An efficient synthesis of a new class of DNA intercalating antitumor 7,10-dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-ones

Article abstract of DOI:10.1055/s-1999-3494

An efficient synthesis of KW-2170 (1), which is a 7,10-dihydroxy-6H- pyrazolo[4,5,1-de]acridin-6-one derivative and a new class of DNA intercalating antitumor agents, is described. The selective monobromination of the methyl group before the cyclization to the pyrazoloacridone is easily carried out. In the improved process, the bromination of the corresponding methyl group is achieved prior to the hydroquinone formation, so the protection of the hydroxy groups is not necessary unlike the original method. In comparison with the original synthetic route of KW-2170 (1), the new route decreases the synthetic steps from 2-bromo-6-methoxybenzoic acid (2) from 15 to 13 and increases the overall yield from 2 to 12%.

Full text of DOI:10.1055/s-1999-3494

PAPER
947
An Efficient Synthesis of a New Class of DNA Intercalating Antitumor 7,10-
Dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-ones
Takashi Mimura,* Nobuyuki Kato, Toru Sugaya, Masanori Ikuta,§ Sachiko Kato, Yukihiro Kuge, Shinji Tomioka,§
Masaji Kasai
Sakai Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., 1-1-53 Takasucho, Sakai, 590–8554, Japan
Fax +81(722)277214; E-mail takashi.mimura@kyowa.co.jp
Received 30 October 1998; revised 4 January 1999
over the development of the large scale synthesis. The im-
Abstract: An efficient synthesis of KW-2170 (1), which is a 7,10-
portant oxidation process to the quinone and the following
reduction to the 7,10-dihydroxy compound were devel-
dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-one derivative and a
new class of DNA intercalating antitumor agents, is described. The
selective monobromination of the methyl group before the cycliza-
oped and the improved preparation of the key intermedi-
tion to the pyrazoloacridone is easily carried out. In the improved ate of the KW-2170 (1) was published.5 However, there
process, the bromination of the corresponding methyl group is
are remaining issues to be resolved for establishing the
achieved prior to the hydroquinone formation, so the protection of
synthetic processes, and also the purification methods
the hydroxy groups is not necessary unlike the original method. In
both of the intermediates and the final product should be
comparison with the original synthetic route of KW-2170 (1), the
improved. Herein, we describe the results of the develop-
new route decreases the synthetic steps from 2-bromo-6-methoxy-
benzoic acid (2) from 15 to 13 and increases the overall yield from
2 to 12%.
ment of the synthetic process and the accomplishment of
a new efficient synthesis of KW-2170.
Key words: antitumor agents, pyrazoloacridone, process improve-
ment
The substrate of the previously reported oxidation using
the hypervalent iodine reagent was the 2-methyl com-
pound 5.5 The resulting 7,10-hydroxy compound was ex-
pected to be easily converted to KW-2170 even on a
practical synthetic scale. However, the selectivity of the
monobromination at the 2-methyl group was low, because
the dibromide was generated and also the unreacted start-
ing material remained. Therefore, it was necessary to use
chromatography for the isolation of the monobromide.
6H-Pyrazolo[4,5,1-de]acridin-6-one (pyrazoloacridone)
derivatives were synthesized as a new class of DNA inter-
calating antitumor agents.1 In researching the structure–
activity relationships, the 7,10-dihydroxy derivatives
showed excellent antitumor activities.2 The most promis-
ing compound, KW-2170 (1; Figure), has superior anti-
proliferactive activity in vitro against HeLa S3 and
excellent antitumor activities in vivo against the murine
sarcoma 180 solid tumor, murine P388 leukemia and adri-
amycin-resistant murine P388 leukemia.2,3 Thus, KW-
2170 (1) was evaluated in pre-clinical studies and is now
in phase I clinical trials.
We first attempted the investigation of the selective
monobromination with N-bromosuccinimide (NBS) at the
2-methyl group of various substrates, i.e., the original syn-
thetic intermediate with protected C-7 and C-10 hydroxy
groups 9,2 the 7-methoxypyrazoloacridone without the C-
10 hydroxy group 4a4 and the ester of the indazole 14b
(Table). The ester 14b was prepared from the correspond-
ing sodium salt 14a4 in 87% yield (Scheme 2). In this
study, 2,2’-azobisisobutyronitrile (AIBN) was tried as the
radical initiator instead of the more explosive benzoyl per-
oxide (BPO). Among those compounds, it was found that
the ester 14b was the best substrate for bromination from
the view point of the monobromination selectivity. The
ratio of monobromide 15 in the reaction mixture was
76.0% (determined by HPLC analysis6). The monobromi-
nated compound 15 was isolated from the reaction mix-
ture with diisopropyl ether. This isolated 15 was
suspended in methanol and the undissolved solid was col-
lected to obtain the purified 15 in 65% yield (93% purity,
determined by HPLC analysis6). The monobrominated es-
ter was then cyclized to pyrazoloacridone with trifluo-
romethansulfonic acid at 100 °C in high yield. At this
stage, the methyl ether at C-7 was partially cleaved and a
mixture of the methyl ether 16a and the phenol 16b (47/
53, determined by HPLC analysis6) was obtained. This
mixture was treated with hydrogen bromide in acetic acid
Figure
As these studies were proceeding, the development of a
synthetic route of KW-2170 (1) that would be amenable to
commercial production was required. The original syn-
thetic route of KW-2170 (1) is depicted in Scheme 1.2
This synthetic route was planned based on the synthetic
method of the 7-hydroxy derivatives.2,4 The synthetic
route for introducing the 10-hydroxy moiety took priority
Synthesis 1999, No. 6, 947–952 ISSN 0039-7881 © Thieme Stuttgart · New York

948
T. Mimura et al.
PAPER
Scheme 1
at 70 °C to accomplish the cleavage of the methyl ether.
Preceding to the study of the oxidation, the protection at
C-8 by bromination was carried out with bromine at 25 °C
and gave the C-8 bromo derivative 17. The overall yield
of 17 from the C-2 monobrominated ester 15 was 80% and
the purity of 17 was 96% based on an HPLC analysis6
(Scheme 3).
Scheme 2
The introduction of the hydroxy group at C-10 of the 2-
bromomethyl derivative 17 was accomplished by a two-
step sequence involving the hypervalent iodine oxidation
method followed by reduction. The reaction of 17 with
[(diacetoxy)iodo]benzene in trifluoroacetic acid (TFA)
and acetic acid (6:4) was heterogeneous, and the oxidation
stopped. However, the reaction was promoted by the ad-
Synthesis 1999, No. 6, 947–952 ISSN 0039-7881 © Thieme Stuttgart · New York

PAPER
A New Class of DNA Intercalating Antitumor 7,10-Dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-ones
949
Scheme 3
dition of tetra-n-butylammonium hydrogen sulfate (n- purified by silica gel chromatography to over 99% based
Bu4NHSO4) as the phase transfer catalyst and the quinone on HPLC analysis.9 This procedure helped to avoid the
18 was obtained in 89% yield. The quinone was incom- HPLC purification of KW-2170 in the final process. The
pletely converted to the hydroquinone with aqueous sodi- amino side chain at C-5 was introduced with propane-1,3-
um hydrosulfite in the same way as already reported.5,7 diamine at 40 °C in chloroform and the compound 21 was
After the investigation of various reducing methods, the purified by recrystallization from a mixture of chloro-
reduction was accomplished in dichloromethane using form, methanol and acetonitrile. The final intermediate 21
tetra-n-butylammonium
triacetoxyborohydride
(n- was obtained from 19 for two steps with a 58% overall
Bu4NBH(OAc)3)8 which was prepared in situ from tetra- yield to keep the purity at 99% or more in the HPLC
n-butylammonium borohydride (n-Bu4NBH4) and acetic analysis10 (Scheme 4).
acid. The hydroquinone 19 was obtained from the phenol
The final step, the removal of N-benzyl and C-8 bromo
17 in 80% overall yield (Scheme 3). In the improved pro-
cess, the bromination of the corresponding methyl group
was achieved prior to the hydroquinone formation. Thus,
there was no necessity for protection of the hydroxy
groups unlike the original method in which those hydroxy
groups should be protected by an acetyl or methyl group
before reacting with NBS. So far, there were neither high-
ly toxic intermediates nor reagents and the steps were car-
groups, was accomplished by hydrogenolysis with 10%
palladium on carbon in a mixture of dilute hydrochloric
acid and ethanol. In the original route, the HPLC purifica-
tion of the final product was indispensable for removing
impurities which were formed at about 3% under the se-
vere reaction conditions needed for the O-demethylation
at C-7 (Scheme 1). In the improved route, however, the
conditions were mild enough to keep the purity high
ried out under common synthetic conditions. The
improved synthetic route has been used for the practical
preparation of a key intermediate 19 of KW-2170.
(>99%),11 so the final KW-2170 was obtained merely by
recrystallization from dilute hydrochloric acid and etha-
nol.
Although the two amino side chains at C-2 and C-5 could
In summary, we have established an efficient synthetic
be introduced as in the original method,1,2 the formation of
route to KW-2170 (1), a new DNA intercalating antitumor
the disubstituted amino compound should be avoided by
the introduction of the C-2 amino side chain. The highly
diluted conditions and the large excess amount of amine
were adopted in the original method and it suffered from
low productivity. Therefore, we attempted to use a sec-
ondary amine, 2-[(phenylmethyl)amino]ethanol (22), to
generate the tertiary amine 20. The benzyl group in the fi-
nal product could be removed at the last step. The intro-
agent. In comparison with the original synthetic route,2
the improved route showed that the synthetic steps from
2-bromo-6-methoxybenzoic acid (2) decreased from 15 to
13 and the overall yield was increased from 2 to 12%. In
addition, the bulk substance of KW-2170 with a high pu-
rity (>99%) has been synthesized by an innovative meth-
odology and provided for clinical trial tests.
duction of 2-[(phenylmethyl)amino]ethanol (22) was
All the reagents and solvents were commercially available and were
used without purification. Mps were determined using a Mettler
FP61 melting point instrument and are uncorrected. NMR spectra
(internal standard TMS) were recorded on a Bruker AC300 (300
MHz) and JEOL LA300 (300 MHz) spectrometer. IR spectra were
recorded using a Shimadzu FTIR-4300 spectrophotometer. Mass
spectra were recorded on a Hitachi M-80B mass spectrometer. Ele-
carried out at 25 °C in N,N-dimethylformamide and the
concentration of the reaction increased from 3 g/L to
200 g/L. The amine 20 was crystallized from the reaction
mixture with methanol. At this stage, the purity of 20 was
97% (determined by HPLC analysis9). To obtain the final
product of KW-2170 in high purity, the compound 20 was
Synthesis 1999, No. 6, 947– 952 ISSN 0039-7881 © Thieme Stuttgart · New York

950
T. Mimura et al.
PAPER
Anal: calcd for C17H15BrN2O3: C, 54.42; H, 4.03; N, 7.47. Found:
C, 54.28; H, 4.01; N, 7.19.
2
OH
10
N
Br
N
O
Methyl 2-(6-Bromo-3-bromomethyl-1H-indazol-1-yl)-6-meth-
oxybenzoate (15)
Br
To a solution of the methyl ester 14b (2,934 g, 7.82 mol) in
ClCH2CH2Cl (58.7 L) was added NBS (1,670 g, 9.38 mol) and
AIBN (257 g, 1.56 mol) at 25 °C. The mixture was refluxed for 1.5
h and the solution was concentrated under reduced pressure. To the
residue was added diisopropyl ether (14.7 L) and the mixture was
cooled to 5 °C, then the precipitated solid was filtered. To the fil-
tered solid was added MeOH (30 L) and the suspension was cooled
to 5 °C. The precipitate was filtered and washed with MeOH (4 L)
to give the bromide 15 (2,319 g, 65%) as colorless solid; mp 133 °C.
1H NMR (CDCl3/TMS): d = 3.64 (s, 3H), 3.93 (s, 3H), 4.81 (s, 2H),
7.05 (d, 1H, J = 8.4 Hz), 7.19 (d, 1H, J = 8.0 Hz), 7.40 (dd, 1H, J =
8.6, 1.5 Hz), 7.55 (t, 1H, J = 8.4 Hz), 7.73 (d, 1H, J = 8.6 Hz), 7.74
(d, 1H, J = 1.5 Hz).
OH
Br
19
OH
22
OH
N
OH
N
N
H
N
O
DMF, 25oC, 4h
70%
Br
OH
OH
Br
20
OH
N
N
H2N(CH2)3NH2
N
O
IR (KBr): ν = 1724, 1609, 1587, 1472, 1285, 1121, 1059, 802 cm–1.
MS (EI): m/z (%) = 452/454/456 (M+, 5/9/5), 373/375 (100/91),
CHCl3
40oC, 1h
Br
83%
343/345 (39/38).
OH
HN
21
NH2
Anal: calcd for C17H14Br2N2O3: C, 44.96; H, 3.11; N, 6.17. Found:
C, 45.07; H, 3.16; N, 5.87.
5-Bromo-2-bromomethyl-7-hydroxy-6H-pyrazolo-
[4,5,1-de]acridin-6-one (16b)
OH
2
H2, Pd/C
dil. HCl, EtOH
60oC, 12h
OH
10
N
N
H
The ester 15 (2,273 g, 5.01 mol) was heated in CF3SO3H (45.5 L) at
100 °C for 4 h. The mixture was poured into MeOH (100 L) and
cooled to 5 °C. Then, the precipitated solid was filtered. To the fil-
tered solid was added MeOH (23 L) and the suspension was stirred
at 5 °C. The precipitate was filtered to give 47/53 mixture (deter-
mined by HPLC analysis6) of 7-methoxy ether 16a and 7-hydroxy
derivative 16b as yellow solid; 1,908 g. To the mixture of 16a and
16b (1,808 g) was added 30% HBr in HOAc (27.1 L) and the mix-
ture was heated at 70 °C for 4 h. After cooling to 15 °C, to the mix-
ture was added MeOH (54.3 L) dropwise and cooled to 5 °C. The
precipitate was filtered and washed with MeOH (40.5 L) to give the
phenol 16b (1,710 g, 88% from the ester 15) as yellow solid; mp
220 °C (dec).
N
O
2 HCl
87%
7
OH
5
HN
NH2
KW-2170 (1)
Scheme 4
mental analyses were performed using a Yanaco MT-3 CHN appa-
ratus. Sodium 2-(6-bromo-3-methyl-1H-indazol-1-yl)-6-methoxy-
benzoate (14a) was prepared from 2-bromo-6-methoxybenzoic acid
(2)4 and 3-methyl-6-nitroindazole (3),12 according to the reported
procedure.4
1H NMR (CDCl3/TMS): d = 4.90 (s, 2H), 6.90 (d, 1H, J = 8.1 Hz),
7.60 (d, 1H, J = 8.2 Hz), 7.67 (t, 1H, J = 8.1 Hz), 7.79 (d, 1H, J =
8.2 Hz), 8.04 (d, 1H, J = 8.2 Hz), 13.20 (s, 1H).
Methyl 2-(6-Bromo-3-methyl-1H-indazol-1-yl)-6-methoxyben-
zoate (14b)
IR (KBr): ν = 1638, 1603, 1589, 1514, 1473, 1339, 1294, 1259,
1225, 1134, 1070, 812 cm–1.
A suspension of sodium 2-(6-bromo-3-methyl-1H-indazol-1-yl)-6-
methoxybenzoate (14a,4 3,800 g, 9.91 mol) in MeOH (5.7 L) and
DMF (5.7 L) was heated at 75 °C for 1 h and cooled to 30 °C. To
this suspension was added K2CO3 (684 g, 9.91 mol) and MeI (3,085
mL, 49.6 mol) and the mixture was stirred at 50 °C for 2 h. After
removing the solvent under reduced pressure, the mixture of MeOH
(3.8 L) and H2O (34.2 L) was added dropwise to the residue. The
precipitate was filtered, washed with H2O (8 L), and the filtered sol-
id was dissolved in CHCl3 (23 L) and dried (MgSO4). The solution
was concentrated under reduced pressure and the residue was crys-
tallized from CHCl3 (5.7 L) and MeOH (34.2 L) to give the methyl
ester 14b (3,231 g, 87%) as colorless solid; mp 164 °C.
MS (EI): m/z (%) = 406/408/410 (M+, 11/20/10), 327/329 (100/
100).
Anal: calcd for C15H8Br2N2O2: C, 44.15; H, 1.98; N, 6.87. Found:
C, 44.21; H, 2.13; N, 6.68.
5,8-Dibromo-2-bromomethyl-7-hydroxy-6H-pyrazolo-
[4,5,1-de]acridin-6-one (17)
To a solution of the phenol 16b (1,613 g, 3.95 mol) in ClCH2CH2Cl
(341 L) was added Br2 (647 g, 4.05 mol) in ClCH2CH2Cl (4 L). The
mixture was stirred at 25 °C for 2 h and the mixture was concentrat-
ed to 48 L under reduced pressure. After cooling, the precipitate was
filtered and washed with i-PrOH (25 L) to give the bromide 17
(1,744 g, 91%) as brown solid; mp 262 °C (dec).
1H NMR (CDCl3/TMS): d = 4.83 (s, 2H), 7.54 (d, 1H, J = 8.7 Hz),
7.79 (d, 1H, J = 8.3 Hz), 7.88 (d, 1H, J = 8.7 Hz), 8.03 (d, 1H, J =
8.3 Hz), 13.97 (s, 1H).
1H NMR (CDCl3/TMS): d = 2.58 (s, 3H), 3.66 (s, 3H), 3.91 (s, 3H),
6.99 (d, 1H, J = 8.4 Hz), 7.18 (d, 1H, J = 8.0 Hz), 7.29 (dd, 1H, J =
8.5, 1.5 Hz), 7.51 (t, 1H, J = 8.3 Hz), 7.54 (d, 1H, J = 8.4 Hz), 7.71
(d, 1H, J = 1.5Hz).
IR (KBr): ν = 1728, 1598, 1516, 1460, 1333, 1256, 1132,
1111, 1096, 1049, 980 cm–1.
IR (KBr): ν = 1636, 1587, 1508, 1448, 1342, 1281, 1256, 1234,
1121, 1049, 814 cm–1.
MS (EI): m/z (%) = 374/376 (M+, 100/99), 343/345 (63/55).
Synthesis 1999, No. 6, 947–952 ISSN 0039-7881 © Thieme Stuttgart · New York

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A New Class of DNA Intercalating Antitumor 7,10-Dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-ones
951
MS (EI): m/z (%) = 484/486/488/490 (M+, 11/31/33/12), 405/407/
409 (52/100/55).
IR (KBr): ν = 1640, 1608, 1590, 1516, 1450, 1290, 1242, 1130,
1090, 810cm–1.
Anal: calcd for C15H7Br3N2O2: C, 37.00; H, 1.45; N, 5.75. Found:
C, 37.13; H, 1.58; N, 5.56.
MS (EI): m/z (%) = 571/573/575 (M+, 17/19/8), 540/542/544 (9/17/
9), 421/423/425 (57/100/53).
Anal: calcd for C24H19Br2N3O4: C, 50.29; H, 3.34; N, 7.33. Found:
C, 50.17; H, 3.34; N, 7.14.
5,8-Dibromo-2-bromomethyl-6H-pyrazolo[4,5,1-de]acridine-
6,7,10-trione (18)
To a suspension of the bromide 17 (300.0 g, 0.616 mol) in the mix-
ture of HOAc (1.2 L) and TFA (1.8 L) was added tetra-n-butylam-
monium hydrogen sulfate (120.0 g, 40%wt) and [(diacetoxy)iodo]-
benzene (597.0 g, 1.85 mol). The mixture was stirred at 40 °C for 2
h, then cooled to 25 °C. To the mixture was added MeOH (6.0 L)
and cooled to 5 °C, then the precipitate was filtered and washed with
MeOH (2.0 L) to give the quinone 18 (274.3 g, 89%) as brown solid;
mp 205 °C (dec).
1H NMR (CDCl3/TMS): d = 4.83 (s, 2H), 7.44 (s, 1H), 7.89 (d, 1H,
J = 8.3 Hz), 8.03 (d, 1H, J = 8.3 Hz).
IR (KBr): ν = 1682, 1622, 1520, 1504, 1201, 853, 572 cm–1.
5-[(3-Aminopropyl)amino]-2-{[(1-benzyl-2-hydroxyethyl)ami-
no]methyl}-8-bromo-7,10-dihydroxy-6H-pyrazolo[4,5,1-
de]acridin-6-one (21)
To a suspension of the amine 20 (97.0 g, 0.169 mol) in CHCl3 (1.5
L) was added propane-1,3-diamine (705 mL, 8.45 mol) at 25 °C and
the mixture was heated at 40 °C for 1 h. After cooling to 25 °C, i-
PrOH (2.9 L) was added and the suspension was cooled to 5 °C. The
precipitate was filtered to give the crude amine 21 (104.2 g). The
crude compound 21 (104.0 g) was purified by recrystallizing from
the mixture of CHCl3 (4.7 L), MeOH (1.6 L) and MeCN (2.1 L) to
give the amine 21 (79.2 g, 83% from 20) as yellow solid; mp 182 °C
(dec).
1H NMR (DMSO-d6/TMS): d = 1.73 (m, 2H, J = 6.7 Hz), 2.57 (m,
2H, J = 6.3 Hz), 2.69 (t, 2H, J = 6.6 Hz), 3.53–3.59 (m, 4H), 3.67
(s, 2H), 4.10 (s, 2H), 6.94 (d, 1H, J = 9.2 Hz), 7.19–7.36 (m, 5H),
7.49 (s, 1H), 8.10 (d, 1H, J = 9.2 Hz).
MS (EI): m/z (%) = 500/502/504/506 (M++2, 15/37/37/12), 421/
423/425 (55/100/52).
Anal: calcd for C15H5Br3N2O3: C, 35.97; H, 1.01; N, 5.59. Found:
C, 36.03; H, 1.23; N, 5.35.
IR (KBr): ν = 1658, 1600, 1580, 1520, 1402, 1290, 1250, 1062,
1030, 820 cm–1.
MS (EI): m/z (%) = 565/567 (M+, 27/18), 415/417 (52/56), 386/388
5,8-Dibromo-2-bromomethyl-7,10-dihydroxy-6H-pyrazolo-
[4,5,1-de]acridin-6-one (19)
To a solution of tetra-n-butylammonium borohydride (168.9 g,
0.656 mol) in CH2Cl2 (1.4 L) was added HOAc (337 mL, 5.33 mol)
under N2 and the mixture was stirred at 25 °C for 30 min. To the sus-
pension of the quinone 18 (274.0 g, 0.547 mol) in CH2Cl2 (1.4 L)
was added the solution of the reducing agent under N2 and the mix-
ture was stirred at 25 °C for 2 h. To the mixture was added MeOH
(3.3 L) dropwise and the suspension was cooled to 5 °C. The pre-
cipitate was filtered and washed with MeOH (1.4 L) to give the hy-
droquinone 19 (246.6 g, 90%) as dark red solid; mp 249 °C (dec).
(37/33), 358/360 (57/54), 337 (100).
Anal: calcd for C27H28BrN5O4: C, 57.25; H, 4.98; N, 12.36. Found:
C, 56.98; H, 4.91; N, 12.15.
5-[(3-Aminopropyl)amino]-7,10-dihydroxy-2-{[(2-hydroxyeth-
yl)amino]methyl}-6H-pyrazolo[4,5,1-de]acridin-6-one
Dihydrochloride (KW-2170, 1)
A mixture of the amine 21 (60.0 g, 0.106 mol) and 10% palladium
on carbon (12.0 g, 20%wt) in a solution of 0.6 mol/L HCl (1.5 L)
and EtOH (1.5 L) was stirred under hydrogen flow at 60 °C for 12
h. After the atmosphere of the reaction vessel was displaced with
N2, the mixture was filtered and the residue was washed with a mix-
ture of H2O and EtOH (1:1, 600 mL). To the filtrate was added conc.
HCl (240 mL) and KW-2170 (1) (500 mg) as seed, and the mixture
was stirred at 25 °C for 20 h and cooled to 5 °C. The precipitate was
filtered to give the crude compound 1 (44.3 g). The crude compound
1 (42.0 g) was purified by recrystallization from the mixture of 1
mol/L HCl (420 mL) and EtOH (840 mL) to give the purified KW-
2170 (1) (41.0 g, 87% from 21) as a yellow crystalline solid; mp
>300 °C.
1H NMR (CDCl3/TMS): d = 4.85 (s, 2H), 7.63 (s, 1H), 7.83 (d, 1H,
J = 8.4 Hz), 8.08 (d, 1H, J = 8.4 Hz), 9.40 (s, 1H), 13.35 (s, 1H).
IR (KBr): ν = 1638, 1605, 1590, 1512, 1290, 1245, 1210, 1130,
1095, 910 cm–1.
MS (EI): m/z (%) = 500/502/504/506 (M+, 14/38/38/13), 421/423/
425 (73/100/48).
Anal: calcd for C15H7Br3N2O3: C, 35.82; H, 1.40; N, 5.57. Found:
C, 36.00; H, 1.56; N, 5.38.
2-{[(1-Benzyl-2-hydroxyethyl)amino]methyl}-5,8-dibromo-
7,10-dihydroxy-6H-pyrazolo[4,5,1-de]acridin-6-one (20)
1H NMR (DMSO-d6/TMS): d = 2.01 (m, 2H), 2.94 (t, 2H, J = 6.8
Hz), 3.15 (t, 2H, J = 5.4 Hz), 3.70 (dd, 2H, J = 7.0, 13.4 Hz), 3.76
(dd, 2H, J = 7.1, 10.5 Hz), 4.75 (s, 2H), 5.29 (t, 1H, J = 4.8 Hz), 6.80
(d, 1H, J = 8.8 Hz), 7.17 (d, 1H, J = 9.2 Hz), 7.37 (d, 1H, J = 8.8
Hz), 8.05 (br s, 3H), 8.35 (d, 1H, J = 9.2 Hz), 9.33 (t, 1H, J = 6.0
Hz), 9.61 (br s, 2H), 9.70 (s, 1H), 13.02 (s, 1H).
13C NMR (DMSO-d6/TMS): d = 26.7, 36.4, 39.5, 41.7, 49.0, 56.4,
99.2, 109.3, 110.1, 110.3, 112.0, 122.4, 123.6, 132.1, 137.4, 137.8,
141.8, 152.9, 154.8, 182.1.
To a suspension of the hydroquinone 19 (246.0 g, 0.489 mol) in
DMF (1.2 L) was 2-[(phenylmethyl)amino]ethanol (155.3 g, 1.03
mol) at 25 °C and the mixture was stirred at 25 °C for 4 h. After add-
ing MeOH (1.2 L) to the mixture, a small amount of 20 (200 mg)
was seeded. The mixture was stirred at 25 °C for 1 h, then MeOH
(2.5 L) was added dropwise and the suspension was cooled to 5 °C.
The precipitate was filtered to give the crude amine 20 (251.0 g).
The crude compound (250.0 g) was purified by silica gel column
chromatography (Wakogel-200,13 2,500 g, CHCl3) and recrystal-
lized from CHCl3 (2.5 L) and MeOH (5.0 L) to give purified 20
(194.8 g, 70% from 19) as dark red solid; mp 160 °C (dec).
IR (KBr): ν = 3404, 2928, 1659, 1626, 1582, 1520, 1286, 1234,
1170, 816 cm–1.
SIMS: m/z = 398 (M++1).
1H NMR (CDCl3/TMS): d = 2.10 (br s, 1H), 2.78 (t, 2H, J = 5.3Hz),
3.67 (t, 2H, J = 5.3Hz), 3.69 (s, 2H), 4.11 (s, 2H), 7.17–7.29 (m,
5H), 7.49 (s, 1H), 7.69 (d, 1H, J = 8.3 Hz), 7.86 (d, 1H, J = 8.3 Hz),
9.49 (s, 1H), 13.26 (s, 1H).
Anal: calcd for C20H23N5O4·2HCl: C, 51.07; H, 5.36; N, 14.89.
Found: C, 50.99; H, 5.38; N, 14.64.
Synthesis 1999, No. 6, 947– 952 ISSN 0039-7881 © Thieme Stuttgart · New York

952
T. Mimura et al.
PAPER
drosulfite. On the other hand, the reduction reaction progres-
sed in the two-phase system of aqueous sodium hydrosulfite
and a large amount (300 v/w based on 18) of CHCl3. This me-
thod was too diluted and very inefficient for the production, so
a new efficient reducing method was required.
References
§
Present address: Yokkaichi Research Laboratories, Kyowa
Yuka Co., Ltd., 2-3 Daikyocho, Yokkaichi, Mie 510–8502,
Japan
(1) Sugaya, T.; Mimura, Y.; Shida, Y.; Osawa, Y.; Matsukuma, I.;
Ikeda, S.; Akinaga, S.; Morimoto, M.; Ashizawa, T.; Okabe,
M.; Ohno, H.; Gomi, K.; Kasai, M. J. Med. Chem. 1994, 37,
1028.
(2) a) Ikeda, S.; Mimura, Y.; Sugaya, T.; Nakamura, A.; Ashiza-
wa, T.; Akinaga, S.; Ohno, H.; Okabe, M.; Gomi, K.; Kasai,
M. In 24th National Medicinal Chemistry Symposium; Ab-
stracts No. 110, Salt Lake City, Utah, USA, 1994.
b) Mimura, Y.; Shida, Y.; Kasai, M.; Ashizawa, T.; Gomi, K.
U.S. Patent 5220026, 1993; Eur. Patent 487097A1, 1992;
Chem. Abstr. 1992, 117, 90167.
(8) Evans, D. A.; Chapman, K. T.; Carreira, E. M. J. Am. Chem.
Soc. 1988, 110, 3560.
(9) HPLC analysis conditions; column: YMC pack AM-312 ODS
6 × 150 mm; mobile phase: MeCN/phosphate buffer (0.1 mol/
L, pH 2.7) = 70/30 and sodium 1-octanesufonate (0.01 mol/L);
flow rate: 1.0 mL/min; detection: UV 254 nm.
(10) HPLC analysis conditions; column: YMC pack AM-312 ODS
6 × 150 mm; mobile phase: MeCN/phosphate buffer (0.1 mol/
L, pH 2.7) = 35/65 and sodium 1-octanesufonate (0.01 mol/L);
flow rate: 1.0 mL/min; detection: UV 254 nm.
(11) HPLC analysis conditions; column: YMC pack AM-312 ODS
6 × 150 mm; mobile phase: MeCN/phosphate buffer (0.1 mol/
L, pH 2.7) = 30/70 and sodium 1-octanesufonate (0.01 mol/L);
flow rate: 1.0 mL/min; detection: UV 254 nm.
(3) Ashizawa, T.; Shimizu, M.; Gomi, K.; Okabe, M. Anti-Cancer
Drugs 1998, 9, 263.
(4) Sugaya, T.; Mimura, Y.; Kato, N.; Ikuta, M.; Mimura, T.;
Kasai, M.; Tomioka, S. Synthesis 1994, 73.
(12) Kovendi, A.; Kiricz, M. Chem. Ber. 1964, 97, 1896.
(13) Wakogel-200 was commercially available silica gel from
Wako Chemical Co.
(5) Kato, N.; Sugaya, T.; Mimura, T.; Ikuta, M.; Kato, S.; Kuge,
Y.; Tomioka, S.; Kasai, M. Synthesis 1997, 625.
(6) HPLC analysis conditions; column: YMC pack AM-312 ODS
6 × 150 mm; mobile phase: MeOH/H2O = 6/1, pH 2.7 adjusted
with 85% phosphoric acid; flow rate: 1.0 mL/min; detection:
UV 254 nm.
Article Identifier:
1437-210X,E;1999,0,06,0947,0952,ftx,en;F08398SS.pdf
(7) In the similar method reported by Kato et al.,5 the quinone 18
was reduced incompletely in MeCN and aqueous sodium hy-
Synthesis 1999, No. 6, 947–952 ISSN 0039-7881 © Thieme Stuttgart · New York