Antitumor agents. 166. Synthesis and biological evaluation of 5,6,7,8-substituted-2-phenylthiochromen-4-ones

Article abstract of DOI:10.1021/jm960008c

As a continuation of our structure-activity relationship study of substituted 2-phenyl-4-quinolones and flavonoids as antitumor and antiviral agents, a series of 5,6,7,8-substituted-2-phenylthiochromen-4-ones has been synthesized by condensation of substi

Full text of DOI:10.1021/jm960008c

J . Med. Chem. 1996, 39, 1975-1980
1975
An titu m or Agen ts. 166. Syn th esis a n d Biologica l Eva lu a tion of
5,6,7,8-Su bstitu ted -2-p h en ylth ioch r om en -4-on es†
Hui-Kang Wang,‡ Kenneth F. Bastow,‡ L. Mark Cosentino,§ and Kuo-Hsiung Lee*,‡
Division of Medicinal Chemistry and Natural Products, School of Pharmacy, University of North Carolina,
Chapel Hill, North Carolina 27599, and Biotech Research Laboratories, Rockville, Maryland 20850
Received J anuary 2, 1996X
As a continuation of our structure-activity relationship study of substituted 2-phenyl-4-
quinolones and flavonoids as antitumor and antiviral agents, a series of 5,6,7,8-substituted-
2-phenylthiochromen-4-ones has been synthesized by condensation of substituted thiophenols
and ethyl benzoylacetates. Target compounds were evaluated for biological activity. Among
them, compounds 7, 10, 12, and 13 displayed significant growth inhibitory action against a
panel of tumor cell lines including human ileocecal carcinoma (HCT-8), murine leukemia (P-
388), human melanoma (RPMI), and human central nervous system tumor (TE671) cells.
Compounds 10, 12, and 19 displayed DNA topoisomerase I inhibitory activity in vitro and
compound 11 was an in vitro, inhibitor of DNA topoisomerase II. Compound 11 was most
active (ED50 value, 0.65 µM) against HIV in acutely infected H9 lymphocytes and had a
therapeutic index of about 5.
As part of our continuing search for potential anti-
cancer drug candidates in the 2-phenyl-4-quinolone
series, we have synthesized a series of substituted
2-phenyl-4-quinolones and related compounds and evalu-
ated them as cytotoxic compounds and as antimitotic
agents interacting with tubulin. Among them, some
compounds (Q1 and Q2, Figure 1) were potent inhibitors
of tubulin polymerization with activities comparable to
those of the antimitotic natural products colchicine,
podophyllotoxin, and combretastatin A-4.2-4 For ex-
ample, compound Q2 totally inhibited the growth of
about half of the NCI tumor cell lines at subnanomolar
concentrations (log TGI < -9.00) and was also a potent
inhibitor of tubulin polymerization with an IC50 value
of 0.44 µM.
In our earlier studies, certain flavonoids (Figure 1)
displayed antitumor activity or anti-HIV activity. For
F igu r e 1. Bioactive 2-phenyl-4-quinolones and flavonoids.
example, two flavonols isolated from Polanisia dode-
candra [5,3′-dihydroxy-3,6,7,8,4′-pentamethoxyflavone
(F 1) and 5,4′-dihydroxy-3,6,7,8,3′-pentamethoxyflavone
(F 2)] displayed interesting antitumor activities.7 [Com-
pound F 1 was also previous isolated as a cytotoxic
principle, centaureidin, from Polymnia fruticosa by
Beutler et al.5 and as an antimitotic agent from Zie-
ridium pseudobtusifolium by Lichius et al.6] Compound
F 1 was cytotoxic in vitro against a panel of cell lines
derived from central nervous system cancers, nonsmall
cell lung cancers, small cell lung cancers, ovarian
cancers, colon cancers, renal cancers, a melanoma, and
leukemia cells, with GI50 values in the low micromolar
to nanomolar concentration range. Compound F 1 also
inhibited tubulin polymerization (IC50 ) 0.83 ( 0.2 µM)
and the binding of radiolabeled colchicine to tubulin
(59% inhibition when present at equimolar concentra-
tion), and appears to be the first example of a flavonol
displaying such bioactivity. Compound F 2 was also
cytotoxic for medulloblastoma tumor cells with an ED50
value of 0.99 µg/mL. On the other hand, apigenin-7-
O-â-D-glucopyranoside (F 3) isolated from Kummerowia
striata8 and chrysin (F 4) isolated from Chrysanthemum
morifolium9 as anti-HIV active principles displayed
EC50 values of 1.8 µg/mL and 5 µM against H9 cell
replication with therapeutic index value of >55.6 and
9, respectively.
Comparison of the structures between the bioactive
2-phenyl-4-quinolones and flavones discussed above
revealed that these two types of compound share similar
skeletons except they possess a different heteroatom at
position 1 of the C ring. Therefore, similar or possible
novel biological activities might be anticipated following
the substitution of the heteroatom by a bioisostere such
as sulfur.
The aim of the present work was to further investi-
gate the structure-activity relationships (SAR) of 2-phen-
yl-4-quinolones and flavonones by replacing the het-
eroatom in the C ring with a sulfur atom. Consequently,
a series of 5,6,7,8-substituted-2-phenylthiochromen-4-
ones and related 2-phenyl-4-quinolones have been syn-
thesized and evaluated for cytotoxic, topoisomerase I
and II inhibitory, and anti-HIV activities.
* To whom correspondence should be addressed.
† For Part 165, see ref 1.
‡ University of North Carolina.
§ Biotech Research Laboratories.
X Abstract published in Advance ACS Abstracts, April 15, 1996.
S0022-2623(96)00008-8 CCC: $12.00 © 1996 American Chemical Society

1976 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 10
Wang et al.
Sch em e 1. Synthesis of 2-Phenylthiochromen-4-one Derivatives 1, 2, 7, 8, 10, 11, and 13
Sch em e 2. Synthesis of 2-Phenylthiochromen-4-one Derivatives 3-6 and 9
Sch em e 3. Synthesis of 2-Phenylthiochromen-4-one Derivatives 14-19
Ch em istr y
shown in Schemes 1-3. Compounds 1-4, 8, 13, and
14-19 were synthesized by the condensation of corre-
sponding substituted thiophenols (compounds A1-A7)
and ethylbenzoyl acetates (B1-B2) in heated (90-100
°C) polyphosphoric acid (PPA) for 2 h.10 Compounds
Compounds 1-19 were synthesized by condensation
of commercially available substituted thiophenols (com-
pounds A1-A7) and ethyl benzoylacetates (B1-B2)
followed by demethylation, acetylation, or oxidation as

Antitumor Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 10 1977
Ta ble 1. In Vitro Cytotoxic Activities of Substituted 2-Phenylthiochromen-4-ones in Various Tumor Cell
ED50 (µg /mL)a
compd no.
R5
R6
R7
R8
R3′
KBb
A-549b
HCT-8b
P-388b
RPMIb
TE671b
1
2
3
4
5
6
7
8
9
10
11
12
13
14
19
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
OCH3
-
-
-
-
-
OCH3
OH
5.5
-
9.16
-
-
-
-
OCH3
OH
-
5.5
-
-
-
-
-
-
-
-
-
-
-
OH
OCH3
-
-
-
0.55
8.89
-
2.87
-
OCH3
OAc
OH
4.23
-
9.33
-
-
-
-
OH
OAc
5.5
5.5
5.5
5.5
-
1.45
-
4.51
4.83
1.18
0.61
7.46
9.31
3.92
5.5
5.5
5.5
5.5
-
5.50
4.58
3.66
-
OAc
5.79
6.35
-
OCH3
OCH3
OCH3
NH2
-
-
NH2
-
-
a
EC50 was the concentration of drug which affords 50% reduction in cell number after a 3-day incubation. For significant activity of
b
the pure compound, an EC50 e 4.0 µg/mL is required. Human epidermoid carcinoma of the nasopharynx (KB), human lung carcinoma
(A-549), human ileocecal carcinoma (HCT-8), murine leukimia (P-388), human melanoma (RPMI), and human CNS tumor (TE671). c “-”
means inactive.
Ta ble 2. Human DNA Topoisomerase I and II Inhibitory Activities of Substituted 2-Phenylthiochromen-4-ones
% inhibition (100 µM)
cellular protein-linked
compd no.
R5
R6
R7
topoisomerase Ia
topoisomerase IIb
DNA breaks (-fold)d
Q2e
10
11
12
19
ND
100
0
100
100
0
0
100
0
ND
1
1
1
1
OH
OAc
OH
OAc
NH2
0
a
Measured as ATP-independent relaxation of supercoiled plasmid DNA compared to enzyme and DNA control reactions. Camptothecin
b
at 100 µm served as the positive inhibitor control. Measured as ATP-dependent unknotting of P4 DNA compared to enzyme and DNA
control reactions. VP-16 (100 µm) completely inhibited the unknotting activity. c Compounds (6, 9, 13-17) were not active at 100 µm.
d
Measured as potassium-SDS precipitable cpm in KB oral carcinoma cell cultures treated in triplicate at 50 µm for 1 h. VP-16 and
camptothecin stimulated levels of protein-DNA complexes by (36 ( 5)- and (21 ( 2)-fold respectively. e See Figure 1; ND ) not tested.
5-7 and 10 were obtained by boron tribromide dem-
ethylation of 2-4 and 8. Compounds 9 and 11 were
generated by acetic anhydride-pyridine acetylation of
5 and 10 at room temperature. Peroxidation of com-
pound 1 afforded compound 12 (Scheme 1). Using
3-methoxythiophenol (A4), both compounds 3 and 4
were generated due to condensation at either the 2- or
the 6-position of the original thiophenol (Scheme 2).
With the aminothiophenols A5-A7, the thiochromen-4-
ones 14, 16, and 18 were accompanied by the corre-
sponding 2-phenyl-4-quinolones 15, 17, and 19, respec-
tively (Scheme 3).
linked DNA breaks, for cytotoxicity in cancer cells, and
for HIV-inhibitory effects were carried out according to
the procedures described previously.11-17
Table 1 shows the in vitro cytotoxic activities of 15
target compounds. Among these compounds, 1, 2, 5, 6,
and 9 were inactive and 3, 4, 8, 11, 14, and 19 were
only weak cytotoxic agents. However, compounds 7, 10,
12, and 13 displayed significant activity (ED50 < 4.0 µg/
mL) against several tumor human cell lines including
ileocecal carcinoma (HCT-8), murine leukemia (P-388),
human melanoma (RPMI), and a central nervous system
(CNS) tumor (TE671), respectively.
Table 2 shows the activity of target compounds
against DNA topoisomerase I and II in vitro. Due to
the insolubilities at the testing concentration, results
for compounds 1-5, 7, 8, and 18 were not obtained.
Resu lts a n d Discu ssion
Assays for the inhibition of mammalian DNA topo-
isomerase I and II, for production of cellular protein-

1978 J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 10
Wang et al.
Ta ble 3. HIV-Inhibitory Effects of Substituted
2-Phenylthiochromen-4-ones on Infected H9 Lymphocytes
isomerase inhibitory effects observed in vitro were
unexpected and the involvement of these enzymes as
biochemical targets for 10, 11, 12, and 19 are currently
being evaluated.
Exp er im en ta l Section
Melting points were determined on a Fisher-J ohn melting
point apparatus and are uncorrected. Elemental analyses
were performed by Atlantic Microlabs, Atlantic, GA; 1H NMR
spectra were measured at 300 MHz on a Bruker 300 spec-
trometer and were recorded in CDCl3, a mixture of CDCl3 and
CD3OD, or DMSO-d6. Chemical shifts are reported in δ (ppm)
units relative to the internal reference Me4Si. Infrared (IR)
spectra were recorded on a Perkin-Elmer IR 400 spectrometer
as KBr pellets.
Gen er a l P r oced u r e for th e Syn th esis of Com p ou n d s
1-4, 8, a n d 13-19. Thirteen grams of phosphorus pentoxide
were added to 7 mL of concentrated phosphoric acid; the
polyphosphoric acid (PPA) generated was heated to 90-100
°C, and the appropriate substituted thiophenol (compounds
A1-A7, 0.01 mol) was added. The ethyl benzoylacetate (B1-
B2, 0.01 mol) was added dropwise over 1.5 h at 90-100 °C,
and the resulting mixture was further stirred for 30 min. After
cooling, water was added and the precipitate was collected.
The crude products were further purified by recrystallization
or by silica gel column chromatographic separation (CHCl3:
CH3OH ) 100:1).
Gen er a l P r oced u r e for th e Syn th esis of Com p ou n d s
5-7 a n d 10. Compound 8 (298 mg) was dissolved in 30 mL of
CH2Cl2, and 2.0 mL of BBr3 (1.0 M in hexane) was added with
ice-cooling. The mixture was further stirred at room temper-
ature for 30 min, and then 5 mL of CH3OH was added slowly
with ice-cooling to quench the reaction. After evaporation, the
residue was washed with water and the precipitate was
collected (95% yield). The product was recrystallized from
CHCl3 and CH3OH to afford pure compound 10. The same
procedure was used for the synthesis of compounds 5-7.
2-P h en ylth ioch r om en -4-on e (1): yield 95%; colorless
needles; mp 121-122 °C; IR (KBr) ν 1610 (shoulder, conjugated
CdO); 1H NMR (CDCl3) δ 7.27 (1H, s, H-3), 7.50-7.55 (3H,
m, H-2′,4′,6′), 7.58 (1H, dd, J ) 2.2, 8.1 Hz, H-8), 7.65 (1H, dt,
J ) 1.4, 8.1 Hz, H-6), 7.68-7.75 (3H, m, H-7,3′,5′), 8.57 (1H,
dd, J ) 1.4, 8.1 Hz, H-5). Anal. (C16H12O2S) C, H.
6-Meth oxy-2-p h en ylth ioch r om en -4-on e (2): yield 96%;
colorless needles; mp 147-148 °C; IR (KBr) ν 1610 (shoulder,
conjugated CdO); 1H NMR (CDCl3) δ 3.97 (3H, s, OCH3), 7.30
(1H, dd, J ) 2.8, 8.8 Hz, H-7), 7.42 (1H, s, H-3), 7.50-7.54
(3H, m, H-2′,4′,6′), 7.63 (1H, d, J ) 8.8 Hz, H-8), 7.71-7.75
(2H, m, H-3′,5′), 8.02 (1H, d, J ) 2.8 Hz, H-5). Anal.
(C16H12O2S) C, H.
compd
no.
IC50
ED50
therapeutic
index
R5
R6
R7
R8 (µg/mL)a (µg /mL)b
4c
6
7
OCH3
7
40
40
45
7
7
9.5
80
0.8
0.65
4
12
7
1.0
5.7
4.2
0.6
2.3
4.6
3.8
2.9
1.1
5.4
2.5
OH
OH
9
OAc
10
11
14
15
16
18
19
OH OH
OAc OAc
1.8
3
15
35
7.5
35
50
NH2
SH
NH2
SH
NH2
6.5
20
a
Concentration which inhibits uninfected growth by 50%; AZT
had an IC50 value of 2000 µM. b Concentration which inhibits virus
replication growth by 50%; AZT had an ED50 value of 0.04 µM.
c Crystals of the test agents were observed at both the 100 and 20
µg/mL concentration with compounds 1, 3, 8, 10, and 14. The cells
were dead at these concentrations. For compounds 5, 6, 7, and 9,
crystals were observed at 100, 20, and 4 µg/mL, but cells were
only dead at the 100 µg/mL concentration. For compounds 2, 11,
15, 16, 18, and 19, crystals were found at the 100 µg/mL
concentration, and the cells were dead at that concentration.
Compounds 13 and 17 did not dissolve in DMSO and could not be
tested.
Compounds 10, 12, and 19 displayed topoisomerase I
inhibitory activities, and compound 11 inhibited topo-
isomerase II activity. It is interesting that compounds
10 and 11 act selectively on different topoisomerases
in vitro. Because the inhibition of neither topoisomerase
I nor II was reported for either 2-phenyl-4-quinolones
or flavonoids, the mechanism of inhibition for 2-phen-
ylthiochromen-4-ones deserves further investigation. A
preliminary study indicated that topoisomerase-DNA
cleavable complexes were not induced in treated cells
(Table 2).
Table 3 shows the HIV-inhibitory effects of the
synthetic compounds. Due to the crystallization or
insolubility of certain target compounds at the testing
concentration, activities of 1-3, 5, 8, 12-13, and 17
could not be evaluated. Compounds 4, 6, 7, 10, 11, 14,
16, and 18 displayed selective antiviral activity (ED50
0.65-9.5 µg/mL) with therapeutic indexes in the range
of 0.6-5.7 against HIV in acutely infected H9 lympho-
cytes. Among the active compounds, 11, bearing ac-
etoxy groups on both the 6- and 7-positions, was most
active (ED50 value, 0.65 µg/mL) with a therapeutic index
of about 5.
Compared with the 2-phenyl-4-quinolones, the 2-phen-
ylthiochromen-4-ones exhibited improved solubility in
organic solvents which made them easy to purify by
crystallization, but it was more difficult to evaluate
them in biological assays. Therefore, improved solubil-
ity of these synthetic compounds and the introduction
of a wider variety of substituents will be necessary
before clear SAR conclusions can be drawn. Despite the
limitations of the current study, it is clear that hetero-
atom replacement by sulfur is tolerable because anti-
tumor or anti-HIV activities, as observed with 2-phenyl-
4-quinolones or flavonoids, were retained. The topo-
7-Meth oxy-2-p h en ylth ioch r om en -4-on e (3): yield 94%;
colorless needles; mp 135 °C; IR (KBr) ν 1610 (shoulder,
conjugated CdO); 1H NMR (CDCl3) δ 3.93 (3H, s, OCH3), 7.07
(1H, d, J ) 2.3 Hz, H-8), 7.11 (1H, dd, J ) 2.3, 8.9 Hz, H-6),
7.18 (1H, s, H-3), 7.50-7.52 (3H, m, H-2′,4′,6′), 7.67-7.70 (2H,
m, H-3′,5′), 8.48 (1H, d, J ) 8.9 Hz, H-5). Anal. (C16H12O2S)
C, H.
5-Meth oxy-2-p h en ylth ioch r om en -4-on e (4): yield 92%;
colorless needles; mp 200-201 °C; IR (KBr) ν 1610 (shoulder,
conjugated CdO); 1H NMR (CDCl3) δ 4.01 (3H, s, OCH3), 6.98
(1H, d, J ) 8.2 Hz, H-8), 7.14 (1H, s, H-3), 7.24 (1H, t, J ) 8.2
Hz, H-7), 7.50-7.52 (3H, m, H-2′,4′,6′), 7.53 (1H, d, J ) 8.2
Hz, H-6), 7.68-7.71 (2H, m, H-3′,5′). Anal. (C16H12O2S) C,
H.
5-Hyd r oxy-2-p h en ylth ioch r om en -4-on e (5): yield 97%;
yellow needles; mp 155-156 °C; IR (KBr) ν 3400 (ÃΗ), 1610
1
(shoulder, conjugated CdO); H NMR (CDCl3) δ 6.97 (1H, br
d, J ) 8.0 Hz, H-8), 7.13 (1H, br d, J ) 8.0 Hz, H-6), 7.14 (1H,
s, H-3), 7.51 (1H, t, J ) 8.0 Hz, H-7), 7.52-7.60 (3H, m,
H-2′,4′,6′), 7.68-7.73 (2H, m, H-3′,5′), 14.01 (1H, s, OH). Anal.
(C15H10O2S) C, H.
7-Hyd r oxy-2-p h en ylth ioch r om en -4-on e (6): yield 95%;
colorless needles; mp 268-269 °C dec; IR (KBr) ν 3400-3200
(OH), 1610 (shoulder, conjugated CdO); 1H NMR (CDCl3:CD3-
OD ) 5:1) δ 6.97 (1H, dd, J ) 2.2, 8.8 Hz, H-6), 7.00 (1H, d, J

Antitumor Agents
J ournal of Medicinal Chemistry, 1996, Vol. 39, No. 10 1979
) 2.2 Hz, H-8), 7.12 (1H, s, H-3), 7.38-7.44 (3H, m, H-2′,4′,6′),
7.57-7.61 (2H, m, H-3′,5′), 8.28 (1H, d, J ) 8.8 Hz, H-5). Anal.
(C15H10O2S) C, H.
7.55-7.57 (3H, m, H-2′,4′,6′), 7.61 (1H, br d, J ) 8.0 Hz, H-7),
7.66 (1H, d, J ) 8.0 Hz, H-8), 7.75-7.77 (2H, m, H-3′,5′), 8.22
(1H, br s, H-5). Anal. (C15H11NOS) C, H.
6-Hyd r oxy-2-p h en ylth ioch r om en -4-on e (7): yield 96%;
yellowish needles; mp 288-289 °C; IR (KBr) ν 3400-3200
(OH), 1610 (shoulder, conjugated CdO); 1H NMR (CDCl3:CD3-
OD ) 5:1) δ 7.05 (1H, s, H-3), 7.08 (1H, dd, J ) 2.8, 8.8 Hz,
H-7), 7.31-7.36 (3H, m, H-2′,4′,6′), 7.45 (1H, d, J ) 8.8 Hz,
H-8), 7.51-7.54 (2H, m, H-3′,5′), 7.68 (1H, d, J ) 2.8 Hz, H-5).
Anal. (C15H10O2S) C, H.
6,7-Dim eth oxy-2-p h en ylth ioch r om en -4-on e (8): yield
92%; colorless needles; mp 171-172 °C; IR (KBr) ν 3400-3200
(OH), 1610 (shoulder, conjugated CdO); 1H NMR (CDCl3) δ
4.03, 4.04 (3H each, both s, OCH3 × 2), 7.04 (1H, s, H-8), 7.24
(1H, s, H-3), 7.51-7.53 (3H, m, H-2′,4′,6′), 7.68-7.71 (2H, m,
H-3′,5′), 7.99 (1H, s, H-5). Anal. (C17H14O3S) C, H.
7-Acetyl-2-p h en ylth ioch r om en -4-on e (9): yield 99%; col-
orless needles; mp 228-229 °C; IR (KBr) ν 1750 (acetyl CdO),
1610 (shoulder, conjugated CdO); 1H NMR (CDCl3) δ 2.37 (3H,
s, OAc), 7.37 (1H, s, H-3), 7.46 (1H, dd, J ) 2.5, 8.8 Hz, H-6),
7.51-7.56 (3H, m, H-2′,4′,6′), 7.71-7.73 (2H, m, H-3′,5′), 7.73
(1H, d, J ) 8.8 Hz, H-5), 8.28 (1H, d, J ) 2.5 Hz, H-8). Anal.
(C17H12O3S) C, H.
5-Mer ca p to-2-p h en yl-4-qu in olon e (18): yield 49%; orange
crystals; mp 141-142 °C; IR (KBr) ν 1610 (shoulder, conju-
gated CdO); 1H NMR (CDCl3) δ 7.12 (1H, s, H-3), 7.15 (1H, d,
J ) 8.0 Hz, H-6), 7.18 (1H, d, J ) 8.0 Hz, H-8), 7.45 (1H, t, J
) 8.0 Hz, H-7), 7.52-7.55 (3H, m, H-2′,4′,6′), 7.67-7.70 (2H,
m, H-3′,5′). Anal. (C15H11NOS) C, H, N.
5-Am in o-2-p h en ylth ioch r om en -4-on e (19): yield 21%;
fine crystals; mp >300 °C dec; IR (KBr) ν 3350 and 3250 (NH2),
1610 (shoulder, conjugated CdO); 1H NMR (DMSO-d6) δ 6.40
(1H, s, H-3), 7.41 (1H, d, J ) 7.5 Hz, H-6), 7.51 (1H, t, J ) 7.5
Hz, H-7), 7.56 (1H, d, J ) 7.5 Hz, H-8), 7.60-7.61 (3H, m,
H-2′,4′,6′), 7.83-7.86 (2H, m, H-3′,5′). Anal. (C15H11NOS) C,
H, N.
Biologica l Assa ys. The in vitro cytotoxicity assay was
carried out according to procedures described in Geran et al.11
and Ferguson et al.12 The assay against KB (nasal pharyngeal
carcinoma), A-549 (human lung cancer), HCT-8 (human colon
carcinoma), PRMI-7951 (human melanoma), TE-671 (human
medulloblastoma), and P-388 (murine leukemia) tumor cells
was based on a method reported in Lee et al.13
Assays for the in vitro inhibition of DNA topoisomerases I
and II and for production of cellular protein-linked DNA breaks
were carried out according to the procedures described previ-
ously.14,15
6,7-Dih yd r oxy-2-p h en ylth ioch r om en -4-on e (10): yield
91%; yellow needles; mp 259-261 °C dec; IR (KBr) ν 3450-
3200 (broad, ÃΗ), 1610 (shoulder, conjugated CdO); 1H NMR
(CDCl3 + CD3OD ) 5 + 1) δ 7.21 (1H, s, H-3), 7.41-7.46 (3H,
m, H-2′,4′,6′), 7.44 (1H, s, H-8), 7.59-7.62 (2H, m, H-3′,5′), 7.79
(1H, s, H-5). Anal. (C15H10O3S) C, H.
HIV inhibition was measured as described previously.16,17
The H9 T cell line was maintained in continuous culture with
complete medium (RPMI 1640 and 10% fetal calf serum) at
5% CO2 and 37 °C and was used in experiments only when in
log phase of growth. The cells were incubated with HIV-1 (IIIB
isolate, TCID50 104 IU/mL, at a multiplicity of infection of 0.1-
0.01 IU/cell) for 1 h at 37 °C and 5% CO2. The cells then were
washed thoroughly to remove unabsorbed virus and resus-
pended at 4 × 105 cells/mL in complete medium. Aliquots (1
mL) were placed in wells of 24-well culture plates containing
an equal volume of test compound (diluted in the culture
medium). After a 4-day incubation at 37 °C, cell density of
uninfected cultures was determined by counting cells in a
Coulter counter to assess toxicity of the test compound. A p24
antigen ELISA assay was used to determine the level of virus
released in the medium of the HIV-infected cultures. The p24
antigen assay uses an HIV-1 anti-p24 specific monoclonal
antibody as the capture antibody coated on 96-well plates.
Following a sample incubation period, rabbit serum containing
antibodies for HIV-1 p24 is used to tag any p24 “captured”
onto the microtiter well surface. Peroxidase conjugated goat
anti-rabbit serum is then used to tag HIV-1 p24 specific rabbit
antibodies that have complexed with captured p24. The
presence of p24 in test samples is then revealed by addition
of substrate. The cutoff for the p24 ELISA assay is 12.5 pg/
mL. P24 in the culture medium was quantitated against a
standard curve containing known amounts of p24. The
effective (EC50) and inhibitory (IC50) concentrations (for anti-
HIV activity and cytotoxicity, respectively) were determined.
6,7-Diacetyl-2-ph en ylth ioch r om en -4-on e (11): yield 98%;
colorless needles; mp 171-172 °C; IR (KBr) ν 1755 (acetyl
1
CdÃ), 1610 (shoulder, conjugated CdO); H NMR (CDCl3) δ
2.36 (6H, s, OAc × 2), 7.26 (1H, s, H-3), 7.50-7.55 (3H, m,
H-2′,4′,6′), 7.61 (1H, s, H-8), 7.68-7.70 (2H, m, H-3′,5′), 8.36
(1H, s, H-5). Anal. (C19H14O5S) C, H.
2-P h en ylth ioch r om en -4-on e 1-oxid e (12): yield 90%.
Compound 1 (100 mg) was dissolved in 5 mL of CH2Cl2, and
m-chloroperbenzoic acid (300 mg) was added. The mixture was
further stirred at room temperature for 12 h. The reaction
solution was introduced directly to a silica gel column (eluent
of CHCl3:MeOH ) 100:1) to afford compound 12 (99 mg, yield
93%): colorless needles; mp 126-127 °C; IR (KBr) ν 1645
1
(conjugated CdO), 1145 (SdO); H NMR (CDCl3) δ 6.85 (1H,
s, H-3), 7.51-7.62 (3H, m, H-2′,4′,6′), 7.79 (1H, t, J ) 8.0 Hz,
H-6), 7.87-7.93 (2H, m, H-3′,5′), 7.90 (1H, d, J ) 8.0 Hz, H-7),
8.13 (1H, d, J ) 8.0 Hz, H-8), 8.24 (1H, d, J ) 8.0 Hz, H-5).
Anal. (C15H10O2S) C, H.
6,7,3′-Tr im eth oxy-2-ph en ylth ioch r om en -4-on e (13): yield
89%; colorless needles; mp 189-190 °C; IR (KBr) ν 1610
(shoulder, conjugated CdO); 1H NMR (CDCl3) δ 3.89, 4.02, 4.04
(3H each, all s, OCH3 × 3), 7.04 (1H, s, H-3), 7.05 (1H, dd, J
) 2.0, 8.0 Hz, H-4′), 7.21 (1H, t, J ) 2.0 Hz, H-2′), 7.25 (1H, s,
H-8), 7.28 (1H, dd, J ) 2.0, 8.0 Hz, H-6′), 7.42 (1H, t, J ) 8.0
Hz, H-5′), 7.98 (1H, s, H-5). Anal. (C18H16O4S) C, H.
8-Am in o-2-p h en ylth ioch r om en -4-on e (14): yield 55%;
colorless needles; mp 254 °C; IR (KBr) ν 3350 and 3250 (NH2),
1610 (shoulder, conjugated CdO); 1H NMR (CDCl3) δ 6.71 (1H,
s, H-3), 7.18 (1H, dt, J ) 1.5, 7.0 Hz, H-6), 7.48 (1H, dd, J )
1.5, 7.0 Hz, H-7), 7.50-7.62 (6H, m, H-5,2′-6′′), 12.15 (1H, br
s, NH). Anal. (C15H11NOS) C, H, N.
8-Mer ca p to-2-p h en yl-4-qu in olon e (15): yield 26%; color-
less needles; mp 105-106 °C; IR (KBr) ν 1610 (shoulder,
conjugated CdO); 1H NMR (CDCl3) δ 7.41 (1H, t, J ) 8.0 Hz,
H-6), 7.52 (1H, s, H-3), 7.50-7.55 (3H, m, H-2′,4′,6′), 7.93 (1H,
d, J ) 8.0 Hz, H-7), 8.13 (1H, d, J ) 8.0 Hz, H-5), 8.11-8.16
(2H, m, H-3′,5′). Anal. (C15H11NOS) C, H, N.
Ack n ow led gm en t . This investigation was sup-
ported by Grant CA 17625 from the National Cancer
Institute awarded to K. H. Lee.
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6-Am in o-2-p h en ylth ioch r om en -4-on e (16): yield 35%;
colorless needles; mp 252-253 °C; IR (KBr) ν 3350 and 3250
1
(NH2), 1610 (shoulder, conjugated CdO); H NMR (CDCl3) δ
6.80 (1H, s, H-3), 7.48 (1H, br d, J ) 7.0 Hz, H-7), 7.49 (1H, d,
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