1
02
Matsumoto et al.
gest that the nitrile group would markedly perturb the elec- effects of FYX-051 may arise from its high affinity for XOR
tronic environment of the molybdopterin cofactor. and stable binding to XOR in vitro. Therefore, FYX-051
In previous in vivo study, allopurinol is immediately con- appears to be a promising candidate drug for the treatment
verted to oxipurinol, which is detected in human plasma and of gout and hyperuricemia.
urine at relatively high concentration (Hosoya et al., 1991),
whereas only a little 2-hydroxy-FYX-051 is detected in hu- Acknowledgments
man urine after the administration of FYX-051 (Nakazawa et
We thank Drs. Tomohiro Matsumura and Teruo Kusano (Nippon
al., 2006). In the present study, we investigated in vitro Medical School, Tokyo, Japan) for data collection of crystallographic
hydroxylation by XO to clarify the difference in metabolic data; Mr. Koichi Omura and Dr. Takahiro Sato (Fuji Yakuhin,
Saitama, Japan) for LC/MS and NMR analyses; officials at the Pho-
behavior between two inhibitors in vivo. These results imply
ton Factory (Tsukuba, Japan) for beam time; and the beam line staff
of NW12A for support.
that 2-hydroxy-FYX-051 enzyme complex is so stable (Fig. 3;
t1/2 ϭ approximately 20 h) that only a little 2-hydroxy-FYX-
0
51 can be dissociated from the complex (Fig. 8). Further-
Authorship Contributions
more, 2-hydroxy-FYX-051 was slightly increased when incu-
bation was continued, and no formation of metabolites with
two or three hydroxyl groups was observed until nonreacted
FYX-051 was completely consumed (data not shown). It is
likely that this reflects the difference in binding affinity
between FYX-051 and 2-hydroxy-FYX-051 for the enzyme. It
is likely that the conversion of allopurinol to oxipurinol con-
Participated in research design: Matsumoto, Okamoto, Ashizawa,
and Nishino.
Conducted experiments: Matsumoto, Okamoto, and Ashizawa.
Contributed new reagents or analytic tools: Matsumoto and
Ashizawa.
Performed data analysis: Matsumoto, Okamoto, and Nishino.
Wrote or contributed to the writing of the manuscript: Matsumoto,
tinues until the oxipurinol-enzyme complex is fully formed, Okamoto, and Nishino.
because allopurinol is a good substrate for XOR. This oxipuri-
nol, which is deposited in the kidney during renal excretion,
sometimes causes severe adverse effects in patients with
renal insufficiency (Hande et al., 1984). In the case of FYX-
Other: Nishino acquired funding for the research.
References
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0
2
51, such an adverse effect would not be expected, because
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9
) arise from its high affinity for the enzyme and stable
binding to the enzyme.
In conclusion, we have studied in detail how FYX-051
interacts with bovine milk XOR. It was found that the 2-hy-
droxy-FYX-051-XOR complex formed is more stable than the
oxipurinol-XOR complex, although FYX-051, as well as allo-
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Ϫ9
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i
allopurinol. Moreover, we found that XOR hydroxylates three
carbons of FYX-051 and finally generates a trihydroxylated
derivative, which forms a covalent linkage to molybdenum
via the nitrile group and brings a distinct spectral change of
the enzyme. However, only a little 2-hydroxy-FYX-051 was
detected, and no other hydroxylation metabolites were ob-
served in an in vivo study (Nakazawa et al., 2006). Dihy-
droxylated and trihydroxylated forms of FYX-051 ap-
peared only when the enzyme-inhibitor complex was
incubated under artificial conditions, that is, under the
condition of excess enzyme. Therefore, the pharmacologi-
cal effects of the dihydroxylated and trihydroxylated me-
tabolites should have no impact in the clinical context.
Moreover, we have demonstrated that FYX-051 exerts a
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