Synthesis and biological evaluation of a novel structural type of serotonin 5-HT3 receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2006.02.006

A series of novel 3-substituted quinoxalin-2-carboxamides were designed as per the pharmacophoric requirement for 5-HT₃ receptor antagonists and prepared by microwave irradiation and also by conventional method. The compounds were characterized

Full text of DOI:10.1016/j.bmcl.2006.02.006

Bioorganic & Medicinal Chemistry Letters 16 (2006) 2769–2772
Synthesis and biological evaluation of a novel structural type
of serotonin 5-HT3 receptor antagonists
Ramachandran Venkatesha Perumala,* and Radhakrishnan Maheshb
aDepartment of Pharmaceutical Chemistry, Periyar College of Pharmaceutical Sciences for Girls,
Trichirappalli-620 021, Tamil Nadu, India
bPharmacy Group, Birla Institute of Technology & Science, Pilani-333 031, Rajasthan, India
Received 30 September 2005; revised 20 January 2006; accepted 2 February 2006
Available online 28 February 2006
Abstract—A series of novel 3-substituted quinoxalin-2-carboxamides were designed as per the pharmacophoric requirement for
5-HT3 receptor antagonists and prepared by microwave irradiation and also by conventional method. The compounds were char-
1
acterized by spectral data (IR, H NMR, and MS) and the purity was ascertained by microanalysis. The synthesized compounds
were evaluated for 5-HT3 antagonisms in longitudinal muscle-myenteric plexus preparation from guinea pig ileum against 5-HT3
agonist, 2-methyl-5-HT. Among the test compounds, N-{3-[(4-methylpiperazin-1-yl)methyl]-4-hydroxyphenyl}-3-methoxyquinoxalin-2-
carboxamide 4e showed most favorable 5-HT3 receptor antagonism.
Ó 2006 Elsevier Ltd. All rights reserved.
Serotonin (5-hydroxytryptamine, 5-HT) is a neurotrans-
mitter involved in various pharmacological effects in
several peripheral and central nervous systems.1 Fifteen
5-HT receptor subtypes belonging to 7 major classes (5-
HT1–5-HT7) have been reported so far.2 Recently,
5-HT3 receptor subtype has gained much attention be-
cause of the clinical use of 5-HT3 receptor antagonists
(RAs) in the treatment of cancer chemotherapy-induced
nausea and vomiting,3 and also in postoperative nausea
and vomiting.4,5 Moreover, a number of preclinical
studies suggest that 5-HT3 RAs can be used in the treat-
Figure 1. Pharmacophore of 5-HT3 receptor antagonists.
ment of various central nervous disorders such as
anxiety, schizophrenia, drug abuse, withdrawal, and
age-associated memory impairments.6,7 Hibert et al.8
proposed the pharmacophore of 5-HT3 RAs, which
(see Fig. 2). In the present paper, we describe the synthe-
sis of a structurally novel series of 3-substituted quinox-
consists of three components: an aromatic ring, a car-
alin-2-carboxamides and evaluated their 5-HT3
bonyl-containing linking moiety, and a basic center in
antagonistic activities in the longitudinal muscle-myen-
a specific spatial arrangement (Fig. 1). Based on this
pharmacophore, a number of molecules have been
reported so far.9,10 In view of the various therapeutic
against 5-HT3 agonist, 2-methyl-5-HT.
teric plexus (LMMP) preparation of guinea pig ileum
implications of 5-HT3 RAs and in continuation of our
interest in developing novel 5-HT3 RAs,11–13 our efforts
The title compounds, 3-substituted quinoxalin-2-carb-
oxamides, were designed according to the pharmaco-
were focused on introducing different substituents in the
3rd position of the quinoxaline moiety of compound 111
phoric requirements (proposed by Hibert et al.8) for
5-HT3 RAs. The pharmacophore consists of three com-
ponents (Fig. 1): an aromatic/heteroaromatic ring, a car-
bonyl-containing linking moiety, and a basic center in
a specific spatial arrangement. In the proposed series,
the least energy conformation of the molecules was
Keywords: 5-HT3 receptor antagonist; Quinoxalin-2-carboxamides.
*
Corresponding author. Tel.: +91 431 2459911; fax: +91 431
2456677; e-mail: vppharma2001@yahoo.co.in
0960-894X/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2006.02.006

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R. Venkatesha Perumal, R. Mahesh / Bioorg. Med. Chem. Lett. 16 (2006) 2769–2772
yields were obtained by both the methods. It was ob-
served that the reaction was accelerated greatly when
carried out in microwave environment. All the synthe-
sized compounds were characterized by spectral (IR,
1H NMR, and MS) and elemental analysis data. IR
spectral analysis of the final compounds (4a–4k and
5a–5k) showed absorption bands at ꢀ3400, ꢀ3200,
and ꢀ1670 cmÀ1 due to OH, NH, and C@O functions,
Figure 2.
1
generated by Tripos-Alchemy 2000 software (Tripos
Associated Inc., St. Louis, USA) and the pharmaco-
phoric distances were measured from the heteroaromat-
ic ring to carbonyl oxygen, carbonyl oxygen to basic
nitrogen of Mannich derivatives, and heteroaromatic
ring to basic nitrogen. The molecules identified for syn-
thesis comply with the pharmacophoric model.8
respectively. In H NMR spectra, OH showed singlet
at d ꢀ 11.5, NH showed singlet at d ꢀ 9.8, and aromatic
protons showed multiplet in the range of d 7.1–8.4.
Elemental analysis indicated that the calculated and
observed values were within the acceptable limits
( 0.4%). Physical data of the final compounds are given
in Table 1.
The synthetic procedures are illustrated in Scheme 1.
The starting material, 2-methoxy-3-methylquinoxaline,
was prepared as per the procedure described by Cheese-
man,14 and 2-ethoxy-3-methylquinoxaline was prepared
as per the procedure described by Newbold and Spring15
The intermediates, Mannich derivatives of p-aminophe-
nol, were prepared as per our previously reported
method.16 2-Methoxy-3-methylquinoxaline, and 2-eth-
oxy-3-methylquinoxaline on oxidation with a mixture
of sodium dichromate and sulfuric acid, as per our
previously reported method,13 afforded 3-methoxyqui-
noxalin-2-carboxylic acid,17 and 3-ethoxyquinoxalin-2-
carboxylic acid,18 respectively. The obtained product
was refluxed with excess thionyl chloride and few drops
of dimethylformamide for about 1 h. Excess thionyl
chloride was removed under reduced pressure to obtain
the crude acid chloride. To the solution of acid chloride
in DMF, a mixture of appropriate hydrochloride salt of
Mannich derivatives of p-aminophenol16 in DMF and
triethylamine was added. This reaction mexture was
then irradiated with microwaves for 2 min at 720 W.
DMF was removed using rotary flask evaporator and
to the residue, water was added. The separated product
was filtered, washed with water, dried, and recrystallized
from ethanol–acetone mixture to give 4 and 5, respec-
tively. When this reaction was carried out by conven-
tional heating (oil bath), the reaction was completed
(monitored by TLC) in 6–8 h. The product obtained
by both methods was identical in all aspects (mp, mixed
mp, co-TLC, and superimposable IR). Almost similar
The Institutional Animal Ethics Committee of the Birla
Institute of Technology & Science, Pilani, India,
approved the experimentation on animals (Protocol
No. IAEC/RES/6, dated 21.04.03). Male Dunkin Hart-
ley guinea pigs (250–300 g; Hissar Agricultural Universi-
ty, Hissar, Haryana, India) were sacrificed by cervical
dislocation. The abdomen was cut open and a length
of ileum was excised about 2 cm from the ileo-caecal
junction. The longitudinal muscle-myenteric plexus
(LMMP), 3–4 cm in length, was prepared and mounted
as described by the literature method.19 The tissue was
equilibrated for 30 min. under a resting tension of
500 mg and constant aeration in a 40 ml organ bath con-
taining Tyrode solution maintained at ca. 37 °C. Non-
cumulative concentrations of 2-methyl-5-HT (Tocris,
UK) were added with a 15 min dosing cycle (to prevent
desensitization) and left in contact with the tissue until
the maximal contraction had developed. To study the
antagonist effect of the test compounds on the response
evoked by 2-methyl-5-HT, the compounds were added
to the organ bath and left in contact with the tissue
for at least 10 min prior to the addition of 2-methyl-5-
HT. The contractions were recorded using a T-305
Force transducer coupled to a Student’s physiograph
(Bio Devices, Ambala, India). Antagonism was ex-
pressed in the form of pA2 values, which were graphical-
ly determined.20 The pA2 values of the test compounds
were compared with the standard antagonist Ondanse-
tron (Natco Pharma, Hyderabad, India). The observed
pharmacological data are represented in Table 1.
Scheme 1.

R. Venkatesha Perumal, R. Mahesh / Bioorg. Med. Chem. Lett. 16 (2006) 2769–2772
Table 1. Physical and pharmacological data of compounds 4a–4k and 5a–5k
2771
Compound
R
R1
R2
Yield (%) Mp (°C) Molecular
Molecular Antagonism to
weightb
formulaa
2-Me-5-HT pA2
c
4a
4b
4c
4d
4e
4f
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
OCH3
Pyrrolidinyl
Piperidinyl
H
61
65
62
67
69
65
56
55
59
156–158 C21H22N4O3 378
170–171 C22H24N4O3 392
3.9
3.5
H
Morpholinyl
Piperazinyl
H
124–126 C21H22N4O4 394
161–163 C21H23N5O3 393
149–150 C22H25N5O3 407
137–138 C23H27N5O3 412
177–178 C26H31N5O3 461
143–145 C26H31N5O5 493
181–183 C26H31N7O3 492
170–172 C28H37N7O3 520
155–156 C30H41N7O3 548
194–196 C22H24N4O3 392
207–208 C23H26N4O3 406
159–160 C22H24N4O4 408
196–197 C22H25N5O3 407
182–183 C23H27N5O3 421
169–170 C24H29N5O3 435
219–220 C27H33N5O3 475
177–179 C27H33N5O5 507
211–212 C27H33N7O3 506
202–203 C29H39N7O3 534
189–190 C31H43N7O3 562
3.0
4.5
H
N-Methyl-piperazinyl
N-Ethyl-piperazinyl
Pyrrolidinyl
H
5.7
4.9
H
4g
4h
4i
Pyrrolidinyl
Morpholinyl
Piperazinyl
<3.0
<3.0
4.0
Morpholinyl
Piperazinyl
4j
N-Methyl-piperazinyl N-Methyl-piperazinyl 60
4.8
4.1
4k
5a
5b
5c
5d
5e
5f
N-Ethyl-piperazinyl
N-Ethyl-piperazinyl
H
58
63
65
63
66
67
65
55
56
58
OC2H5 Pyrrolidinyl
OC2H5 Piperidinyl
3.5
3.1
H
OC2H5 Morpholinyl
OC2H5 Piperazinyl
H
<3.0
3.4
H
OC2H5 N-Methyl-piperazinyl
OC2H5 N-Ethyl-piperazinyl
OC2H5 Pyrrolidinyl
OC2H5 Morpholinyl
OC2H5 Piperazinyl
H
4.6
3.9
H
5g
5h
5i
Pyrrolidinyl
Morpholinyl
Piperazinyl
<3.0
<3.0
<3.0
<3.0
<3.0
5j
5k
OC2H5 N-Methyl-piperazinyl N-Methyl-piperazinyl 54
OC2H5 N-Ethyl-piperazinyl N-Ethyl-piperazinyl 57
a Elemental (C, H, and N) analysis indicated that the calculated and observed values were within the acceptable limits ( 0.4%).
b Molecular weight determination by mass spectral analysis.
c Results are means from three separate experiments except for those pA2 values <3.0, which are means of two separate experiments. SE was less than
10% of the mean.
In the present study, we have demonstrated the synthesis
and 5-HT3 receptor antagonistic activity of novel 3-
substituted quinoxalin-2-carboxamides in the isolated
guinea pig ileum. Ethoxy group (5a–5k) in the 3rd posi-
tion of the quinoxaline moiety showed lesser 5-HT3
antagonism than the methoxy group (4a–4k), which is
lesser than the chloro group (compound 113). Pipera-
zines attached with methylene group of p-aminophenol
(4d–4f, 4i–4k, and 5d–5f) exhibit higher 5-HT3 antago-
nism than other cyclic secondary amines (4a–4c, 4g,
4h, and 5a–5c). These results clearly indicate that N4
piperazine is essential for activity. Compounds 4d, 5d
(with no substitution at N4 piperazine) showed pA2 4.5
and 3.4, respectively; with increased lipophilicity (i.e.,
methyl group, 4e, 5e) activity increased (pA2 5.7 and
4.6, respectively). Further increase in lipophilicity (i.e.,
ethyl group, 4f, 5f) decreased the activity (pA2 4.9 and
3.9, respectively). It has been observed that when the
side chain of the 3-substituted quinoxalin-2-carboxa-
mides is attached with bis Mannich derivatives of p-ami-
nophenol (4g–4k and 5g–5k) the antagonism decreased
when compared to mono Mannich derivatives of p-ami-
nophenol (4a–4f and 5a–5f). Most of the reported 5-HT3
RAs contain aromatic/heteroaromatic esters or amides
having bridged bicyclic amines like azabicyclo[2.2.2]
octane,21 azabicyclo[3.2.1]octane22 and azabicy-
clo[3.3.1]nonane21,22 for the basic nitrogen in the phar-
macophore and moreover the bicyclic amine is directly
attached to the amide/ester moiety. In contrary, we have
the monocyclic amines are attached to the amide moiety
through a benzyl group.
Acknowledgments
This research was supported by grants from the Univer-
sity Grants Commission, New Delhi, India. We are
thankful to the CSIR, New Delhi, India, for awarding
SRF to one of the authors (R.V.P.). We are grateful
to the Birla Institute of Technology & Science, Pilani,
for providing necessary facilities to carry out this re-
search and the RSIC, Punjab University, Chandigarh,
India, for providing spectral and analytical data.
References and notes
1. Boess, F. G.; Martin, I. L. Neuropharmacology 1994, 33,
275.
2. Hoger, D.; Martin, G. Neuropharmacology 1997, 36, 419.
3. Karim, F.; Roerig, S. C.; Saphier, D. Biochem. Pharmacol.
1996, 52, 685.
4. Negus, B.; Markocic, S. Acute Pain 2003, 5, 63.
5. Gardner, C.; Perren, M. Neuropharmacology 1998, 37,
1643.
6. Jones, B. J.; Blackburn, T. P. Pharmacol. Biochem. Behav.
2002, 71, 555.
7. Barnes, J. M.; Barnes, N. M.; Cooper, S. J. Neurosci.
Biobehav. Rev. 1993, 16, 107.
8. Hibert, M. F.; Hoffmann, R.; Miller, R. C.; Carr, A. A.
J. Med. Chem. 1990, 33, 1594.
developed
novel
molecules
which
contain
heteroaromatic amide having monocyclic amine viz.,
pyrrolidine, piperidine, morpholine, piperazine, and 4-
substituted piperazines for the basic nitrogen and also
9. Clark, R. D.; Miller, A. B.; Berger, J.; Repke, D.
B.; Weinhardt, K. K.; Kowalczyk, B. A.; Eglen, R.
M.; Bonhaus, D. W.; Lee, C.-H.; Michel, A. D.;

2772
R. Venkatesha Perumal, R. Mahesh / Bioorg. Med. Chem. Lett. 16 (2006) 2769–2772
Smith, W. L.; Wong, E. H. F. J. Med. Chem. 1993,
36, 2645.
10. Ohta, M.; Suzuki, T.; Koide, T.; Matsuhisa, A.; Furuya,
T.; Miyata, K.; Yanagisawa, I. Chem. Pharm. Bull. 1996,
44, 991.
11. Mahesh, R.; Venkatesha Perumal, R.; Vijaya Pandi, P.
Biol. Pharm. Bull. 2004, 27, 1403.
12. Mahesh, R.; Venkatesha Perumal, R.; Vijaya Pandi, P.
Bioorg. Med. Chem. Lett. 2004, 14, 5179.
OCH3), 7.71 (d, 2H, H6 and H7 quinoxaline), 8.17 (d, 2H,
H5 & H8 quinoxaline), 12.60 (s, 1H, CO2H). Anal. Calcd
for C10H8N2O3: C, 55.04; H, 3.95; N, 13.72. Found. C,
55.31; H, 4.17; N, 13.51.
18. Yield 73%; mp 120–121 °C. IR (KBr) (cmÀ1): 3310,
1706, 1585. 1H NMR (300 MHz) (CDCl3) (d) ppm:
1.31 (t, 3H, CH3), 3.97 (q, 2H, CH2), 7.67 (d, 2H, H6
and H7 quinoxaline), 8.10 (d, 2H, H5 and H8 quinox-
aline), 12.67 (s, 1H, CO2H). Anal. Calcd for
C11H10N2O3: C, 60.55; H, 4.62; N, 12.84. Found. C,
60.31; H, 4.40; N, 13.02.
13. Mahesh, R.; Venkatesha Perumal, R.; Vijaya Pandi, P.
Pharmazie 2004, 60, 411.
14. Cheeseman, G. W. H. J. Chem. Soc. 1945, 1, 1804.
15. Newbold, G. T.; Spring, F. S. J. Chem. Soc. 1948, 519.
16. Mahesh, R.; Venkatesha Perumal, R. Indian J. Chem.
2004, 43B, 1012.
19. Paton, W. D. M.; Zar, A. M. J. Physiol. 1968, 194, 13.
20. MacKay, D. J. Pharm. Pharmacol. 1978, 30, 312.
21. Lopez-Rodriguez, M. L.; Morcillo, M. J.; Benhamu, B.;
Riaguas, D. Bioorg. Med. Chem. Lett. 1996, 6, 1195.
22. Coldwell, M. C.; Gadre, A.; Jerman, J.; King, F. D.; Nash,
D. Bioorg. Med. Chem. Lett. 1995, 5, 39.
17. Yield 76%; mp 101–102 °C. IR (KBr) (cmÀ1): 3303, 1701,
1
1594. H NMR (300 MHz) (CDCl3) (d) ppm: 3.71 (s, 3H,