3600
L. Gong et al. / Bioorg. Med. Chem. Lett. 13 (2003) 3597–3600
23a–27c are constrained in the chair conformation. As
seen in Table 1, compound 18, the boat conformation is
less active (IC50=5.3 mM) than the original lead
compound 1 (IC50=0.5 mM). The chair conformation
of 23a also did not look promising for this series
(IC50=1.8 mM, Table 2). Possible explanations are
unfavorable steric interactions due to the ethylene
bridge or that the central scaffold is now overly rigid.
However, this chair scaffold does result in more active
compounds when paired with the urea linker as in 24c,
25b, 26b, and 27c. The combination of side chain
modification [MeCH(OH) vs Me] and urea head sub-
stitution (trimethoxy vs methylsufonyl) ultimately gives
the most potent CCR3 antagonist, 24c (IC50=0.0082
mM). A similar trend is also observed in the chemotaxis
assay where 24c exhibited an IC50 of 0.0024 mM.
2. Oppenheim, J.; Feldman, M. Cytokine Reference; Aca-
demic: London, 2001; Vol. 1, p 3.
3. Daugherty, B. L.; Siciliano, S.; Demartino, J. A.; Mal-
kowitz, L.; Sirontina, A.; Springer, M. S. J. Exp. Med. 1996,
183, 2349.
4. Sallusto, F.; Mackay, C. R.; Lanzavecchia, A. Science
1997, 277, 2005.
5. Ochi, H; Hirani, W. M.; Yuan, Q.; Friend, D. S.; Austen,
K. F; Boyce, J. A. J. Exp. Med. 1999, 190, 267.
6. Bryan, S. A.; Ponath, P. D.; Wilhelm, R. S. In New Drugs
for Asthma, Allergy and COPD; Hansel, T. T., Barnes, P. J.,
Eds.; Karger: Basel, 2001; Vol. 31, p 288.
7. Naya, A; Sagara, Y.; Ohwaki, K.; Saeki, T.; Ichikawa, D.;
Iwasawa, Y.; Noguchi, K.; Ohtake, N. J. Med. Chem. 2001,
44, 1429.
8. Dhanak, D.; Christmann, L. T.; Darcy, M. G.; Jurewicz,
A. J.; Keenan, R. M.; Lee, J.; Sarau, H. M.; Widdowson,
K. L.; White, J. R. Bioorg. Med. Chem. Lett. 2001, 11, 1441.
9. Naya, A.; Kobayashi, K.; Ishikawa, M.; Ohwaki, K.; Saeki, T.;
Noguchi, K.; Ohtake, N. Bioorg. Med. Chem. Lett. 2001, 11, 1219.
10. Kato, M.; Ito, K.; Nishino, S.; Yamakuni, H; Takasugi,
H. Chem. Pharm. Bull. 1995, 43, 1351.
The pharmacokinetic profiles of several compounds in
Table 2 were also evaluated. The iv data in rats of
compound 23a demonstrated high clearance (7.06 l/h/
kg) coupled with a high volume of distribution (30.1 l/
kg) and t1/2 of 3.5 h. This is characteristic of lipophilic
compounds with basic nitrogen and the PK profile of
23a is similar throughout this series. A subsequent po
study in rats (30mpk) for more potent compounds 24c,
25b and 27c gave exposure values [AUC (0–24 h, mg/mL
h)] of 0.19, 0.28, and 0.71, respectively. The low po
exposure levels of these compounds are indicative of a
high volume of distribution, which was quantified in the
iv study with 23a.
11. The small molecule inhibitors were characterized by inhi-
biting the binding of 125I human eotaxin (Amersham) to
butyrate-treated hCCR3 L1.2 transfectants. The compounds
in DMSO were diluted with binding buffer (50mM HEPES, 1
mM CaCl2, 5 mM MgCl2, 0.5% BSA, 0.02% sodium azide,
pH 7.24). 25 mL of the test solution was added to each well of
a 96-well polypropylene plate, followed by 25 mL of 125I-
eotaxin (100 pmol) and 1.5Â105 of the CCR3 L1.2 transfected
cells in 25 mL binding buffer. After incubation for 1 h at rt, the
reaction was terminated by filtering through polyethyenimine
treated Packard Unifilter GF/C filter plate. The filters were
washed 5 times with ice cold wash buffer containing 10mM
HEPES and 0.5 M NaCl (pH 7.2) and dried at 65 ꢀC for 30
min. 25 mL/well of Microscint-20TM scintillation fluid was
added and the radioactivity retained on the filters was deter-
We designed and synthesized a novel class of CCR3
antagonists by modifying the lead compound 1. Func-
tional antagonism is demonstrated by their ability to
inhibit a chemotactic response. The potencies of these
compounds appear to be dependent on the presence of a
basic nitrogen, which presumably interacts with the
glutamic acid in TM7 of CCR3. Optimization of these
di-substituted piperidines, based on the aryl urea 24c,
will be the subject of future communications.
mined by using the Packard TopCountTM
.
12. Mirzadegan, T.; Diehl, F.; Ebi, B.; Bhakta, S.; Polsky, I.;
McCarley, D.; Mulkins, M; Weatherhead, G. S.; Lapierre,
J. M.; Dankwardt, J.; Morgans, D., Jr.; Wilhelm, R.;
Jarnagin, K. J. Biol. Chem. 2000, 275, 25562.
13. Manuscript in preparation.
14. The CCR3 antagonist activity can also be determined by
measuring the inhibition of eotaxin mediated chemotaxis of
the CCR3 L1.2 transfectant cells using the method described
by: Ponath, P. D.; Qin, S.; Ringler, D. J.; Clark-Lewis, I.;
Wang, J.; Kassam, N.; Smith, H.; Shi, X.; Gonzalo, J.; New-
man, W.; Gutierrez-Ramos, J.; Mackay, C. R. J. Clin. Invest.
1996, 97, 604.
References and Notes
1. Busse, W. W.; Lamanke, R. F., Jr. N. Engl. J. Med. 2001,
344, 350.