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the compounds in this series, two compounds 11 and 25 exhibited
References and notes
not only good inhibitory activity against HCV but also low cytotox-
icity, resulting in high selectivity index values (SI = 21 and 25,
respectively).
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With preliminary SAR data in hand, we explored R1 and R2 sub-
stitution effects with various amines. The results are summarized
in Table 3. Interestingly, compounds 29–32 bearing basic diamine
moiety apparently demonstrated good antiviral activity (EC50
=
0.28–1.0 M) and low cytotoxicity (CC50 = 8.8–22 M), resulting
l
l
in a high selectivity index (SI = 18–41). In addition, these molecules
contain three carbons between two nitrogens. However, methylpi-
perazine (33) and compounds containing mono-amine (34–39)
slightly reduced anti-HCV activity. Piperidine–carboxamide com-
pound 40 and 41 even further deteriorated inhibitory activity.
Among the compounds tested, isopropyl piperazine 32 turned
out to be the most promising in this series.
By using the infectious HCV cell culture system for primary
screening which allows targeting the entire viral life cycle, the
mechanism of action (MoA) and the targets of potent hit com-
pounds are initially unknown. In order to identify which part of
the viral life cycle is inhibited, we devised strategies to elucidate
the MoA of the 7-APP scaffold. To discriminate between viral RNA
replication and HCV entry mediated by the viral envelope pro-
teins E1 and E2, we conducted experiments with the HCV repli-
con (HCVrp) and the HCV pseudoparticle (HCVpp) system,
respectively. HCV subgenomic replicons express only the non-
structural protein NS3 to NS5B,27 whereas HCVpp are pseudo-
typed retroviral particles expressing HCV E1 and E2.28,29 The
inhibitory effect of 7-APP on viral RNA replication was examined
in HCVrp cells replicating genotype 2 subgenomes and in the
HCVpp system expressing HCV genotype 1 envelope proteins (Ta-
ble 4). The 7-APPs were inactive in the HCVrp or in the HCVpp as-
say, suggesting that 7-APP is neither a viral RNA replication
inhibitor nor a HCV E1/E2-mediated entry inhibitor. In fact, the
result that 7-APP is inactive against HCV replicon corroborates
the previous report.26
Subsequently, pharmacological properties including solubility
and microsomal stability were evaluated. The metabolic stability
assay indicated that compound 32 exhibited long half-life in both
human and rat (t1/2 > 60 min), whereas compound 29 showed rel-
atively shorter half-life in rat microsomes (t1/2 = 22.4 min). Both
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Corsi, M.; Capelli, A. M. Neuropharmacology 2011, 61, 957.
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compounds showed good solubility (76 and 73
in neutral buffer conditions.
lM, respectively)
In conclusion, we described the discovery and characterization
of a novel class of compounds with anti-HCV activity. This series
of compounds appears to inhibit HCV life cycle, but does not inter-
fere with either HCV genotype 1 E1/E2-mediated entry or genotype
2 RNA replication of viral. Although the exact mechanism of action
of this series remains unclear, the 7-APP scaffold can be utilized as
a new tool compound for further anti-HCV studies.
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Acknowledgments
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McKeating, J. A. Proc. Natl. Acad. Sci. U.S.A. 2003, 100, 7271.
This work was supported by the National Research foundation
of Korea (NRF) grant funded by the Korea government (MEST)
(No.2011-00244 and 2012-00011), Gyeonggi-do and KISTI.