Synthesis of new thiophene and benzo[b]thiophene hydrazide derivatives as human NPY Y5 antagonists

Article abstract of DOI:10.1016/j.bmcl.2003.11.070

Neuropeptide Y is one of the most potent appetite stimulating hormones known. Novel thiophene and benzo[b]thiophene hydrazide derivatives were synthetized and evaluated biologically as NPY Y₁ and Y₅ receptor subtype antagonists. They were found to have nanomolar binding affinities for human NPY Y₅ receptor, obtaining the lead compound, trans-N-4-[N′-(thiophene-2-carbonyl)hydrazinocarbonyl]cyclohexylmethyl-4- bromobenzenesulfonamide, which binds with a 7.70 nM IC₅₀ to the hY₅ receptor.

Full text of DOI:10.1016/j.bmcl.2003.11.070

Bioorganic & Medicinal Chemistry Letters 14 (2004) 597–599
Synthesis of new thiophene and benzo[b]thiophene hydrazide
derivatives as human NPY Y5 antagonists
Silvia Galiano, Oihana Erviti, Silvia Perez, Antonio Moreno, Laura Juanenea,
Ignacio Aldana* and Antonio Monge
´
Unidad en Investigacion y Desarrollo de Medicamentos, Centro de Investigacion en Farmacobiologıa Aplicada (CIFA),
´
´
Universidad de Navarra, c/Irunlarrea s/n, 31080 Pamplona, Spain
Received 4 July 2003; revised 3 November 2003; accepted 25 November 2003
Abstract—Neuropeptide Y is one of the most potent appetite stimulating hormones known. Novel thiophene and benzo[b]thio-
phene hydrazide derivatives were synthetized and evaluated biologically as NPY Y1 and Y5 receptor subtype antagonists. They
were found to have nanomolar binding affinities for human NPY Y5 receptor, obtaining the lead compound, trans-N-4-[N0-(thio-
phene-2-carbonyl)hydrazinocarbonyl]cyclohexylmethyl-4-bromobenzenesulfonamide, which binds with a 7.70 nM IC50 to the hY5
receptor.
# 2003 Elsevier Ltd. All rights reserved.
Neuropeptide Y (NPY), a 36 amino acid peptide, is
widely expressed in both the peripheral and central ner-
vous system.1 NPY regulates a variety of physiological
functions, such as food intake, intestinal motility, vaso-
constriction, blood pressure, nasal congestion, anxiety,
depression, pain, motor and sexual behaviour.2,3 Five
distinct types of G-protein coupled NPY receptors (Y1,
Y2, Y4, Y5 and Y6) have been cloned.4 NPY Y1 and Y5
receptor subtypes have been implicated in mediating the
orexigenic effects of NPY and in regulating food intake
and body weight. On the basis of the potent orexigenic
effects of NPY in vivo,5,6 a number of papers have been
published with NPY receptor-specific ligands as a target
for developing antagonists for the treatment of obesity.7
Diverse structural series of arylsulfonamidomethylcy-
clohexyl derivatives as CGP71683A8 and Synaptic
Pharmaceutical’s tetralin derivative 19 (Fig. 1) have
been reported to antagonize the NPY Y5 receptor,
reducing food intake in ob/ob mice and Zucker obese rat
models.10 Consequently, the aim of this work was the
synthesis and biological evaluation of new thiophene
and benzo[b]thiophene hydrazide derivatives, structu-
rally related to the previous arylsulfonamidomethyl-
cyclohexyl derivatives, as part of a program to discover
a novel antiobesity agent with antagonistic activity for
human NPY Y5 receptor subtype.
Synthesis of new thiophene and benzo[b]thiophene
hydrazide derivatives have been carried out following
the synthetic route shown in Scheme 1.
The formation reaction of the primary sulfonamides 2a–
g consists of a nucleophilic attack on the part of amine 1
(1.00 equiv) against different sulfonyl chlorides (1.50
equiv),11 following a SN2mechanism, by means of
NaOH 2M (150 mL) is used as a solvent. The reaction is
Figure 1.
* Corresponding author. Tel.:+34-948-425653; fax: +34-948-425652; e-mail: ialdana@unav.es
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.11.070

598
S. Galiano et al. / Bioorg. Med. Chem. Lett. 14 (2004) 597–599
Scheme 1.
stirred at room temperature for 48 h. Sulfonamide deri-
vatives are obtained by precipitation, adding HCl (c)
until pH 1–2and then washing with H 2O and n-hexane.
Table 1. Y1 and Y5 receptor binding affinities of compounds 5a–g
The sulfonamides 2a–g are maintained at 0 ꢀC, for
1 h, in dry CH2Cl2 (150 mL), under N2 atmosphere, and
0
.
1-ethyl - 3- (3 -dimethylaminopropyl)carbodiimide HCl
(EDC) (1.13 equiv)12 is used for the activation of the
carboxylic group (1.00 equiv). Next, hydrazide deriva-
tives 3 or 4 (1.13 equiv) is added and the reaction is
stirred at room temperature for 24 h. The solvent is
evaporated and the residue is washed with H2O (2 0 mL)
and diethyl ether (5 mL), yielding the corresponding
thiophene and benzo[b]thiophene hydrazide derivatives
5a–g, 6a–g. All of the compounds were chemically
characterized by thin layer chromatography (TLC),
melting point, infrared, nuclear magnetic resonance (1H
NMR), elemental microanalysis and HPLC.
Compd
Ar1–SO2–NH–
Y1 IC50 (nM)
Y5 IC50 (nM)
32.2
5a
>104
5b
5c
5d
>104
>104
>104
7.70
48.4
20.4
5e
>104
47.7
Binding assays for both receptors NPY1 and NPY5 were
carried out as described by Duhault et al.13 For the
human Y1 receptor binding assay, using iodinated pep-
tide YY (NEN), incubations were performed at 30 ꢀC
for 90 min with various competitors concentrations in
Buffer A (Hepes/NaOH 20 mM, pH 7.4, NaCl 10 mM,
KH2PO4 220 mM, CaCl2 1.26 mM, MgSO4 0.81 mM
and bovine serum albumin 0.1%) with SK-N-MC cell
membranes (50 mg of protein/mL of assay) in a total
volume of 500 mL. Non-specific binding was determined
in the presence of 1 mM NPY. The reaction was then
stopped by filtration. The filters (GF/B, Whatman, pre-
coated in 0.3% PEI) were extensively washed with buf-
fer A, and counted in a gamma counter (Packard). For
human Y5 receptor binding assay, the binding was car-
ried out with iodinated peptide YY (NEN) as follows:
COS cells transfected with the human Y5 NPY receptor
5f
>104
>104
12.5
147
5g
were lysed and the membranes were prepared by differ-
ential centrifugation. These membranes contained about
2pmol per mg of protein of this receptor. Incubations
were perfomed in 500 mL comprising, 20 pM final of
[
125I]PYY in 50 mL, 400 mL of membrane suspension
(0.15 mg/mL) and competitor dilutions in 50 mL, at
30 ꢀC for 2h. The reaction was stopped by filtration
through GF/C filters (Whatman) and the results were
expressed in IC50 (Tables 1 and 2).

S. Galiano et al. / Bioorg. Med. Chem. Lett. 14 (2004) 597–599
599
Table 2. Y1 and Y5 receptor binding affinities of compounds 6a–g
We are continuing our search for in vivo assays to that
will allow us to clearly understand the SAR trends and
the absorption and permeability to brain.
In summary, we have developed a new series of thio-
phene and benzo[b]thiophene derivatives as potent and
selective antagonists at the human NPY Y5 receptor.
Based on the obtained data, we propose a more thor-
ough research program regarding the determination of
the importance of the substituents Ar1 with in vivo
assays, the biological significance of the results
obtained, and the role played by neuropeptide Y in food
intake response.
Compd
6a
Ar1–SO2–NH–
Y1 IC50 (nM)
>104
Y5 IC50 (nM)
175
6b
>104
28.4
6c
6d
>104
>104
279
42.9
Acknowledgements
We would like to thank Carmen Elizalde for her colla-
boration in this study.
6e
>104
142
6f
>104
>104
11.5
92.2
References and notes
1. Hipskind, P.; Gehlert, D. Annu. Rep. Med. Chem. 1996,
31, 1.
6g
2. Kalra, S. P.; Dube, M. G.; Fournier, A.; Karla, P. S.
Physiol. Behav. 1992, 50, 5.
3. Gehlert, D. Proc. Soc. Exp. Biol. Med. 1998, 218, 7.
4. Blomqvist, A. G.; Herzog, H. Trends Neurosci. 1997, 20,
294.
5. Clark, J. T.; Kalra, P. S.; Crowley, W. R.; Kalra, S. P.
Endocrinology 1984, 115, 427.
6. Stanley, B. G.; Leibowitz, S. F. Life Sci. 1984, 35, 2635.
7. Islam, I.; Dhanoa, D.; Finn, J.; Du, P.; Walker, M. W.;
Salon, J. A.; Zhang, J.; Gluchowski, C. Bioorg. Med.
Chem. Lett. 2002, 12, 1767.
Fourteen new compounds, derivatives of the thiophene
and benzobthiophene hydrazide, have been synthetized.
The NPY Y1 and Y5 receptor binding affinities for these
compounds are reported in Tables 1 and 2.
The results of the in vitro evaluation of the antagonist
activity on the NPY Y5 receptor have shown that all the
compounds present good affinity on said receptor lead-
ing compound 5b with an IC50 value of 7.7 nM and
selectivity as neither of them were found to antagonize
the NPY Y1 receptor (IC50104 nM).
8. Rueguer, H.; Schmidlin, T.; Rigollier, P.; Yamaguchi, Y.;
¨
Tintelnot-Blomley, L.; Schilling, W.; Criscione, L.; Mah,
R. PCT WO 97/20823.
9. Islam, I.; Dhanoa, D. S.; Finn, J. M.; Du, P.; Gluchowski,
C.; Jean, Y.T. PCT WO 97/19682.
10. Hofbauer, K. G.; Schaffhauser, A. O.; Batzl-Hartmann,
C.; Stricker-Krongrad, A.; Whitebread, S.; Cumin, F.;
Rigolier, P.; Yamaguchi, Y.; Chiesi, M.; Levens, N.;
Structure–activity relationships have been studied,
introducing different substituents Ar1 of the molecule’s
general structure. The results showed that the thiophene
derivatives (5a–f) present better affinity than their
benzo[b]thiophene analogues (6a–f). Three compounds,
5b, 5f and 6f with IC50 values of 7.7, 12.5 and 11.5 nM,
showed the best activity. All of these potent hY5
antagonists possess a tiophene ring in their structures,
as either an Arl substituent or a hydrazide rest. The
presence of the ring of thiophene could be determinant
in their link to the receptor.
Schilling, W.; Walker, M. W.; Gerald, C.; Rueger, H.;
¨
Criscione, L. Regul. Pept. 1997, 71, 211.
11. Smith, M. B., March, J., Eds. March’s Advanced Chem-
istry, 5th Ed; Wiley-Interscience: New York, 2001; p 574.
12. Sheehan, J.; Preston, J.; Cruickshank, P. A. J. Am. Chem.
Soc. 1965, 87, 2492.
13. Duhault, J.; Boulanger, M.; Chamorro, S.; Boutin, J. A.;
Della Zuana, O.; Douillet, E.; Fauchere, J. L.; Feletou,
M.; Germain, M.; Husson, B.; Renard, P.; Tisserand, F.
Can. J. Biochem. Physiol. 2000, 78, 173.