Syntheses and bioactivities of substituted 9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrones. Unusual reactivities with amines

Article abstract of DOI:10.1021/jo010958s

A number of substituted 9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrones have been synthesized and their anticancer and antimalarial activities evaluated. A one-pot synthesis of 2,5,8-trimethoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4-dione (4) was achieved by heating a mixture of 1,4-dimethoxyanthracene, methoxyhydroquinone, silver oxide, and zinc iodide in toluene. Regioselective bromination of 4 and 2-methoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrone (7) with N-bromosuccinimide provided 2-bromo-3,5,8-trimethoxy-9,10-dihydro-9,10-[1,2]-benzenoanthracene-1,4-dione and 2-bromo-3-methoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrone (1), respectively. The reactions of 1 with aliphatic primary amines and secondary amines, respectively, produced different products, a result most likely attributed to the different basicities (or nucleophilicities) and steric effects of the two kinds of amines. The structure of the displacement product, 2-bromo-3- [2-(tert-butoxycarbonyl)ethylamino]-9,10-dihydro-9,10-[1,2] benzenoanthracene-1,4,5,8-tetrone, from the reaction of 1 with tert-butyl 3-aminopropanoate was unequivocally determined by a single-crystal X-ray analysis. IC₅₀ values of triptycene bisquinones for the inhibition of L1210 leukemia cell viability are in the 0.11-0.27 μM range and for the inhibition of Plasmodium falciparum 3D7 are in the 4.7-8.0 μM range.

Full text of DOI:10.1021/jo010958s

J . Org. Chem. 2002, 67, 2907-2912
2907
Syn th eses a n d Bioa ctivities of Su bstitu ted
9,10-Dih yd r o-9,10-[1,2]ben zen oa n th r a cen e-1,4,5,8-tetr on es. Un u su a l
Rea ctivities w ith Am in es
Duy H. Hua,* Masafumi Tamura, Xiaodong Huang, Heidi A. Stephany, Brian A. Helfrich,
Elisabeth M. Perchellet,† Bonnie J . Sperfslage,† J ean-Pierre Perchellet,† Suping J iang,‡
Dennis E. Kyle,‡ and Peter K. Chiang‡
Department of Chemistry, Kansas State University, Manhattan, Kansas 66506, Department of Biology,
Kansas State University, Manhattan, Kansas 66506, and Division of Experimental Therapeutics,
Walter Reed Army Institute of Research, Silver Spring, Maryland 20910-7500
duy@ksu.edu
Received September 27, 2001
A number of substituted 9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrones have been
synthesized and their anticancer and antimalarial activities evaluated. A one-pot synthesis of 2,5,8-
trimethoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4-dione (4) was achieved by heating a
mixture of 1,4-dimethoxyanthracene, methoxyhydroquinone, silver oxide, and zinc iodide in toluene.
Regioselective bromination of 4 and 2-methoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-
tetrone (7) with N-bromosuccinimide provided 2-bromo-3,5,8-trimethoxy-9,10-dihydro-9,10-[1,2]-
benzenoanthracene-1,4-dione and 2-bromo-3-methoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-
1,4,5,8-tetrone (1), respectively. The reactions of 1 with aliphatic primary amines and secondary
amines, respectively, produced different products, a result most likely attributed to the different
basicities (or nucleophilicities) and steric effects of the two kinds of amines. The structure of the
displacement product, 2-bromo-3-[2-(tert-butoxycarbonyl)ethylamino]-9,10-dihydro-9,10-[1,2]ben-
zenoanthracene-1,4,5,8-tetrone, from the reaction of 1 with tert-butyl 3-aminopropanoate was
unequivocally determined by a single-crystal X-ray analysis. IC50 values of triptycene bisquinones
for the inhibition of L1210 leukemia cell viability are in the 0.11-0.27 µM range and for the
inhibition of Plasmodium falciparum 3D7 are in the 4.7-8.0 µM range.
I. In tr od u ction
ticularly substituted analogues,4 show potent anticancer5
and antimalarial activities. Herein are reported the
synthesis of 2-bromo-3-methoxy-9,10-dihydro-9,10-[1,2]-
benzenoanthracene-1,4,5,8-tetrone (1) (a substituted trip-
tycene bisquinone), its unusual addition reactions with
aliphatic primary and secondary amines, and a summary
of its biological activities.
The unique shape and spectral properties of triptycenes
and triptycene quinones have attracted a number of
studies on their synthesis and application in new de-
vices,1 but only limited reports on biological activities.2
In our studies of beltenes3 and their nanotubes and
biological activity, triptycene bisquinones were required.
Fortuitously, a number of triptycene bisquinones, par-
II. Resu lts a n d Discu ssion
† Department of Biology, Kansas State University.
In the reported synthesis of 5,8-disubstituted trip-
tycene monoquinones,1 a sequence of three reactions was
carried out: Diels-Alder reaction of 1,4-dimethoxyan-
thracene (2)6 and p-benzoquinone followed by isomeriza-
tion1c of the adducts (stereoisomers) with potassium
hydroxide and then oxidation with silver oxide. We have
successfully combined these three reactions into one
simple operation, viz. heating a mixture of anthracene 2
(derived from the reduction of 1,4-anthracenedione5 with
sodium hydrosulfite followed by sodium hydride and
iodomethane), 1.4 equiv of methoxyhydroquinone (3), 2.7
‡ Walter Reed Army Institute of Research.
(1) (a) Bartlett, P. D.; Ryan, M. J .; Cohen, S. G. J . Am. Chem. Soc.
1942, 64, 2649. (b) Skvarchenko, V. R.; Shalaev, V. K.; Klabunovskii,
E. I. Russ. Chem. Rev. 1974, 43, 951. (c) Iwamura, H.; Makino, K. J .
Chem. Soc., Chem. Commun. 1978, 720. (d) Quast, H.; Fuchsbauer,
H.-L. Chem. Ber. 1986, 119, 1016. (e) Quast, H.; Fuchsbauer, H.-L.
Chem. Ber. 1986, 119, 2414. (f) Patney, H. K. Synthesis 1991, 694.
Charge-transfer complexes: (f) Lipczynska-Kochany, E.; Iwamura, H.
Chem. Lett. 1982, 1075. (g) Daub, J .; J akob, L.; Salbeck, J . Chem. Ber.
1988, 121, 2187. Radical Anions: (h) Russell, G. A.; Suleman, N. K.;
Iwamura, H.; Webster, O. W. J . Am. Chem. Soc. 1981, 103, 1560.
Liquid crystals: (i) Norvez, S. J . Org. Chem. 1993, 58, 2414.
(2) Driscoll, J . S.; Hazard, G. F. J r.; Wood, H. B., J r.; Goldin, A.
Cancer Chemother. Rep. 1974, P2, 4(2), 1.
(3) Reviews: (a) Schroder, A.; Mekelburger, H.-B.; Vogtle, F. Top.
Curr. Chem. 1994, 172, 179. (b) Kohnke, F. H.; Mathias, J . P.; Stoddart,
J . F. Top. Curr. Chem. 1993, 165, 1. (c) Stoddart, J . F. J . Inclusion
Phenom. Mol. Recog. Chem. 1989, 7, 227. (d) Kohnke, F. H.; Slawin,
A. M. Z.; Stoddart, J . F.; Williams, D. J . Angew. Chem., Int. Ed. Engl.
1987, 26, 892. Calculation of beltenes and cyclacenes: (e) Alder, R.
W.; Sessions, R. B. J . Chem. Soc., Perkin Trans. 2 1985, 1849. (f) Angus,
R. O., J r.; J ohnson, R. P. J . Org. Chem. 1988, 53, 314. (g) Choi, H. S.;
Kim, K. S. Angew. Chem., Int. Ed. 1999, 38, 2256 and references
therein. For other synthetic studies: (h) Kintzel, O.; Luger, P.; Weber,
M.; Schluter, A.-D. Eur. J . Org. Chem. 1998, 99. (i) Cory, R. M.;
McPhail, C. L. Tetrahedron Lett. 1996, 37, 1987. (j) Graham, R. J .;
Paquette, L. A. J . Org. Chem. 1995, 60, 5770. (k) Alder, R. W.; Allen,
P. R.; Edwards, L. S.; Fray, G. I.; Fuller, K. E.; Gore, P. M.; Hext, N.
M.; Perry, M. H.; Thomas, A. R.; Turner, K. S. J . Chem. Soc., Perkin
Trans. 1 1994, 3071.
(4) A preliminary report was presented at the 35th Midwest
Regional Meeting of the American Chemical Society, St. Louis, MO;
Oct 2000, Abstract No. 166.
(5) Our previously reported anti-cancer studies: Perchellet, E. M.;
Magill, M. J .; Huang, X.; Brantis, C. E.; Hua, D. H.; Perchellet, J . P.
Anti-cancer Drugs, 1999, 10, 749.
(6) Criswell, T. R.; Klanderman, B. H. J . Org. Chem. 1974, 39, 770.
10.1021/jo010958s CCC: $22.00 © 2002 American Chemical Society
Published on Web 04/06/2002

2908 J . Org. Chem., Vol. 67, No. 9, 2002
Hua et al.
Sch em e 1
Sch em e 2
nucleophilicity of the adjacent double-bonded carbon
resulting in the bromination.
To permit an examination of the reactivities of trip-
tycene bromoquinone 1 and the synthesis of new ana-
logues, 1 was treated with primary and secondary amines
(Scheme 2).8 Surprisingly, compound 1 reacts with pri-
mary amines such as methylamine and secondary amines
such as dimethylamine to give different regioisomeric
products. Hence, treatment of bromide 1 with methyl-
amine in THF at 0 °C for 20 min gave a 66% yield of
displacement product 8 and a 21% recovery of 1. On the
other hand, when 1 was treated with dimethylamine in
THF at 0 °C, regioisomers 9 and 10 (1:1 ratio) were
isolated in 96% yield and separated by silica gel column
chromatography. The regiochemistry of 9 and 10 has not
been determined.9 The presence of a bromine atom in the
products was indicated by their mass spectra in which
the M + 2 peaks (81Br isotope) were almost equal in
intensity to those of the molecular (EI) or quasimolecular
(CI) ions. No other byproducts were identifiable in these
reactions. These unusual addition reactions are unprec-
edented. It has been reported that p-benzoquinone un-
dergoes addition reactions10 with aliphatic primary amines
to give a mixture of products including mono- and
diadducts from the 1,4-addition reactions, while tetra-
chloro-1,4-benzoquinone and 2,3-dichloro-1,4-naphtho-
quinone react with secondary amines to give displace-
ment products11 (addition to the double bond followed by
elimination of chlorine). Contrary to these literature
results, bromide 1 reacts with primary amines to undergo
displacement of the methoxy group instead of bromine11c
and with secondary amines exclusively undergoes simple
1,4-addition on the unsubstituted quinone ring followed
by oxidation. Presumably, a less basic primary amine,
like methylamine,12 undergoes 1,4-addition reaction on
the more electron-deficient bromoquinone ring. Since the
C2-bromine of 1 is an electron-withdrawing group, the
resulting anion from the addition of an amine on C3
equiv of silver oxide, and 0.2 equiv of zinc iodide in
toluene under reflux for 4 days gave a 70% yield of
triptycene monoquinone 4 (Scheme 1). Presumably, silver
oxide oxidizes methoxyhydroquinone to methoxy-p-ben-
zoquinone, which undergoes a Diels-Alder reaction with
anthracene 2 to give adducts 5 (two stereoisomers).
Compound 5 then undergoes oxidation with silver oxide
to give quinone 4. A catalytic amount of zinc iodide was
added to facilitate the Diels-Alder reaction. No other
regioisomers (such as isomers with the methoxy sub-
stituent attached at C4a) were detected. To verify the
reaction sequence, methoxyquinone 6, obtained from the
oxidation of 3 with silver oxide and potassium carbonate
in benzene (98% yield), was treated with anthracene 2
in toluene at 150 °C in a sealed tube to give adducts 5
(48% yield). Isomerization of ketones 5 with KOH in
p-dioxane and water1c followed by oxidation with silver
oxide gave monoquinone 4 (95% overall yield). Oxidation
of monoquinone 4 with ceric ammonium nitrate1c afforded
a 94% yield of triptycene bisquinone 7. Regioselective
bromination of 7 was achieved by the treatment with
N-bromosuccinimide (NBS) in DMF at 25 °C for 10 h to
give bromoquinone 1 (45% yield). Alternatively, com-
pound 1 was also obtained from the bromination of
monoquinone 4 with NBS in DMF at 40 °C followed by
oxidation with ceric ammonium nitrate (59% overall
yield). Apparently, the reactivity of the methoxyquinone
moiety toward NBS is greater than that of quinone and
1,4-dimethoxyphenyl moieties.7 It is likely that the
methoxy substituent of 7 or 4 (at C2) enhances the
(8) Nucleophilic reactions of quinones have been reviewed: Kutyrev,
A. Tetrahedron 1991, 47, 8043. Nucleophilic reactions of triptycene
quinones have not been reported.
(9) Recrystallization of compounds 9 or 10 from different organic
solvents such as ethyl acetate, diethyl ether, diisopropyl ether, ethanol,
etc. failed to provide single crystals suitable for X-ray analysis.
(10) Ott, R.; Pinter, E.; Kajtna, P. Monatsh. Chem. 1980, 111, 813.
(11) (a) Gauss, W. Heitzer, H.; Petersen, S. Liebigs Ann. Chem. 1972,
764, 131. Displacement of 2-methoxy-1,4-naphthoquinone11a and 6-meth-
oxy-5,8-quinolinequinone has also been reported: (b) Pratt, Y. T.;
Drake, N. L. J . Am. Chem. Soc. 1955, 77, 37. Displacement of 7-bromo-
6-methoxy-5,8-quinolinedione with ammonia: (c) Liao, T. K.; Nyberg,
W. H.; Cheng, C. C. Angew. Chem., Int. Ed. Engl. 1967, 6, 82.
(12) The pKb values of methylamine and dimethylamine are 3.38
and 3.23, respectively: Hand C. W.; Blewitt, H. L. Acid-Base
Chemistry; Macmillan: New York, 1986; pp 253, 254.
(7) It should be noted that 2-methyl-9,10-dimethoxyanthracene
reacts with NBS to give 2-methyl-9,10-anthraquinone: (a) J iang, H.;
Xu, H.; Ye, J . J . Chem. Soc., Perkin Trans. 2 2000, 925. Under our
reaction conditions, no oxidation of the 1,4-dimethoxyphenyl ring
moiety of 4 with NBS was observed. Bromination of juglone (5-hydroxy-
1,4-naphthoquinone) with NBS has been reported: (b) Wurm, G.;
Geres, U. Arch. Pharm. 1989, 322, 155.

9,10-Dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrones
J . Org. Chem., Vol. 67, No. 9, 2002 2909
Sch em e 3
F igu r e 1. ORTEP drawing of X-ray structure of 12.
Ta ble 1. Cytotoxicities of Tr ip tycen e Bisqu in on e
An a logs in th e L1210 Leu k em ic Cell System in Vitr o
compd
IC50 (µΜ)
compd
IC50 (µΜ)
1
7
8
9
0.135 ( 0.009
0.125 ( 0.012
0.270 ( 0.015
0.116 ( 0.010
11
12
13
14
0.110 ( 0.008
0.176 ( 0.014
0.694 ( 0.042
2.151 ( 0.093
romethane gave a quantitative yield of acid 13. The
water-soluble sodium salt 14 was obtained by the treat-
ment of 13 with 1 equiv of sodium hydroxide.
would be more stabilized than that from the addition on
C2. On the other hand, a more basic secondary amine,
like dimethylamine (a more reactive amine than methyl-
amine),12 is less affected by the electronic effect of the
quinone ring and is more affected by steric effects, and
consequently prefers to add on the unsubstituted (less
hindered) quinone moiety to provide compounds 9 and
10.
Due to the potent anticancer activity of amine 8 (vide
infra) and the need for water-soluble analogues, aliphatic
primary amines containing an ester function were used
to synthesize various triptycene quinones, and their
biological activities were studied (Scheme 3).
Treatment of bromide 1 with ethyl 2-aminopropanoate
in THF at -40 °C for 6 h afforded a 46% yield of
displacement product 11 along with 30% recovered
starting material 1. No other regioisomers were detected.
Similarly, reaction of 1 with tert-butyl 2-aminopropanoate
at -40 °C for 8 h gave a 62% yield of 12. Recrystallization
of 12 in hexane/ethyl acetate (5:1) afforded single crystals
whose structure was unequivocally shown by X-ray
analysis (Figure 1).13 Interestingly, in the stacking of
crystals, the three different bridges (rings) of triptycene
12 stack with the same groups of three other molecules
with an average distance of ∼3.7 Å. That is, the phenyl
ring of a molecule stacks with the phenyl ring of another
molecule, unsubstituted quinone ring stacks with an
unsubstituted quinone ring, and the substituted quinone
ring stacks with a substituted quinone ring (C2 of one
molecule is on top of C4 of another molecule). A hydrogen
bond was present between the NH and the C4-oxygen (a
distance of 2.186 Å). Deprotection of the tert-butyl ester
of 12 by treatment with trifluoroacetic acid in dichlo-
Anticancer and antimalarial activities of triptycene
bisquinones have been evaluated. Table 1 summarizes
IC50 values (the concentrations of drugs required to
inhibit by 50% the viability of L1210 leukemic cells at
day 4) of various triptycene bisquinones. In general, the
IC50 values are in the 0.11-0.27 µM range with the
exception of carboxylic acid 13 and its sodium salt 14.
Possibly, the ability of sodium salt 14 to pass through
the cell membrane may decrease as the water solubility
increases. This would, in turn, decrease the ability of this
compound to destroy cancer cells. The IC50 value of
daunomycin, a known anticancer drug, under similar
treatment conditions is 0.030 µM.5 Triptycene bisquino-
nes induce DNA cleavage and inhibit nucleoside trans-
port. In contrast, daunomycin is a DNA topoisomerase
II inhibitor that does not block nucleoside transport.
Moreover, triptycene bisquinones 7-9 and ritonavir14
inhibit Plasmodium falciparum 3D7 (a malaria strain)15a
with IC50 values of 8.0, 4.7, 5.6, and 9.7 µM, respectively.
Malaria protease plasmepsin II15b-d was also inhibited
by compounds 8, 9, and ritonavir,14 and IC50 values of
these compounds are 9.7, 23.6, and 0.10 µM, respectively.
Presumably, these reactive triptycene bisquinones, such
as 8 and 9, undergo addition reactions with lysine,16
tryptophan, histidine, and cysteine17 residues of proteins
to produce cross-linked proteins.
(14) Ritonavir,
a protease inhibitor, was used for comparison.
Danner, S. A.; Carr, A.; Leonard, J . M.; Lehman, L. M.; Gudiol, F.;
Gonzales, J .; Raventos, A.; Rubio, R.; Bouza, E.; Pintado, V.; Aguado,
A. G.; Garcia de Lomas, J .; Delgado, R.; Borleffs, J . C. C.; Hsu, A.;
Valdes, J . M.; Boucher, C. A. B.; Cooper, D. A. N. Engl. J . Med. 1995,
333, 1528.
(15) The protocol for bioassay for P. falciparum 3D7: (a) Opsenica,
D.; Pocsfalvi, G.; J uranic, Z.; Tinant, B.; Declercq, J .-P.; Kyle, D. E.;
Milhous, W. K.; Solaja, B. A. J . Med. Chem. 2000, 43, 3274. Bioassay
for plasmepsin inhibition: (b) J iang, S.; Prigge, S. T.; Wei, L.; Gao, Y.;
Hudson, T. H.; Gerena, L.; Dame, J . B.; Kyle, D. E. Antimicrobial
Agents Chemother. 2001, 45, 2577. (c) Carroll, C. D.; Orlowski, M. Adv.
Exp. Med. Biol. 1998, 436, 375. (d) Carroll, C. D., T. O. J ohnson, S.
Tao, G. Lauri, M. Orlowski, I. Y. Gluzman, D. E. Goldberg, and R. E.
Dolle. Bioorg. Med. Chem. Lett. 1998, 8, 3203.
(13) The author has deposited atomic coordinates for this structure
with the Cambridge Crystallographic Data Centre as no. CCDC
175022. The coordinates can be obtained on request, from the Director,
Cambridge Crystallographic Data Centre, 12 Union Road, Cambridge,
CB2 1EZ, U.K. R factor of the X-ray analysis is 0.0379 and the crystals
have a space group of C2/c. It should be noted that only one single-
crystal X-ray structure of triptycene monoquinone has been reported:
Wiehe, A.; Senge, M. O.; Kurreck, H. Liebigs Ann. Chem. 1997, 1951.

2910 J . Org. Chem., Vol. 67, No. 9, 2002
Hua et al.
compound 2. Crystallization from ether/hexane (1:1) gave 0.89
g (78% yield) of yellow solids: mp 132-133 °C (lit.6 134-136
III. Con clu sion s
1
°C); MS (FAB) m/z 239 (M + 1), 238 (M+); H NMR δ 8.70 (s,
A one-pot synthesis of 2,5,8-trimethoxy-9,10-dihydro-
9,10-[1,2]benzenoanthracene-1,4-dione (4) was accom-
plished in high yield by heating a mixture of 1,4-
dimethoxyanthracene (2), methoxyhydroquinone (3), silver
oxide, and zinc iodide in toluene. A regioselective bromi-
nation of triptycene bisquinone 7 with NBS was found
to provide 2-bromo-3-methoxy-9,10-dihydro-9,10-[1,2]-
benzenoanthracene-1,4,5,8-tetrone (1). Under similar
reaction conditions, monoquinone 4 was also brominated
with NBS to provide the C2-bromo derivative. Unusual
displacement and addition reactions of 1 with aliphatic
primary and secondary amines, respectively, provided a
number of potent anticancer agents. Other nucleophilic
reagents such as thiols and aminocarbohydrates may be
suitable for the above reactions. The potent anticancer
and antimalarial activities provide lead compounds for
further investigation in new drug discovery. The afore-
mentioned reactions and biological studies are being
undertaken.
2 H, C9,10 H), 7.97 (dd, J ) 6.1, 3.6 Hz, 2 H, C5,8 H), 7.40
(dd, J ) 6.6, 3.2 Hz, 2 H, C6,7 H), 6.55 (s, 2 H, C2,3 H), 3.97
(s, 6 H, OCH3); 13C NMR δ 149.5 (s, C8a,10a), 131.5 (s, C4a,9a),
128.5 (d, C9,10), 125.5 (d, C5,8), 120.7 (d, C6,7), 100.9 (d, C2,3),
55.6 (s, OCH3).
2,5,8-Tr im et h oxy-9,10-d ih yd r o-9,10-[1,2]b en zen oa n -
th r a cen e-1,4-d ion e (4). A mixture of 2.00 g (8.40 mmol) of
1,4-dimethoxyanthracene (2), 1.411 g (10.1 mmol) of meth-
oxyhydroquinone (3), 3.90 g (16.8 mmol) of silver oxide, and
0.536 g (1.68 mmol) of zinc iodide in 30 mL of toluene under
argon was refluxed for 3 d. To the mixture were added 0.25 g
(1.79 mmol) of 3 and 1.30 g (5.60 mmol) of silver oxide, and
the reaction mixture was refluxed for another day. The
reaction mixture was cooled, diluted with 200 mL of dichlo-
romethane, and filtered through Celite, and the filtrate was
washed with aqueous NH4Cl and brine, dried (MgSO4),
concentrated, and column chromatographed on silica gel using
a gradient mixture of hexane and ethyl acetate as solvent to
give 2.21 g (70% yield) of 4: mp 108-110 °C; 1H NMR δ 7.43
(dd, J ) 5.5, 3 Hz, 2 H), 7.00 (dd, J ) 5.5, 3 Hz, 2 H), 6.52 (s,
2 H, C6,7 H), 6.25 (s, 1 H), 6.23 (s, 1 H), 5.71 (s, 1 H, C3 H),
3.79 (s, 6 H, OCH3), 3.72 (s, 3 H, OCH3); 13C NMR δ 183.5
(CdO), 178.2 (CdO), 158.4, 153.8, 150.9, 150.5, 149.6, 149.5,
144.3, 144.1, 133.6, 133.5, 125.2 (2 C), 124.5, 124.4, 109.5,
105.6, 56.4, 56.38, 56.3, 41.5, 41.2. Anal. Calcd for C23H18O5:
C, 73.79; H, 4.85. Found: C, 73.51; H, 5.07.
2,5,8-Tr im eth oxy-4a ,9,9a ,10-tetr a h yd r o-9,10-[1,2]ben -
zen oa n th r a cen e-1,4-d ion e (5). A mixture of 1.00 g (7.10
mmol) of methoxyhydroquinone (3), 2.50 g (10.7 mmol) of silver
oxide, and 1.20 g (8.50 mmol) of K2CO3 in 50 mL of benzene
was stirred under argon at 25 °C for 3 h, and the mixture was
filtered through Celite and washed with 5 mL of dichlo-
romethane. The filtrate was concentrated to give 0.970 g (99%
yield) of methoxybenzoquinone (6):19 1H NMR δ 6.72 (s, 2 H),
5.95 (s, 1 H), 3.84 (s, 3 H); 13C NMR δ 187.4, 181.6, 137.1,
134.4, 111.5, 107.6, 56.2. This material was used in the next
step without purification.
IV. Exp er im en ta l Section
Gen er a l Meth od s. NMR spectra were obtained at 400 MHz
for 1H and 100 MHz for 13C in CDCl3, unless otherwise
indicated. IR spectra are reported in wavenumbers (cm-1). FAB
spectra were taken by using Xe beam (8 KV) and m-nitrobenzyl
alcohol as matrix. High-resolution mass spectra were obtained
using the MALDI technique and CHCl3/MeOH (1:1) as a
matrix. Davisil silica gel, grade 643 (200-425 mesh), was used
for the flash column chromatographic separation.
1,4-Dim eth oxya n th r a cen e (2). To a cold (0 °C) methanol
(20 mL) solution of 1.00 g (0.0042 mol) of quinizarin under
argon was added 0.638 g (0.0168 mol) of sodium borohydride.
The resulting mixture was stirred at 0 °C for 1 h. To it, was
added 11 mL of 6 N HCl dropwise at 0 °C over a period of 10
min. The precipitated orange solids were collected, washed
several times with distilled water, dried under vacuum, and
recrystallized from acetone-ether to give 0.83 g (95% yield)
of anthracene-1,4-dione18 as yellow crystals: mp 204-206 °C;
1H NMR δ 8.60 (s, 2 H, C9,10 H), 8.10 (dd, J ) 6.4, 3.2 Hz, 2
H, C5,8 H), 7.70 (dd, J ) 6.4, 3.2 Hz, 2 H, C6,7 H), 7.10 (s, 2
H, C2,3 H); 13C NMR δ 184.7 (s, CdO), 140.1 (d), 134.8 (s),
130.2 (d), 129.6 (d), 128.9 (d), 128.4 (s).
To 2.00 g (1.00 mmol) of 1,4-anthracenedione was added a
solution of 6.68 g (38.0 mmol) of sodium hydrosulfite in 50 mL
of water and 50 mL of 1,4-dioxane. The resulting mixture was
stirred at 25 °C for 3 h and added to 100 mL of water. The
mixture was cooled over an ice-water bath, and the precipi-
tated dark green solid was collected by filtration, washed twice
with water, and dried under vacuum to give 1.75 g (87% yield)
of 1,4-dihydroxyanthracene: mp 167-169 °C; 1H NMR δ 8.70
(s, 2 H, C9,10 H), 8.05 (m, 2 H, C5,8 H), 7.50 (s, 2 H, C6,7 H),
6.60 (s, 2 H, C2,3 H); 13C NMR δ 145.4, 130.5, 128.4, 125.3,
125.2, 120.6, 105.6.
To a 0.275 g (11.0 mmol) of oil-free sodium hydride under
argon were added 1.00 g (4.80 mmol) of 1,4-dihydroxy-
anthracene, 0.75 mL (12.0 mmol) of iodomethane, and 10 mL
of dry DMF. The solution was stirred at 25 °C for 1.5 h, diluted
with 20 mL of water, and acidified with 6 N HCl (pH ∼2). The
mixture was extracted three times with ethyl acetate, and the
combined extract was washed twice with water and brine,
dried (MgSO4), and concentrated to give 1.07 g (94% yield) of
A solution of 0.70 g (2.90 mmol) of 2 and 1.00 g (7.20 mmol)
of 6 in 10 mL of toluene in a sealed tube was heated at 150 °C
for 1 d, cooled to 25 °C, concentrated to dryness, and column
chromatographed on silica gel using a gradient mixture of
hexane and ethyl acetate as solvent to give 0.52 g (48% yield)
of compounds 5 as a mixture of stereoisomers (1:1) along with
0.62 g of 6. The stereoisomers were partially separated by silica
gel column chromatography; the stereochemistry has not been
assigned. Compound 5, less polar isomer: MS m/z 376 (M+),
1
375; H NMR δ 7.24-7.18 (m, 2 H), 7.08-7.04 (m, 2 H), 6.65
(s, 2 H, C6,7 H), 5.61 (s, 1 H), 5.33 (bs, 2 H), 3.82 (s, 6 H,
OCH3), 3.48 (s, 3 H, OCH3), 3.12 (d, J ) 9 Hz, 1 H), 3.06 (d, J
) 9 Hz, 1 H); 13C NMR δ 197.4 (CdO), 193.5 (CdO), 162.0,
148.7, 141.9, 140.4, 139.8, 131.6, 131.4, 129.3, 126.6, 125.3,
125.0, 113.7, 109.5, 109.3, 56.7, 56.4, 49.3, 48.6, 42.7, 42.6, 42.4.
More polar isomer: mp 175-176 °C; MS m/z 376 (M+), 375;
1H NMR δ 7.40 (m, 2 H), 7.16 (m, 2 H), 6.60 (d, J ) 8.8 Hz, 1
H), 6.56 (d, J ) 8.8 Hz, 1 H), 5.65 (s, 1 H, dCH), 5.33 (bs, 2 H,
C9,10 H), 3.76 (s, 3 H, OCH3), 3.73 (s, 3 H, OCH3), 3.51 (s, 3
H, OCH3), 3.13 (d, J ) 9 Hz, 1 H), 3.08 (d, J ) 9 Hz, 1 H); 13
C
NMR δ 197.0 (CdO), 193.2 (CdO), 162.0, 149.7, 149.3, 142.1,
141.9, 130.1, 129.3, 126.7, 126.6, 124.2, 124.17, 113.4, 110.2,
109.8, 56.7, 56.6, 56.2, 49.6, 48.7, 42.7, 42.5. Anal. Calcd for
C
23H20O5: C, 73.39; H, 5.36. Found: C, 73.11; H, 5.68.
1,4-Dih yd r oxy-2,5,8-tr im eth oxy-9,10-d ih yd r o-9,10-[1,2]-
ben zen oa n th r a cen e. To a solution of 0.77 g (2.0 mmol) of 5
(a mixture of stereoisomers) in 30 mL of 1,4-dioxane and 30
mL of water was added 1.12 g (20 mmol) of KOH. The solution
was stirred at 25 °C for 1 h, acidified with 1 N HCl, and
extracted three times with dichloromethane. The combined
extract was washed with brine, dried (MgSO4), and concen-
(16) Compound 1 reacted with L-lysine to give the expected C-3
displacement product. These data along with a full account of biological
studies will be reported in due course.
(17) Quinones have been proposed to react with the thiol group of
cdc25 phosphatase: Ham, S. W.; Park, H. J .; Lim, D. H. Bioorg. Chem.
1997, 25, 33, and references therein.
(18) Bedworth, P. V.; Perry, J . W.; Marder, S. R. Chem. Commun.
1997, 1353.
(19) Cavill, G. W. K.; Quinn, R. J . Aust. J . Chem. 1973, 26, 595 and
references therein.

9,10-Dihydro-9,10-[1,2]benzenoanthracene-1,4,5,8-tetrones
J . Org. Chem., Vol. 67, No. 9, 2002 2911
trated to give 0.77 g (100% yield) of 1,4-dihydroxy-2,5,8-
ether, washed with water and brine, dried (MgSO4), concen-
trated, and column chromatographed on silica gel using a
gradient mixture of hexane and ether as eluant to give 60 mg
(59% yield) of 1.
trimethoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene: mp 257-
1
259 °C; MS m/z 376 (M+); H NMR δ 7.43 (dd, J ) 5.5, 3 Hz,
1 H), 7.38 (dd, J ) 5.5, 3 Hz, 1 H), 6.96 (dd, J ) 5.5, 3 Hz, 2
H), 6.51 (s, 2 H, C6,7 H), 6.29 (s, 1 H, C3 H), 6.09 (s, 1 H),
5.98 (s, 1 H), 5.3 (s, 1 H, OH), 4.96 (bs, 1 H, OH), 3.81 (s, 3 H,
OCH3), 3.805 (s, 3 H, OCH3), 3.66 (s, 3 H, OCH3); 13C NMR
(DMSO-d6) δ 148.5, 148.3, 146.4, 146.0, 145.7, 144.1, 135.5,
135.0, 134.6, 132.6, 124.6, 124.4, 123.5, 123.3, 123.1, 109.0,
108.8, 97.4, 56.1 (2 C), 56.0, 40.4, 39.8. Anal. Calcd for
2-Br om o-3-(m eth yla m in o)-9,10-d ih yd r o-9,10-[1,2]ben -
zen oa n th r a cen e-1,4,5,8-tetr on e (8). A solution of 0.200 g
(0.473 mmol) of 1 and 0.47 mL (0.946 mmol) of methylamine
(2.0 M in THF) in 1 mL of THF was stirred under argon at 0
°C for 1 h. The solution was concentrated to dryness and
column chromatographed on silica gel using a gradient mixture
of hexane and ethyl acetate as solvent to give 0.132 g (66%
yield) of 8 and 41 mg (21% recovery) of 1. Compound 8: mp
>350 °C (polymerized); MS (CI) m/z 424, 422 (∼1:1; M + 1);
1H NMR δ 7.48 (dd, J ) 5, 3 Hz, 1 H), 7.45 (dd, J ) 5, 3 Hz,
1 H), 7.07 (dd, J ) 5, 3 Hz, 2 H), 6.64 (s, 2 H, C6,7 H), 6.28 (s,
1 H), 6.10 (s, 1 H), 5.88 (bs, 1 H, NH), 3.33 (d, J ) 5.6 Hz, 3
H, CH3N); 13C NMR (acetone-d6) δ 183.33, 183.3, 178.2, 170.9,
154.4, 152.3, 152.0, 148.0, 147.0, 143.7, 143.5, 136.4, 136.3,
126.5 (2 C), 126.0, 125.7, 125.0, 44.3, 42.9, 33.2. Anal. Calcd
for C21H12BrNO4: C, 59.74; H, 2.86. Found: C, 59.74; H, 2.76.
2-Br om o-3-m eth oxy-6-(d im eth yla m in o)-9,10-d ih yd r o-
9,10-[1,2]ben zen oan th r acen e-1,4,5,8-tetr on e (9) an d 2-Br o-
m o-3-m eth oxy-7-(dim eth ylam in o)-9,10-dih ydr o-9,10-[1,2]-
ben zen oa n th r a cen e-1,4,5,8-tetr on e (10). To a solution of
0.200 g (0.473 mmol) of 1 in 2 mL of THF under argon at 0 °C
was added 0.24 mL (0.473 mmol) of dimethylamine (2.0 M in
THF). After being stirred at 0 °C for 2 h, the reaction solution
was concentrated to dryness and column chromatographed on
silica gel using a mixture of benzene and ethyl acetate (10:1)
as solvent to give 0.099 g (48% yield) of 9 (less polar isomer;
the regiochemistry has not been determined) and 0.098 g (48%
yield) of 10 (more polar).
C
23H20O5: C, 73.39; H, 5.36. Found: C, 73.45; H, 5.11.
Oxid a tion of 1,4-Dih yd r oxy-2,5,8-tr im eth oxy-9,10-d i-
h yd r o-9,10-[1,2]ben zen oa n th r a cen e to 4. To a mixture of
0.77 g (2.0 mmol) of 1,4-dihydroxy-2,5,8-trimethoxy-9,10-
dihydro-9,10-[1,2]benzenoanthracene and 0.60 g (4.2 mmol) of
sodium sulfate (anhydrous) in 15 mL of dried acetone under
argon at 25 °C was added 0.56 g (4.0 mmol) of silver oxide.
The mixture was heated under reflux for 6 h, cooled to 25 °C,
diluted with dichloromethane, and filtered through Celite. The
filtrate was concentrated to give 0.77 g of the crude product.
Column chromatography on silica gel using a gradient mixture
of hexane and ethyl acetate as solvent gave 0.73 g (95% yield)
of 4.
2-Meth oxy-9,10-d ih yd r o-9,10-[1,2]ben zen oa n th r a cen e-
1,4,5,8-tetr on e (7). To a solution of 0.77 g (2.0 mmol) of 4 in
16 mL of acetonitrile, 26 mL of 1,4-dioxane, and 10 mL of water
was added 2.0 g (3.6 mmol) of ceric ammonium nitrate at 25
°C. The solution was stirred for 12 h, diluted with dichlo-
romethane, and washed with water. The organic layer was
dried (MgSO4) and concentrated to give the crude product.
Crystallization from ether gave 0.65 g (95% yield) of 7: MS
(CI) m/z 345 (M + 1), 317 (-CO); 1H NMR δ 7.48 (dd, J ) 5.5,
3 Hz, 2 H), 7.07 (dd, J ) 5.5, 3 Hz, 2 H), 6.65 (s, 2 H, C6,7 H),
6.2 (s, 1 H), 6.18 (s, 1 H), 5.78 (s, 1 H, C3 H), 3.78 (s, 3 H,
OCH3); 13C NMR δ 182.2, 182.17, 182.0, 176.9, 158.5, 152.5,
151.6, 151.5, 149.6, 142.2, 142.0, 135.4, 135.3, 125.8 (2 C),
125.4, 125.3, 105.7, 56.6 (OCH3), 42.2, 41.9. Anal. Calcd for
Less polar isomer: MS (CI) m/z 468, 466 (∼1:1; M + 1); 1H
NMR δ 7.47-7.43 (m, 2 H, Ar-H), 7.05-7.02 (m, 2 H, Ar-H),
6.22 (s, 1 H), 6.14 (s, 1 H), 5.38 (s, 1 H, C7 H), 3.80 (s, 3 H,
OCH3), 3.11 [s, 6 H, (CH3)2N]; 13C NMR δ 181.4 (2 C), 181.2 (2
C), 153.7, 150.4, 149.3, 144.1, 143.9, 142.2, 141.8, 139.1, 127.1,
125.5, 125.3, 124.7, 124.2, 102.5, 61.1, 42.8, 42.7, 42.0, 41.4;
HRMS m/z 466.0285 (466.0290, calcd for C23H17BrNO5, M-H+).
More polar isomer: MS (CI) m/z 468, 466 (∼1:1; M + 1); 1H
NMR δ 7.48-7.42 (m, 2 H, Ar-H), 7.04-7.01 (m, 2 H, Ar-H),
6.24 (s, 1 H), 6.16 (s, 1 H), 5.39 (s, 1 H, C7 H), 3.79 (s, 3 H,
OCH3), 3.10 [s, 6 H, (CH3)2N]; 13C NMR δ 181.6 (2 C), 181.4 (2
C), 154.0, 150.7, 149.6, 144.9, 144.0, 142.6, 141.5, 139.8, 127.2,
125.6 (2 C), 125.0, 124.3, 102.4, 61.2, 43.0 (2 C), 42.0, 41.9;
HRMS m/z 466.0285 (466.0290, calcd for C23H17BrNO5, M -
H+).
C
21H12O5: C, 73.25; H, 3.51. Found: C, 73.01; H, 3.80.
2-Br om o-3-m eth oxy-9,10-d ih yd r o-9,10-[1,2]ben zen oa n -
th r a cen e-1,4,5,8-tetr on e (1). To a solution of 0.30 g (0.87
mmol) of 7 in 20 mL of DMF under argon at 25 °C was added
0.16 g (0.87 mmol) of N-bromosuccinimide. After the solution
was stirred for 10 h, it was diluted with water and extracted
twice with ethyl acetate. The combined extract was washed
with brine, dried (MgSO4), concentrated, and column chro-
matographed on silica gel using a gradient mixture of hexane
and ether as solvent to give 0.29 g (45% yield) of 1: mp 207-
210 °C; MS EI m/z 424 and 422 (1:1, M+), 344 (M - Br), 300,
2-Br om o-3-[2-(eth oxyca r bon yl)eth yla m in o]-9,10-d ih y-
d r o-9,10-[1,2]ben zen oa n th r a cen e-1,4,5,8-tetr on e (11). To
a mixture of 5.70 mg (0.236 mmol) of oil-free NaH in 2 mL of
DMF under argon at 0 °C was added 36.3 mg (0.236 mmol) of
â-alanine ethyl ester hydrochloride (ethyl 3-aminopropanoate
hydrochloride). The solution was stirred at 0 °C for 30 min
and cooled to -46 °C, and a solution of 100 mg (0.236 mmol)
of 1 in 1 mL of DMF was added via cannula. The solution was
stirred at -46 °C for 6 h, diluted with a mixture of ethyl
acetate and benzene (1:1), washed twice with water and brine,
dried (Na2SO4), concentrated, and column chromatographed
on silica gel using a gradient mixture of hexane and ethyl
acetate as solvent to give 55 mg (46% yield) of 11 and 30 mg
(30% recovery) of 1. Compound 11: mp 119-121 °C; MS m/z
1
287, 232, 152, 126; H NMR δ 7.48 (dd, J ) 5.5, 3 Hz, 2 H),
7.09 (dd, J ) 5.5, 3 Hz, 2 H), 6.66 (s, 2 H, C6,7 H), 6.23 (s, 1
H), 6.16 (s, 1 H), 4.17 (s, 3 H, OCH3); 13C NMR δ 182.2 (s,
CO), 182.0 (s, CO), 176.2 (s, CO), 175.5 (s, CO), 156.2, 151.6,
151.4, 150.2, 141.8, 135.5, 135.4, 126.2, 126.1, 126.07, 126.0,
125.6, 125.4, 117.2, 61.7 (OCH3), 43.0, 42.1. Anal. Calcd for
C
21H11BrO5: C, 59.60; H, 2.62. Found: C, 59.33; H, 2.87.
Syn t h esis of 1 fr om Br om in a t ion of 4 F ollow ed b y
Oxid a tion . To a solution of 90 mg (0.24 mmol) of 4 in 5 mL of
DMF under argon at 25 °C was added 52 mg (0.30 mmol) of
N-bromosuccinimide (NBS). The solution was stirred at 40 °C
for 12 h, diluted with diethyl ether, washed twice with water
and once with brine, dried (MgSO4), and concentrated to give
109 mg (quantitative yield) of 2-bromo-3,5,8-trimethoxy-9,10-
dihydro-9,10-[1,2]benzenoanthracene-1,4-dione: MS (EI) m/z
1
510, 508 (∼1:1; M + 1), 422, 420 (1:1); H NMR δ 7.45-7.42
(m, 2 H, Ar-H), 7.08-7.04 (m, 2 H, Ar-H), 6.65 (s, 2 H, C6,7
H), 6.3 (bs, 1 H, NH), 6.27 (s, 1 H), 6.10 (s, 1 H), 4.16 (q, J )
7 Hz, 2 H, OCH2), 4.03 (q, J ) 6 Hz, 2 H, CH2N), 2.64 (t, J )
6 Hz, 2 H, CH2CO), 1.26 (t, J ) 7 Hz, 3 H, CH3); 13C NMR δ
184.0, 183.6, 182.6, 182.3, 177.3, 171.7, 154.5, 151.8, 151.7,
147.4, 144.3, 142.4, 142.2, 135.8, 135.5, 126.2, 126.1, 125.9,
1
454, 452 (1:1, M+); H NMR δ 7.44 (dd, J ) 5.5, 3 Hz, 2 H),
7.01 (dd, J ) 5.5, 3 Hz, 2 H), 6.54 (s, 2 H, C6,7 H), 6.29 (s, 1
H), 6.22 (s, 1 H), 4.14 (s, 3 H, OCH3), 3.81 (s, 3 H, OCH3), 3.79
(s, 3 H, OCH3); 13C NMR δ 177.7, 177.0, 156.4, 153.2, 151.6,
149.8, 149.7, 144.14, 144.1, 133.4, 125.7, 125.6 (2 C), 124.8,
124.7, 117.6, 109.9, 109.8, 61.8, 60.6, 56.6, 42.5, 41.6. This
material was used in the next step without purification.
To a solution of 0.109 g (0.240 mmol) of 2-bromo-3,5,8-
trimethoxy-9,10-dihydro-9,10-[1,2]benzenoanthracene-1,4-di-
one in 5 mL of acetonitrile, 6 mL of 1,4-dioxane, and 2 mL of
water was added 0.660 g (1.20 mmol) of ceric ammonium
nitrate. The solution was stirred at 25 °C for 6 h, diluted with
125.3, 61.3, 43.7, 42.2, 40.5, 35.4, 14.4. Anal. Calcd for C25H18
BrNO6: C, 59.07; H, 3.57. Found: C, 58.69; H, 3.72.
-
2-Br om o-3-[2-(ter t-b u t oxyca r b on yl)et h yla m in o]-9,10-
dih ydr o-9,10-[1,2]ben zen oan th r acen e-1,4,5,8-tetr on e (12).
To a mixture of 12.0 mg (0.496 mmol) of oil-free NaH in 2 mL
of DMF under argon at 0 °C was added 90.0 mg (0.496 mmol)
of â-alanine tert-butyl ester hydrochloride. The solution was

2912 J . Org. Chem., Vol. 67, No. 9, 2002
Hua et al.
stirred at 0 °C for 30 min and cooled to -46 °C, and a solution
of 201 mg (0.496 mmol) of 1 in 1 mL of DMF was added via
cannula. The solution was stirred at -46 °C for 8 h, diluted
with a mixture of ethyl acetate and benzene (1:1), washed twice
with water and brine, dried (Na2SO4), concentrated, and
column chromatographed on silica gel using a mixture of
hexane and ethyl acetate (10:1) as solvent to give 165 mg (62%
yield) of 12: mp 171-173 °C; MS m/z 538, 536 (∼1:1; M + 1),
482, 480 (1:1); 1H NMR δ 7.45-7.42 (m, 2 H, Ar-H), 7.08-
7.04 (m, 2 H, Ar-H), 6.64 (s, 2 H, C6,7 H), 6.27 (s, 1 H), 6.22
(bs, 1 H, NH), 6.10 (s, 1 H), 4.00 (q, J ) 6.7 Hz, 2 H, CH2N),
2.55 (t, J ) 6.7 Hz, 2 H, CH2CO), 1.44 (s, 9 H, CH3); 13C NMR
δ 182.3, 182.0, 177.0, 173.5, 170.6, 154.1, 151.4, 151.37, 147.0,
144.2, 142.1, 141.9, 135.4, 135.2, 128.8, 125.8, 125.76, 125.5,
125.0, 81.5, 43.5, 41.9, 40.5, 36.2, 28.0 (3 C). Anal. Calcd for
percentage of the net absorbance of MTS/formazan after
bioreduction by vehicle-treated control tumor cells at day 4
(A490nm ) 1.837 ( 0.125, 100 ( 7%). Blank values (A490nm
)
0.325 at day 4) for cell-free culture medium supplemented with
MTS/PMS reagent were subtracted from the results. IC50
values were calculated from linear regression of the slopes of
the log-transformed concentration-survival curves.
An tim a la r ia l Activity Assa y. The reported protocol15a was
followed.
P la sm ep sin Assa y. The substrate used for the plasmepsin
assay (BACHEM) is a synthetic peptide (Dabcyl-Glu-Arg-Nle-
Phe-Leu-Ser-Phe-Pro-Edans) designed to mimic the cleavage
site present in hemoglobin. The kinetic constants for the
substrate are Km ) 0.78 s-1 and Kcat. ) 0.10 µM for P.
falciparum plasmepsin and Km ) 0.69 s-1 and Kcat. ) 0.16 µM
for P. vivax plasmepsin. The substrate is conjugated with the
fluorescent donor EDANS and the quencher DABCYL.21
Fluorescence is only detectable when the EDANS group is
separated from the DABCYL group by cleavage of the sub-
strate.22 Compounds were manually added to 96-well plates
followed by the addition of assay buffer (15 mM NaCl, 100 mM
formate, pH 4.4) using an automated dilutor. After thorough
mixing and dilution, the contents of the plates were transferred
to test plates, and plasmepsin enzyme solution was added with
the dilutor. After a 10-min incubation at 37 °C, background
fluorescence was measured with a fluorescence plate reader.
Finally, the substrate was added (final concentration of 10 µM),
and the reaction mixture was incubated for 30 min at 37 °C
followed by fluorescence detection. Each compound in this
prescreen was tested in triplicate at the concentration of 10
µg/mL. Compounds that reduced the activity of plasmepsin by
50% or more at this concentration were selected for a second
screen to determine IC50 values.
C
27H22BrNO6: C, 60.46; H, 4.13. Found: C, 60.75; H, 4.39.
N-(2-Br om o-9,10-d ih yd r o-1,4,5,8-tetr a oxo-9,10-[1,2]ben -
zen oa n th r a cen e-3-yl)-3-a m in op r op a n oic Acid (13). A so-
lution of 0.100 g (0.187 mmol) of 12 and 0.2 mL (2.60 mmol)
of trifluoroacetic acid in 5 mL of dichloromethane was stirred
at 0 °C for 1 h and then at 25 °C for 2 h. The solution was
concentrated to dryness and crystallized from benzene to give
0.090 g (100% yield) of purple solid: mp 149-151 °C; MS m/z
1
482, 480 (∼1:1; M + 1), 422, 420 (1:1); H NMR δ 7.49-7.46
(m, 2 H), 7.08-7.06 (m, 2 H), 6.64 (s, 2 H), 6.27 (s, 1 H), 6.22
(bs, 1 H, NH), 6.10 (s, 1 H), 4.04 (q, J ) 6 Hz, 2 H, CH2N),
3.40 (bs, 1 H, OH), 2.73 (t, J ) 6 Hz, 2 H, CH2); 13C NMR
(CD3OD) δ 182.8, 182.77, 177.2, 174.2, 174.1, 151.6, 151.4,
147.6, 145.6, 142.9, 142.6, 137.9, 135.4, 129.0, 128.9, 128.2,
128.18, 126.1, 125.2, 43.8, 42.4, 42.3, 34.8; HRMS m/z 483.0304
(483.0317, calcd for C23H18BrNO6, M - H+; bisquinone rings
were reduced under the mass spectrum measuring conditions).
Anal. Calcd for C23H14BrNO6: C, 57.52; H, 2.94. Found: C,
56.72; H, 2.88.
Cell Via bility Assa y.5 L1210 lymphocytic leukemia cells
were seeded in triplicate at an initial density of 9375 cells/mL
of RPMI 1640 medium supplemented with 8.25% fortified
bovine calf serum and penicillin (100 IU/mL)-streptomycin
(100 µg/mL) and incubated at 37 °C in 48 well Costar cell
culture plates for 4 d in the presence or absence (control) of
increasing concentrations of the triptycene bisquinones. Dauno-
mycin was tested under similar conditions for comparison. The
viability of drug-treated cells was assessed from their ability
to bioreduce the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymeth-
oxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) reagent in
the presence of phenazine methosulfate (PMS) into a water-
soluble formazan product which absorbs at 490 nm.20 After 4
d in culture, cell samples (about 106/0.5 mL/well for controls)
were further incubated at 37 °C for 3 h in the dark in the
presence of 0.1 mL of MTS/PMS (2:0.1) reagent, and their
relative cell viability was estimated by recording the absor-
bance at 490 nm, using an automatic microplate reader.5 Cell
viability results (means ( SD, n ) 3) were expressed as the
Ack n ow led gm en t. We gratefully acknowledge fi-
nancial support from the National Institutes of Health
(National Cancer Institute, CA86842 and Center of
Biomedical Research Excellence, No. 1P20RR15563),
National Science Foundation (No. CHE-0078921), Ameri-
can Heart Association, Heartland Affiliate (Nos.
0051658Z, 9951177Z), NASA-BioServe Space Technolo-
gies (No. NAGW-1197), Great Plains Diabetes Research,
Inc, Howard Hughes Medical Institute (Biological Sci-
ences Education Grant), and Kansas State University
(Center for Basic Cancer Research).
Su p p or tin g In for m a tion Ava ila ble: X-ray details for
compound 12. This material is available free of charge via the
Internet at http://pubs.acs.org.
J O010958S
(21) Matayoshi, E. D.; Wang, G. T.; Krafft, G. A.; Erickson, J . Science
1990, 247, 954.
(20) Cory, A. H.; Owen, T. C.; Barltrop, J . A.; Cory, J . G. Cancer
Commun. 1991, 3, 207.
(22) Luker, K. E.; Francis, S. E.; Gluzman, I. Y.; Goldberg, D. E.
Mol. Biochem. Parasitol. 1996, 79, 71.