Antitumor agents. Part 226: Synthesis and cytotoxicity of 2-phenyl-4-quinolone acetic acids and their esters

Article abstract of DOI:10.1016/S0960-894X(03)00624-3

2-Phenyl-4-quinolone acetic acids and their esters were synthesized and evaluated for interaction with tubulin and for cytotoxicity against a panel of human tumor cell lines. 2-Phenyl- and 2-(2′-fluorophenyl)-4-quinolone-8- acetic acids (11 and 12) displa

Full text of DOI:10.1016/S0960-894X(03)00624-3

Bioorganic & Medicinal Chemistry Letters 13 (2003) 2891–2893
Antitumor Agents. Part 226:y Synthesis and Cytotoxicity of
2-Phenyl-4-quinolone Acetic Acids and Their Esters
Yi Xia,a Zheng-Yu Yang,a Peng Xia,a Kenneth F. Bastow,a Yuka Nakanishi,a
Priya Nampoothiri,b Ernest Hamel,b Arnold Brossia and Kuo-Hsiung Leea,*
aNatural Products Laboratory, School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA
bScreening Technologies Branch, Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis,
National Cancer Institute at Frederick, National Institutes of Health, Frederick, MD 21702, USA
Received 5 March 2003; accepted 15 May 2003
Abstract—2-Phenyl-4-quinolone acetic acids and their esters were synthesized and evaluated for interaction with tubulin and for
cytotoxicity against a panel of human tumor cell lines. 2-Phenyl- and 2-(20-fluorophenyl)-4-quinolone-8-acetic acids (11 and 12)
displayed potent cytotoxicity with ED50 values at nanomolar concentrations, but had minimal activity against tubulin polymeri-
zation. 2-(20-Fluorophenyl)-4-quinolone-6-acetic acid (3) and 2-(20-fluorophenyl)-4-quinolone-8-acetic acid methyl ester (10) mod-
erately inhibited tubulin polymerization.
# 2003 Elsevier Ltd. All rights reserved.
Flavone acetic acid (FAA, Fig. 1) is a synthetic flavone
with a unique pattern of antitumor activity. Unlike
conventional antitumor agents, it causes rapid tumor
necrosis with little resultant toxicity in normal tissues.
FAA has demonstrated excellent activity against murine
colon adenocarcinoma 38 and a broad spectrum of
slow-growing solid tumors that are usually insensitive to
most cytotoxic drugs.2,3 In contrast to its solid tumor
activity, FAA shows poor activity against murine leu-
kemia cell lines (P388 and L1210). Because of its unique
pre-clinical solid tumor activity, FAA has been eval-
uated in clinical trials.4 FAA’s precise mechanism of
anticancer action in experimental animals is poorly
understood,5,6 but undoubtedly is novel. Modification
of FAA is continuing7 as the unusual profile of FAA
and its unclear mode of action make the development of
structural analogues of great interest.
values in the low micromolar to nanomolar concen-
tration range. In general, a good correlation was found
between cytotoxicity and inhibition of tubulin poly-
merization.8ꢀ11 Thirteen compounds in this series have
been selected for in vivo xenograft testing, and to date,
compound 1 (NSC 656158) is active in vivo. In the
xenograft ovarian OVCAR-3 model, treated mice demon-
strated a 130% increase in life span.
In our continuing studies, we designed, synthesized and
evaluated a series of 2-phenyl-4-quinolone acetic acids
with an acetic acid side chain at different positions of
the Aring. These compounds combine the skeleton of
the 2-phenyl-4-quinolones with the acetic acid side chain
of FAA. 2-Phenyl-4-quinolone-8-acetic acid also is a
bioisostere of FAA, in which NH replaces the O in the
pyrone ring.
In our previous studies, numerous substituted 2-phenyl-
4-quinolones, which are amino analogues of flavonoids
(NH replacing O), were synthesized and evaluated for
antimitotic and antitumor activities. Most compounds
in this series showed promising in vitro activity in the
NCI’s human tumor cell lines (HTCL) assay with GI50
Scheme 1 shows the preparation of 2-phenyl-4-quino-
lone 5-, 6-, and 7-acetic acids. The 6-acetic acid (3) was
*Corresponding author. Tel.: +1-919-962-0066; fax: +1-919-966-
3893; e-mail: khlee@unc.edu
yFor Part 225, see ref 1.
Figure 1.
0960-894X/03/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00624-3

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Y. Xia et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2891–2893
made and condensed immediately with ethyl benzoyl-
acetates 7 and 8, to give esters 915 and 10, which then
were hydrolyzed with aq NaOH to the corresponding
acids 11 and 12.
Compounds 2–4 and 9–12 were evaluated in a tubulin
polymerization assay. As shown in Table 1,16,17 5- and
8-acetic acid substituted compounds were essentially
inactive (2, 11, and 12) or showed little activity (9) in
this assay. However, compound 10, which is the methyl
ester of analogue 12, had moderate activity. In addition,
the 6-substituted compound (3) also had moderate
activity similar to that of compound 10 and the 7-acetic
acid (4) had modest activity. Compounds 2–4 and 9–10
displayed moderate cytotoxicity in vitro against seven
human tumor cell lines, including epidermoid, bone,
ovarian, glioblastoma, melanoma, lung, and breast
cancer cell lines. Among these five compounds, 10, one
of the two most potent compounds in the tubulin assay,
was more active than the 5-, 6-, or 7-acetic acid quino-
lones. In addition, 10 showed selective activity against
ovarian cancers. Interestingly, 2-phenyl-4-quinolone-8-
acetic acid (11) and 2-(20-fluorophenyl)-4-quinolone-8-
acetic acid (12), which were inactive in the tubulin assay,
were extremely potent in the cytotoxicity assay. The
ED50 values of these two compounds were less than
0.1 mg/mL against most tested cell lines. Atarget other
than tubulin is probably involved as the cause of the
activities of 11 and 12.
Scheme 1.
Scheme 2.
efficiently prepared by condensation of commercially
available 4-aminophenylacetic acid and ethyl 20-fluoro-
benzoylacetate when heated (90–100 ꢁC) in polyphos-
phoric acid (PPA) for 2.5 h. The 5-acetic acid (2) and
the 7-acetic acid (4) were obtained by condensation of
3-aminophenyl acetic acid and ethyl 2-fluorobenzoyl-
acetate in PPA; the two isomers were separated by
column chromatography.14
In summary, these findings with the 2-phenyl-4-quino-
lone acetic acid derivatives13 showed unique and
interesting results. 2-Phenyl-4-quinolone-8-acetic acid
and 2-(20-fluorophenyl)-4-quinolone-8-acetic acid are
the most structurally similar to FAA and were the
only highly active antitumor compounds in this
series. Unlike most 2-phenyl-4-quinolones, they were
not inhibitors of tubulin polymerization. 2-(20-Fluoro-
phenyl)-4-quinolone-6-acetic acid (3) and 2-(20-fluoro-
phenyl)-4-quinolone-8-acetic acid methyl ester (10)
showed moderate inhibition of tubulin polymerization.
Mechanism studies and further SAR evaluation are
ongoing.
As shown in Scheme 2, the unsubstituted and 2-(20-
fluorophenyl)-4-quinolone-8-acetic acids (11 and 12)
were prepared from the key intermediate, methyl 2-
aminophenylacetate (6), and ethyl benzoylacetate (7)
and ethyl 2-fluorobenzoylacetate (8). Compound 6 was
synthesized by esterification of commercially available
(o-nitrophenyl)acetic acid followed by reduction of the
nitro group. However, 6 readily undergoes an intramol-
ecular cyclization to the 2-oxindole via neutral and base
catalyzed proton transfer.12 Therefore, it was freshly
Table 1. Biological activities of 2-phenyl-4-quinolone acetic acids and their esters13
Compd
Cell linea ED50 (mg/mL)b
ITPc IC50 (mM)ꢂSD
KB
HOS
1A9
U87-MG
SKMEL-2
A549
MCF-7
2
3
4
9
10
11
12
NANA NA NA
>20 (36)d
>20 (15)
>20 (9)
>20 (17)
>20 (14)
>20 (12)
>10 (11)
0.06
>20 (8)
>20 (24)
>20 (21)
NA
>10 (22)
0.11
0.06
>20 (7)
>20 (16)
>20 (13)
>20 (15)
>10 (14)
0.44
>40
5.2ꢂ0.5
10ꢂ2
30ꢂ9
5.5ꢂ0.4
>40
>20 (37)
>20 (15)
>20 (30)
7.50
>20 (9)
NA
NA
NA
0.12
0.07
>20 (8)
>20 (8)
>10 (29)
0.04
NA
>20 (7)
>10 (18)
0.07
0.04
0.02
0.03
0.04
0.05
0.12
>40
aCell lines include human epidermoid carcinoma of the nasopharynx (KB), bone carcinoma (HOS), human ovarian cancer (1A9), glioblastoma
carcinoma (U-87-MG), human melanoma cancer (SKMEL-2), human lung cancer (A549), and human breast cancer (MCF-7).
bCytotoxicity, ED50 for each cell line, is the concentration of compound that causes a 50% reduction in adsorbance at 562 nm relative to untreated
cells using the SRB assay.20
cTubulin polymerization was evaluated as described in ref 11. Aminimum of two independent experiments was performed with each compound. The
IC50 value is defined as the concentration that inhibits the extent of assembly by 50% after 20 min at 26 ꢁC.
dNumbers in parentheses represent the percentage of inhibition at tested concentration.

Y. Xia et al. / Bioorg. Med. Chem. Lett. 13 (2003) 2891–2893
2893
Acknowledgements
R. M.; Tosini, S.; Skehan, P.; Scudiero, P. A.; Monks, A.;
Boyd, M. R. J. Natl. Cancer Inst. 1990, 82, 1113.
14. Melting points were determined on a Fisher-Johns melting
point apparatus without correction. 1H NMR spectra were
measured on a Bruker AC-300 spectrometer with TMS as
internal reference and CDDl3 or DMSO-d6 as solvent. Flash
chromatography was performed on silica gel (mesh 25–150
mm).
This investigation was supported by grant CA-17625
from the National Cancer Institute awarded to K. H.
Lee.
References and Notes
15. General procedure for the synthesis of 2-phenyl-4-quinolone
acetic acids and their esters. Methyl 2-aminophenylacetate (6)
(1.65 g, 10 mmol) was suspended in 8 g of polyphosphoric acid
(PPA). The mixture was warmed at 90–100 ꢁC, and 1.92 g (10
mmol) of ethyl benzoylacetate (7) was added dropwise. The
resulting mixture was further stirred for 1 h. After cooling,
water was added, then aqueous NaOH (10%) was added
slowly until pH=6 and the solution extracted with CHCl3.
The organic layer was dried over sodium sulfate and con-
centrated in vacuo. Chromatography using CHCl3/CH3OH
(30:1) as eluant afforded 2.93 g of compound 9, yield 65.5%;
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mp 76–78 ꢁC; H NMR (CDCl3) d 3.77 (s, 3H, CH3), 3.95 (s,
1
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3H, H-30, H-40, and H-50), 7.82 (m, 2H, H-20, H-60), 8.34 (d,
1H, J=8.0 Hz, H-5), 10.1 (s, 1H, NH).
6. Murray, J. C.; Smith, K. A.; Stern, D. M. Eur. J. Cancer
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13. Tubulin polymerization assays were performed as descri-
bed in ref 11. Cytotoxic assays were performed as described in:
Rubinstein, L. V.; Shoemaker, R. H.; Paull, K. D.; Simo,
16. 2-Phenyl-4-quinolone-8-acetic acid (11). 2-Phenyl-4-quino-
lone-8-methyl acetate (9) (150 mg, 0.51 mmol) was suspended
in 50% aq EtOH (10 mL) containing NaOH (100 mg). The
mixture was heated under reflux for 2 h. After cooling, the
solution was slowly acidified with aqueous HCl. The pre-
cipitate was collected, and washed with water to provide 140
mg of 11, yield 98%; mp 238–240 ꢁC; H NMR (DMSO-d6) d
1
4.17 (s, 2H, CH2), 7.40 (s, 1H, H-3), 7.56 (m, 2H, H-6, H-7),
7.61–8.01 (m, 5H, aromatic), 8.14 (d, 1H, J=8.0 Hz, H-5),
11.9 (s, 1H, NH), 12.9 (s, 1H, COOH).
17. 2-(20-Fluorophenyl)-4-quinolone-8-acetic acid (12). Obtained
by hydrolysis of 10 with aqueous 50% EtOH containing
NaOH using the same synthetic procedure as for 11, yield
78.9%; mp 243–245 ꢁC; H NMR (DMSO-d6) d 4.07 (s, 2H,
1
CH2), 7.35 (s, 1H, H-3), 7.35–7.40 (m, 4H, H-6, H-7, H-30 and
H-50), 7.53–7.63 (m, 2H, H-40, H-60), 8.08 (d, 1H, J=9.0 Hz,
H-5), 12.0 (br s, 1H, NH), 12.4 (S, 1H, COOH).