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3.0E+05
2.0E+05
1.0E+05
0.0E+00
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PBS Veh
10
1
Veh
10
3
1
--------------
AMG487
(mg/mL)
------------------------
Compound 21
(mg/mL)
Exposure (nM)
AMG487
205
37
11. (a) Knight, R. L.; Allen, D. R.; Birch, H. L.; Chapman, G. A.; Galvin, F. C.; Jopling, L.
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Compound 21
291
182
86
Figure 3. Compound 21 was efficacious in mouse lung bleomycin model. *p <0.005
as determined by Student’s t-tests.
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pump18 on the same day of bleomycin challenge for six days. On
day six, the number of cells collected in BAL fluid was counted.
Plasma samples were also collected at the end of the experiment.
The efficacy observed with AMG 487 in this experiment is consis-
tent with previous reports. Compound 21 inhibited infiltration of
cells into the lung at a steady state plasma concentration of
P86 nM.
In conclusion, substitution alpha to the left ring nitrogen of the
fused hetero-bicyclic cores such as the 8-azaquinazolinone, the
imidazo-pyrimidine, and the imidazo-pyrazine core, generally im-
proved the potency of the CXCR3 antagonists. In particular, the
imidazo-pyrazines had superior pharmacokinetic properties in
addition to its potency. Compound 21 demonstrated in vivo effi-
cacy in blocking the leukocyte migration into the lung in response
to bleomycin.
14. See Ref. 13b for assay protocol.
15. All compounds were characterized by 1H NMR and LC/MS and were >95% in
purity by reverse phase HPLC. Chiral purity was analyzed with ChiralTech AD
column. The enantiomeric purity was higher than 95% ee.
16. This assay measures cell-surface CXCR3 occupancy by FACS analysis of
fluorescent using
a fluorescently-tagged anti-CXCR3 antibody specific to
CXCR3. Addition of the CXCR3 ligand, ITAC, results in the loss of fluorescent
signal of antibody due to the competing ligand binding. The antagonist blocks
ITAC binding against receptor, but does not compete against anti-CXCR3
antibody binding to CXCR3 receptor. The fluorescent signal of the antibody was
restored with the addition of the antagonists. An ITAC concentration of 500 ng/
ml was used in the assay.
References and notes
17. It has been shown that CXCR3 plays an important role in noninfectious lung
injury induced by bleomycin instilled intratracheally into CXCR3-deficient
C57BL/6J mice background and WT littermate controls. Examination of the
inflammatory response after bleomycin-induced lung injury revealed an
insignificant reduction in total inflammatory cells in the bronchoalveolar
lavage (BAL) fluid from CXCR3-deficient animals as compared with WT
animals. Differential counts of the BAL cells revealed fewer total lymphocytes
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CXCR3: a Review of Current Progress. In Chemkine Biology—Basic Research and
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Publishers, 2007; Vol. II, p 79. 2.
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in CXCR3-deficient mice after bleomycin-induced lung injury at day
7
compared with WT controls. See: Jiang, D.; Liang, J.; Hodge, J.; Lu, B.; Zhu,
Z.; Yu, S.; Fan, J.; Gao, Y.; Yin, Z.; Homer, R.; Gerard, C.; Noble, P. W. J. Clin.
Invest. 2004, 114, 291.
18. The 10 mg/ml group used 4.8 mg/kg/day of compound; 3 mg/ml used 1.44 mg/
kg/day of compound; and 1 mg/ml used 0.48 mg/kg/day of compound. For
more information of Alzet osmotic mini-pump, please visit http://