Synthesis and biological evaluation of some polymethoxylated fused pyridine ring systems as antitumor agents

Article abstract of DOI:10.1002/ardp.200900062

A series of 3,5-bis(arylidene)-4-piperidones like chalcone analogues carrying variety of methoxylated aryl groups, pyrazolo[4,3-c]pyridines, pyrido[4,3-d]pyrimidines, and pyrido[3,2-c]pyridines, carrying an arylidene moiety, and some pyrano[3,2-c]pyridine

Full text of DOI:10.1002/ardp.200900062

584
Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
Full Paper
Synthesis and Biological Evaluation of Some
Polymethoxylated Fused Pyridine Ring Systems as Antitumor
Agents
Sherif A. F. Rostom1, Ghada S. Hassan2, and Hussein I. El-Subbagh2
1 Department of Pharmaceutical Chemistry, Faculty of Pharmacy, King Abdulaziz University, Jeddah, Saudi
Arabia
2 Department of Pharmaceutical Chemistry, College of Pharmacy, King Saud University, Riyadh, Saudi Arabia
A series of 3,5-bis(arylidene)-4-piperidones like chalcone analogues carrying variety of methoxy-
lated aryl groups, pyrazolo[4,3-c]pyridines, pyrido[4,3-d]pyrimidines, and pyrido[3,2-c]pyridines,
carrying an arylidene moiety, and some pyrano[3,2-c]pyridines, like flavone and coumarin iso-
steres, were synthesized and screened for their in-vitro antitumor activity at the National Cancer
Institute (NCI, USA). The tested compounds 7, 9, 10, 12, 13, 15, 17, and 19 exhibited a broad spec-
trum of antitumor activity. Compounds belonging to the pyrazolo[4,3-c]pyridine series proved to
be more active than those of the pyrido[3,2-c]pyridine and pyrano[3,2-c]pyridine analogues, in
which the monomethoxylated derivatives showed better antitumor activity when compared
with their corresponding dimethoxylated congeners. Compound 7 is considered to be the most
active member identified in this study with a broad spectrum of activity against 22 different
tumor cell lines belonging to the nine subpanels employed, and a particular effectiveness
against the breast cancer T-47D cell line (GI 54.7%). The pyrano[3,2-c]pyridine heterocyclic system
19 proved to be the most active antitumor agent among the six-membered fused pyridines, with
variable activity against 18 different tumor cell lines, and special activity against the non-small
cell lung cancer Hop-92 and ovarian cancer OVCAR-4 cell lines (GI values 63.9 and 48.5%, respec-
tively).
Keywords: Antitumor activity / Chalcones / Fused pyridines / Synthesis /
Received: March 22, 2009; accepted: June 15, 2009
DOI 10.1002/ardp.200900062
tems have attracted great attention as potential chemo-
Introduction
therapeutic agents [1–7]. In particular, the cytotoxic
activity of (E)-3,5-bis(benzylidene)-4-piperidones and their
specificity toward leukemia cell lines with IC50 values less
than 10 lM have been reported [8–11]. This type of com-
pounds is a combination of cyclic a,b-unsaturated ketone
(chalcone) and b-amino ketone in one structure and is
known as ”curcuminoids” being structurally related to
curcumin (1,7-bis-(4-hydroxy-3-methoxyphenyl)-1,6-hepta-
diene-3,5-dione), a hydrophobic polyphenol derived from
the rhizome (turmeric) of the herb Curcuma longa [12].
Several studies revealed that curcumin and its congeners
manifest antiproliferative activity against various can-
cers, however, the actual mechanism of the antitumor
potential of curcuminoids are still far from being under-
Cancer poses a serious human health problem despite
much progress in understanding its biology and pharma-
cology. Consequently, the design of new lead structures
employed as antitumor agents is one of the most urgent
research areas in contemporary medicinal chemistry.
Among the wide range of pharmacologically active het-
erocycles, pyridines and some pyridine fused ring sys-
Correspondence: Sherif A. F. Rostom, Ph.D. Department of Pharma-
ceutical Chemistry, Faculty of Pharmacy, King Abdulaziz University, P.O.
Box 80260, Jeddah 21589, Saudi Arabia.
E-mail: sherifrostom@yahoo.com
Fax: +9662-6408404
i 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim

Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
Polymethoxylated Fused Pyridines as Antitumor Agents
585
stood [13]. With this in mind, it was reported that such
compounds of Michael acceptor-type could inhibit can-
cer cell proliferation in vitro and in vivo via promotion of
apoptosis in cancer cells by suppressing cyclin D and
endothelial growth factor receptor (EGFR) function,
while simultaneously decreasing levels of phosphokinase
B (AKT), c-myc, and phospho-AKT (pAKT) [14]. In one of the
more significant findings on the anticancer activity of
compounds inspired by curcumin, Adams et al. [15]
announced the superior activity of 2,6-bis(2-fluorobenzy-
lidene)piperidone (EF24) in anti-angiogenesis, cell cycle
arrest, and apoptosis of cancer cells. These authors
observed that the bis-benzylidenepiperidone, pyrone,
and cyclohexanone derivatives, containing the a,b-unsa-
turated ketone unit, exhibit much greater anticancer
and anti-angiogenesis activities than curcumin, with its
1,3-diketone unit [15].
Scheme 1. Synthesis of compounds 4–14.
During our ongoing studies aimed at the discovery of
new heterocycles endowed with antitumour activity, we
have reported on the synthesis and antitumor activities
of a series of (E)-3,5-bis(arylidene)-4-piperidones carrying a
variety of aryl and heteroaryl groups [16]. Some 2-amino-
quinazolines, their aza analogues, and the other fused
pyridine analogues such as 5-deazaaminopetrin [17] and
quinolones [18] interfere with the folic acid synthesis.
These findings rationalized the design and synthesis of
some pyrazolo[4,3-c]pyridine, pyrido[4,3-d]pyrimidine,
pyrano[3,2-c]pyridine, and pyrido[3,2-c]pyridine targets
carrying an arylidene moiety to be evaluated as antitu-
mor agents which may exert their activity through folic
acid synthesis inhibition. The results revealed that com-
pounds exhibited a broad spectrum of antitumor activ-
ity. In addition, some members proved to be of moderate
selectivity toward leukemia cell lines. Among the investi-
gated heterocycles, the pyrano[3,2-c]pyridine heterocy-
clic system proved to be the most active antitumor agents
[16]. On the other hand, we have identified some related
members belonging to the same fused heterocyclic ring
systems as potential antioxidants with remarkable oxy-
gen free-radical scavenger activity [19]. Careful literature
survey revealed that incorporation of alkoxy substituents
(methoxy and / or aryloxy moieties) results in significant
enhancement of several biological activities due to the
magnification of the compounds' lipophilicity [20–23].
Motivated by the above-mentioned findings, the
present study deals with the synthesis of a series of (E)-3,5-
bis(arylidene)-4-piperidones and some derived pyra-
zolo[4,3-c]pyridine, pyrido[4,3-d]pyrimidine, pyrano[3,2-
c]pyridine, and pyrido[3,2-c]pyridine ring systems carry-
ing methoxylated aryl groups. The variation in the
nature and size of substituents at such structures was
thought to be of interest representing variable elec-
tronic, lipophilic, and steric environment that would
influence the anticipated biological activity. The in-vitro
antitumor activity of the newly synthesized compounds
was evaluated according to the current one-dose protocol
of the National Cancer Institute (NCI) in-vitro disease-ori-
ented human cells screening panel assay.
Results and discussion
Chemistry
The designed target compounds depicted in Schemes 1
and 2 were obtained by reacting the starting material 1-
methyl-4-piperidone with variety of methoxylated aro-
matic aldehydes 1–3 under aldol condensation condi-
tions to produce the diarylidene ketone analogues 4–6
according to a previously described procedure [16]. These
chalcones were employed as key intermediates for fur-
ther synthesis of the other target molecules. They were
subjected to cycloaddition condensation reactions using
4-fluoro- or 4-sulfamylphenylhydrazine hydrochlorides
to give the corresponding 2-(4-fluorophenyl or 4-amino-
sulfonylphenyl)pyrazolo[4,3-c]pyridines 7–11. Analo-
gously, condensation of the same diarylidene derivatives
4–6 with 2-hydrazinoethanol in the presence of sodium
metal afforded the corresponding 2-(2-hydroxyethyl)pyr-
azolo[4,3-c]pyridines 12-14 (Scheme 1). On the other
hand, the chalcones 5 and 6 were further utilized for
another cyclocondensation reactions using either malo-
nonitrile or ethyl cyanoacetate in the presence of ammo-
nium acetate in refluxing ethanol to afford the 2-imino-
pyrido[3,2-c]pyridine and pyrido[3,2-c]pyridin-2-one deriv-
atives 15, 16 and 17, 18, respectively. It is worth mention-
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Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
files against most of the tested compounds. Among these,
the non-small cell lung cancer Hop-92 cell line exhibited
a wide range of sensitivity towards all of the tested ana-
logues, particularly compound 19 (GI 63.9%). Moreover,
compounds 7, 12, 15, and 17 showed moderate growth
inhibitory activity against the same cell line with GI val-
ues of 40.8, 49.5, 46.3, and 46.0%, respectively. Further-
more, the growth of the breast cancer T-47D cell line was
variably affected by the presence of all the tested com-
pounds except the analog 10. Particular effectiveness
against this cell line was shown by compound 7 with a GI
value of 54.7%, whereas the analogues 15 and 17 exhib-
ited moderate activity against the same cell line (GI 34.7
and 38.6%, respectively). Among the ovarian cancer sub-
panel, the OVCAR-4 cell line showed moderate sensitivity
towards compound 19 (GI 48.5%), whereas the MCF7 cell
line was mildly affected by the presence of seven out of
Scheme 2. Synthesis of compounds 15–22.
ing that reaction of 5 and 6 with malononitrile in reflux- the tested compounds with a GI range of 15.0 to 29.2%.
ing n-butanol produced the pyrano[3,2-c]pyridine ana- Regarding the activity against the leukemia subpanel,
logues 19 and 20, respectively. Finally, the targeted pyr- the growth of the RPMI-8226 cell line was found to be
ido[4,3-d]pyrimidine derivatives 21 and 22 could be suc- affected by the presence of the eight tested compounds
cessfully obtained via the reaction of 5 and 6 with thio- with a reliable sensitivity to compounds 7 and 9 (GI 43.1
urea in the presence of sodium hydroxide in refluxing and 52.0%, respectively). Meanwhile, the leukemia MOLT-
ethanol (Scheme 2).
4 cell line showed moderate to weak sensitivity towards
five of the tested compounds especially the compounds 9
and 10 (GI 39.2 and 37.7%, respectively). Additionally,
Preliminary in-vitro anticancer screening
Out of the newly synthesized compounds, eight deriv- compounds 12 and 19 showed noticeable activity against
atives 7, 9, 10, 12, 13, 15, 17, and 19 were selected by the the leukemia SR cell line with GI values of 43.9 and
National Cancer Institute (NCI) in-vitro disease-oriented 37.5%, respectively. Referring to the renal cancer subpa-
human cells screening panel assay to be evaluated for nel, the UO-31 cell line showed moderate to mild sensitiv-
their in-vitro antitumor activity. The effective one-dose ity to all the tested compounds, especially compound 7
assay has been added to the NCI 60 cell screen in order to (GI 43.6%). On the other hand, the growth of the mela-
increase the compound throughput and reduce data noma SK-MEL-2 and CNS cancer (SF-268, SNB-75 and
turnaround time to suppliers while maintaining effi- U251) cell lines were mildly affected by the presence of
cient identification of active compounds [24–26]. In this most of the tested compounds with a GI range of 11.0 to
protocol, all compounds submitted to the NCI 60 Cell 26.3%. It is worth-mentioning that only compounds 7
screen, are tested initially at a single high dose (10 lM) in and 9 were able to inhibit the growth of the prostate can-
the full NCI 60 cell panel including leukemia, non-small cer PC-3 cell line with GI values of 25.0 and 23.3%, respec-
cell lung, colon, CNS, melanoma, ovarian, renal, pros- tively.
tate, and breast cancer cell lines. Only compounds which
Structurally, the biologically active compounds belong
satisfy pre-determined threshold inhibition criteria to two series: five-membered fused pyridines namely pyr-
would progress to the five-dose screen. The threshold azolo[4,3-c]pyridines 7, 9, 10, 12, and 13 and six-mem-
inhibition criteria for progression to the five-dose screen bered fused pyridines namely pyrido[3,2-c]pyridines (15,
were designed to efficiently capture compounds with 17) and a pyrano[3,2-c]pyridine (19). In general, com-
antiproliferative activity and are based on careful anal- pounds belonging to the first series proved to be biologi-
ysis of historical Development Therapeutic Program cally more active than those of the second one. Among
(DTP) screening data. The data are reported as a mean the derivatives of the first series, the monomethoxylated
graph of the percent growth of treated cells, and pre- compounds 7, 9, and 12 exerted better antitumor activity
sented as percentage growth inhibition (GI%) caused by when compared with their corresponding dimethoxy-
the test compounds (Table 1).
lated analogues 10 and 13, besides, the 4-fluorophenyl
The obtained data revealed that some of the tested sub- moiety seemed to be the most favorable substituent. In
panel tumor cell lines exhibited variable sensitivity pro- this view, compound 7 (R = OCH3) is considered to be the
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Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
Polymethoxylated Fused Pyridines as Antitumor Agents
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Table 1. In-vitro percentage growth inhibition (GI%) caused by the test compounds against some selected tumor cell lines at the
single-dose assay.a)
Cpd No.
NSC No.
Panel
Subpanel tumor cell lines (% growth inhibitory activity)
7
746578
Non-Small Cell Lung Cancer
Colon Cancer
HOP-92 (40.8)
HCC-2998 (22.9), HCT-116 (31.1)
Breast Cancer
HS-578T (23.1), MCF7 (29.2), T-47D (54.7), MDA-MB-231/ATCC (31.4)
OVCAR-5 (17.3)
Ovarian Cancer
Leukemia
CCRF-CEM (26.6), K-562 (15.0), RPMI-8226 (43.1), SR (13.7)
786-0 (27.4), ACHN (26.4), CAKI-1 (26.8), UO-31 (43.6)
LOX-IMVI (19.2), MALME-3M (29.4), SK-MEL-2 (16.2)
PC-3 (25.0)
Renal Cancer
Melanoma
SE-268 (24.8), U251(19.8)
Prostate Cancer
CNS Cancer
9
746577
Non-Small Cell Lung Cancer
Breast Cancer
HOP-92 (18.3)
MCF7 (15.0), T-47D (24.3)
Leukemia
CCRF-CEM (18.0), HL-60 (TB) (49.4), K-562 (17.8), MOLT-4 (39.2),
RPMI-8226 (52.0)
CAKI-1 (46.4), UO-31 (29.6), ACHN (12.6)
SK-MEL-2 (20.0)
Renal Cancer
PC-3 (23.3)
Melanoma
SE-268 (19.8), SNB-75 (23.4), U251(24.3)
Prostate Cancer
CNS Cancer
10
12
13
746572
746579
746573
Non-Small Cell Lung Cancer
Breast Cancer
Leukemia
HOP-92 (15.7)
MCF7 (16.9), T-47D (15.3)
K-562 (34.3), MOLT-4 (37.7), RPMI-8226 (21.0)
UO-31 (37.9)
LOX IMVI (19.7)
U251 (21.5)
Renal Cancer
Melanoma
CNS Cancer
Non-Small Cell Lung Cancer
Breast Cancer
Leukemia
HOP-92 (49.5)
MCF7 (17.4), T-47D (16.2)
MOLT-4 (20.2), RPMI-8226 (30.9), SR (43.9)
CAKI-1 (34.7), UO-31 (33.5)
MALME-3M (16.0), SK-MEL-2 (20.7)
SNB-75 (14.2), U251 (13.7)
Renal Cancer
Melanoma
CNS Cancer
Non-Small Cell Lung Cancer
Breast Cancer
Ovarian Cancer
Leukemia
HOP-92 (25.1)
T-47D (22.6)
OVCAR-3 (11.6), OVCAR-5 (12.9)
MOLT-4 (25.2), RPMI-8226 (20.1)
786-0 (14.6), ACHN (13.3), UO-31 (25.8)
MALME-3M (16.5), SK-MEL-2 (16.2)
SNB-75 (12.2)
Renal Cancer
Melanoma
CNS Cancer
15
746575
746574
Non-Small Cell Lung Cancer
Breast Cancer
HOP-92 (46.3)
HS-578T (16.9), MCF7 (17.2), T-47D (34.7)
RPMI-8226 (29.3), SR (19.9)
CAKI-1 (28.2), UO-31 (28.8)
MALME-3M (26.5), SK-MEL-2 (20.7)
SF-268 (26.0), SNB-75 (26.3), U251 (12.8)
Leukemia
Renal Cancer
Melanoma
CNS Cancer
Non-Small Cell Lung Cancer
Breast Cancer
17
HOP-92 (46.0)
HS-578T (25.1), MCF7 (15.1), T-47D (38.6)
CCRF-CEM (18.2), HL-60 (TB) (21.0), K-562 (20.0), MOLT-4 (18.2),
RPMI-8226 (11.9)
Leukemia
UO-31 (18.0)
LOX IMVI (12.1), SK-MEL-2 (17.2)
SF-268 (15.4), SNB-75 (11.0), U251 (16.6)
Renal Cancer
Melanoma
CNS Cancer
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Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
Table 1. Continued.
Cpd No.
NSC No.
Panel
Subpanel tumor cell lines (% growth inhibitory activity)
19
746576
Non-Small Cell Lung Cancer
Breast Cancer
HOP-92 (63.9)
HS-578T (39.3), MCF7 (16.5), T-47D (27.5)
OVCAR-3 (21.3), OVCAR-4 (48.6),
OVCAR-5 (22.6)
Ovarian Cancer
RPMI-8226 (11.6), SR (37.5)
Leukemia
Renal Cancer
786-0 (21.0), ACHN (19.2), CAKI-1 (16.0), UO-31 (37.0)
LOX IMVI (15.5), SK-MEL-2 (24.3)
SF-268 (28.4), SNB-75 (26.0), U251 (16.5)
Melanoma
CNS Cancer
a)
The data obtained from NCI's in-vitro disease-oriented human tumor cell screen at 10 lM conc.
Table 2. Physicochemical and analytical data of compounds 7–22.
Compound
R
M.p. (8C)
Yield
(%)
Molecular Formula
(Mol. Weight)b)
(Cryst. solv.)a)
7
8
9
4-OCH3-C6H4
3,4-di-OCH3-C6H3
4-OCH3-C6H4
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
4-OCH3-C6H4
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
3,4-di-OCH3-C6H3
3,4,5-tri-OCH3-C6H2
248-9 (E)
183-5 (E)
60
54
86
84
73
48
83
14
20
11
21
13
31
50
27
14
C28H28FN3O2 (457.54)
C30H32FN3O4 (517.59)
C28H30N4O4S (518.63)
C30H34N4O6S (578.68)
C32H38N4O8S (638.73)
C24H29N3O3 (407.51)
C26H33N3O5 (467.56)
C28H37N3O7 (527.61)
C27H28N4O4 (472.54)
C29H32N4O6 (532.59)
C27H27N3O5 (473.52)
C29H31N3O7 (533.57)
C27H29N3O5 (475.54)
C29H33N3O7 (535.59)
C25H29N3O4S (467.58)
C27H33N3O6S (527.63)
202-3 (E / B)
264-4 (E / B)
148-9 (E)
125-6 (E / W)
86-8 (E / W)
133-35 (M / W)
246-8 (DMF / W)
174-6 (DMF / W)
273-5 (E)
157-9 (E)
168-9 (E)
160-2 (E / PE)
180-2 (E)
161-3 (E / W)
10
11
12
13
14
15
16
17
18
19
20
21
22
a)
Crystallization solvent(s): B: benzene, E: ethanol, DMF: N,N-dimethylformamide, M: methanol, PE: petroleum ether (60:80), W:
water.
b)
The found values were within l 0.4% of the theoretical values.
most active member identified in this study with a broad the non-small cell lung cancer Hop-92 and leukemia SR
spectrum of activity against about 22 different tumor cell lines (GI values 49.5 and 43.9%, respectively). Shifting
cell lines belonging to the nine subpanels employed, to the second series (Scheme 2), the most impressive
with particular effectiveness against the breast cancer T- growth inhibitory potential was displayed by the pyrano-
47D cell line (GI 54.7%). Replacement of the fluorine pyridine analog 19 which was active against 18 different
atom with a sulfonamide functional group resulted in tumor cell lines with special remarkable activity against
two less active compounds 9 and 10, of which the mono- the non-small cell lung cancer Hop-92 and ovarian cancer
methoxylated analog (9, R = OCH3) exhibited a better OVCAR-4 cell lines (GI values 63.9 and 48.5%, respec-
potential and spectrum of activity especially against the tively).
leukemia subpanel tumor cell lines. On the other hand,
These heterocycles could be considered as useful tem-
substitution of the pyrazole moiety with a hydroxyethyl plates for future development and further derivatization
fragment (as in 12 and 13) resulted in an obvious reduc- or modification to obtain more potent and selective anti-
tion in activity. Among these, the monomethoxylated tumor agents. The physicochemical and analytical data
analog 12 (R = OCH3) showed noticeable activity against of compounds 7–22 are presented in Table 2.
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Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
Polymethoxylated Fused Pyridines as Antitumor Agents
589
The authors are deeply thankful to the staff members of the (E)-2-(2-Hydroxyethyl)-3-aryl-5-methyl-7-arylidene-
Department of Health and Human Services, National Cancer 3,3a,4,5,6,7-hexahydro-2H-pyrazolo[4,3-c] pyridines 12–
Institute (NCI), Bethesda, MD, USA for carrying out the anti- 14
A solution of the appropriate diarylidene derivative 4–6
(0.01 mol) in ethanol (20 mL) was heated under reflux with 2-
cancer screening of the newly synthesized compounds.
hydrazinoethanol (1.52 g, 0.02 mol) and Na metal (0.5 g,
The authors have declared no conflict of interest.
0.022 mol) for 8–10 h. The volume of the reaction mixture was
concentrated to half and the separated solid product was fil-
tered, washed with cold ethanol, and recrystallized. Physico-
Experimental
chemical and analytical data are recorded in Table 2.
1H-NMR (CDCl3, ppm) d:
Chemistry
12: 2.17 (s, 3H, NCH3), 2.95 (m, 4H, CH2CH2), 3.0–3.09 (m, 2H, N-
CH2), 3.16–3.25 (m, 2H, CH2-N), 3.89 (s, 3H, OCH3), 3.93 (s, 3H,
OCH3), 4.74 (d, J = 8 Hz, 1H, CH-CH), 4.88 (d, J = 8 Hz, 1H, CH-CH),
6.04 (brs, 1H, OH), 6.81 (s, 1H, CH=C), 6.96–7.18 (m, 8H, ArH).
13: 2.19 (s, 3H, NCH3), 2.99 (m, 4H, CH2CH2), 3.06–3.11 (m, 2H,
N-CH2), 3.17–3.26 (m, 2H, CH2-N), 3.92 (s, 6H, 2 OCH3), 3.97 (s, 6H,
2 OCH3), 4.77 (d, J = 8 Hz, 1H, CH-CH), 4.90 (d, J = 8 Hz, 1H, CH-CH),
6.09 (brs, 1H, OH), 6.78 (s, 1H, CH=C), 7.08–7.23 (m, 6H, ArH).
14: 2.20 (s, 3H, NCH3), 2.97 (m, 4H, CH2CH2), 3.03–3.11 (m, 2H,
N-CH2), 3.19–3.24 (m, 2H, CH2-N), 3.88 (s, 9H, 3 OCH3), 3.95 (s, 9H,
3 OCH3), 4.75 (d, J = 8 Hz, 1H, CH-CH), 4.91 (d, J = 8 Hz, 1H, CH-CH),
5.96 (brs, 1H, OH), 6.79 (s, 1H, CH=C), 7.05–7.21 (m, 4H, ArH).
Melting points were determined in open glass capillaries on a
Gallenkamp melting point apparatus (Weiss-Gallenkamp, Lon-
don, UK) and were uncorrected. The 1H-NMR spectra were
recorded on a Varian EM 360 spectrometer using tetramethylsi-
lane as the internal standard (Varian Inc., Palo Alto, CA, USA;
Chemical shifts in d, ppm). Splitting patterns were designated as
follows: s: singlet; brs: broad singlet; d: doublet; m: multiplet.
Elemental analyses were performed at the Microanalytical Unit,
Faculty of Science, King Abdul-Aziz University, Jeddah, Saudi
Arabia, and the found values were within l 0.4% of the theoreti-
cal values. Follow up of the reactions and checking the homoge-
neity of the compounds were made by TLC on silica gel-protected
aluminum sheets (Type 60 F254, Merck, Germany) and the spots
were detected by exposure to UV-lamp at k = 254 nm.
General procedure for preparation of (E)-3-cyano-4-aryl-
2-imino-6-methyl-8-arylidene-5,6,7,8-tetrahydro-1H-
pyrido[3,2-c]pyridines 15, 16 and (E)-3-cyano-4-aryl-6-
methyl-8-arylidene-5,6,7,8-tetrahydro-1H-pyrido[3,2-
General procedure for preparation of (E)-2-(4-
fluorophenyl)-3-aryl-5-methyl-7-arylidene-3,3a,4,5,6,7-
hexahydro-2H-pyrazolo[4,3-c]pyridines 7, 8 and (E)-2-(4-
aminosulfonylphenyl)-3-aryl-5-methyl-7-arylidene-
3,3a,4,5,6,7-hexahydro-2H-pyrazolo[4,3-c]pyridines
c]pyridin-2-ones 17, 18
A mixture of the diarylidene compound 5 or 6 (0.01 mol), ammo-
nium acetate (4.7 g, 0.06 mol) and malononitrile (for 15, 16) or
ethyl cyanoacetate (for 17, 18) (0.01 mol) was refluxed in ethanol
(50 mL) for 4–8 h. Solvent was removed under reduced pressure
and the obtained residue was triturated with water, filtered,
dried, and recrystallized. Physicochemical and analytical data
are recorded in Table 2.
9–11
A mixture of the appropriate diarylidene derivative 4–6 (0.01
mol), and the corresponding 4-substituted phenylhydrazine
hydrochloride (0.015 mol) in EtOH (30 mL) was heated under
reflux for 8–10 h. The solvent was then removed under reduced
pressure and the remaining residue was triturated with water,
filtered, dried, and recrystallized. Physicochemical and analyti-
cal data are recorded in Table 2.
1H-NMR (DMSO-d6, ppm) d:
15: 2.19 (s, 3H, NCH3), 2.49–2.55 (m, 4H, CH2NCH2), 3.86 (s, 6H,
2 OCH3), 3.90 (s, 6H, 2 OCH3), 6.87 (s, 1H, CH=C), 7.07–8.09 (m,
8H, ArH + 2 NH).
16: 2.18 (s, 3H, NCH3), 2.47–2.52 (m, 4H, CH2NCH2), 3.85 (s, 9H,
3 OCH3), 3.89 (s, 9H, 3 OCH3), 6.87 (s, 1H, CH=C), 7.21–8.13 (m,
6H, ArH + 2 NH).
17: 2.22 (s, 3H, NCH3), 2.58–2.64 (m, 4H, CH2NCH2), 3.79 (s, 6H,
2 OCH3), 3.84 (s, 6H, 2 OCH3), 6.98 (s, 1H, CH=C), 7.23–8.12 (m,
7H, ArH + NH).
1H-NMR (CDCl3, ppm) d:
7: 2.23 (s, 3H, N-CH3), 2.97–3.06 (m, 2H, N-CH2), 3.18–3.21 (m,
2H, CH2-N), 3.87 (s, 3H, OCH3), 3.90 (s, 3H, OCH3), 4.67 (d, J = 8 Hz,
1H, CH-CH), 4.78 (d, J = 8 Hz, 1H, CH-CH), 6.79 (s, 1H, CH=C), 6.89–
7.27 (m, 12H, ArH).
8: 2.20 (s, 3H, NCH3), 3.02–3.11 (m, 2H, N-CH2), 3.20–3.27 (m,
2H, CH2-N), 3.90 (s, 6H, 2 OCH3), 3.96 (s, 6H, 2 OCH3), 4.71 (d, J = 8
Hz, 1H, CH-CH), 4.77 (d, J = 8 Hz, 1H, CH-CH), 6.69 (s, 1H, CH=C),
7.04–7.31 (m, 10H, ArH).
9: 2.15 (s, 3H, NCH3), 2.91–2.96 (m, 2H, N-CH2), 3.04–3.12 (m,
2H, CH2-N), 3.79 (s, 3H, OCH3), 3.86 (s, 3H, OCH3), 4.63 (d, J = 8 Hz,
1H, CH-CH), 4.68 (d, J = 8 Hz, 1H, CH-CH), 6.67 (s, 1H, CH=C), 6.81–
7.19 (m, 12H, ArH), 10.64 (brs, 2H, NH2).
18: 2.23 (s, 3H, NCH3), 2.51–2.57 (m, 4H, CH2NCH2), 3.81 (s, 9H,
3 OCH3), 3.88 (s, 9H, 3 OCH3), 6.94 (s, 1H, CH=C), 7.19–8.09 (m,
5H, ArH + NH).
(E)-2-Amino-3-cyano-4-aryl-6-methyl-8-arylidene-
5,6,7,8-tetrahydro-4H-pyrano[3,2-c]pyridines 19, 20
To a solution of the diarylidene compound 5 or 6 (0.01 mol) in n-
butanol (50 mL), was added malononitrile (0.7 g, 0.01 mol), and
the reaction mixture was heated under reflux for 5–6 h. After
cooling, the precipitated product was filtered off and recrystal-
lized. Physicochemical and analytical data are recorded in Table
2.
10: 2.16 (s, 3H, NCH3), 2.93–2.97 (m, 2H, N-CH2), 3.03–3.08 (m,
2H, CH2-N), 3.81 (s, 6H, 2 OCH3), 3.89 (s, 6H, 2 OCH3), 4.65 (d, J = 8
Hz, 1H, CH-CH), 4.74 (d, J = 8 Hz, 1H, CH-CH), 6.73 (s, 1H, CH=C),
6.89–7.26 (m, 10H, ArH), 10.5 (brs, 2H, NH2).
11: 2.13 (s, 3H, NCH3), 2.96–2.99 (m, 2H, N-CH2), 3.04–3.10 (m,
2H, CH2-N), 3.87 (s, 9H, 3 OCH3), 3.94 (s, 9H, 3 OCH3), 4.71 (d, J = 8
Hz, 1H, CH-CH), 4.78 (d, J = 8 Hz, 1H, CH-CH), 6.70 (s, 1H, CH=C),
6.81–7.25 (m, 8H, ArH), 10.57 (brs, 2H, NH2).
1H-NMR (DMSO-d6, ppm) d:
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590
S: A. F. Rostom et al.
Arch. Pharm. Chem. Life Sci. 2009, 342, 584–590
19: 2.16 (s, 3H, NCH3), 2.68–2.81 (m, 4H, CH2NCH2), 3.79 (s, 6H,
2 OCH3), 3.84 (s, 6H, 2 OCH3), 4.19 (s, 1H, pyran-C4-H), 6.83 (brs,
2H, NH2), 6.90 (s, 1H, CH=C), 7.12–7.27 (m, 6H, ArH).
[3] A. Perez-Rebolledo, J. D. Ayala, G. M. de Lima, N. Marchini,
et al., Eur. J. Med. Chem. 2005, 40, 467–472.
[4] X. Liu, M.-L. Go, Bioorg. Med. Chem. 2006, 14, 153–163.
20: 2.14 (s, 3H, NCH3), 2.65–2.79 (m, 4H, CH2NCH2), 3.80 (s, 9H,
3 OCH3), 3.87 (s, 9H, 3 OCH3), 4.21 (s, 1H, pyran-H), 6.75 (brs, 2H,
NH2), 6.92 (s, 1H, CH=C), 7.15–7.31 (m, 4H, ArH).
[5] S. Huang, R. Lin, Y. Yu, Y. Lu, et al., Bioorg. Med. Chem. Lett.
2007, 17, 1243–1245.
[6] A. Bazgira, M. M. Khanaposhtani, A. A. Soorki, Bioorg. Med.
Chem. Lett. 2008, 18, 5800–5803.
(E)-4-Aryl-6-methyl-8-arylidene-3,4,5,6,7,8-hexahydro-
1H-pyrido[4,3-d]pyrimidine-2-thiones 21, 22
[7] V. Nair, G. Chi, Q. Shu, J. Julander, D. F. Smee, Bioorg. Med.
Chem. Lett. 2009, 19, 1425–1427.
A mixture of the diarylidene derivative 5 or 6 (0.01 mol), thio-
urea (0.8 g, 0.01 mol), and NaOH (0.4 g, 0.01 mol) in ethanol (50
mL), was heated under reflux for 8–10 h. After cooling to room
temperature, water (20 mL) was added and the mixture was neu-
tralized to pH 6 using 10% HCl solution. The separated solid was
filtered, washed with water, dried, and recrystallized. Physico-
chemical and analytical data are recorded in Table 2.
1H-NMR (DMSO-d6, ppm) d:
21: 2.27 (s, 3H, NCH3), 3.07–3.15 (dd, J = 8.0 Hz, 2H, NCH2),
3.59–3.71 (m, 2H, CH2N), 3.86 (s, 6H, 2 OCH3), 3.90 (s, 6H, 2 OCH3),
5.19 (s, 1H, CH-NH), 6.68 (s, 1H, CH=C), 7.13–7.42 (m, 6H, ArH),
8.13 (brs, 1H, NH), 8.44 (brs, 1H, NH).
[8] J. R. Dimmock, V. K. Arora, M. J. Duffy, R. S. Reid, et al.,
Drug Des. Discov. 1992, 8, 291–299.
[9] J. R. Dimmock, S. K. Raghavan, B. M. Logan, G. E. Bigam,
Eur. J. Med. Chem. 1983, 18, 248–254.
[10] U. Das, J. Alcorn, A. Shrivastav, R. K. Sharma, et al., Eur. J.
Med. Chem. 2007, 42, 71–80.
[11] H. N. Pati, U. Das, S. Das, B. Bandy, et al., Eur. J. Med. Chem.
2009, 44, 54–62.
[12] G. Liang, L. Shao, Y. Wang, C. Zhao, et al., Bioorg. Med.
Chem. 2009, 17, 2623–2631.
[13] P. Anand, S. G. Thomas, A. B. Kunnumakkara, C. Sun-
22: 2.25 (s, 3H, NCH3), 3.11–3.24 (dd, J = 8.0 Hz, 2H, NCH2),
3.55–3.69 (m, 2H, CH2N), 3.84 (s, 9H, 3 OCH3), 3.91 (s, 9H, 3 OCH3),
5.12 (s, 1H, CH-NH), 6.64 (s, 1H, CH=C), 7.08–7.36 (m, 4H, ArH),
8.08 (brs, 1H, NH), 8.29 (brs, 1H, NH).
daram, et al., Biochem. Pharmacol. 2008, 76, 1590–1611.
[14] M. W. Amolins, L. B. Peterson, B. S. J. Blagg, Bioorg. Med.
Chem. 2009, 17, 360–367.
[15] B. K. Adams, J. Cai, J. Armstrong, M. Herold, et al., Anti-
cancer Drugs 2005, 16, 263–275.
Preliminary in-vitro anticancer screening
[16] H. I. El-Subbagh, S. M. Abu-Zaid, M. A. Mahran, F. A. Badria,
Out of the newly synthesized compounds, eight derivatives,
namely 7, 9, 10, 12, 13, 15, 17, and 19 were selected by the
National Cancer Institute (NCI) for the in-vitro disease-oriented
human cells screening panel assay to be evaluated for their in-
vitro anticancer activity. A primary in-vitro one-dose anticancer
assay was performed using the full NCI 60 cell panel in accord-
ance with the current protocol of the Drug Evaluation Branch,
NCI, Bethesda, MD, USA [24–26]. These cell lines were incubated
with one concentration (10 lM) for each tested compound. A 48
h continuous drug exposure protocol was used, and a sulpho-
rhodamine B (SRB) protein assay was employed to estimate cell
viability or growth.
A. M. Al-Obaid, J. Med. Chem. 2000, 43, 2915–2921.
[17] C. Temple, R. Elliot, J. Montogomery, J. Org. Chem. 1982, 47,
761–764.
[18] L. Li, H. Wang, S. Kuo, T. Wu, et al., J. Med. Chem. 1994, 37,
3400–3407.
[19] M. A. Al-Omar, K. M. Youssef, M. A. El-Sherbeny, S. A. Awa-
dalla, H. I. El-Subbagh, Arch. Pharm. 2005, 338, 175–180.
[20] D. J. Guerin, D. Mazeas, M. S. Musale, F. N. M. Naguib, et
al., Bioorg. Med. Chem. Lett. 1999, 9, 1477–1480.
[21] C. Lin, J. Yang, C. Chang, S. Kuo, et al., Bioorg. Med. Chem.
2005, 13, 1537–1544.
[22] L. Barboni, G. Giarlo, R. Ballini, G. Fontana, Bioorg. Med.
Chem. Lett. 2006, 16, 5389–5391.
[23] T. Walle, Semin. Cancer Biol. 2007, 17, 354–362.
References
[24] M. R. Grever, S. A. Schepartz, B. A. Chabner, Semin. Oncol.
1992, 19, 622–638.
[1] K. Poreba, A. Opolski, J. Wietrzyk, M. Kowalska, Arch.
Pharm. 2001, 334, 219–223.
[25] M. R. Boyd, K. D. Paull, Drug Rev. Res. 1995, 34, 91–109.
[2] A. Gangjee, O. Adaira, S. F. Queener, Bioorg. Med. Chem.
2001, 9, 2929–2935.
[26] A. Monks, D. Scudiero, P. Skehan, R. Shoemaker, et al., J.
Natl. Cancer Inst. 1991, 83, 757–766.
i 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
www.archpharm.com