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metabolic stability over 7d with a slight decrease in activity. With
28, a lipophilic region around the 2,3,4-position of the pyridine
ring of type A might contribute to these unfavorable profiles. Lead
compound 7d and compound 25b, one of the extremely balanced
derivatives, were selected for evaluation in a rat PK model. A sum-
mary of key properties is shown in Table 4.
The half-life of 25b was increased at 4.1 h compared to 7d at
1.8 h, and the plasma exposure of 25b at 1.0 mg/kg, po was almost
equal to that of 7d at 3.0 mg/kg, po. Further, the oral bioavailability
of 25b was markedly increased to 78% compared to 27% with 7d.
We believe that proper adjustment of lipophilicity brought about
these PK improvements. In addition, 25b neither blocked the hu-
man ether a-go-go-related gene (hERG)-encoded potassium chan-
nel nor showed phototoxicity (data not shown).
6. Yamamoto, T.; Yamada, H.WO2004/089412.
7. Cayatte, A. J.; Rupin, A.; Oliver-Krasinski, J.; Maitland, K.; Sansilvestri-Morel, P.;
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9. Inhibitory activity against ROS production derived from NADPH was carried
out as follows: HUVECs were seeded at a density of 3 ꢂ 104 cells/well (100
lL)
into a 96-well plate coated with collagen; culture medium was MCDB131 (10%
fetal bovine serum, 2 mM glutamine, 100 U/mL penicillin, 100 U/mL
streptomycin, and 10 ng/mL recombinant human basic-FGF). After overnight
incubation, the culture medium was removed via aspirator, and test
compounds were added along with 100
lL/well of phosphate buffered saline
containing 25 mM glucose, 200 M NADPH, and 0.2% nitrotetrazolium blue,
l
In conclusion, we synthesized a series of quinolone derivatives
using a variety of substituents as a new class of inhibitors of ROS
production in HUVECs. Generally, the inhibitory activity of these
compounds was extremely potent compared to DPI, and addition
of appropriate substituents to the 2-position of the quinolone ring
induced obvious improvement in CYP2D6 inhibitory activity, solu-
bility, and metabolic stability in vitro compared with our seed
compound 1. In particular, compound 25b, one of the most bal-
anced compounds with respect to the above-mentioned issues
and ROS production inhibitory activity, showed excellent in vivo
PK profiles in SD rats. We believe that these derivatives, including
25b, may be useful in treating various diseases related to excessive
ROS production, and results of pharmacological studies on these
derivatives and their inhibition mechanisms will be published in
due course.
followed by culturing at 37 °C. Nitrotetrazolium blue reacts with intracellularly
produced ROS and turns into a water-insoluble blue pigment. After 2 h, the
supernatant was discarded, and the wells were washed three times with
phosphate buffered physiological saline. DMSO water (90%, 100 lL/well)
containing 0.04 M sodium hydroxide was added to each well to completely
dissolve the pigment, and absorbance was measured at 715 nm. ROS
production inhibitory rate was calculated using the following equation:
Inhibition rateð%Þ ¼ ðA ꢀ BÞ=ðA ꢀ CÞ ꢂ 100
where A is absorbance with no addition of a test compound, B is absorbance
with addition of a test compound, and C is absorbance with no addition of glu-
cose, NADPH, and a test compound.
10. Tatsuta, K.; Tamura, T. J. Antibiot. 2000, 53, 418.
11. CYP2D6 inhibition test was carried out in accordance with following reference.
Sato, I.; Morihira, K.; Inami, H.; Kubota, H.; Morokata, T.; Suzuki, K.; Iura, Y.;
Nitta, A.; Imaoka, T.; Takahashi, T.; Takeuchi, M.; Ohta, M.; Tsukamoto, S.
Bioorg. Med. Chem. 2008, 16, 8607; Solubility was tested as following reference.
Sugaya, Y.; Yoshiba, T.; Kajima, T.; Ishihama, Y. Yakugaku Zasshi 2002, 123, 237;
In vitro liver microsomal stabilities were evaluated using below method
(0.2 mg/mL protein, 0.2 lM substrate concentration). Sato, I.; Morihira, K.;
Inami, H.; Kubota, H.; Morokata, T.; Suzuki, K.; Ohno, K.; Iura, Y.; Nitta, A.;
Imaoka, T.; Takahashi, T.; Takeuchi, M.; Ohta, M.; Tsukamoto, S. Bioorg. Med.
Chem. 2009, 17, 5989.
Supplementary data
Supplementary data associated with this article can be found, in
12. Tanaka, M.; Okita, M.; Miyamoto, M.; Kaneko, T.; Kawahara, T.; Akamatsu, K.;
Chiba, K.; Obaishi, H.; Sakurai, H.; Abe, S.; Kobayashi, S.; Yamanaka, T. WO1996/
36608. Tatsuta, K. JP2001/233859.
13. Aldridge, G. R.; Jaffe, E. E.; Matrick, H. US 3334102.
References and notes
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