MCF7 cell line in a similar assay.16 High growth
inhibitory activity was also obtained for naphthalene-substituted
derivatives 1-2n. A tert-butyl substituent introduced at the para
position, as in compounds 1-2d, displayed better activity than
compounds bearing a methyl group at the same position (1-2b).
Meanwhile, compounds containing an electron-withdrawing
group at the para position such as an ester (1-2i) were potent for
the three cell lines with GI50 ˂ 10 μM whereas compounds
containing an electron-donating group on the aromatic ring at
either the ortho or the para position were selective for MCF7 cell
line with, for instance, GI50 = 35.3 μM (1e) and GI50 = 10.1 μM
(1k). In these cases, the linker length appeared to have an
important effect on the selectivity with better results being
observed for the shorter linker (n = 1). A similar phenomenon
was observed for compound 1l and 1m bearing respectively at
the ortho position a chlorine and a fluorine atom. The ethyl esters
(1i and 2i) showed a 3-fold improvement over the methyl esters
(1h and 2i). These results are in line with previous observations
suggesting that bulkier substituents are beneficial for the
antiproliferative activity.
Scheme 4. Synthesis of 1a-n and 2a-n.
The antiproliferative activity of the compounds 1a-n and 2a-
n was evaluated on three human tumour cell lines, namely, HT-
29 colon carcinoma cells, M21 skin melanoma, and MCF7
estrogen-dependent breast adenocarcinoma. Cell growth
inhibition was assessed according to the NCI/NIH
Developmental Therapeutics Program.15 The results are
summarized in Table 1 and expressed as the concentration, in
μM, of drug inhibiting the cell growth by 50% (GI50).
Table 1. Growth inhibition activity of compounds 1a-n and
2a-n on three different human cancer cell lines.
Antiproliferative activity of the MCF7 selective compounds
1a, 1e, 1k, and 1l was evaluated on four additional human
invasive breast ductal carcinoma cell lines, namely, MDA-MB-
231, MDA-MB-468, SKBR3, and T47D.17 The GI50 results are
presented in Table 2. Among the four compounds tested, 1a, 1e,
and 1l showed activity against only one cell line (T47D) with
GI50 = 24.1, 24.5, and 65.5 μM respectively, while 1k had no
antiproliferative activity. Both MCF7 and T47D are accepted
models for ER+ tumors,17 hence, compounds 1a, 1e and 1l may
function as antiestrogens while the mechanism of action of 1k
might be different. Further experiments should be conducted to
confirm this hypothesis.
GI50 (M)a
Compd
HT-29
>100
66.5
33.9
52.3
15.2
13.3
8.9
M21
>100
>100
71.0
57.6
47.2
15.1
19.3
18.4
>100
69.8
50.3
45.2
43.1
MCF7
34.3
53.2
30.6
47.5
21.8
11.3
19.7
13.1
35.3
56.6
25.5
19.7
22.5
1a
2a
1b
2b
1c
2c
1d
2d
1e
2e
1f
10.3
>100
72.3
37.7
30.5
29.1
2f
1g

Table 2. Growth inhibition activity of compounds 1a, 1e, 1k, and
1l on four different human cancer cell lines.
5.
(a) Fabbro, D.; Ruetz, S.; Buchdunger, E.; Cowan-Jacob, S. W.;
Fendrich, G.; Liebetanz, J.; Mestan, J.; O’Reilly, T.; Traxler, P.;
Chaudhuri, B.; Fretz, H.; Zimmermann, J.; Meyer, T.; Caravatti,
G.; Furet, P.; Manley, P. W. Pharmacol. Ther. 2002, 93, 79. (b)
Tavares, F. X.; Boucheron, J. A.; Dickerson, S. H.; Griffin, R. J.;
Preugschat, F.; Thomson, S. A.; Wang, T. Y.; Zhou, H.-Q. J. Med.
Chem. 2004, 47, 4716. (c) Selvam, T. P.; James, C. R.; Dniandev,
P. V.; Valzita, S. K. Res. Pharm. 2012, 2, 1.
Reviews: (a) Winum, J.-Y.; Scozzafava, A.; Montero, J.-L.;
Supuran, C. T. Med Res Rev. 2006, 26, 767. (b) Reitz, A. B.;
Smith, G. R.; Parker, M. H. Expert Opin Ther Pat. 2009, 19, 1449.
For selected recent examples of bioactive sulfamides, see (a) Dou,
D.; Tiew, K.-C.; He, G.; Mandadapu, S. R.; Aravapalli, S.;
Alliston, K. R.; Kim, Y.; Chang, K.-O.; Groutas, W. C. Bioorg.
Med. Chem. 2011, 19, 5975. (b) Suthagar, K.; Watson, A. J. A.;
Wilkinson, B. L.; Fairbanks, A. J. Eur. J. Med. Chem. 2015, 102,
153. (c) Villalba, M. L.; Enrique, A. V.; Higgs, J.; Castaño, R. A.;
Goicoechea, S.; Taborda, F. D.; Gavernet, L.; Lick, I. D.; Marder,
M.; Bruno Blanch, L. E. Eur. J. Pharmacol. 2016, 774, 55. (d)
Villalba, M. L.; Palestro, P.; Ceruso, M.; Gonzales Funes, J. L.;
Talevi, A.; Bruno Blanch, L.; Supuran, C. T.; Gavernet, L. Bioorg.
Med. Chem. 2016, 24, 894.
GI50 (M)a
Compd
MDA-MB-231
>100
MDA-MB-468
>100
SKBR3
>100
>100
>100
>100
T47D
24.1
1a
1e
1k
1l
>100
>100
24.5
>100
>100
>100
65.5
>100
>100
6.
7.
a Values are means of three experiments and the deviation from the mean
is <10% of the mean value.
Finally, we evaluated the toxicity of the MCF7 selective
compounds against HaCat (human spontaneously transformed
aneuploid immortal keratinocytes) and HDFn (normal human
dermal fibroblasts-neonatal) cells by measuring their growth
inhibition activity. All the compounds showed no effect, i.e., GI50
>100 M, for HaCat. For HDFn, compounds 1a and 1k had no
influence (GI50 >100 M) whereas compound 1e and 1l exhibited
limited inhibition with GI50 values of 77 M and 93 M
respectively.
In summary, the synthesis of two series of novel substituted
pyrimidine derivatives bearing a sulfamide group (1 and 2) have
been described and their in vitro cancer growth inhibition
activities have been evaluated against three human tumour cell
lines (HT-29, M21, and MCF7). In general, growth inhibition
activity has been enhanced by the introduction of a bulky
substituent on the aromatic ring with the best compound (2n)
having GI50 < 6 μM for all the human tumour cell lines. In
addition, the results revealed that a shorter linker from the
piperidine to the sulfamide group was beneficial for selective
activity. The MCF7 selective compounds were evaluated on four
additional human invasive breast ductal carcinoma cell lines
(MDA-MB-231, MDA-MB-468, SKBR3, and T47D) and were
selective against T47D cell line in all cases except one,
suggesting a potential antiestrogen activity, which may help to
address the problem of cancer therapy resistance.
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9.
Shayhidin, E. E.; Forcellini, E.; Boulanger, M.-C.; Mahmut, A.;
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Acknowledgments
This work was supported by the Natural Sciences and
Engineering Research Council of Canada, the Fonds de recherche
sur la nature et les technologies (FRQNT), FRQNT Research
Network on Protein Function, Structure and Engineering
(PROTEO) and the Université Laval.
Supplementary Material
Supplementary data
(experimental procedures
and
spectroscopic characterizations of the compounds) associated
with this article can be found, in the online version, at
http://dx.doi.org/10.1016/...
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