J. Liu et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6840–6844
6843
The substitution effect on the phenyl ring of the 1-N benzoyl
moiety is shown in Table 5. Compounds with methyl substitution
at the para and meta positions (36 and 35) displayed better po-
tency than the ortho-substituted analog (34). 3,4-Dimethyl com-
pound (37) was similar in potency to that of mono para or meta-
methyl compounds (36 and 37). In general, substitutions at the
para-position afforded compounds with good CRTH2 binding affin-
ity regardless of electronic and steric effects (36, 38–42). Some
substitutions, such as phenoxy (42), methoxy (38) and trifluoro-
methoxy (41), at the para-position yielded more potent com-
pounds compared to the unsubstituted compound (1).
plasma, these (2S,4R) enantiomers displayed strong inhibitory
activity for the CRTH2 receptor.
In addition to having high affinity for the CRTH2 receptor, these
compounds are also potent functional antagonists. Compounds 1a,
9a, 38a and 41a were potent inhibitors of PGD2-mediated human
eosinophil shape change28 (Table 7). In particular, compound 9a
had an IC50 of 0.77 nM. The affinity of these compounds for the
DP receptor was evaluated with a 3H-PGD2 displacement assay
using 293 cells stably transfected with hDP receptor. These com-
pounds were found to be selective for the CRTH2 receptor over DP.
The pharmacokinetics properties of compound 41a were evalu-
ated in male Sprague Dawley rats following IV (0.7 mg/kg) and oral
(2.0 mg/kg) dosing. The total body clearance and the terminal half-
life were 0.73 L/h/kg and 5.1 h, respectively. After oral administra-
tion the compound showed good bioavailability (38%) following
administration of a solution formulation (1 mL/kg; 10% etha-
nol:90% PEG400).
Based partially on their binding affinity, compounds 9, 38 and
41 were selected and resolved by chiral HPLC and each enantiomer
was evaluated (Table 6). Like compound 1, the (2S,4R) enantiomers
had greater affinity for the CRTH2 receptor. Even in the presence of
In summary, we have discovered and optimized a series of tet-
rahydroquinoline derivatives as potent CRTH2 antagonists with
selectivity over DP. Furthermore, we have identified compound
41a as a potent CRTH2 antagonist with good pharmacokinetic
properties rendering it a useful tool compound for in vivo studies
of CRTH2 functions.
Table 6
O
chiral
N
4
R1
N2 CH3
O
References and notes
R2
CRTH2 IC50 in
1. Lewis, R. A.; Soter, N. A.; Diamond, P. T.; Austen, K. F.; Oates, J. A.; Roberts, L. J. J.
Immunol. 1982, 129, 1627.
2. Holgate, S. T.; Burns, G. B.; Robinson, C.; Church, M. K. J. Immunol. 1984, 133,
2138.
3. Gundel, R. H.; Kinkade, P.; Torcellini, C. A.; Clarke, C. C.; Watrous, J.; Desai, S.;
Homon, C. A.; Farina, P. R.; Wegner, C. D. Am. Rev. Respir. Dis. 1991, 144, 76.
4. Nagata, K.; Hirai, H.; Tanaka, K.; Ogawa, K.; Aso, T.; Sugamura, K.; Nakamura,
M.; Takano, S. FEBS Lett. 1999, 459, 195.
5. Nagata, K.; Tanaka, K.; Ogawa, K.; Kemmotsu, K.; Imai, T.; Yoshie, O.; Abe, H.;
Tada, K.; Nakamura, M.; Sugamura, K.; Takano, S. J. Immunol. 1999, 162, 1278.
6. Cosmi, L.; Annunziato, F.; Galli, M. I. G.; Maggi, R. M. E.; Nagata, K.; Romagnani,
S. Eur. J. Immunol. 2000, 30, 2972.
7. Miadonna, A.; Tedeschi, A.; Brasca, C.; Folco, G.; Sala, A.; Murphy, A. J. Allergy
Clin. Immunol. 1990, 85, 906.
8. Turner, N. C.; Fuller, R. W.; Jackson, D. M. J. Lipid. Mediators Cell Signalling 1995,
11, 93.
9. Hirai, H.; Tanaka, K.; Yoshie, O.; Ogawa, K.; Kenmotsu, K.; Takamori, Y.;
Ichimasa, M.; Sugamura, K.; Nakamura, M.; Takano, S.; Nagata, K. J. Exp. Med.
2001, 193, 255.
a
a
Compd
R1
R2
CRTH2 IC50 in
buffer (
l
M)
plasma (lM)
9 racemic
–CO(CH2)2CO2H
–CO(CH2)2CO2H
–CO(CH2)2CO2H
CH3
CH3
CH3
CH3
CH3
CH3
H
H
H
OMe
OMe
OMe
OCF3
OCF3
OCF3
0.005
0.003
0.40
0.028
0.015
0.40
0.026
0.025
0.217
0.028
0.009
>10
0.13
0.039
>10
0.18
0.106
>10
9a (2S,4R)b
9b (2R,4S)b
38 racemic
38a (2S,4R)b
38b (2R,4S)b
41 racemic
41a (2S,4R)b
41b (2R,4S)b
Displacement of 3H-PGD2 from the CRTH2 or DP receptors expressed on 293
cells. Assay run in buffer containing 0.5% BSA or in 50% plasma. See Ref. 20 for assay
protocol. Values are means of three experiments, standard deviation is 30%.
Stereochemistry assigned based on the retention times of chiral HPLC compared
to 1a and 1b and CRTH2 activities, ee >99%.
a
b
10. Sugimoto, H.; Shichijo, M.; Iino, T.; Manabe, Y.; Watanabe, A.; Shimazaki, M.;
Gantner, F.; Bacon, K. B. J. Pharmacol. Exp. Ther. 2003, 305, 347.
11. Monneret, G.; Gravel, S.; Diamond, M.; Rokach, J.; Powell, W. S. Blood 2001, 98,
1942.
12. Gosset, P.; Bureau, F.; Angeli, V.; Pichavant, M.; Faveeuw, C.; Tonnel, A. B.;
Trottein, F. J. Immunol. 2003, 170, 4943.
Table 7
13. Liu, J.; Fu, Z.; Wang, Y.; Schmitt, M.; Huang, A.; Marshall, D.; Tonn, G.; Seitz, L.;
Sullivan, T.; Tang, H. L.; Collins, T.; Medina, J. Bioorg. Med. Chem. Lett. 2009.
14. Ghosh, S.; Elder, A. M.; Carson, K. G.; Sprott, K.; Harrison, S. WO Patent 032848,
2004.
O
chiral
N
N
R1
15. Ghosh, S.; Elder, A. M.; Carson, K. G.; Sprott, K.; Harrison, S. J.; Hicks, F. A.;
Renou, C. C.; Reynolds, D. WO Patent 100321, 2005.
CH3
O
16. Kuhn, C.; Feru, F.; Bazin, M.; Awad, M.; Goldstein, S. W. EP Patent 1413306,
2004.
17. Kotera, O.; Oshima, E.; Ueno, K.; Ikemura, T.; Manabe, H.; Sawada, M.; Mimura,
H.; Miyaji, H.; Nonaka, H. WO Patent 052863, 2004.
R2
Compd R1
R2
Eosinophil Shape Changea DP IC50
IC50 (nM)
(lM)
b
18. Inman, W.; Liu, J.; Medina, J. C.; Miao, S. WO Patent 007094, 2005.
19. Mimura, H.; Ikemura, T.; Kotera, O.; Sawada, M.; Tashiro, S.; Fuse, E.; Ueno, K.;
Manabe, H.; Ohshima, E.; Karasawa, A.; Miyaji, H. J. Pharmacol. Exp. Ther. 2005,
314, 244.
20. The CRTH2 radioligand binding assay was performed on 293 cells stably
expressing human CRTH2. To measure binding, [3H]-PGD2 was incubated
together with 293(hCRTH2) cells in the presence of increasing concentrations
of compounds. After washing, the amount of [3H]-PGD2 that remained bound
to the cells was measured by scintillation counting and the concentration of
compounds required to achieve a 50% inhibition of [3H]-PGD2 binding (the
IC50) was determined. The binding buffer contains either 0.5% BSA (buffer
binding) or 50% human plasma (plasma binding).
1ac
CH3
H
H
141
0.77
>10
>10
>10
>10
9ad
–CO(CH2)2CO2H
38ad
41ad
CH3
CH3
OMe 49.3
OCF3 48.8
a
PGD2-mediated human eosinophil shape change assay. See Ref. 28 for assay
protocol. Values are means of two experiments.
Displacement of 3H-labeled PGD2 from the DP receptors expressed on 293 cells.
b
Assay run in buffer containing 0.5% BSA. See Ref. 29 for assay protocol. Values are
means of three experiments, standard deviation is 30%.
Stereochemistry determined by synthesis (Scheme 2), ee >99%.
Stereochemistry assigned based on the retention times of chiral HPLC compared
to 1a and 1b and CRTH2 activities., ee >99%.
21. CRTH2 mediated cell migration was analyzed in a transwell migration assay
using hCRTH2 stably transfected CEM cells (a T lymphoblast cell line). The cells
were incubated with increasing concentrations of compounds for 3 h in a 96-
well migration chamber on top of a transwell filter and the number of cells
c
d