Structural development studies of anti-hepatitis C virus agents with a phenanthridinone skeleton

Article abstract of DOI:10.1016/j.bmc.2010.02.057

A phenanthridinone skeleton was derived from our previous researches on thalidomide and retinoids as a multi-template for generation of anti-viral lead compounds. Structural development studies focusing on anti-hepatitis C virus activity afforded 5-butyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)phenanthridin-6(5H)-one (10) and 5-butylbenzo[b]phenanthridin-6(5H)-one (39), which showed EC₅₀ values of approximately 3.7 and 3.2 μM, respectively.

Full text of DOI:10.1016/j.bmc.2010.02.057

Bioorganic & Medicinal Chemistry 18 (2010) 2402–2411
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Structural development studies of anti-hepatitis C virus agents
with a phenanthridinone skeleton
Masahiko Nakamura a, Atsushi Aoyama a, Mohammed T. A. Salim b, Mika Okamoto b, Masanori Baba b,
,
*
Hiroyuki Miyachi c, Yuichi Hashimoto a, Hiroshi Aoyama a,
*
a Institute of Molecular & Cellular Biosciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-0032, Japan
b Division of Antiviral Chemotherapy, Center for Chronic Viral Diseases, Graduate School of Medical and Dental Sciences, Kagoshima University, 8-35-1 Sakuragaoka, Kagoshima
890-8544, Japan
c Division of Pharmaceutical Sciences, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 1-1-1 Tsushima-Naka, Kita-ku, Okayama
700-8530, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
A phenanthridinone skeleton was derived from our previous researches on thalidomide and retinoids as a
multi-template for generation of anti-viral lead compounds. Structural development studies focusing on
anti-hepatitis C virus activity afforded 5-butyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)phenan-
thridin-6(5H)-one (10) and 5-butylbenzo[b]phenanthridin-6(5H)-one (39), which showed EC50 values
Received 10 February 2010
Revised 24 February 2010
Accepted 25 February 2010
Available online 2 March 2010
of approximately 3.7 and 3.2 lM, respectively.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Anti-HCV agents
Phenanthridinone
Thalidomide
Am80
Am580
1. Introduction
thalidomide (1) and/or Am80 (2)/Am580 (3) as multi-templates
to develop anti-viral agents and/or anti-proliferative agents for
virus-infected cells, that is, anti-bovine viral diarrhea virus (anti-
BVDV) agents, including SK3M4M5M (4) derived from thalidomide
(1)14,15 and adult T-cell leukemia (ATL) cell-selective proliferation
inhibitors, including TMN(COCH3) (5) and TMN(OH)(COCH3) (6)
derived from Am80 (2)/Am580 (3) (Fig. 1).4 We next aimed to de-
velop anti-hepatitis C virus (HCV) agents.
HCV infection is thought to be a major cause of human hepati-
tis,16,17 and it is estimated that at least 170 million people world-
wide are chronically infected with this virus.18 Most infections
become persistent and about 60% of cases progress to chronic liver
disease, which in turn can lead to development of cirrhosis, hepa-
tocellular carcinoma, and liver failure.19,20 Currently, no vaccine is
available against HCV infection, and the standard treatment for
The efficient identification of small-molecular scaffolds for the
development of biologically active compounds is very important
in chemical genetics and medicinal chemistry. As one approach,
we have been utilizing the multi-template hypothesis,1–5 based
on the idea that the number of protein fold structure types that
comprise all the domains occurring in natural proteins is quite lim-
ited, in spite of the huge number of natural proteins.6–8 A given fold
structure might be characteristic of many natural proteins, and
therefore, ignoring physical/chemical interactions, one might ex-
pect that a template/scaffold structure which is spatially comple-
mentary to one fold structure might serve as a multi-template
for structural development of ligands that would interact specifi-
cally with many different natural proteins. As candidate multi-
template structures, we have focused particularly on thalidomide
(1) and retinoids, including synthetic retinoids Am80 (2) and
Am580 (3) (Fig. 1).1,9–13 All of these compounds 1–3 elicit a wide
range of biological activities, and thalidomide (1) is well-estab-
lished to be multi-target drug.1,9–13 In fact, we recently applied
chronic hepatitis C consists of pegylated interferon (IFN)-
a in
combination with the nucleoside analog ribavirin (1-b- -ribofur-
D
anosyl-1,2,4-triaxole-3-carboxamide). However, the virus cannot
be eliminated from approximately half of infected patients treated
with these agents.21 In addition, the side effects of these agents are
sometimes serious and unacceptable to patients. Therefore, alter-
native agents for the treatment and prevention of HCV infection
are urgently needed.
* Corresponding authors. Tel.: +81 3 5841 7848; fax: +81 5841 8495 (H.A.); tel.:
+81 99 275 5930; fax: +81 99 275 5932 (M.B.).
E-mail addresses: m-baba@vanilla.ocn.ne.jp (M. Baba), aoyama@iam.u-tokyo.
ac.jp (H. Aoyama).
As mentioned above, we have been succeeded in the develop-
ment of potent anti-BVDV agents, including SK3M4M5M (4), which
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2010.02.057

M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
2403
Figure 1. Armchair structural development of thalidomide (1) and retinoids (2, 3) to the first scaffold 8.
have an EC50 value of 3.5 nM.15 Although BVDV belongs to the Fla-
viviridae family, as HCV does,22 and is thought to be a surrogate
model for HCV,23–25 SK3M4M5M (4) showed only a very weak
activity against HCV. Therefore, we tried to develop another scaf-
fold for the development of anti-HCV agents, as shown in Figure 1.
ing of teterahydrotetramethylnaphthalene moiety and N-ethylphe-
nanthridinone (7) would afford other scaffold candidate structures,
as shown in Figure 1.
Compound 8 was prepared as shown in Scheme 1.26,27 Briefly,
N-ethylaniline was treated with hexafluoroacetone to give the
1,1,1,3,3,3-hexafluoro-2-hydroxypropyl derivative, which was cou-
pled with 2-iodobenzoyl chloride. The resulting anilide was cy-
clized to give compound 8. The anti-HCV activity of compound 8
was determined in the established HCV RNA replicon cells.28
Briefly, NNC #2 cells carrying full-genomic HCV RNA replicons
were cultured in the presence of various concentrations of the test
compounds for 3 days. The cells were examined for HCV RNA and
glyceraldehyde-3-phosphate dehydrogenase (GAPDH) RNA levels
by real-time reverse transcription (RT)-PCR. The anti-HCV activity
and cytotoxicity of test compounds were expressed as 50% effec-
tive concentration (EC50) and 50% cytotoxic concentration (CC50),
defined in terms of decrease of HCV RNA and GAPDH RNA levels
to 50% of the respective control levels. As shown in Figure 2, com-
pound 8 showed apparent anti-HCV activity (EC50 and CC50: 14.1
2. Results and discussion
2.1. Armchair structural development leading to phenanthrid-
inone derivatives
First, we applied armchair structural development to phenylph-
thalimide, which was itself developed from thalidomide (1). The
phenylphthalimide skeleton is a superior multi-template, and we
have developed various biologically active phenylphthalimide
derivatives, including tumor necrosis factor-
tors, tubulin polymerization inhibitors, dipeptidylpeptidase type
IV inhibitors, liver X receptor antagonists, -glucosidase inhibitors,
a production regula-
a
and so on.1,9–12 The armchair ring opening of phenylphthalimide
gave N-ethylbenzanilide, whose recyclization should afford N-ethyl-
phenanthridinone (7) (Fig. 1). On the other hand, synthetic retinoids
Am80 (2) and Am580 (3) both possess a benzoic acid moiety.
Because our previous studies on anti-BVDV agents suggested that
carboxyl acid derivatives are not suitable as lead structures, we con-
sidered that a bioisosteric functional group, the 1,1,1,3,3,3-hexaflu-
oropropan-2-ol-substituted phenyl moiety, might be introduced
into N-ethylphenanthridinone (7) to give the first scaffold structure,
5-ethyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl) phenan-
thridin-6(5H)-one (8) (Fig. 1). In addition, both Am80 (2) and
Am580 (3) possess a teterahydrotetramethylnaphthalene moiety,
which has been established to be a useful core structure to develop
ATL cell-selective proliferation inhibitors, including TMN(COCH3)
(5) and TMN(OH)(COCH3) (6).4 We therefore considered that dock-
and >40 lM, respectively), suggesting that compound 8 could be
a lead compound for structural development of anti-HCV agents.
2.2. Effects of N-substituents
Since compound 8 showed anti-HCV activity, we examined the
effects of N-substituents. Several N-alkylated derivatives of 8 (9–
14) were prepared as shown in Scheme 2, and their anti-HCV activ-
ity was measured as described above (Figs. 2 and 3).
As shown in Figures 2 and 3, introduction of three fluorine
atoms at the terminal methyl group (9) did not improve the activ-
ity. However, introduction of a longer-chain alkyl group, n-butyl
(10), n-hexyl (11), or n-nonyl (12), resulted in enhancement of
anti-HCV activity, though at the same time, the cytotoxicity was
increased. The anti-HCV activity of these compounds decreased

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M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
Scheme 1.
log (14) showed moderate anti-HCV activity (EC50: 7.7 and 6.7
respectively) with weaker cytotoxicity (CC50: 18.2 and 14.4
l
l
M,
M,
respectively). Cytotoxicity was not affected by the length of the
introduced alkyl group (CC50: 10.2–12.5 M). Among this series,
compound 10 showed the most potent anti-HCV activity (EC50
3.7 M). Therefore, we selected the N-butylphenanthridinone skel-
l
:
l
eton (corresponding to compound 10) as a scaffold structure for
further structural development studies.
2.3. Regioisomers of methyl-substituted N-butylphenanthrid-
inone
Figure 2. Effects of N-substituents on anti-HCV activity. EC50
concentration, based on the decrease of the amount of HCV RNA. CC50
:
50% effective
50%
:
cytotoxic concentration, based on the decrease of the amount of GAPDH RNA.
Based on our previous structure–activity relationship studies of
anti-BVDV
c-carboline analogs, which indicated that regio-selec-
tive methyl-substitution dramatically influenced the anti-viral
activity,14,15 the effect of methyl-substitution was investigated.
For this purpose, we synthesized all the regioisomers of methyl-
in the order of: 10 > 11 > 12. On the other hand, their cytotoxicity
tended to decrease in the reverse order, though the differences
were small. The benzyl analog (13) and the cyclohexylmethyl ana-
Scheme 2.
Figure 3. Dose-dependency curves of anti-HCV and cytotoxic activity elicited by compounds 8 and 10–14.

M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
2405
substituted N-butylphenanthridinone (Fig. 4) by the method
shown in Scheme 3.
As shown in Figure 4, N-butylphenanthridinone itself (an ex-
tracted structure from compound 10) was completely inactive.
However, methyl-substitution at position 2, that is, the position
at which the 1,1,1,3,3,3-hexafluoropropan-2-ol group was attached
in compound 10, resulted in the appearance of anti-HCV activity
(EC50: 42.0 lM). All the other regioisomers, except the 8-methyl
analog (21), that is, compounds 16, 18–20, 22, and 23, were inac-
tive, and the anti-HCV activity elicited by 21 was very weak. There-
fore, position 2 seems to be the best position at which to introduce
a substituent.
2.4. Effect of 2-subsituents on anti-HCV activity
The results described above prompted us to examine the effect
of 2-substitutents, and we prepared various derivatives (24–33) as
shown in Figure 5. Although some derivatives, including com-
pounds 28–33, showed improved anti-HCV activity compared to
the 2-methyl analog 17, their activity was weaker than that of
10. As already mentioned, the unsubstituted analog 15 was inac-
tive. Its fluoro analog 24 was also inactive, suggesting that a
mono-atomic substituent is inappropriate to improve the activity.
Figure 5. Anti-HCV activity of 2-substituted N-butylphenanthridinone, 15, 17 and
24–33.
Compounds 28–33 showed moderate anti-HCV activity (EC50
:
10.6–21.8 M). Among the 2-alkylated derivatives, the activity de-
l
creased in the order of t-butyl (31) > i-propyl (28) > ethyl
(27) > methyl (17), suggesting that the hydrophobicity of the 2-
substituent contributes to the activity, at least in part. On the other
hand, introduction of a hydroxyl group into the ethyl (30) or
methyl (29) group of 27 and 17, respectively, seemed to slightly
Though the 2-methyl analog 17 showed some activity (EC50
:
35.8 M), its trifluoromethyl analog 26 was inactive. On the con-
l
trary, its hydroxyl analog 25 showed activity comparable to that
of the 2-methyl analog 17. The 2-ethyl analog 27 also showed
activity comparable to that of 17/25.
enhance the activity, that is, 27 (EC50: 34.7
lM) versus 30 (EC50
:
21.8 M) and 17 (EC50: 35.8 M) versus 29 (EC50: 21.5 l
l
l
M). The
2-acetyl derivative 32 showed slightly more potent activity than
30.
2.5. Tetrahydrotetramethylnaphthalene-related analogs and
benzophenanthridinone analogs
Although the structure–activity relationships of 2-substituted
N-butylphenanthridinone were clearly interpretable, hydrophobic-
ity around the 2-position also seemed to contribute to the activity.
This and the armchair structural development shown in Figure 1
prompted us to prepare compounds 34–38 (Fig. 6). We also pre-
pared benzophenanthridinone derivatives 39–44 (Fig. 7). Among
the tetrahydrotetramethylnaphthalene-related analogs 34–38,
only the N-butyl and N-benzyl derivatives (35 and 36, respectively)
were moderately active, as shown in Figure 6. The regioisomer of
35, that is, compound 38, was inactive.
Figure 4. Anti-HCV activity of methyl-substituted regioisomers of N-butylphe-
nanthridinone, 16–23.
Scheme 3.

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M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
Figure 6. Anti-HCV activity of tetrahydrotetramethylnaphthalene-related analogs, 34–38.
Figure 7. Anti-HCV activity of benzophenathridinone derivatives, 39–44.
Benzophenanthridinone derivatives 39–44 were prepared as
shown in Scheme 4. All the benzophenanthridinone derivatives,
except 43, showed potent or moderate anti-HCV activity. The activ-
ity decreased in the order of: benzo[b] (39) > benzo[j] (40) > ben-
zo[c] (41) > benzo[a] (42) > benzo[i] (44) >> benzo[k] (43) as
shown in Figures 7 and 8. Although compound 39 showed the most
Scheme 4.

M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
2407
Figure 8. Dose-dependency curves of anti-HCV and cytotoxic activity elicited by compounds 39 and 40.
potent anti-HCV activity (EC50: 3.2
this paper, its selectivity index (SI = CC50/EC50) was low (SI: 1.97).
In terms of SI, compound 40 (EC50: 11.0 lM) seems to be the best
(SI: >3.64).
l
M) among the compounds in
carbonate (2 equiv) and N,N-dimethylacetamide were added palla-
dium (II) acetate (10 mol %) and tricyclohexylphosphine tetrafluo-
roborate (0.15 equiv), and the mixture was heated to 130 °C. The
catalyst was filtered off and washed several times with ethyl
acetate. The combined organic layers were washed with water
and brine successively, dried over anhydrous magnesium sulfate,
and concentrated. The residue was purified by silica gel column
chromatography (eluent; n-hexane/ethyl acetate) to afford 5-
ethyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)phenanthri-
din-6(5H)-one (8) as a white solid. Mp 208.0–209.0 °C. 1H NMR
(500 MHz, CDCl3) d 8.68 (d, J = 1.5 Hz, 1H), 8.56 (dd, J = 7.9,
1.5 Hz, 1H), 8.31 (d, J = 7.9 Hz, 1H), 7.85 (d, J = 9.4 Hz, 1H), 7.80
(td, J = 7.9, 1.3 Hz, 1H), 7.63 (t, J = 7.7 Hz, 1H), 7.50 (d, J = 9.0 Hz,
1H), 4.48 (q, J = 7.3 Hz, 2H), 3.71 (s, 1H), 1.44 (t, J = 7.3 Hz, 3H).
HRMS (FAB) calcd for C18H14F6NO2 390.0929; found: 390.0918
(M+H)+.
3. Conclusion
Based on a phenanthridinone skeleton derived by armchair
structural development of thalidomide and retinoids, we devel-
oped candidate anti-HCV agents, 10 (EC50: 3.7
lM,), 39 (EC50:
3.2 M,), and 40 (EC50: 11 M, CC50: >40 M). Further structural
l
l
l
development may yield highly potent and selective drug
candidates.
4. Experimental
4.3. 2-(1,1,1,3,3,3-Hexafluoro-2-hydroxypropan-2-yl)-5-(2,2,2-
trifluoroethyl)phenanthridin-6(5H)-one (9)
4.1. General comments
Melting points were determined by using a Yanagimoto hot-
stage melting point apparatus and are uncorrected. 1H NMR spec-
tra were recorded on a JEOL JNM-GX500 (500 MHz) spectrometer.
Chemical shifts are expressed in d (ppm) values with tetramethyl-
silane (TMS) as an internal reference. The following abbreviations
are used: s = singlet, d = doublet, t = triplet, m = multiplet, dd =
double doublet, dt = double triplet, br = broad. Mass spectra (MS)
and high-resolution mass spectra (HRMS) were recorded on a JEOL
JMS-DX303 spectrometer. Elemental analysis was performed with
a Yanagimoto MT-6 elemental analyser.
The title compound was prepared by a method similar to that
described for the synthesis of 8, using aniline as a starting material,
with slight modifications. 2-(4-Aminophenyl)-1,1,1,3,3,3-hexaflu-
oropropan-2-ol (prepared from aniline and 1,1,1,3,3,3-hexafluoro-
acetone)29 was coupled with 2-iodobenzoyl chloride, followed by
protection of the hydroxyl group with 2-methoxylethoxymethyl
chloride. It was then N-alkylated by the use of 2,2,2-trifluoroethyl
triflate. The 2-methoxyethoxymethyl group of the obtained benza-
nilide derivative was removed by treatment with titanium tetra-
chloride, and the deprotected benzanilide derivative was cyclized
by the method used for the synthesis of 8. White solid. Mp 76.1–
77.8 °C. 1H NMR (500 MHz, CDCl3) d 8.70 (d, J = 1.1 Hz, 1H), 8.56
(dd, J = 8.1, 1.1 Hz, 1H), 8.33 (d, J = 8.1 Hz, 1H), 7.87 (dd, J = 7.7,
1.1 Hz, 1H), 7.85 (dd, J = 7.7, 1.1 Hz, 1H), 7.66 (td, J = 8.1 Hz, 1H),
7.51 (d, J = 9.0 Hz, 1H), 5.13 (s, 2H), 3.78 (s, 1H). HRMS (FAB) calcd
for C18H10F9NO2 444.0646; found: 444.0653 (M+H)+.
4.2. 5-Ethyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)-
phenanthridin-6(5H)-one (8)
To a solution of 5 mmol of N-ethylaniline and 25 mL of toluene
were added hexafluoroacetone trihydrate (1.2 equiv) and p-TsOH
(0.1 equiv). The mixture was stirred for 22 h at 120 °C and the
solvent was evaporated in vacuo. The residue was purified by silica
gel column chromatography (eluent; n-hexane/ethyl acetate) to
afford 2-(4-ethylaminophenyl)-1,1,1,3,3,3-hexafluoropropan-2-ol.
The obtained compound was dissolved in dichloromethane
(0.1 mmol/mL), and then triethylamine (1:40 v/v) and 2-iodoben-
zoyl chloride (1.2 equiv) were added. The mixture was stirred for
15 h at room temperature and the solvent was evaporated in va-
cuo. The residue was purified by silica gel column chromatography
(eluent; n-hexane/ethyl acetate) to afford benzanilide derivatives.
To a solution of the obtained benzanilide derivative, cesium
4.4. 5-Butyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)-
phenanthridin-6(5H)-one (10)
The title compound was prepared by a method similar to that
described for the synthesis of 9 using 1-iodobutane instead of
2,2,2-trifluoroethyl triflate. White solid. Mp 166.0–166.6 °C. 1H
NMR (500 MHz, CDCl3) d 8.67 (s, 1H), 8.55 (d, J = 7.9 Hz, 1H), 8.30
(d, J = 7.9 Hz, 1H), 7.84 (d, J = 7.9 Hz, 1H), 7.79 (dd, J = 7.9, 7.3 Hz,
1H), 7.62 (dd, J = 7.9, 7.3 Hz, 1H), 7.47 (d, J = 7.9 Hz, 1H), 4.39 (t,
J = 7.9 Hz, 2H), 3.81 (s, 1H), 1.83–1.76 (m, 2H), 1.56–1.50 (m, 2H),

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M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
1.03 (t, J = 7.3 Hz, 3H). HRMS (FAB) calcd for C20H18F6NO2
1H, J = 7.4, Hz), 7.53 (ddd, 1H, J = 8.5, 7.4, 1.2 Hz), 7.40 (d, 1H,
J = 8.5 Hz), 7.30 (dd, 1H, J = 8.0, 7.4 Hz), 4.38 (t, 2H, J = 7.9, Hz),
1.78 (td, 2H, J = 7.9, 7.3 Hz), 1.52 (sextet, 2H, J = 7.3 Hz), 1.00 (t,
3H, J = 7.3, Hz). HRMS (FAB) calcd for C17H17NO 252.1388; found:
252.1349 (M+H)+.
418.1242; found: 418.1262 (M+H)+.
4.5. 2-(1,1,1,3,3,3-Hexafluoro-2-hydroxypropan-2-yl)-5-hexyl-
phenanthridin-6(5H)-one (11)
The title compound was prepared by a method similar to that
described for the synthesis of compound 9, using 1-iodohexane in-
stead of 2,2,2-trifluoroethyl triflate. White solid. Mp 132.0–
132.2 °C. 1H NMR (500 MHz, CDCl3) d 8.67 (d, J = 1.8 Hz, 1H), 8.54
(d, J = 7.9, 1.8 Hz, 1H), 8.29 (d, J = 7.9 Hz, 1H), 7.84 (d, J = 8.5 Hz,
1H), 7.78 (dd, J = 8.5, 7.3 Hz, 1H), 7.62 (dd, J = 8.5, 7.3 Hz, 1H),
7.46 (d, J = 8.5 Hz, 1H), 4.37 (t, J = 7.9 Hz, 2H), 3.91 (s, 1H), 1.84–
1.76 (m, 2H), 1.53–1.47 (m, 2H), 1.42–1.32 (m, 4H), 0.91 (t,
J = 7.3 Hz, 3H). HRMS (FAB) calcd for C22H22F6NO2 446.1555;
found: 446.1516 (M+H)+.
4.10. 5-Butyl-1-methylphenanthridin-6(5H)-one (16)
The title compound was prepared by a method similar to that
described for the synthesis of 15, using 2-iodo-3-methylaniline
as a starting material. Benzoyl chloride was used instead of 2-iodo-
benzoyl chloride. Pale brown oil. 1H NMR (500 MHz, CDCl3) d 8.64
(d, 1H, J = 8.0 Hz), 8.44 (d, 1H, J = 8.0 Hz), 7.73 (t, 1H, J = 8.0 Hz),
7.59 (t, 1H, J = 8.0 Hz), 7.42 (t, 1H, J = 8.0 Hz), 7.33 (d, 1H,
J = 8.0 Hz), 7.17 (d, 1H, J = 8.0 Hz), 4.40 (t, 2H, J = 7.6 Hz), 2.96 (s,
3H), 184–1.77 (m, 2H), 158–1.48 (m, 2H), 1.02 (t, 3H, J = 7.6 Hz).
HRMS (FAB) calcd for C18H19NO 266.1545; found: 266.1568
(M+H)+.
4.6. 2-(1,1,1,3,3,3-Hexafluoro-2-hydroxypropan-2-yl)-5-nonyl-
phenanthridin-6(5H)-one (12)
4.11. 5-Butyl-2-methylphenanthridin-6(5H)-one (17)
The title compound was prepared by a method similar to that
described for the synthesis of compound 9, using 1-bromononane
instead of 2,2,2-trifluoroethyl triflate. White solid. Mp 98.1–
99.0 °C. 1H NMR (500 MHz, CDCl3) d 8.67 (d, J = 1.8 Hz, 1H), 8.54
(d, J = 7.9, 1.8 Hz, 1H), 8.29 (d, J = 7.9 Hz, 1H), 7.84 (d, J = 8.5 Hz,
1H), 7.79 (dd, J = 7.9, 7.3 Hz, 1H), 7.62 (dd, J = 7.9, 7.3 Hz, 1H),
7.46 (d, J = 8.5 Hz, 1H), 4.37 (t, J = 7.9 Hz, 2H), 3.87 (s, 1H), 1.84–
1.76 (m, 2H), 1.53–1.46 (m, 2H), 1.43–1.36 (m, 2H), 1.33–1.24
(m, 8H), 0.88 (t, J = 7.3 Hz, 3H). HRMS (FAB) calcd for C25H28F6NO2
488.2024; found: 488.1981 (M+H)+.
The title compound was prepared by a method similar to that
described for the synthesis of 15, using 4-methylaniline as a start-
ing material. 2-Bromobenzoyl chloride was used instead of 2-iodo-
benzoyl chloride. Colorless oil. 1H NMR (500 MHz, CDCl3) d 8.55
(dd, 1H, J = 8.0, 1.2 Hz), 8.28 (d, 1H, J = 8.0 Hz), 8.10 (s, 1H), 7.74
(td, 1H, J = 8.0, 1.2 Hz), 7.57 (m, 1H), 7.37–7.30 (d, 1H, J = 7.6 Hz),
4.38 (t, 2H, J = 7.6 Hz), 2.49 (s, 3H), 1.82–1.75 (m, 3H), 1.56–1.48
(m, 2H), 1.01 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for C18H19NO
266.1545; found: 266.1584 (M+H)+.
4.7. 5-Benzyl-2-(1,1,1,3,3,3-hexafluoro-2-hydroxypropan-2-yl)-
phenanthridin-6(5H)-one (13)
4.12. 5-Butyl-3-methylphenanthridin-6(5H)-one (18)
The title compound was prepared by a method similar to that
described for the synthesis of 16, using 2-bromo-5-methylaniline
as a starting material. Pale brown oil. 1H NMR (500 MHz, CDCl3)
d 8.53 (d, 1H, J = 8.0 Hz), 8.24 (d, 1H, J = 8.0 Hz), 8.80 (d, 1H,
J = 8.0 Hz), 7.73 (t, 1H, J = 8.0 Hz), 7.55 (t, 1H, J = 8.0 Hz), 7.20 (s,
1H), 7.13 (d, 1H, J = 8.0 Hz), 4.39 (t, 2H, J = 7.5 Hz), 2.52 (s, 3H),
1.83–1.76 (m, 2H), 1.57–1.49 (m, 2H), 1.03 (t, 3H, J = 7.5 Hz). HRMS
(FAB) calcd for C18H19NO 266.1545; found: 266.1512 (M+H)+.
The title compound was prepared by a method similar to that
described for the synthesis of compound 9, using benzyl bromide
instead of 2,2,2-trifluoroethyl triflate. White solid. Mp 80.0–
80.9 °C. 1H NMR (500 MHz, CDCl3) d 8.67 (d, J = 1.3 Hz, 1H), 8.63
(d, J = 8.1, 1.3 Hz, 1H), 8.34 (d, J = 8.1 Hz, 1H), 7.84 (td, J = 7.2,
1.3 Hz, 1H), 7.68 (q, J = 8.3 Hz, 2H), 7.38 (d, J = 9.4 Hz, 1H), 7.34–
7.23 (m, 5H), 5.66 (s, 2H), 3.63 (s, 1H). HRMS (FAB) calcd for
C23H15F6NO2 452.1085; found: 452.1057 (M+H)+. Anal. Calcd for
C23H15F6NO2ꢀ1/3 H2O: C, 60.40; H, 3.45; N, 3.06. Found: C, 60.46;
H, 3.47; N, 3.08.
4.13. 5-Butyl-4-methylphenanthridin-6(5H)-one (19)
The title compound was prepared by a method similar to that
described for the synthesis of 17, using 2-methylaniline as a start-
ing material. Colorless oil. 1H NMR (500 MHz, CDCl3) d 8.50 (dd, 1H,
J = 8.0, 1.2 Hz), 8.22 (d, 1H, J = 8.0 Hz), 8.13 (d, 1H, J = 8.0 Hz), 7.20
(td, 1H, J = 7.6, 1.4 Hz), 7.55 (t, 1H, J = 8.0 Hz), 7.30 (d, 1H,
J = 7.6 Hz), 7.21 (t, 1H, J = 7.6 Hz), 4.49 (t, 2H, J = 7.6 Hz), 2.66 (s,
3H), 1.66–1.60 (m, 2H), 1.20–1.19 (m, 2H), 0.86 (t, 3H, J = 7.3 Hz).
HRMS (FAB) calcd for C18H19NO 266.1545; found: 266.1557
(M+H)+.
4.8. 5-(Cyclohexylmethyl)-2-(1,1,1,3,3,3-hexafluoro-2-hydroxy-
propan-2-yl)phenanthridin-6(5H)-one (14)
The title compound was prepared by a method similar to that
described for the synthesis of compound 9, using cyclohexylmethyl
bromide instead of 2,2,2-trifluoroethyl triflate. White solid. Mp
200.9–201.7 °C. 1H NMR (500 MHz, CDCl3) d 8.67 (d, J = 1.8 Hz,
1H), 8.55 (d, J = 8.5, 1.8 Hz, 1H), 8.30 (d, J = 8.5 Hz, 1H), 7.83 (d,
J = 8.5 Hz, 1H), 7.79 (dd, J = 8.5, 7.3 Hz, 1H), 7.62 (dd, J = 8.5,
7.3 Hz, 1H), 7.46 (d, J = 8.5 Hz, 1H), 4.30 (br s, 2H), 3.75 (s, 1H),
1.98–1.88 (m, 1H), 1.76–1.64 (m, 5H), 1.24–1.17 (m, 5H). HRMS
(FAB) calcd for C23H22F6NO2 458.1555; found: 458.1569 (M+H)+.
4.14. 5-Butyl-7-methylphenanthridin-6(5H)-one (20)
The title compound was prepared by a method similar to that
described for the synthesis of 16, using 2-bromoaniline as a start-
ing material. 2-Methylbenzoyl chloride was used instead of
benzoyl chloride. White amorphous solid. 1H NMR (500 MHz,
CDCl3) d 8.27 (d, 1H, J = 7.5 Hz), 8.17 (d, 1H, J = 7.5 Hz), 7.58 (t,
1H, J = 7.5 Hz), 7.51 (t, 1H, J = 7.5 Hz), 7.35 (t, 1H, J = 7.5 Hz), 7.26
(t, 1H, J = 7.5 Hz), 4.31 (t, 2H, J = 8.0 Hz), 2.98 (s, 3H), 1.81–1.74
(m, 2H), 1.58–1.49 (m, 2H), 1.02 (t, 3H, J = 7.3 Hz). HRMS (FAB)
calcd for C18H19NO 266.1545; found: 266.1573 (M+H)+.
4.9. 5-Butylphenanthridin-6(5H)-one (15)
The title compound was prepared by a method similar to that
described for the synthesis of 9, using aniline as a starting material,
with slight modifications, that is, 1-iodobutane was used in place
of 2,2,2-trifluoroethyl triflate. Colorless oil. 1H NMR (500 MHz,
CDCl3) d 8.54 (dd, 1H, J = 7.9, 1.3 Hz), 8.29 (dd, 1H, J = 8.6, 1.3 Hz),
8.27 (d, 1H, J = 8.6 Hz), 7.74 (ddd, 1H, J = 8.0, 7.4, 1.2 Hz), 7.57 (t,

M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
2409
4.15. 5-Butyl-8-methylphenanthridin-6(5H)-one (21)
J = 8.0, 1.5 Hz), 7.34 (d, 1H, J = 8.0 Hz), 4.39 (t, 2H, J = 8.0 Hz), 2.79
(q, 2H, J = 8.0 Hz), 1.83–1.76 (m, 2H), 1.57–1.49 (m, 2H), 1.34 (t,
3H, J = 7.6 Hz), 1.02 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for
C19H21NO 280.1701; found: 280.1696 (M+H)+.
The title compound was prepared by a method similar to that
described for the synthesis of 20. 3-Methylbenzoyl chloride was
used instead of 2-methylbenzoyl chloride. White amorphous solid.
1H NMR (500 MHz, CDCl3) d 8.36 (s, 1H), 8.28 (d, 1H, J = 8.0 Hz),
8.18 (d, 1H, J = 8.0 Hz), 7.58 (dd, 1H, J = 8.0, 1.8 Hz), 7.52 (t, 1H,
J = 8.0 Hz), 7.41 (d, 1H, J = 8.0 Hz), 7.30 (t, 1H, J = 8.0 Hz), 4.40 (t,
2H, J = 8.0 Hz), 2.52 (s, 3H), 1.83–1.76 (m, 2H), 1.56–1.50 (m, 2H),
1.02 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for C18H19NO 266.1545;
found: 266.1588 (M+H)+.
4.21. 5-Butyl-2-isopropylphenanthridin-6(5H)-one (28)
The title compound was prepared by a method similar to that
described for the synthesis of 15, using 4-isopropylaniline as a
starting material. Colorless oil. 1H NMR (500 MHz, CDCl3) d 8.54
(dd, 1H, J = 8.0, 1.2 Hz), 8.30 (d, 1H, J = 8.0 Hz), 8.12 (d, 1H,
J = 1.9 Hz), 7.74 (ddd, 1H, J = 7.9, 7.4, 1.2 Hz), 7.56 (dd, 1H, J = 8.0,
7.4 Hz), 7.41 (dd, 1H, J = 8.6, 1.9 Hz), 7.34 (d, 1H, J = 8.6 Hz), 4.37
(t, 2H, J = 8.0 Hz), 3.05 (septet, 1H, J = 7.4 Hz), 1.77 (quintet, 2H,
J = 8.0 Hz), 1.51 (sextet, 2H, J = 7.4 Hz), 1.34 (d, 6H, J = 7.4 Hz),
1.00 (t, 3H, J = 7.4 Hz). HRMS (FAB) calcd for C20H23NO 294.1858;
found: 294.1851 (M+H)+.
4.16. 5-Butyl-9-methylphenanthridin-6(5H)-one (22)
The title compound was prepared by a method similar to that
described for the synthesis of 20. 4-Methylbenzoyl chloride was
used instead of 2-methylbenzoyl chloride. White solid. Mp 70.0–
73.0 °C. 1H NMR (500 MHz, CDCl3) d 8.44 (d, 1H, J = 7.5 Hz), 8.30
(d, 1H, J = 7.5 Hz), 8.07 (s, 1H7.5), 7.53 (td, 1H, J = 7.5 Hz), 7.40 (d,
2H, J = 7.5 Hz), 7.30 (dd, 1H, J = 7.5 Hz), 4.39 (t, 2H, J = 8.0 Hz),
2.57 (s, 3H), 1.81.75 (m, 2H), 1.51.49 (m, 2H), 1.02 (t, 3H,
J = 7.3 Hz). HRMS (FAB) calcd for C18H19NO 266.1545; found:
266.1527 (M+H)+.
4.22. 5-Butyl-2-tert-butylphenanthridin-6(5H)-one (31)
The title compound was prepared by a method similar to that
described for the synthesis of 17 using 4-tert-burylaniline as a
starting material. Pale brown oil. 1H NMR (500 MHz, CDCl3) d
8.56 (dd, 1H, J = 8.0, 1.5 Hz), 8.32 (d, 1H, J = 8.0 Hz), 8.31 (d, 1H,
J = 1.5 Hz), 7.78–7.74 (m, 1H), 7.60 (dd, 1H, J = 8.0, 1.5 Hz), 7.57
(d, 1H, J = 8.0 Hz), 7.37 (d, 1H, J = 8.0 Hz), 4.39 (t, 2H, J = 7.6 Hz),
1.83–1.76 (m, 2H), 1.56–1.50 (m, 2H), 1.44 (s, 9H), 1.02 (t, 3H,
J = 7.3 Hz). HRMS (FAB) calcd for C21H25NO 308.2014; found:
308.2008 (M+H)+.
4.17. 5-Butyl-10-methylphenanthridin-6(5H)-one (23)
The title compound was prepared by a method similar to that
described for the synthesis of 20. Colorless oil. 1H NMR
(500 MHz, CDCl3) d 8.53 (d, 1H, J = 8.0 Hz), 8.70 (dd, 1H, J = 8.0,
1.2 Hz), 7.61 (d, 1H, J = 8.0 Hz), 7.56–7.52 (m, 1H), 7.49 (d, 1H,
J = 8.0 Hz), 7.46 (dd, 1H, J = 8.0, 1.2 Hz), 7.32–7.27 (m, 1H), 4.40
(t, 2H, J = 7.3 Hz), 2.97 (s, 3H), 1.85–1.78 (m, 2H), 1.50–1.50 (m,
2H), 1.02 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for C18H19NO
266.1545; found: 266.1553 (M+H)+.
4.23. 5-Butyl-2-hydroxyphenanthridin-6(5H)-one (25)
The title compound was prepared by a method similar to that de-
scribed for the synthesis of 8, using 4-tert-butyldimethylsilyloxyan-
iline (prepared from p-nitrophenol) as a starting material, with
slight modifications. 4-tert-Butyldimethylsilyloxyaniline was acyl-
ated with butyryl chloride in the presenceof triethylamine in dichlo-
romethane, and then hydrogenated with lithium aluminum hydride
in tetrahydrofuran. Obtained N-butylaniline was coupled with 2-io-
dobenzoyl chloride, and then cyclized in the presence of palladium
(II) acetate, tricyclohexylphosphine tetrafluoroborate and potas-
sium carbonate in N,N-dimethylacetamide. White solid. Mp 187.0–
192.0 °C. 1H NMR (500 MHz, CDCl3) d 8.55 (d, 1H, J = 7.9 Hz), 8.10
(d, 1H, J = 7.9 Hz), 7.78 (d, 1H, J = 2.4 Hz), 7.67 (t, 1H, J = 7.3 Hz),
7.55 (t, 1H, J = 7.3 Hz), 7.28 (d, 1H, J = 9.2 Hz), 7.14 (dd, 1H, J = 9.2,
2.4 Hz), 6.68 (br s, 1H), 4.37 (t, 2H, J = 7.3 Hz), 1.77 (quintet, 2H,
J = 7.3 Hz), 1.49 (sextet, 2H, J = 7.3 Hz), 0.97 (t, 3H, J = 7.3 Hz). HRMS
(FAB) calcd for C17H17NO2 268.1338; found: 268.1344 (M+H)+.
4.18. 5-Butyl-2-fluorophenanthridin-6(5H)-one (24)
The title compound was prepared by a method similar to that de-
scribed for the synthesis of 17, using 4-fluoroaniline as a starting
material. White solid. FAB-MS m/z 270 (M+H)+. Mp 114.0–
117.5 °C. 1H NMR (500 MHz, CDCl3) d 8.56 (dd, 1H, J = 8.0, 1.5 Hz),
8.17 (d, 1H, J = 8.0 Hz), 7.90 (dd, 1H, J = 9.7, 3.0 Hz), 7.77 (t, 1H,
J = 8.0 Hz), 7.63 (t, 1H, J = 8.0 Hz), 7.37 (dd, 1H, J = 9.0, 4.5 Hz),
7.29–7.24 (m, 1H), 4.38 (t, 2H, J = 8.0 Hz), 1.82–1.75 (m, 2H), 1.57–
1.48 (m, 2H), 1.02 (t, 3H, J = 7.3 Hz). Anal. Calcd for C17H16NFO: C,
75.82; H, 5.99; N, 5.20. Found: C, 76.22; H, 6.24; N, 5.24.
4.19. 5-Butyl-2-trifluoromethylphenanthridin-6(5H)-one (26)
The title compound was prepared by a method similar to that de-
scribed for the synthesis of 17, using 4-trifluoromethylaniline as a
starting material. White solid. FAB-MS m/z 320 (M+H)+. Mp 111.0–
112.0 °C. 1H NMR (500 MHz, CDCl3) d 8.56 (d, 1H, J = 8.0 Hz), 8.53
(s, 1H), 8.30 (d, 1H, J = 8.0 Hz), 7.82 (t, 1H, J = 8.0 Hz), 7.77 (dd, 1H,
J = 8.0, 1.8 Hz), 7.65 (t, 1H, J = 8.0 Hz), 7.00 (d, 1H, J = 8.0 Hz), 4.41
(t, 2H, J = 7.9 Hz), 1.83–1.75 (m, 2H), 1.58–1.49 (m, 2H), 1.03 (t, 3H,
J = 7.3 Hz). Anal. Calcd for C18H16NF3O: C, 67.70; H, 5.05; N, 4.39.
Found: C, 67.91; H, 5.33; N, 4.38.
4.24. 5-Butyl-2-hydroxymethylphenanthridin-6(5H)-one (29)
White solid. FAB-MS m/z 282 (M+H)+. Mp 150.0–153.0 °C. 1H
NMR (500 MHz, CDCl3) d 18.54 (d, 1H, J = 8.0 Hz), 8.28–8.26 (m,
2H), 7.75 (t, 1H, J = 8.0 Hz), 7.59 (t, 1H, J = 8.0, 1.8 Hz), 7.52 (dd, 1H,
J = 8.0, 1.8 Hz), 7.36 (d, 1H, J = 8.0 Hz), 4.83 (d, 2H, J = 6.0 Hz), 4.37
(t, 2H, J = 7.5 Hz), 1.99–1.94 (m, 1H), 1.81–1.74 (m, 2H), 1.55–1.48
(m, 2H), 1.01 (t, 3H, J = 7.5 Hz). Anal. Calcd for C18H19NO2: C,
76.84; H, 6.81; N, 4.98. Found: C, 76.94; H, 6.71; N, 5.01.
4.20. 5-Butyl-2-ethylphenanthridin-6(5H)-one (27)
4.25. 5-Butyl-2-(10-hydroxyethyl)phenanthridin-6(5H)-one (30)
The title compound was prepared by a method similar to de-
scribed that for the synthesis of 17, using 4-ethylaniline as a start-
ing material. Pale brown oil. 1H NMR (500 MHz, CDCl3) d 8.55 (dd,
1H, J = 8.0, 1.5 Hz), 8.30 (d, 1H, J = 8.0 Hz), 8.12 (d, 1H, J = 1.5 Hz),
7.75 (td, 1H, J = 8.0, 1.5 Hz), 7.58 (t, 1H, J = 8.0 Hz), 7.39 (dd, 1H,
The title compound was prepared by a method similar to that
described for the synthesis of 25, using 1-(4-aminophenyl)ethanol
as a starting material. Pale yellow solid. Mp 95.0–99.0 °C. 1H NMR
(500 MHz, CDCl3) d 8.51 (dd, 1H, J = 7.9, 1.2 Hz), 8.23 (d, 1H,
J = 8.5 Hz), 8.22 (d, 1H, J = 1.8 Hz), 7.72 (td, 1H, J = 7.3, 1.2 Hz),

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M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
7.56 (dd, 1H, J = 7.9, 7.3 Hz), 7.49 (dd, 1H, J = 8.5, 1.8 Hz), 7.29 (d,
1H, J = 8.5 Hz), 5.03 (q, 1H, J = 6.7 Hz), 4.33 (t, 2H, J = 7.9 Hz), 1.74
(quintet, 2H, J = 7.9 Hz), 1.57 (d, 3H, J = 6.7 Hz), 1.49 (sextet, 2H,
J = 7.3 Hz), 0.99 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for
C19H21NO2 296.1651; found: 296.1661 (M+H)+.
1.94–1.89 (m, 1H), 1.78 (s, 4H), 1.76 (t, 4H, J = 13.00 Hz), 1.41 (s,
6H), 1.38 (s, 6H), 1.29–1.15 (m, 6H). HRMS (FAB) calcd for
C28H35NO 402.2797; found: 402.2791 (M+H)+.
4.31. 5-Butyl-8,9,10,11-tetrahydro-8,8,11,11-tetramethylbenzo-
[2,3-j]phenanthridin-6(5H)-one (38)
4.26. 5-Butyl-2-acetylphenanthridin-6(5H)-one (32)
The title compound was prepared by a method similar to that
described for the synthesis of 20. 5,6,7,8-Tetrahydro-5,5,8,8-tetra-
methyl-2-naphthoyl chloride (prepared from 5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene-2-carboxylic acid)4 was used
instead of 2-methylbenzoyl chloride. White solid. FAB-MS m/z
362 (M+H)+. Mp 103.0–109.0 °C. 1H NMR (500 MHz, CDCl3) d
8.51 (s, 1H), 8.29 (d, 1H, J = 7.9 Hz), 8.21 (s, 1H), 7.50 (t, 1H,
J = 7.9 Hz), 7.38 (d, 1H, J = 7.9 Hz), 7.29 (t, 1H, J = 7.3 Hz), 4.38 (t,
2H, J = 7.6 Hz), 1.81–1.74 (m, 6H), 1.56–1.50 (m, 2H), 1.42 (s, 6H),
1.40 (s, 6H), 1.02 (t, 3H, J = 7.3 Hz). Anal. Calcd for C25H31NOꢀ1/7
H2O: C, 82.47; H, 8.66; N, 3.85. Found: C, 82.64; H, 8.76; N, 3.71.
The title compound was prepared by the same method as de-
scribed for the synthesis of 30. The title compound was generated
by partial oxidization of the alcoholic hydroxy group at the final
cyclization step. White solid. Mp 114.0–118.0 °C. 1H NMR
(500 MHz, CDCl3) d 8.92 (d, 1H, J = 1.8 Hz), 8.53 (d, 1H, J = 7.3 Hz),
8.37 (d, 1H, J = 7.9 Hz), 8.10 (dd, 1H, J = 9.2, 1.8 Hz), 7.79 (t, 1H,
J = 7.3 Hz), 7.61 (dd, 1H, J = 7.9, 7.3 Hz), 7.44 (d, 1H, J = 9.2 Hz),
4.39 (t, 2H, J = 7.9 Hz), 2.69 (s, 3H), 1.77 (quintet, 2H, J = 7.9 Hz),
1.52 (sextet, 2H, J = 7.3 Hz), 1.01 (t, 3H, J = 7.3 Hz). HRMS (FAB)
calcd for C19H19NO2 294.1494; found: 294.1501 (M+H)+.
4.27. 5-Butyl-2-phenethylphenanthridin-6(5H)-one (33)
4.32. 8,9,10,11-Tetrahydro-8,8,11,11-tetramethylbenzo[2,3-b]-
phenanthridin-5(6H)-one (34)
Compound 29 was oxidized with manganese oxide in dichloro-
methane to give an aldehyde derivative, which was coupled by
Wittig reaction to give a styryl derivative. This product was hydro-
genated in the presence of palladium–carbon under a hydrogen
atmosphere to give the title compound. Colorless oil. 1H NMR
(500 MHz, CDCl3) d 8.55 (dd, 1H, J = 8.0, 1.2 Hz), 8.22 (d, 1H,
J = 8.0 Hz), 8.05 (s, 1H), 7.74 (ddd, 1H, J = 8.0, 6.5, 1.2 Hz), 7.58 (t,
1H, J = 8.0 Hz), 7.37–7.29 (m, 4H), 7.24–7.21 (m, 3H), 4.38 (t, 2H,
J = 7.9 Hz), 3.09–2.99 (m, 4H), 1.82–1.76 (m, 2H), 1.55–1.49 (m,
2H), 1.02 (t, 3H, J = 7.3 Hz). HRMS (FAB) calcd for C25H26NO
356.2014; found: 356.1995 (M+H)+.
The title compound was prepared by a method similar to that
described for the synthesis of 35, using 4-methoxybenzyl bromide
instead of 1-iodobutane, followed by deprotection of the 4-
methoxybenzyl group with trifluoroacetic acid. White solid. Mp
280 °C (decomp). 1H NMR (500 MHz, CDCl3) d 9.27 (s, 1H), 8.51
(dd, 1H, J = 7.8, 1.3 Hz), 8.28 (t, 1H, J = 7.8 Hz), 8.14 (s, 1H), 7.78
(td, 1H, J = 7.8, 1.3 Hz), 7.57 (t, 1H, J = 7.8 Hz), 7.11 (s, 1H), 1.76
(s, 4H), 1.39 (s, 6H), 1.38 (s, 6H). HRMS (FAB) calcd for C21H23NO
306.1858; found: 306.1817 (M+H)+.
4.33. 6-Butylbenzo[b]phenanthridin-5(6H)-one (39)
4.28. 6-Butyl-8,9,10,11-tetrahydro-8,8,11,11-tetramethylbenzo-
[2,3-b]phenanthridin-5(6H)-one (35)
The title compound was prepared by a method similar to that
described for the synthesis of 8, using 2-butylaminonaphthalene
as a starting material. 2-Butylaminonaphthalene was prepared
from 2-bromonaphthalene by coupling reaction with n-butylamine
in the presence of tris(dibenzylidenacetone)dipalladium, sodium
tert-butoxide and n-butylamine. 2-Bromobenzoyl chloride was
used instead of 2-iodobenzoyl chloride. White solid. Mp 101.0–
104.0 °C. 1H NMR (500 MHz, CDCl3) d 8.90 (s, 1H), 8.57 (dd, 1H,
J = 8.0, 1.8 Hz), 8.46 (d, 1H, J = 8.0 Hz), 7.98 (d, 1H, J = 8.0 Hz),
7.92 (d, 1H, J = 8.0 Hz), 7.82–7.78 (m, 1H), 7.73 (s, 1H), 7.62 (t,
1H, J = 7.3 Hz), 7.55 (t, 1H, J = 7.3 Hz), 7.48 (t, 1H, J = 7.3 Hz), 4.50
(t, 3H, J = 7.9 Hz), 1.92–1.85 (m, 2H), 1.63–1.52 (m, 2H), 1.06 (t,
3H, J = 7.3 Hz). HRMS (FAB) calcd for C21H19NO 302.1545; found:
302.1555 (M+H)+.
The title compound was prepared by a method similar to that de-
scribed for the synthesis of 17, using 2-amino-5,6,7,8-tetrahydro-
5,5,8,8-tetramethylnaphthalene as a starting material. Colorless
oil. 1H NMR (500 MHz, CDCl3) d 8.52 (d, 1H, J = 8.0 Hz), 8.26 (d, 1H,
J = 8.0 Hz), 8.21 (s, 1H), 7.73 (t, 1H, J = 8.0 Hz), 7.54 (t, 1H,
J = 8.0 Hz), 7.31 (s, 1H), 4.39 (t, 2H, J = 7.3 Hz), 1.84–1.75 (m, 6H),
1.59–1.50 (m, 2H), 1.40 (s, 6H), 1.38 (s, 6H), 1.04 (t, 3H, J = 7.3 Hz).
HRMS (FAB) calcd for C25H31NO 362.2484; found: 362.2473 (M+H)+.
4.29. 6-Benzyl-8,9,10,11-tetrahydro-8,8,11,11-tetramethylbenzo-
[2,3-b]phenanthridin-5(6H)-one (36)
The title compound was prepared by a method similar to that
described for the synthesis of 35. N-Alkylation was performed by
using benzyl bromide. White amorphous solid. 1H NMR
(500 MHz, CDCl3) d 8.60 (dd, 2H, J = 8.0, 1.5 Hz), 8.29 (d, 1H,
J = 8.0 Hz), 8.18 (s, 1H), 7.79–7.75 (m, 1H), 7.58 (t, 1H, J = 8.0 Hz),
7.37–7.28 (m, 4H), 7.23 (s, 1H), 7.22 (t, 1H, J = 8.0 Hz), 5.65 (s,
2H), 1.69 (d, 4H, J = 1.2 Hz), 1.36 (s, 6H), 1.14 (s, 6H). HRMS (FAB)
calcd for C28H29NO 396.2327; found: 396.2296 (M+H)+.
4.34. 5-Butylbenzo[j]phenanthridin-6(5H)-one (40)
The title compound was prepared by a method similar to that
described for the synthesis of 20. 2-Naphthoyl chloride was used in-
stead of 2-methylbenzoyl chloride. White solid. Mp 111.0–115.0 °C.
1H NMR (500 MHz, CDCl3) d 9.14 (s, 1H), 8.74 (s, 1H), 8.48 (dd, 1H,
J = 8.0, 1.5 Hz), 8.09 (d, 1H, J = 8.0 Hz), 8.04 (d, 1H, J = 8.0 Hz), 7.63
(t, 1H, J = 8.0 Hz), 7.57 (d, 1H, J = 8.0 Hz), 7.54 (dd, 1H, J = 8.0,
1.5 Hz), 7.42 (d, 1H, J = 8.0 Hz), 7.35 (t, 1H, J = 8.0 Hz), 4.43 (t, 2H,
J = 7.6 Hz), 1.87–1.80 (m, 2H), 1.58–1.52 (m, 2H). HRMS (FAB) calcd
for C21H19NO 302.1545; found: 302.1591 (M+H)+.
4.30. 6-Cyclohexylmethyl-8,9,10,11-tetrahydro-8,8,11,11-tetra-
methylbenzo[2,3-b]phenanthridin-5(6H)-one (37)
The title compound was prepared by a method similar to that
described for the synthesis of 35. N-Alkylation was performed by
using cyclohexylmethyl bromide. White solid. Mp 148.0–
4.35. 5-Butylbenzo[c]phenanthridin-6(5H)-one (41)
153.0 °C. 1H NMR (500 MHz, CDCl3)
d 8.52 (dd, 2H, J = 8.0,
The title compound was prepared by a method similar to that
1.2 Hz), 8.27 (d, 1H, J = 8.0 Hz), 8.21 (s, 1H), 7.73 (td, 1H, J = 8.0,
described for the synthesis of 17, using 1-aminonaphthlene as a
1.2 Hz), 7.54 (td, 1H, J = 8.0, 1.2 Hz), 7.29 (s, 1H), 4.30 (br s, 2H),

M. Nakamura et al. / Bioorg. Med. Chem. 18 (2010) 2402–2411
2411
starting material. White amorphous solid. 1H NMR (500 MHz,
CDCl3) d 8.55 (dd, 1H, J = 8.0, 1.2 Hz), 8.29–8.22 (m, 3H), 7.92–
7.89 (m, 1H), 7.81–7.77 (m, 1H), 7.73 (d, 1H, J = 8.0 Hz), 7.60 (t,
3H, J = 8.0 Hz), 7.56–7.50 (m, 2H), 4.59 (t, 2H, J = 7.3 Hz), 1.93–
1.86 (m, 2H), 1.26–1.18 (m, 2H), 0.83 (t, 3H, J = 7.3 Hz). HRMS
(FAB) calcd for C21H19NO 302.1545; found: 302.1574 (M+H)+.
Gene Expression Cell-to-CT™ Kit. Expression levels of HCV RNA
and GAPDH RNA were measured with the kit, according to the
manufacturer’s instructions. The RNA levels were quantified by
real-time RT-PCR using an ABI 7500 Real-Time PCR System (Ap-
plied Biosystems). The anti-HCV activity and cytotoxicity of test
compounds were expressed as EC50 and CC50 determined from
the decrease of HCV RNA and GAPDH RNA levels, respectively, as
described above.
4.36. 6-Butylbenzo[a]phenanthridin-5(6H)-one (42)
The title compound was prepared by the same method as de-
scribed for the synthesis of 39. The title compound was fraction-
ated by means of HPLC. Colorless oil. 1H NMR (500 MHz, CDCl3) d
8.80 (d, 1H, J = 8.5 Hz), 8.68 (d, 1H, J = 8.5 Hz), 8.64 (dd, 1H,
J = 8.5, 1.5 Hz), 7.95 (t, 2H, J = 8.5 Hz), 7.79 (td, 1H, J = 8.5, 1.5 Hz),
7.65–7.60 (m, 3H), 7.52 (t, 1H, J = 8.5 Hz), 4.50 (t, 2H, J = 7.5 Hz),
1.88–1.80 (m, 2H), 1.58–1.53 (m, 2H), 1.03 (t, 3H, J = 7.3 Hz). HRMS
(FAB) calcd for C21H19NO 302.1545; found: 302.1534 (M+H)+.
Acknowledgments
The work described in this paper was partially supported by
Grants-in Aid for Scientific Research from the Science and Technol-
ogy Incubation Program in Advanced Regions, Japan Science and
Technology Agency (JST), Japan and The Ministry of Education, Cul-
ture, Sports, Science and Technology, Japan, and a Grant from the
Japan Society for the Promotion of Science.
4.37. 5-Butylbenzo[k]phenanthridin-6(5H)-one (43)
References and notes
The title compound was prepared by the same method as de-
scribed for the synthesis of 40. The title compound was fraction-
ated by means of HPLC. White solid. Mp 54.0–60.0 °C. 1H NMR
(500 MHz, CDCl3) d 8.88 (d, 1H, J = 7.3 Hz), 8.66 (d, 1H, J = 8.5 Hz),
8.51 (d, 1H, J = 8.5 Hz), 8.01 (dd, 2H, J = 7.3, 3.0 Hz), 7.95 (d, 1H,
J = 8.5 Hz), 7.70–7.63 (m, 3H), 7.59 (t, 1H, J = 7.3 Hz), 7.53 (d, 2H,
J = 8.5 Hz), 7.36 (t, 2H, J = 8.5 Hz), 4.45 (t, 2H, J = 7.9 Hz), 1.90–
1.83 (m, 3H), 1.57–1.51 (m, 2H), 1.03 (t, 3H, J = 7.5 Hz). HRMS
(FAB) calcd for C21H19NO 302.1545; found: 302.1567 (M+H)+.
1. Hashimoto, Y. Arch. Pharm. Life Sci. 2008, 341, 536.
2. Hosoda, S.; Matsuda, D.; Tomoda, H.; Hashimoto, Y. Mini-Rev. Med. Chem. 2009,
9, 572.
3. Hosoda, S.; Aoyama, H.; Goto, Y.; Salim, M. T. A.; Okamoto, M.; Hashimoto, M.;
Baba, M.; Hashimoto, Y. Bioorg. Med. Chem. Lett. 2009, 19, 3157.
4. Nakamura, M.; Hamasaki, T.; Tokitou, M.; Baba, M.; Hashimoto, Y.; Aoyama, H.
Bioorg. Med. Chem. 2009, 17, 4740.
5. Hosoda, S.; Matsuda, D.; Tomoda, H.; Hashimoto, M.; Aoyama, H.; Hashimoto,
Y. Bioorg. Med. Chem. Lett. 2009, 19, 4228.
6. Koonin, E. V.; Wolf, Y. I.; Karev, G. P. Nature 2002, 420, 218.
7. Grishin, N. V. J. Struct. Biol. 2001, 134, 167.
8. Koch, M. A.; Wittenberg, L.-O.; Basu, S.; Jeyaraj, D. A.; Gourzoulidou, E.;
Reinecke, K.; Odermatt, A.; Waldmann, H. Proc. Natl. Acad. Sci. U.S.A. 2004, 101,
16721.
4.38. 6-Butylbenzo[i]phenanthridin-5(6H)-one (44)
9. Hashimoto, Y. Curr. Med. Chem. 1998, 5, 163.
10. Hashimoto, Y. Bioorg. Med. Chem. 2002, 10, 461.
11. Hashimoto, Y. Mini-Rev. Med. Chem. 2002, 2, 543.
12. Hashimoto, Y.; Tanatani, A.; Nagasawa, K.; Miyachi, H. Drugs Future 2004, 29,
383.
13. Hashimoto, Y. Cell Biol. Rev. 1991, 25, 209.
14. Sako, K.; Aoyama, H.; Sato, S.; Hashimoto, Y.; Baba, M. Bioorg. Med. Chem. 2008,
16, 3780.
15. Aoyama, H.; Sako, K.; Sato, S.; Nakamura, M.; Miyachi, H.; Baba, M.; Hashimoto,
Y. Heterocycles 2009, 77, 779.
16. Liang, T. J.; Rehermann, B.; Seeff, L. B.; Hoofnagle, J. H. Ann. Intern. Med. 2000,
132, 296.
17. Hayashi, P. H.; Di Bisceglie, A. M. Med. Clin. North Am. 2005, 89, 371.
18. Memon, M. I.; Memon, M. A. J. Viral Hepat. 2002, 9, 84.
19. Echevarria-Mayo, J. M. Enferm. Infecc. Microbiol. Clin. 2006, 24, 45.
20. Bosch, F. X.; Ribes, J.; Cleries, R.; Diaz, M. Clin. Liver Dis. 2005, 9, 191.
21. McHutchison, J. G.; Gordon, S. C.; Schiff, E. R.; Shiffman, M. L.; Lee, W. M.;
Rustgi, V. K.; Goodman, Z. D.; Ling, M. H.; Cort, S.; Albrecht, J. K. N. Eng. J. Med.
1998, 339, 1485.
22. Tan, S. L.; Pause, A.; Shi, Y.; Sonenberg, N. Nat. Rev. Drug Disc. 2002, 1, 867.
23. Buckwold, V. E.; Beer, B. E.; Donis, R. O. Antiviral Res. 2003, 60, 1.
24. Buckwold, V. E.; Wei, J.; Wenzel-Mathers, M.; Russell, J. Antimicrob. Agents
Chemother. 2003, 47, 2293.
25. Yanagida, K.; Baba, C.; Baba, M. Antiviral Res. 2004, 64, 195.
26. Aoyama, A.; Aoyama, H.; Dodo, K.; Makishima, M.; Hashimoto, Y.; Miyachi, H.
Heterocycles 2008, 76, 137.
27. Miyachi, H.; Aoyama, A.; Hashimoto, Y. MedChemNews 2009, 19(2), 30.
28. Ishii, N.; Watashi, K.; Hishiki, T.; Goto, K.; Inoue, D.; Hijikata, M.; Wakita, T.;
Kato, N.; Shimotohno, K. J. Virol. 2006, 80, 4510.
The title compound was prepared by a method similar to that
described for the synthesis of 40. 1-Naphthoyl chloride was used
instead of 2-naphthoyl chloride. White amorpous solid. 1H NMR
(500 MHz, CDCl3) d 10.30 (d, 1H, J = 9.0 Hz), 8.44 (dd, 1H, J = 8.0,
1.2 Hz), 8.39 (d, 1H, J = 9.0 Hz), 8n.17 (d, 1H, J = 9.0 Hz), 7.94 (dd,
1H, J = 8.0, 1.2 Hz), 7.77–7.73 (m, 1H), 7.65–7.60 (m, 2H), 7.49 (d,
1H, J = 8.0 Hz), 7.36 (t, 1H, J = 8.0 Hz), 4.50 (t, 2H, J = 7.6 Hz),
1.90–1.83 (m, 2H), 1.63–1.53 (m, 2H), 1.06 (t, 3H, J = 7.3 Hz). HRMS
(FAB) calcd for C21H19NO 302.1545; found: 302.1540 (M+H)+.
4.39. Bioassay
NNC #2 cells carrying full-genomic HCV RNA replicons were
maintained in high-glucose Dulbecco’s modified Eagle’s medium
(DMEM) in the presence of 10% (v/v) fetal bovine serum (FBS)
and 1 mg/mL G418 (Sigma). Cells at 70% confluence were collected
after treatment with trypsin and resuspended in the same medium
(5 ꢁ 104 cells/mL). One hundred microliters of the cell suspension
was transferred to each well of a 96-well plate and cultured at
37 °C for 24 h. Then the medium was removed, and 200 lL of
DMEM supplemented with 10% FBS containing various concentra-
tions of test compound was added to each well. After incubation
for 3 days, the cells were treated with lysis buffer of a TaqMan
29. Li, L.; Liu, J.; Zhu, L.; Cutler, S.; Hasegawa, H.; Shan, B.; Medina, J. C. Bioorg. Med.
Chem. Lett. 2006, 16, 1638.