1-Toluene-sulfonyl-3-[(3′-hydroxy-5′-substituted)-γ-butyrolactone]-indoles: Synthesis, COX-2 inhibition and anti-cancer activities

Article abstract of DOI:10.1016/j.bmcl.2007.11.010

Indoles carrying a cyclic ester (γ-butyrolactone) at C-3 position have been synthesized by the allylation of 3-indoleglyoxylate followed by iodocyclisation and the nucleophilic replacement of the iodo-group. Screening of these molecules for COX-2 inhibiti

Full text of DOI:10.1016/j.bmcl.2007.11.010

Available online at www.sciencedirect.com
Bioorganic & Medicinal Chemistry Letters 18 (2008) 85–89
1-Toluene-sulfonyl-3-[(30-hydroxy-50-substituted)-c-butyrolactone]-
indoles: Synthesis, COX-2 inhibition and anti-cancer activities
Palwinder Singh,a,* Anu Mittal,a Atul Bhardwaj,a
Satwinderjeet Kaurb and Subodh Kumara
aDepartment of Chemistry, Guru Nanak Dev University, Amritsar 143005, India
bDepartment of Botanical & Environmental Sciences, Guru Nanak Dev University, Amritsar 143005, India
Received 16 September 2007; revised 12 October 2007; accepted 2 November 2007
Available online 6 November 2007
Abstract—Indoles carrying a cyclic ester (c-butyrolactone) at C-3 position have been synthesized by the allylation of 3-indolegly-
oxylate followed by iodocyclisation and the nucleophilic replacement of the iodo-group. Screening of these molecules for COX-2
inhibition and anti-cancer activities has identified compounds 10 and 11 as highly potent and selective for COX-2 as well as showing
remarkable anti-cancer activities (better than that of indomethacin).
Ó 2007 Elsevier Ltd. All rights reserved.
The critical role of indole moiety in indole acetic acid1 (a
plant growth regulator hormone), in tryptophan1 (an
amino acid) and as a part of a number of alkaloids2
has subjected it to several derivatisations for procuring
medicinally important molecules. The development of
indomethacin3 (1) as a non-steroidal anti-inflammatory
drug (NSAID) (non-selective COX-2 inhibitor) has been
followed by the modifications in the different parts of
the molecule, especially in the C-3 substituent, for
increasing its efficacy4 as well as for transforming it into
selective COX-2 inhibitors (1, n = 2, 3, branched alkyl
groups, R = NH2, alkyl).5
The synthesis of the target molecules has been achieved
by the indium mediated diastereoselective allylation of
3-indoleglyoxylate, followed by diastereoselective iodo-
cyclisation7 and nucleophilic replacement of the iodo-
group. Commercially available 3-indoleglyoxylate (3)
was N-alkylated by reaction with p-toluene-sulfonyl
chloride to get indole substituted a-keto ester 4 (Scheme
1). A solution of 4, allyl bromide and indium metal (sus-
pension) (1:1.5:1) in THF–H2O (2:1) on stirring at 0 °C
for 6–8 h, after usual workup and chromatography, pro-
vided compound 5 (87%) as a thick liquid [M+ m/z 399]
(Scheme 1). The participation of Cram’s chelation mod-
el in this reaction explains the diastereoselective forma-
tion of compound 5.7
In our ongoing research programme for developing
COX-2 inhibitors, it has been planned to introduce a
c-butyrolactone at C-3 and a tolylsulfonyl group at N-
1 of indole (2). The c-butyrolactone moiety (cyclic ester)
mimics the furanone moiety present in rofecoxib6 (a
selective COX-2 inhibitor), while tolylsulfonyl is a key
pharmacophore in most of the COX-2 inhibitors. The
rationally designed molecules (2) have been synthesized
and evaluated for their COX-1, COX-2 inhibitory activ-
ities and also screened for anti-cancer properties over 59
human tumour cell lines (Fig. 1).
Stirring of solution of 5 in C2H5OH–THF (2:1) with 2%
NaOH for 2 h, after acidification and extraction with
OR
HO
( )n
X
H3CO
O
CH3
O
N
O
N
O
S
O
O
Cl
1, Indomethacin; n=1, R=H
H3C
Keywords: Indole; Butyrolactone; Synthesis; COX-2 inhibition; Doc-
kings; Anti-cancer activities.
2
*
Figure 1.
Corresponding author. E-mail: palwinder_singh_2000@yahoo.com
0960-894X/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.11.010

86
P. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 85–89
O
H
H
X
HO
HO
OCH3
CH2I
CH2
O
O
O
O
OCH3
O
O
N
S
N
S
N
S
O
a, b, c
a
O
O
O
O
O
O
N
H
CH3
4
3
b
CH3
CH3
7
9-11 9, X = SC2H5
10, X = CN
OH
HO
N
11, X = SCN
OH
O
CH3
Scheme 3. Reagents and conditions: (a) ethanthiol, CH3CN, K2CO3,
stir, rt (for 9); (b) NaCN, CH3CN, stir, rt (for 10); (c) KSCN, CH3CN,
stir, rt (for 11).
O
O
c
N
O
S
O
O
O
S
groups like CH2SC2H5, CH2SCN, and CH2CN at C-50
of butyrolactone ring. Nucleophilic substitutions of 7
with ethanthiol, NaCN and KSCN in CH3CN gave
compounds 9, 10 and 11 possessing, respectively,
SC2H5, CN and SCN groups in place of iodine in 7
(Scheme 3).
CH3
6 (70%)
CH3
5 (87 %)
Scheme 1. Reagents and conditions: (a) p-tolunenesulfonyl chloride,
NaH, CH3CN; (b) allyl bromide, In, THF/H2O (2:1), 0 °C, stir; (c)
ethanol–THF, 2% NaOH, stir.
Compounds 7, 9, 10 and 11 have been evaluated for
in-vitro COX-1, COX-2 inhibitory activities using
COX (ovine) inhibitor screening assay kit (Cayman
Chemicals, cat. No. 560101). The results of these
investigations are given in Table 1. Out of these four
compounds, 10 and 11 with, respectively, CH2CN
and CH2SCN groups at C-50 show the best results
with 90% and 85% inhibition of COX-2 at 10ꢀ8 M
concentration and IC50 values <0.01 lM. 60 and
65% COX-1 inhibitions at 10ꢀ5 M concentrations have
been observed for compounds 10 and 11, respectively.
Significantly, the results of these two compounds are
better than that of indomethacin and rofecoxib. The
COX-2 inhibition by compound 9 is also considerable
(65% at 10ꢀ8 M) while compound 7 exhibits poor inhi-
bition. The results of these investigations indicate that
along with other components, the C-50 substituent sig-
nificantly controls the COX-2 inhibitory activities of
these compounds.
ethyl acetate, gave the corresponding acid 6 (70%)
[M+ꢀOH m/z 368] (Scheme 1). Compound 6 on treat-
ment with I2 (3 equiv) and NaHCO3 (3 equiv) in dry
CH3CN for 3–4 h after usual workup gave a mixture
of two diastereomers in the ratio 3:1 (1H NMR integra-
tion) (84%) (Scheme 2).8 In higher Rf component 7;
61%, mp 130 °C; the observation of NOE between C-
50H and aromatic protons and no NOE between CH2I
unit and aromatic protons indicates that C-50H and in-
dole moiety are placed on the same face of furanone and
helps in assigning stereochemistry 30S*, 50R* at the two
chiral centers. In case of lower Rf component 8, 18%,
NOE has been observed at ArHs when CH2I protons
were irradiated and accordingly stereochemistry 30S*,
50S* has been assigned at the two chiral centres. Com-
pound 6 also underwent iodocyclisation with I2 in the
presence of silica gel (for TLC) in ethyl acetate-metha-
nol to give a mixture of 7 and 8 (85%) in the diastereo-
meric ratio 3:1.
In order to investigate the interactions of these com-
pounds in the active site of COX-2 and to rationalize
the role of C-50 substituent towards COX-2 inhibition,
we carried out the dockings of these molecules in the
Based upon the previous results,9 docking studies and
the literature reports,10 it was planned to introduce the
H
H
4'
5'
HO
HO
HO
N
CH2I
CH2I
OH
O
O
3'
1'
2'
O
O
O
N
S
N
S
a/ b
O
O
+
O
O
S
O
O
CH3
CH3
CH3
7
8
6
Scheme 2. Reagents and conditions: (a) I2, NaHCO3, CH3CN, 0 °C, stir; (b) I2, silica, ethyl acetate-methanol, stir, rt.

P. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 85–89
Table 1. In-vitro COX-2 inhibitory activities of indole derivatives
87
Compound
% Inhibition
IC50 (lM)
COX-2 selectivity*
COX-2
COX-1
10ꢀ5 M
COX-2
COX-1
10ꢀ5 M
10ꢀ6 M
10ꢀ7 M
10ꢀ8 M
7
30
65
90
85
32
62
87
90
35
67
88
89
44
75
40
35
>10
>10
>10
<10
<10
0.05
40
9
10
<0.001
<0.001
<0.001
0.75
<1000
>1000
>1000
0.067
95
92
60
65
11
Indomethacin5d
Rofecoxib
97
100
100
75
75
0.3
ꢁ133
* COX-2 selectivity = IC50 (COX-1)/IC50 (COX-2).
active site of COX-2.11 The crystal structure of COX-2
with indomethacin shows the interactions of the car-
boxyl group of indomethacin with the guanidine moiety
of R120, amino acid active during the turnover phase of
the enzyme. Figure 2 validates our programme of dock-
ing, where indomethacin docked in the active site of
COX-2 nearly overlaps with the one present in the crys-
tal structure of the enzyme (rms deviation 0.88).
Docking of compound 10 in the active site of COX-2
indicates that this molecule fits in the COX-2 active site
in the same fashion as indomethacin. In compound 10,
the oxygen of C-30 OH group is present at a distance
˚
of 2.10 A from the H of guanidine moiety of R120
and interacts through H-bond formation while the CN
group present at C-50 position approaches the guanidine
Figure 3. Compound 10 docked in the active site of COX-2. OH and
CN groups present on the lactone ring approach R120 at a distance of
˚
2.1 and 2.43 A, respectively. Hs are omitted for clarity.
˚
moiety at a distance of 2.43 A (Fig. 3). The aromatic
part of the tosyl group present at N-1 of compound 10
fits in the hydrophobic pocket formed by F518, W387
and Y385 residues. While similar types of interactions
are observed during the docking of compound 11 in
the active site of COX-2, compounds 7 and 9 are devoid
of such interactions.
compounds for their anti-cancer activities. The screen-
ing for anti-cancer properties was performed on 59 hu-
man tumour cell lines at NIH, Bethesda, USA, using
the standard procedure.13 Out of the four compounds
(7, 9–11), compounds 10 and 11 with CH2CN and
CH2SCN group at C-50 exhibit remarkable anti-cancer
activities with average GI50 over all the cell lines as 1.9
and 9.1 lM, respectively, which is much better than
the average GI50 value of indomethacin (64.3 lM)14
(Table 2). Compound 10 is highly specific for all the cell
lines of leukaemia, MALME-3M and M14 of mela-
noma, RXF393 of renal cancer, PC-3 of prostate cancer
and MCF7, MDA-MB-435 of breast cancer where it
exhibits GI50 in sub-micromolar concentration (0.1–
0.8 lM). Moreover, the relatively high LC50 values of
10 and 11 show their non-toxicities. However, com-
pounds 7 and 9, investigated at 10ꢀ5 M concentration
only, do not show much anti-cancer activity. Therefore,
in parallel with the COX-2 inhibitory activities, com-
pounds 10 and 11 are also highly effective towards var-
ious cancer cell lines of human tumour cell panels.
The role of COX-2 in the propagation of cancer12 and
the evaluations of several COX-2 inhibitors for reducing
cancer propagation led us to investigate the present
Table 2. Anti-cancer activities of compounds 10 and 11 (average GI50
)
Compound
GI50 (lM)
LC50 (lM)
10
11
1.9
9.1
95
74
Figure 2. Indomethacin docked in the active site of COX-2 exactly
overlaps with the one present in the crystal structure of the enzyme.
Indomethacin
64.3
100

88
P. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 85–89
Table 3. Lipinski values for compounds 7, 9–11
5. After the identification of COX-2 as causing inflamma-
tion during arachidonic acid metabolism (a) Black, W.
C.; Bayly, C.; Belley, M.; Chan, C.-C.; Charleson, S.;
Denis, D.; Gauthier, J. Y.; Gordon, R.; Guay, D.;
Kargman, S.; Lau, C. K.; Leblanc, Y.; Mancini, J.;
Ouellet, M.; Percival, D.; Roy, P.; Skorey, K.; Tagari,
P.; Vickers, P.; Wong, E.; Xu, L.; Prasit, P. Bioorg.
Med. Chem. Lett. 1996, 6, 725; (b) Leblanc, Y.; Black,
W. C.; Chan, C. C.; Charleson, S.; Delorme, D.; Denis,
D.; Gauthier, J. Y.; Grimm, E. L.; Gordon, R.; Guay,
D.; Hamel, P.; Kargman, S.; Lau, C. K.; Mancini, J.;
Ouellet, M.; Percival, D.; Roy, P.; Skorey, K.; Tagari,
P.; Vickers, P.; Wong, E.; Xu, L.; Prasit, P. Bioorg.
Med. Chem. Lett. 1996, 6, 731; (c) Kalgutkar, A. S.;
Crews, B. C.; Rowlinson, S. W.; Marnett, A. B.; Kozak,
K. R.; Remmel, R. P.; Marnett, L. J. Proc. Natl. Acad.
Sci. 2000, 97, 925; (d) Kalgutkar, A. S.; Marnett, A. B.;
Crews, B. C.; Remmel, R. P.; Marnett, L. J. J. Med.
Chem. 2000, 43, 2860; (e) Palomer, A.; Cabre, F.;
Pascual, J.; Campos, J.; Trujillo, M. A.; Entrena, A.;
Gallo, M. A.; Garcia, L.; David, M.; Espinosa, A.
J. Med. Chem. 2002, 45, 1402; (f) Olgen, S.; Nebioglu,
D. Il Farmaco 2002, 57, 677; (g) Kalgutkar, A. S.;
Crews, B. C.; Saleh, S.; Prudhomme, D.; Marnett, L. J.
Bioorg. Med. Chem. 2005, 13, 6810; (h) Khanna, S.;
Madan, M.; Vangoori, A.; Banerjee, R.; Thaimattam,
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M. Bioorg. Med. Chem. 2006, 14, 4820.
2
TPSA (A )
˚
Compound
logP
nON
nOHNH
7
9
2.49
2.38
0.94
1.78
85.6
85.6
6
6
7
7
1
1
1
1
10
11
109.4
109.4
Calculations of Lipinski values of these compounds
(Table 3)15 indicate a difference in log P and Total Polar
Surface Area (TPSA) of compounds 10 and 11 from 7
and 9 which might be contributing towards the differ-
ence in the bioactivities of these compounds.
In conclusion, we have constructed a c-lactone moiety at
C-3 of indole by the allylation of 3-indoleglyoxylate fol-
lowed by iodocyclisation. Replacement of iodo-group
with nucleophiles like CN, SCN, SC2H5 and their inves-
tigations for COX-2 inhibition have identified com-
pounds 10 and 11 as highly potent and selective for
COX-2. Along with COX-2 inhibition, the remarkable
anti-cancer activities of 10 and 11 in comparison to
indomethacin enable them to be used as leads for further
investigations and also support the design of these
molecules.
6. Prasit, P.; Wang, Z.; Brideau, C.; Chan, C.-C.; Charleson,
S.; Cromlish, W.; Ethier, D.; Evans, J. F.; Ford-Hutch-
inson, A. W.; Gauthier, J. Y.; Gordon, R.; Guay, J.;
Gresser, M.; Kargman, S.; Kennedy, B.; Leblanc, Y.;
Leger, S.; Mancini, J.; O’Neill, G. P.; Ouellet, M.;
Percival, M. D.; Perrier, H.; Riendeau, D.; Rodger, I.;
Tagari, P.; Therien, M.; Vickers, P.; Wong, E.; Xu, L.-J.;
Young, R. N.; Zamboni, R.; Boyce, S.; Rupniak, N.;
Forrest, M.; Visco, D.; Patrick, D. Bioorg. Med. Chem.
Lett. 1999, 9, 1773.
Acknowledgments
The financial assistances by DST, New Delhi, UGC,
New Delhi, and CSIR, New Delhi, have been gratefully
acknowledged. Anu thanks CSIR, New Delhi, for SRF.
A.B. thanks DST, New Delhi, for JRF. Authors are also
thankful to Dr. V.L. Narayanan and his team at NCI,
NIH, Bethesda, USA, for screening anti-cancer
activities.
7. (a) Kaur, P.; Singh, P.; Kumar, S. Tetrahedron 2005, 61,
8231; (b) Singh, P.; Mittal, A.; Kaur, P.; Kumar, S.
Tetrahedron 2006, 62, 1063.
8. Selected data for compound 7. 1H NMR (300 MHz, CDCl3)
d 2.36 (s, 3H, CH3), 2.53 (dd, 1H, 2J = 13.2 Hz, 3J = 9 Hz,
1H of 40-H), 3.04 (dd, 1H, 2J = 13.2 Hz, 3J = 5.4 Hz, 1H of
40-H), 3.14 (br s 1H, OH, exchanges with D2O), 3.37 (dd,
References and notes
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1H, 2J = 10.2 Hz, J = 7.5 Hz, 1H of CH2I), 3.48 (dd, 1H,
2J = 10.2 Hz, 3J = 5.1 Hz, 1H of CH2I), 4.29–4.38 (m, 1H,
50-H), 7.25–7.32 (m, 3H, ArH), 7.39 (t, 1H, J = 7.5 Hz,
ArH), 7.53 (s, 1H, ArH), 7.78 (t, 3H, J = 8.1 Hz, ArH), 8.01
(d, 1H, J = 8.7 Hz, ArH). The multiple signals at d 4.29–4.38
on decoupling converted all the four double doublets into
doublets. MS (FAB) 511 [M+].
1
Compound 8. H NMR (300 MHz, CDCl3) d 1.60 (br s,
1H, OH, exchanges with D2O), 2.32 (dd, 1H, 2J = 14.1 Hz,
3J = 8.1 Hz, 1H of 40-H), 2.36 (s, 3H, CH3), 3.04 (dd, 1H,
2J = 14.1 Hz, 1H of 40-H), 3.26 (dd, 1H, 2J = 10.5 Hz,
3J = 7.5 Hz, 1H of CH2I), 3.41 (dd, 1H, 2J = 10.5 Hz,
3J = 4.5 Hz, 1H of CH2I), 4.77–4.86 (m, 1H, 50-H), 7.24–
7.30 (m, 3H, ArH), 7.38 (d, 1H, J = 8.4 Hz), 7.65 (d, 1H,
J = 7.8 Hz, ArH), 7.76 (s, 1H, ArH), 7.81 (d, 2H,
J = 8.4 Hz, ArH), 8.02 (d, 1H, J = 8.1 Hz, ArH). MS
(FAB) 511 (M+).
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11. Crystal structure of COX-2 with indomethacin in the
active site was downloaded from protein data bank (pdb

P. Singh et al. / Bioorg. Med. Chem. Lett. 18 (2008) 85–89
89
ID 4COX) and refined as per the usual procedure. The
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15. Lipinski values were calculated using molinspiration.