Synthesis of novel 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine derivatives and their cytotoxic activity

Article abstract of DOI:10.1016/j.bmcl.2013.12.107

A series of novel 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine derivatives 3 and 4 were prepared respectively starting from 6-phenyl-4-(trifluoromethyl)- 1H-pyrazolo[3,4-b]pyridin-3-amine 1 via selective N-propargylation, followed by reaction with divers

Full text of DOI:10.1016/j.bmcl.2013.12.107

Bioorganic & Medicinal Chemistry Letters 24 (2014) 746–749
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Bioorganic & Medicinal Chemistry Letters
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Synthesis of novel 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine
derivatives and their cytotoxic activity
Chavva Kurumurthy a, Banda Veeraswamy a, Pillalamarri Sambasiva Rao a, Gautham Santhosh Kumar a,
Pamulaparthy Shanthan Rao a, Velaturu Loka Reddy b, Janapala Venkateswara Rao b, Banda Narsaiah a,
⇑
a Fluoro Organic Division, CSIR-Indian Institute of Chemical Technology, Hyderabad 500 607, India
b Biology division, CSIR-Indian Institute of Chemical Technology, Tarnaka, Hyderabad 500 607, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine derivatives 3 and 4 were prepared respec-
tively starting from 6-phenyl-4-(trifluoromethyl)-1H-pyrazolo[3,4-b]pyridin-3-amine 1 via selective N-
propargylation, followed by reaction with diverse substituted alkyl/perfluoroalkyl/aryl/aryl amide azides
under Sharpless conditions. All the synthesized compounds 3 and 4 were screened for cytotoxic activity
against four human cancer cell lines such as U937, THP-1, HL60 and B16-F10. Compounds 3e, 4g, 4i and 4j
which showed promising activity have been identified.
Received 14 August 2013
Revised 4 December 2013
Accepted 26 December 2013
Available online 3 January 2014
Keywords:
Pyrazolo[3,4-b]pyridine
Propargylation
Ó 2013 Elsevier Ltd. All rights reserved.
1,2,3-Triazoles
Sharpless conditions
Cytotoxic activity
According to the statistics, cancer is the second most common
cause of death after heart disease.1 During the past decade, cancer
continued to be a worldwide killer.2 The medicinal chemists have
paid great attention to the development of novel lead molecule
from natural source and the demand is growing towards new ther-
apies. Although chemotherapy is the mainstay for cancer treat-
ment, the use of available chemotherapeutics is often limited due
to undesirable side effects. It is important to identify new agents
and new targets for the treatment of cancer. Pyrazole and pyridine
derivatives attracted organic chemists very much due to their bio-
logical and chemotherapeutic importance.
Pyrazolopyridine and related fused heterocycles are of interest as
potential bioactive molecules.3–5 The pyrazolo[3,4-b]pyridine ring
system has proved to be an interesting class in heterocyclic chemis-
try and some of its derivatives are important as anticancer agents
with low toxicity (Fig. 1),6 GSK-3 inhibitors (Fig. 1),7 as anti-inflam-
matory agents,8 as blood platelet aggregation inhibitors,9 as bone
metabolism improvers,10 as adenosine antagonists11,12 and as con-
trolling herbicides.13 In recent years, 1,2,4-triazine compounds have
been reported to possess biological activities as anti-AIDS,14 anti-
cancer15 and antimicrobial activities.16–18 Similarly, 1,2,3-triazoles
are useful building blocks in organic synthesis because of their wide
range of biological activity,19,20 and are stable to moisture, oxygen,
light and also metabolism in the body. Moreover, these moieties
can be tuned to form powerful pharmacophores and also plays an
important role in bio-conjugation. Introduction of pyrazolo[3,4-
b]pyridine moiety to the triazole core will increase the hydrophilic-
ity of the whole molecule, which promotes biological activity. Based
on the importance, and in continuation of our interest towards the
synthesis of heteroring fusedpyridines as potential molecules,21–23
we initiated our synthetic strategy towards a series of novel 1,2,3-
triazole tagged pyrazolo[3,4-b] pyridine derivatives and screened
for anticancer activity against four cancer cell lines such as U937,
THP-1, HL60 and B16-F10. The promising compounds 3e, 4g, 4i
and 4j have been identified and reporting here for the first time.
The 6-phenyl-4-(trifluoromethyl)-1H-pyrazolo[3,4-b]pyridin-3-
amine 124 was reacted with propargyl bromide in acetone using
potassium carbonate as a base at reflux temperature and obtained
exclusively N-propargylated pyrazolo[3,4-b]pyridin-3-amine 2.
Compound 2 was identified based on IR spectrum in which charac-
teristic absorption bands at 3301, 3325 cmÀ1 for NH2 group and 1H
NMR spectrum in which NH2 appeared as broad signal at d 4.41
ppm, disappearance of –NH peak at d 12.22 ppm and N–CH2– pro-
tons appeared as doublet at d 5.24 ppm. Compound 2 was further
reacted with diverse substituted alkyl/perfluoroalkyl /aryl/aryl
amide azides in THF, using Cu(I) catalyzed [3+2] azide–alkyne
cycloaddition reaction (CuAAC) under Sharpless conditions,25,26
as a result an exclusive formation of 1,4-disubstituted-1,2,3-tria-
zole tagged pyrazolo[3,4-b]pyridine derivatives 3, 4 are formed
respectively. The aryl amide azide-containing building blocks were
synthesized in one-pot procedure27 (Scheme 2), in which different
⇑
Corresponding author. Tel.: +91 40 27193630; fax: +91 40 27160387.
E-mail address: narsaiah@iict.res.in (B. Narsaiah).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.12.107

C. Kurumurthy et al. / Bioorg. Med. Chem. Lett. 24 (2014) 746–749
747
generate the corresponding azides. The sequence of reactions out-
lined in Scheme 1–3 and products are tabulated in Tables 1 and 2.
All the final compounds 3a–q, 4a–j were evaluated for their
anticancer activity against U937 (human monocytic leukemia),
THP-1 (human acute monocytic leukemia), HL-60 (Human promy-
elocytic leukemia) and B16-F10 (Mouse Melanoma) cancer cell
lines using Etoposide as standard. IC50 values of the test com-
pounds for 24 h on each cell line was calculated and presented in
Tables 3 and 4. It is evident from the bioactivity assay results that
most of the test compounds have shown significant cytotoxic
activity on all the cell lines tested in a concentration-dependent
manner. Among all the compounds 3e, 4g, 4i and 4j showed prom-
ising activity against four cancer cell lines and compounds 3p, 4f
exhibited cytotoxic activity against three cell lines (THP1, U937 &
Figure 1. Bio-active compounds on pyrazole[3,4-b]pyridine derivatives.
CF3
CF3
N
NH2
N
NH2
N
i
HL60) at below 100 lg/ml concentration. It is apparent from the
N
N
Br
Ph
N
Ph
results that, these derivatives are found to be more active on leu-
kemia cell lines (THP1, U937 & HL60) than the mouse melanoma
(B16-F10) cell line. Among all the compounds 3e was found to be
H
2
1
more potent with IC50 concentration <30 lg/ml on U937 and
Scheme 1. Synthesis of N-propargylated pyrazole[3,4-b]pyridine. Reagents and
conditions: (i) Acetone, Nal, K2CO3, reflux, 24 h.
HL60 cell lines. Derivatives 3a, 3c, 3i, 4a, 4c, 4e and 4h are active
only on two cell lines (THP-1, U937) and 3k, 3m, 3n, 3q, 4b, 4d
are showed activity only on one cell line (THP-1). Remaining deriv-
atives are not active at 100 lg/ml concentration. Structure activity
Bromo
O
O
relationship studies revealed that the presence of amide (–CONH–)
group in between triazole, phenyl rings and decyltriazole tag in 1st
position of pyrazolo[3,4-b]pyridine derivatives promote high activ-
ity as experienced in compounds 4a, 4b, 4c, 4d, 4e and 3e com-
pared to phenyl ring directly attached to triazole ring against
THP1 cell lines. The activity data shows that the most of the
pyrazolo[3,4-b]pyridine derivatives are effective on THP1 cells fol-
lowed by U937, HL60 and B16-F10 cells. The activity order of
acetyl bromide
NaN3/DMF
N3
R'
Br
R'
R' NH2
N
H
N
H
Scheme 2. Synthesis of aryl amide azide-containing building blocks.
types of amines were reacted with bromoacetyl bromide in N,N-di-
methyl formamide followed by SN2 reaction with sodium azide to
CF3
NH2
N
N
Ph
N
2
O
R
N3
i
N3
CF3
N
i
HN
R'
CF3
N
NH2
N
NH2
N
N
N
Ph
Ph
HN R'
O
4
N
3
R
N
N
N
N
N
4a) = C6H5, 4b) = 4-OCH3 C6H4-,
4c) = 4-F C6H4-, 4d) = 3-Cl C6H4-,
3a) = CH3-(CH2)2-CH2-, 3b) = CH3-(CH2)4-CH2-,
3c) = C6F13-CH2-CH2-, 3d) = C8F17-CH2-CH2-,
3e
3f
) = CH3-(CH2)8-CH2-, ) = CH3-(CH2)6-CH2-,
4e) = 4-CH3C6H4-, 4f) = 3-F C6H4-,
4g) = 2-F,6-F C6H3-, 4h) = 2-Br C6H4-,
3g) = C6H5-, 3h) = 4-OCH3 C6H4-,
4i) = 2-Br, 4-F C6H3-, 4j) = Cyclohexyl
3i) = 3-Cl,4-F C6H3-, 3j) = 3-CF3 C6H4-,
3k
3l
) = 3-Cl C6H4-, ) = 4-F C6H4-,
3m) = 4-NO2 C6H4-, 3n) = 4-CH3 C6H4-,
3o) = 3-F,4-Br C6H3-, 3p) = 3-NO2,4-OH C6H3-,
3q
) = 2-CH3,3-Br C6H3-
Scheme 3. Synthesis of 1,2,3-triazole tagged pyrazole[3,4-b]pyridine derivatives. Reagents and conditions: Dry THF, Cul, rt, 24 h.

748
C. Kurumurthy et al. / Bioorg. Med. Chem. Lett. 24 (2014) 746–749
Table 1
Table 4
Synthesis of 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine derivatives 2 and 3a–q
In vitro cytotoxicity of 4a–j against four cancer cell lines
a
Compound no.
R
Mp (°C)
Yield (%)
Compound no.
IC50 SE
2
3a
3b
3c
3d
3e
3f
3g
3h
3i
3j
3k
3l
3m
3n
3o
3p
3q
—
131–133
143–145
148–150
135–137
146–148
112–114
140–142
211–213
193–195
267–269
217–219
215–217
198–200
230–232
207–209
229–231
132–134
187–189
93
90
89
93
91
87
85
90
92
88
90
86
89
91
87
84
81
83
THP1
U937
HL60
B16-F10
CH3–(CH2)2–CH2–
CH3–(CH2)4–CH2–
C6F13–CH2–CH2–
C8F17–CH2–CH2–
CH3–(CH2)8–CH2–
CH3–(CH2)6–CH2–
C6H5–
4-OCH3 C6H4–
3-Cl,4-F C6H3–
3-CF3 C6H4–
4a
4b
4c
4d
4e
4f
4g
4h
54.64 1.21
56.70 2.76
49.55 0.75
37.02 1.26
41.47 1.46
99.48 2.12
43.27 1.24
95.27 1.61
65.63 1.15
64.83 1.02
6.34 1.09
—
—
—
—
96.34 3.56
—
44.77 1.16
—
—
—
—
—
—
—
65.26 2.19
43.52 1.71
30.63 2.12
—
98.34 3.75
88.99 3.15
6.7 0.32
—
63.92 2.83
32.76 0.87
27.2 0.56
63.65 2.10
42.95 1.88
2.13 0.86
41.57 0.91
—
98.02 2.6
99.12 2.32
3.8 0.23
4i
4j
3-Cl C6H4–
4-F C6H4–
Etoposideb
4-NO2 C6H4–
4-CH3 C6H4–
3-F,4-Br C6H3–
3-NO2,4-OH C6H3–
2-CH3,3-Br C6H3–
Exponentially growing cells were treated with different concentrations of 1,2,3-
triazole tagged pyrazolo[3,4-b]pyridine derivatives for 24 h and cell growth inhi-
bition was analyzed through MTT assay.28
a
IC50 is defined as the concentration, which results in a 50% decrease in cell
number as compared with that of the control cultures in the absence of an inhibitor
and were calculated using the respective regression analysis. The values represent
the mean SE of three individual observations.
b
Etoposide was employed as positive control. — indicates that the derivatives are
not active at 100 lg/ml concentration.
Table 2
Synthesis of N-arylacetamide 1,2,3-triazole tagged pyrazolo[3,4-b]pyridine deriva-
tives 4a–j
Compound no.
R
Mp (°C)
Yield (%)
cytotoxic activity. The bioactivity assay results showed that the
compounds 3e, 4g, 4i and 4j exhibited cytotoxic activity against
four human cancer cell lines. Among all the compounds, 3e is con-
sidered to be more potent. Based on the present results, it is war-
ranted that these derivatives to be further evaluated on other
cancer cell lines. Slight structural modification of these active
derivatives may yield as prospective anti-cancer drugs.
4a
4b
4c
4d
4e
4f
4g
4h
4i
C6H5–
4-OCH3 C6H4–
4-F C6H4–
3-Cl C6H4–
4-CH3 C6H4–
3-F C6H4–
2-F,6-F C6H3–
2-Br C6H4–
2-Br,4-F C6H3–
Cyclohexyl
224–226
206–208
212–214
206–208
199–201
180–182
142–144
208–210
217–219
200–202
91
87
90
89
86
88
85
86
83
89
Acknowledgments
4j
Authors are thankful to the Council of Scientific & Industrial Re-
search, New Delhi, India for the financial support through XII five
year plan project DITSF code: CSC-0204. Aauthors, C. Kurumurthy,
B. Veera Swamy, P. Sambasiva Rao, G. Santhosh Kumar are also
thankful to CSIR, India for providing financial assistance in the form
of Senior Research Fellowship and contingency grant.
Table 3
In vitro cytotoxicity of 3a–q against four cancer cell lines
a
Compound no.
IC50 SE (
l
g/ml)
THP1
U937
HL60
B16-F10
3a
3b
46.07 2.16
—
—
—
36.72 2.76
—
—
—
Supplementary data
3c
—
98.23 2.10
97.56 3.14
—
3d
3e
3f
3g
—
—
—
—
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.12.
107.
48.68 1.25
—
—
27.11 2.12
—
—
29.90 2.12
—
—
84.90 3.12
—
—
3h
—
—
—
—
3i
3j
—
—
99.12 1.89
—
35.04 2.12
—
—
—
References and notes
3k
3l
3m
3n
—
—
—
32.40 2.12
—
98.78 3.67
—
—
—
—
—
—
—
—
—
—
—
—
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