Solvent effect on copper-catalyzed azide-alkyne cycloaddition (CuAAC): Synthesis of novel triazolyl substituted quinolines as potential anticancer agents

Article abstract of DOI:10.1016/j.bmcl.2012.03.091

A regioselective route to novel mono triazolyl substituted quinolines has been developed via copper-catalyzed azide-alkyne cycloaddition (CuAAC) of 2,4-diazidoquinoline with terminal alkynes in DMF. The reaction provided bis triazolyl substituted quinolines when performed in water in the presence of Et₃N. A number of the compounds synthesized showed promising anti-proliferative properties when tested in vitro especially against breast cancer cells.

Full text of DOI:10.1016/j.bmcl.2012.03.091

Bioorganic & Medicinal Chemistry Letters 22 (2012) 3455–3459
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Bioorganic & Medicinal Chemistry Letters
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Solvent effect on copper-catalyzed azide–alkyne cycloaddition (CuAAC):
Synthesis of novel triazolyl substituted quinolines as potential anticancer agents
Amarender Reddy Ellanki a,e, Aminul Islam a, Veera Swamy Rama a, Ranga Prasad Pulipati a,
D. Rambabu b,c, G. Rama Krishna d, C. Malla Reddy d, K. Mukkanti e, G.R. Vanaja f, Arunasree M. Kalle f,
K. Shiva Kumar b, Manojit Pal b,
⇑
a Chemistry-Technology Development Center, Dr. Reddy’s Laboratories Limited, Bollaram Road, Miyapur, Hyderabad 500049, Andhra Pradesh, India
b Institute of Life Sciences, University of Hyderabad Campus, Gachibowli, Hyderabad 500046, India
c Department of Chemistry, K. L. University, Vaddeswaram, Guntur 522502, Andhra Pradesh, India
d Department of Chemical Sciences, Indian Institute of Science Education and Research, Kolkata, West Bengal 741252, India
e Chemistry Division, Institute of Science and Technology, JNT University, Kukutpally, Hyderabad 500072, Andhra Pradesh, India
f Department of Animal Sciences, School of Life Sciences, University of Hyderabad, Hyderabad 500046, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A regioselective route to novel mono triazolyl substituted quinolines has been developed via copper-cat-
alyzed azide–alkyne cycloaddition (CuAAC) of 2,4-diazidoquinoline with terminal alkynes in DMF. The
reaction provided bis triazolyl substituted quinolines when performed in water in the presence of
Et3N. A number of the compounds synthesized showed promising anti-proliferative properties when
tested in vitro especially against breast cancer cells.
Received 5 February 2012
Revised 19 March 2012
Accepted 26 March 2012
Available online 30 March 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
Click chemistry
Azide
Alkyne
Quinoline
Cancer
Nitrogen hetrocycles play a central role both in medicinal
chemistry and drug discovery. The development of newer syn-
thetic methods useful for the assembly of molecules containing
nitrogen heterocycles therefore has attracted particular attention
both in academia and industrial organizations. Quinoline deriva-
tives that belong to this class have shown wide applications as
drugs and pharmaceuticals1–6 in addition to their widespread
occurrences in natural products for example, Camptothecin2 and
Luotonin A.3 Quinoline fused with tetrazole has also exhibited
notable pharmacological properties for example, 4-(1H-tetrazol-
5-yl)tetrazolo[1,5-a]quinoline (A, Fig. 1) and 4-argio-6-(tetrazol-
o[1,5-a]quinolin-4-yl)-5,6-dihydropyrimidin-2(1H)-one has shown
antiallergic7a and anti-inflammatory activities,7b respectively. The
4-triazolyl quinoline derivatives on the other hand have been re-
ported as inhibitors of human tumor cells growth.8 Due to our
longstanding interest in the synthesis of quinoline derivatives9 of
potential pharmacological significance we choose to prepare novel
triazolyl substituted quinoline derivatives for assessing their anti-
cancer properties in vitro. Herein we report our initial findings on
R
R
N
N
N
N
N
N
H
N
H
N
N
N
N
N
H
N
N
N
N
N
N
R
N
N
N
A
B
C
Figure 1. General structure of proposed small molecule library (B and C) derived
from A.
N3
N3
N
N
N
N
N
N3
E
D
⇑
Corresponding author. Tel.: +91 40 6657 1500; fax: +91 40 6657 1581.
E-mail address: manojitpal@rediffmail.com (M. Pal).
Scheme 1. Tautomeric forms of 2,4-diazidoquinoline.
0960-894X/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2012.03.091

3456
A. R. Ellanki et al. / Bioorg. Med. Chem. Lett. 22 (2012) 3455–3459
Table 1
Optimization of the reaction conditionsa
N
N
N
N
N
N
N
N3
N
CuI
+
N
+
Solvent
N
N
N
N
N
N3
N
2a
1
3a
4a
Entry
Cu catalyst
Solvent
Time (h)
Yield (%)
3a
4a
1
2
3
4
5
6
CuI
CuI
CuI
CuI
CuI
DMSO
DMF
DMF–H2O (1:1)
H2O
H2O
12
10
10
12
12
20
75
85
50
n.d.
n.d.
n.d.
10
n.d.
30
60b
89b,c
n.d.
Sodium ascorbate–CuSO4Á5H2O
t-BuOH/H2O (1:1)
n.d. = Not detected.
a
All the reactions were carried out by using compound 1 (1.0 equiv), hept-1-yne 2a (1.5 equiv), and CuI (0.01 equiv) at 60–70 °C.
The reaction was carried out using 3.0 equiv of Et3N.
The reaction was carried out using 3.0 equiv of 2a.
b
c
the synthesis and pharmacological evaluation of a series of com-
initially to establish the optimized reaction conditions and the
results are summarized in Table 1. When the compound 1
(1.0 equiv) and alkyne 2a (1.5 equiv) were reacted in DMSO in
the presence of CuI at 60–70 °C for 12 h a mixture of products that
is, mono triazolyl substituted compound (3a) as major and bis
triazolyl substituted compound (4a) as minor product were ob-
tained (Table 1, entry 1). However, changing the solvent to DMF in-
creased the yield of 3a to 85% (Table 1, entry 2) and suppressed the
formation of 4a. While the use of DMF–H2O provided almost 1:1
mixture of 3a and 4a (Table 1, entry 3) the compound 4a was iso-
lated as the only product in water in the presence of Et3N (Table 1,
entry 4). The yield of 4a was improved when 3.0 equiv of alkyne 2a
was used (Table 1, entry 5). These observations suggested that the
compound 1 existed predominantly in form E (Scheme 1) in DMF
leading to the mono triazolyl substituted product 3a. The protic
solvent water in presence of Et3N favored the form D (perhaps
via H-bonding) thereby allowing compound 1 to yield bis triazolyl
substituted product 4a. Nevertheless, while CuI was used as a cat-
alyst in all these reactions the use of other catalysts for example,
the combination of sodium ascorbate–CuSO4Á5H2O was also exam-
ined in t-BuOH/H2O (1:1) but found to be ineffective under the
condition employed (Table 1, entry 6). Additionally, the CuI medi-
ated reaction in H2O/tBuOH (1:1) provided a mixture of mono-tri-
azole 3a and bis-triazole 4a. Thus, the best reaction condition for
the preparation of 3a without generating any side products was
found to be CuI in DMF at 60–70 °C.
Having the optimized reaction condition for the preparation of
mono triazolyl substituted compound in hand we then examined
the generality and scope of this methodology. Thus a variety of ter-
minal alkynes (2) were reacted with 2,4-diazidoquinoline (1) in the
presence of CuI in DMF at 60–70 °C and results are presented in Ta-
ble 2. The reaction proceeded well in all these cases and the sub-
stituents like aryl, alkyl, hydroxy alkyl or chloro present in the
alkyne were well tolerated. The formation of no side product was
observed and the desired product 3 was isolated in good yield in
each case. All the compounds synthesized were well characterized
by spectral (NMR, MS and IR) data. Additionally, the molecular
pound B and C related to A as potential anti-cancer agents
(Fig. 1). The substituent ‘R’ was introduced to generate a library
of small molecules.
Since the report of click chemistry10a by Kolb, Finn, and Sharp-
less in 2001 there has been an explosive growth in this area of
chemistry. The copper-catalyzed azide–alkyne cycloaddition
(CuAAC)10b,c on the other hand represents the best paradigm of
click chemistry. Traditionally, the construction of triazole ring via
Cu(I)-catalyzed 1,3-cycloaddition of an azide to a terminal alkyne
generally involves the use of a mixture of CuSO4 and sodium ascor-
bate as a precatalyst system which generates the Cu(I) species
in situ in the reaction mixture. Recently, the use of a number of
alternative Cu-salts for the generation of Cu(I) species in situ have
been reported which include Cu(OTf)2,11 Cu2O,12 CuBr,13 [(ICy)2-
Cu]PF6,14 tris(triazolyl)methanol-Cu(I) structure15 etc. Although
all these approaches have their own merits and synthetic values
some of them however suffer from few disadvantages for example,
the use of expensive catalysts and ligands and utilization of harsh
reaction conditions. Moreover, the microwave assisted synthesis of
3-triazolyl-2(1H)-quinoline and 4-triazolyl-2(1H)-quinolines that
involved the 1,3-dipolar cycloaddition of organic azides and termi-
nal acetylene in the presence of CuSO4Á5H2O, sodium ascorbate in
DMF at 110 °C for 30 min has been reported.16a While the use of
CuI in click chemistry has been explored earlier its use in the prep-
aration of triazolyl substituted quinoline derivatives has not been
reported.16b Herein we report CuI-mediated efficient and mild syn-
thesis of novel triazolyl substituted quinoline derivatives without
using ascorbic acid or its salt.
For the generation of quinoline based small molecule library
2,4-diazidoquinoline was chosen as the starting compound that
is known to exist in two tautomeric forms for example, 2,4-diazi-
doquinoline (D) and 5-azidotetrazolo[1,5-a]quinoline (E) (Scheme
1). The equilibrium between these two tautomeric forms17–19
depend on several factors, such as the nature of substituents pres-
ent,17 solvent,18 and temperature.17 Nevertheless, the reaction of
2,4-diazidoquinoline19 (1) with hept-1-yne (2a), was examined

A. R. Ellanki et al. / Bioorg. Med. Chem. Lett. 22 (2012) 3455–3459
3457
Table 2
Table 2 (continued)
Synthesis of triazolyl substituted tetrazolo quinoline (3) via Cu mediated click
Entry
Alkynes (2)
Productsb (3)
Yieldc (%)
reaction between 2,4-diazidoquinoline and different alkynesa
OH
R
N
HO
N
N
N
2f
N
N3
N3
N
N
CuI, DMF
6
86
85
80
R
+
60-70 °C
8-10 h
N
N
N
N
N
N3
N
N
N
N
N
N
N
2
3
1
3f
Entry
Alkynes (2)
Productsb (3)
Yieldc (%)
OH
(CH2)4CH3
N
HO
N
N
2a
2g
N
N
N
N
7
1
85
N
N
N
N
N
N
N
3a
3g
Cl
N
N
Cl
N
2h
2b
N
N
N
8
N
2
82
N
N
N
N
N
N
N
3h
3b
(CH2)3CH3
N
N
2i
N
N
2c
9
85
N
N
3
88
N
N
N
N
3i
N
N
N
N
(CH2)5CH3
N
N
2j
3c
N
N
10
87
2d
N
N
N
N
N
4
83
N
3j
All the reactions were carried out by using compound 1 (1.0 equiv), terminal
a
alkyne 2 (1.5 equiv), and CuI (0.01 equiv) at 60–70 °C.
b
Identified by 1H NMR, IR and MS.
Isolated yields.
N
N
N
N
c
3d
structure of a representative compound 3d was established unam-
biguously by single crystal X-ray diffraction (Fig. 2)20 which con-
firmed the presence of a fused tetrazole moiety.
OH
N
N
HO
2e
N
N
Since bis triazolyl substituted compound (4a) was obtained
exclusively when the reaction was performed in water in the pres-
ence of Et3N and 3.0 equiv of terminal alkyne (Table 1, entry 5)
hence we examined the further scope of this methodology. Thus
a number of unprecedented bis triazolyl derivatives21 (4) were pre-
pared in good yields by using this methodology (Table 3). While
the role of Et3N in the present reaction was not clearly understood,
5
78
N
N
N
3e

3458
A. R. Ellanki et al. / Bioorg. Med. Chem. Lett. 22 (2012) 3455–3459
Figure 2. ORTEP representation of the compound 3d (Thermal ellipsoids are drawn at 50% probability level).
the formation of bis-triazolyl derivatives however seemed to
proceed via stepwise formation of mono-triazole ring initially fol-
lowed by the bis-triazole ring. Once formed the mono-triazolyl
derivative would be less soluble in water due to the presence of
hydrophobic side chain. However, the presence of Et3N perhaps in-
creased its solubility in water thereby facilitating the second step.
Thus, the role of Et3N is possibly twofold (i) it helped in generating
the Cu-acetylide intermediate which is well known in the litera-
ture (ii) it increased miscibility of the reactants with water.
Many of the triazolyl substituted tetrazolo quinolines (3) syn-
thesized were tested for their anticancer properties in vitro. Cancer
is the second leading cause of death worldwide after cardiovascu-
lar diseases, according to WHO.22 We were particularly interested
in evaluating our compounds against leukemia (a type of cancer of
the blood or bone marrow characterized by an abnormal increase
of white blood cells) and breast cancer due to the growing inci-
dences of these types of cancers all over the world. The test com-
pounds were examined at two different concentrations in a MTT
(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)
assay23 and results are summarized in Table 4. Harmine, a member
of b-carboline family of compounds showed cytotoxicity against
HL60 and K562 cell lines24 was used as a reference compound.
While most of the compounds showed reasonable inhibition of hu-
studies clearly suggest that triazolyl substituted tetrazolo quino-
line moiety could be an attractive template for the identification
of novel and potential anticancer agents. Since breast cancer
caused 458,503 deaths worldwide (13.7% of cancer deaths in wo-
men) in 2008 hence there is a need for the identification of new
Table 3
Synthesis of bis triazolyl substituted quinoline (4) via CuI-mediated click reaction
between 2,4-diazidoquinoline (1) and different alkynesa
R
N
N
N
CuI, H2O, Et3N
2
1
+
60-70 °C
8-10 h
N
N
R
N
N
4
Entry
Alkynes (2)
Productsb (4)
(CH2)4CH3
Yieldc (%)
N
2a
N
man chronic myeloid leukemia cell growth at 10
tries 1–8) the compound 3g however was found to be active (ꢀ47%
at 1 M) when tested at lower concentrations (Table 4, entry 5). On
lM (Table 4, en-
N
N
l
1
89
the other hand most of the tested compounds were found to be ac-
tive against breast cancer cell lines across all the concentrations
N
N
N
and 3h was found to be the most potent (ꢀ60% at 1
them. In a dose response study, when tested at 10, 1, 0.5, 0.1 and
0.01 M compound 3g showed 52%, 47%, 30%, 22% and 9% inhibi-
tion against K562 cells (IC50 ꢀ4.6 M) whereas 70%, 60%, 46%,
24% and 11% inhibition was observed for compound 3h against
MDA-MB231 cells (IC50 ꢀ0.8 M). Notably, Harmine showed IC50
value of 45 and 54 M when tested against K562 and MDA-
MB231 cells respectively. Thus, compound 3g and 3h were found
to be promising. In general, an aryl side chain directly attached
to the triazole moiety of 3 (e.g. 3b and 3c vs 3d) was found to be
marginally less potent than others whereas a haloalkyl or hydroxy
alkyl long chain (e.g. 3g and 3h) was found to be superior than a
simple alkyl group (e.g. 3i and 3j). We also examined the bis triaz-
olyl substituted quinolines (4) for their potential cytotoxicities
against K562 and MDA-MB231 cells. However, none of them
lM) among
(CH2)4CH3
Cl
4a
l
l
N
N
Cl
l
2h
N
N
l
2
78
N
N
N
Cl
4b
showed inhibition >30% when tested at 10 lM. Nevertheless, our

A. R. Ellanki et al. / Bioorg. Med. Chem. Lett. 22 (2012) 3455–3459
3459
Table 3 (continued)
Entry Alkynes (2)
Supplementary data
Productsb (4)
(CH2)3CH3
Yieldc (%)
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2012.03.091.
N
N
2i
N
N
References and notes
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91
N
N
N
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(CH2)3CH3
4c
(CH2)5CH3
N
N
2j
N
N
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a
All the reactions were carried out by using compound 1 (1.0 equiv), terminal
alkyne 2 (1.5 equiv), Et3N (3.0 equiv) and CuI (0.01 equiv) at 60–70 °C.
b
Identified by 1H NMR, IR and MS.
Isolated yields.
c
Table 4
The %inhibition of growth of cancer cell lines by triazolyl substituted tetrazolo
quinoline derivatives (3)a
Entry
Compounds
K562 (leukemia)
10
MDA-MB231 (breast)
10
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l
M
1
l
M
lM
1 lM
1
2
3
4
5
6
7
8
3b
3c
3d
3e
3g
3h
3i
29.9
44.1
38.1
36.9
52.4
35.2
34.1
37.4
20.6
20.6
26.7
21.8
47.1
19.9
28.2
19.1
56.0
54.0
60.8
46.8
46.1
70.1
57.1
53.4
52.5
47.7
50.5
45.8
42.0
60.2
55.3
49.7
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a
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agents to treat this deadly disease. The triazolyl substituted tetraz-
olo quinolines presented here therefore have medicinal value.
In conclusion, we have developed a regioselective method lead-
ing to mono triazolyl substituted quinolines via copper-catalyzed
azide–alkyne cycloaddition (CuAAC) of 2,4-diazidoquinoline with
terminal alkynes in DMF. CuI was identified as an effective catalyst
for this purpose and the methodology does not require the use of
ascorbic acid or its salt. The use of water in presence of Et3N pro-
vided bis triazolyl substituted quinolines in good yields. A number
of compounds showed promising anti-proliferative properties
when tested against breast cancer cells in vitro. The triazolyl
substituted tetrazolo quinoline moiety therefore could be an
attractive template for the identification of novel and potential
anticancer agents. The simple and general methodology presented
here would find application in the preparation of diversity based
library of small molecules related to triazolyl substituted quinoline
of potential pharmacological interest.
19. Steinschifter, W.; Stadlhauer, W. J. Prakt. Chem. 1994, 336, 311.
20. Crystal data of 3d: molecular formula = C19H15N7, formula weight = 341.14,
crystal system = Monoclinic, space group = P2(1)/n, a = 12.242(2) Å,
b = 5.554(9)Å, c = 23.671(4) Å, V = 1591.5(4) Å3, T = 296(2) K, Z = 4,
Dc = 1.212 mg m–3
,
l
(Mo-K
a
) = 0.08 mm–1, 14992 reflections measured, 3463
independent reflections, 2917 observed reflections [I > 2.0
r(I)], R1_obs = 0.025,
Goodness of fit = 1.063. Crystallographic data (excluding structure factors) for
3d have been deposited with the Cambridge Crystallographic Data Centre as
supplementary publication number CCDC 851296.
21. For some recent reports on the preparation of bis-triazolyl compounds of
pharmacological interest, see: (a) He, X.-P.; Li, C.; Jin, X.-P.; Song, Z.; Zhang, H.-
L.; Zhu, C.-J.; Shen, Q.; Zhang, W.; Sheng, L.; Shi, X.-X.; Tang, Y.; Li, J.; Chen, G.-
R.; Xie, J. New J. Chem. 2011, 35, 622; (b) He, X.-P.; Li, C.; Wang, Z. Z.; Gao, L. X.;
Shi, X. X.; Tang, Y.; Xie, J.; Li, J.; Chen, G. R.; Chen, K. Glycoconj. J. 2011, 28, 493.
22. Parkin, D. M.; Bray, F.; Ferlay, J.; Pisani, P. Cancer J. Clin. 2005, 55, 74.
23. MTT assay: Cell viability was determined by (3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium bromide (MTT) assay. Cells (5 Â 103 cells/well) were
seeded to 96-well culture plate and cultured with or without compounds at
1 and 10
the medium was removed and 20
fresh medium. After 2 h incubation at 37 °C, 100
l
M concentration for 24 h in a final volume of 200
l of MTT (5 mg/ml in PBS) was added to the
l of DMSO was added to each
ll. After treatment,
l
l
well and plates were agitated for 1 min. Absorbance was read at 570 nm on a
multi-well plate reader (Synergy Mx, Biotek Inc., USA). Percent inhibition of
proliferation was calculated as a fraction of control (without compound) (see
supporting information for further details).
Acknowledgments
Mr. A.R.E. thanks Dr. Vilas Dahanukar for encouragement and
the analytical group of DRL for spectral support.
24. Jahaniani, F.; Ebrahimi, S. A.; Rahbar-Roshandel, N.; Mahmoudian, M.
Phytochemistry 2005, 66, 1581.