Antitumor agents. 211. Fluorinated 2-phenyl-4-quinolone derivatives as antimitotic antitumor agents

Article abstract of DOI:10.1021/jm0101085

Fluorinated 2-phenyl-4-quinolone derivatives were synthesized and evaluated in National Cancer Institute's 60 human tumor cell line in vitro screen. From the results, the ketone moiety plays an essential role in activity. Among the compounds tested, 2′-fl

Full text of DOI:10.1021/jm0101085

3932
J . Med. Chem. 2001, 44, 3932-3936
An titu m or Agen ts. 211. F lu or in a ted 2-P h en yl-4-qu in olon e Der iva tives a s
An tim itotic An titu m or Agen ts1
Yi Xia,† Zheng-Yu Yang,† Peng Xia,† Torben Hackl,‡ Ernest Hamel,‡ Anthony Mauger,§ J iu-Hong Wu,| and
Kuo-Hsiung Lee*,†
Division of Medicinal Chemistry and Natural Products, School of Pharmacy, University of North Carolina, Chapel Hill,
North Carolina 27599-7360, Laboratory of Drug Discovery Research and Development, Developmental Therapeutics Program,
Division of Cancer Treatment and Diagnosis, National Cancer Institute, Frederick Cancer Research and Development Center,
Frederick, Maryland 21702, Screening Technologies Branch, Developmental Therapeutics Program, Division of Cancer
Treatment, National Cancer Institute, Frederick Cancer Research and Development Center, Bethesda, Maryland 20892, and
Department of Pharmacy, 306 Hospital of PLA, Beijing 100101, China
Received March 9, 2001
Fluorinated 2-phenyl-4-quinolone derivatives were synthesized and evaluated in National
Cancer Institute’s 60 human tumor cell line in vitro screen. From the results, the ketone moiety
plays an essential role in activity. Among the compounds tested, 2′-fluoro-6-pyrrol-2-phenyl-
4-quinolone (13) exhibited the most potent cytotoxic activities (log GI50 < -8.00) against renal
and melanoma tumor cell lines. Compound 13 was also a potent inhibitor of tubulin
polymerization (IC50 ) 0.46 µM) and of radiolabeled colchicine binding to tubulin, with activities
comparable to those of the potent antimitotic natural products colchicine, podophyllotoxin, and
combretastatin A-4.
In tr od u ction
Most importantly, 1 demonstrated good in vivo activity
against the OVCAR-3 ovarian cell line, prolonging the
life span of mice bearing the tumor by 130%. Thus,
because 1 was the only compound to show such good in
vivo activity and that contained a fluorine atom at the
2′-position, synthesis of additional fluorinated quinolo-
nes and further characterizations of pharmacophores
were warranted. It is well-known that fluorinated drugs
often show unique pharmacological properties. Here, we
describe the synthesis of enol ether (2-5), thioketone
(6), and carbamate (7) derivatives of 1.
Microtubules are an important subcellular target for
development of anticancer chemotherapeutic agents.
Vinca alkaloids2 and taxoids3 are well-known examples
of antimitotic agents that are widely used clinically to
treat different cancers. Colchicine4,5 (Figure 1) is an-
other well-known agent that inhibits microtubule as-
sembly. Although colchicine has limited utility for
cancer therapy, the drug is an important tool in studies
of microtubule structure and function. The vinca alka-
loids, taxoids, and colchicine each interact with tubulin
by a unique mechanism, probably involving distinct
binding sites on the protein.
Two functional moieties are present in 1: a ketone
and an amine. Ketone oxygen and amine hydrogen
atoms are frequently involved in drug-receptor binding;
however, the importance of these two functional moi-
eties has not been well explored with the 2-phenyl-4-
quinolone class. On the other hand, 2-phenyl-4-quino-
lones can tautomerize into a 4-hydroxy-2-phenyl-
quinolone enol form, as seen by the absence of an
infrared carbonyl stretching frequency in CHCl3 solu-
tion. Although the keto-enol equilibrium usually lies
far in favor of the keto form, the enol form can, under
proper conditions, be trapped by alkylation of the
hydroxyl group. The enol ether derivatives may undergo
ready hydrolysis with liberation of the free enol in vivo,
which then reverts to the ketone form. By appropriate
selection of the alkyl group, enol ether derivatives can
be obtained that have varying hydrolysis rates as well
as varying lipophilicity/aqueous solubility ratios.9 In
addition, this modification can determine whether activ-
ity is maintained with both the ketone form and the enol
form.
Previously, we reported the synthesis and biological
evaluation of a series of 2-phenyl-4-quinolones as a new
class of antimitotic antitumor agents.6-8 They were
evaluated in the National Cancer Institute’s (NCI’s) 60
human tumor cell line (HTCL) in vitro screen and in a
tubulin polymerization inhibition assay. Most com-
pounds showed promising cytotoxicity in the HTCL
assay with GI50 values in the low micromolar to nano-
molar concentration range. In general, a good correla-
tion was found between cytotoxicity and inhibition of
tubulin polymerization. SAR studies led to the discovery
of 2′-fluoro-6,7-methylenedioxy-2-phenyl-4-quinolone (1),7
which showed potent cytotoxicity with an average
log GI50 value of -6.47 (log of the concentration that
reduced cell growth by 50%) in the HTCL screen.
Compound 1 (Figure 1) was also a potent inhibitor of
tubulin polymerization with an IC50 value of 0.85 µM.
* To whom correspondence should be addressed. Phone: 919-962-
0066. Fax: 919-966-3893. E-mail: khlee@email.unc.edu.
† University of North Carolina.
‡ Laboratory of Drug Discovery Research and Development, National
Cancer Institute.
Therefore, we first converted the ketone moiety of 1
to various enol ethers (2-5) (Scheme 1). Different chain
lengths should result in compounds with varying hy-
§ Screening Technologies Branch, National Cancer Institute.
| 306 Hospital of PLA.
10.1021/jm0101085 CCC: $20.00 © 2001 American Chemical Society
Published on Web 09/28/2001

Antitumor Agents
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 23 3933
F igu r e 1.
Sch em e 1a
a
(i) NaH/DMF, ClCH2COOEt; (ii) NaH/DMF, ClCH2CH2CH2COOEt; (iii) 10% NaOH; (iv) Lawessen reagent-toluene, reflux; (v) Boc-
CH2Cl2, room temp.
drophobicity and steric hindrance. Hydrolysis of the
esters (2, 4) to the carboxylic acids (3, 5) allowed the
preparation of water-soluble salts. These compounds
could circumvent the solubility problem in the 2-phenyl-
4-quinolone class of compounds, which have only mod-
erate solubility even in DMSO. We also synthesized the
thioketone derivative (6) of 1. Interchanging such groups
in bioactive molecules often results in different solubility
and bioavailability properties.10 Second, we explored the
importance of the amine hydrogen atom by replacing
this atom with tert-butoxycarbonyl (Boc), a frequently
used amino protecting group, in compound 7. Com-
pounds 2-7 were synthesized to explore which phar-
macophores of the lead compound are essential for its
antitumor activity as well as its interaction with tubu-
lin.
Finally, in our previous studies of 2-phenyl-4-quino-
lones, compounds with a heterocyclic ring at the 6-posi-
tion showed increased activity in in vitro cytotoxicity
and tubulin-based assays.8 These results prompted us
to synthesize the fluorinated 6-heterocyclic quinolone
13 (Scheme 2).

3934 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 23
Xia et al.
Sch em e 2a
a
(i) HNO3(f)-H2SO4; (ii) pyrrolidine; (iii) 10% Pd/C; (iv) 2′-fluorobenzoyl chloride-Et3N, THF; (v) t-BuOK-t-BuOH.
Ta ble 1. Inhibition of in Vitro Tumor Cell Growtha by Fluorinated Quinolone Derivatives
cytotoxicity log GI50 (M)b
compd
K562
NCI-H226
HCT116
OVCAR-3
RXF-393
SK-Mel5
SF-268
SF-295
mean log GI50
1
2
3
4
5
6
7
13
c
-6.35
>-4.00
>-4.00
-4.31
-7.22
-4.14
-7.09
c
Nt
-7.68
-4.14
>-4.00
-4.24
-5.40
-5.39
c
-5.64
>-4.00
>-4.00
>-4.00
-4.79
-4.70
c
-7.26
>-4.00
>-4.00
-4.21
-6.87
-4.10
>-4.00
-4.32
-5.33
-4.81
-6.23
-7.62
-4.42
>-4.00
-4.41
-5.68
-4.25
>-4.00
-7.49
>-4.00
>-4.00
-4.60
-5.11
-4.53
>-4.00
<-8.00
>-4.00
>-4.00
-5.40
>-4.00
-4.82
-5.63
-4.67
-7.49
-7.35
-5.07
-5.56
-4.78
-5.26
-4.93
-7.60
-7.35
-6.93
-7.51
-7.47
<-8.00
-7.41
-7.73
a
Data obtained from NCI’s in vitro disease-oriented human tumor cell screen. K-562, leukemia cell line; NCI-H226, non-small-cell
lung cancer cell line; HCT-116, colon cancer cell line; OVCAR-3, ovarian cancer cell line; RXF-393, renal cancer cell line; RXF-393, renal
b
cancer cell line; SK-Mel5, melanoma; SF-286 and SF-295, CNS tumor cell lines. The log concentrations that reduced cell growth by
50%. c Not tested.
Ta ble 2. Antitubulin Effects of Fluorinated Quinolone
Derivatives
Syn th esis a n d Biologica l Assa ys
ICBb (%)
ITPa (%)
IC50 (µM) ( SD
Synthesis of 2′-fluoro-6,7-methylenedioxy-2-phenyl-4-
quinolone (1) was previously reported.7 The syntheses
of enol ether derivatives (2-5) are shown in Scheme 1.
Treatment of 1 with NaH in DMF followed by alkylation
with ethyl chloroacetate or ethyl 4-chlorobutyrate af-
forded 2 and 4. Hydrolysis of 2 and 4 with 10% NaOH
in MeOH gave carboxylic acids 3 and 5, respectively.
Treating 1 with Lawesson’s reagent in toluene gave
thioketone 6. The nitrogen atom in 1 was converted to
the carbamate (7) with tert-butoxycarbonyl (Boc) chlo-
ride in CH2Cl2 at room temperature.
compd
5 µMc
1 ( µMc
1
2
3
4
5
6
7
0.68 ( 0.02
>40
>20
>40
12 ( 3
24 ( 2
3.2 ( 1
0.46 ( 0.003
7.3 ( 1
>40
14 ( 0.9
>40
5.0 ( 0.6
0.80 ( 0.07
0.46 ( 0.02
0.53 ( 0.05
39 ( 2
13
93 ( 1
76 ( 5
14d
15d
16e
17e
18e
Synthesis of 13 (Scheme 2) was based on a literature
method.7 Nitration of 3′-chloroacetophenone (8) gave 2′-
nitro-5′-chloroacetophenone. Nucleophilic displacement
of the 5′-chloro group by pyrroline followed by hydro-
genation gave 11. The biarylamide (12) was formed from
condensation of 11 and 2-fluorobenzoyl chloride in THF.
Cyclization of 12 in the presence of potassium tert-
butoxide (t-BuOK) gave 13.
colchicinef
podophyllotoxinf
combretastatin A-4f
92 ( 3
88 ( 0.4
a
b
ITP ) inhibition of tubulin polymerization. ICB ) inhibition
of colchicine binding, evaluated only when polymerization IC50
e
1.0 µM. c In the colchicine binding experiments, these values refer
to the inhibitor concentration used. The [3H]colchicine concentra-
d
tion was 5 µM, and the tubulin concentration was 1 µM. Data
from ref 6. e Data from ref 16. f Data from ref 15.
Compounds 1-7 and 13 were tested in National
Cancer Institute’s HTCL screen.11,12 This assay involves
determination of a test agent’s effect on growth param-
eters against a panel of approximately 60 human tumor
cell lines, mostly derived from solid tumors. The cyto-
toxic effects of each compound were expressed as
log GI50 values, which represents the log molar drug
concentration required to cause 50% inhibition for
selected tumor cell lines. These compounds were also
assayed as inhibitors of tubulin polymerization, and the
most active compounds were assayed as inhibitors of
[3H]colchicine binding to tubulin.
Resu lts a n d Discu ssion
The cytotoxic activities of 1-7 and 13 are summarized
in Table 1 and effects on tubulin-based assays in Table
2. The results showed that the cytotoxicity decreased
about 100-fold when the ketone form of 1 was converted
to an enol ether (2-5). Compounds 2-5 lack both
functional moieties in the B ring (the amine H and the
ketone O), and these compounds had little or no effect
on tubulin polymerization. Reduced cytotoxic activity
was also found with 6, where a thioketone moiety
replaced the ketone group. Thioketone 6, like the enol

Antitumor Agents
J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 23 3935
ethers, had minimal effect on tubulin polymerization.
These observations suggest that the ketone moiety plays
a crucial role in the interaction of 2-phenyl-4-quinolone
derivatives with tubulin and in the inhibition of cell
growth that results from this interaction. Although the
exact reason is uncertain, possible factors are steric and
electronic influences or reduced H-bonding ability be-
tween drug and target protein.
importantly, the ketone form of the B ring appear to be
involved in the binding of this class of compound to
tubulin. The amine hydrogen of the B ring also may be
important for maximal antitubulin activity, suggested
by the reduced activity of 7, but steric factors remain
to be excluded as the explanation for the reduced
activity of this compound. Additional studies on the
mechanism of action are ongoing, including the synthe-
sis of additional analogues.
When the Boc protecting group replaced the amine
hydrogen, the resulting compound (7) showed interest-
ing cytotoxic data. Compound 7 was less active than 1
against the SF-295 CNS tumor cell line, equipotent with
1 against the HCT-116 colon, and OVCAR-3 ovarian
cancer cell lines, and almost 20-fold more active than 1
against the NCI-H226 non-small-cell lung cancer. These
cells are among those that are exceptionally sensitive
to antitubulin agents,13 and 7 retained moderate activity
as an inhibitor of tubulin polymerization. It is also
possible that 7 undergoes intracellular conversion to a
more active compound. The reduced interaction of 7
relative to 1 with tubulin could be derived from either
steric factors (bulky tert-butyl group) or loss of the amine
hydrogen (altering H-bonding interactions with tubulin).
The loss of antitubulin activities of N-methylquinolone
(15 vs 14, Figure 1)6 and of 2-phenylquinazolinone (17
vs 16)16 supports the idea of a requirement for the N
hydrogen. However, activity was substantially enhanced
when the phenyl group of inactive compound 17 was
replaced by a styryl group (compound 18).16 This result
supports the hypothesis that steric factors account for
the loss of activity in 7 vs 1. However, it is also possible
that the binding site for 2-phenyl-4-quinolones and
quinazolinones (phenyl C ring directly attached to the
B ring) does not completely overlap the binding site of
2-styrylquinazolin-4(3H)-one (18), which has a linker
between the phenyl C ring and the B ring.
Exp er im en ta l Section
Melting points were determined on a Fisher-J ohns melting
point apparatus without correction. Elemental analyses were
performed by Atlantic Microlabs, Atlanta, GA. Optical rota-
tions were determined with a DIP-1000 polarimeter. 1H NMR
spectra were measured on a Bruker AC-300 spectrometer with
TMS as an internal reference and CDCl3 as the solvent. Flash
chromatography was performed on silica gel (mesh 25-150
µm) using a mixture of hexanes-ethyl acetate as eluant.
2′-F lu or o-6,7-(m eth ylen ed ioxy)-2-p h en yl-4-qu in olon e
(1).7 2-Acetyl-4,5-(methylenedioxy)aniline (3.0 mmol) was dis-
solved in 20 mL of THF and 10 mL of triethylamine. The
mixture was cooled in an ice bath. A solution of 2-fluorobenzoyl
chloride (3.0 mmol) was added dropwise. After 30 min at 0
°C, the mixture was stirred at room temperature overnight
and poured onto 50 mL of ice-water. The precipitate was
collected and washed successively with water and MeOH. The
solid was dried under vacuum and then suspended in 20 mL
of tert-butyl alcohol. Potassium tert-butoxide (1.17 g, 10.5
mmol) was added, and the mixture was heated under N2 at
70 °C for 24 h. The mixture was cooled and poured into 30
mL of aqueous NH4Cl solution. The solid was collected and
washed successively with water and a mixture of CHCl3 and
MeOH (10:1). The crude product was recrystallized from a
mixture of CHCl3 and MeOH (20:1). 1H NMR (DMSO-d6): δ
6.20 (s, 1 H, H-3), 6.17 (s, 2 H, OCH2O), 7.09 (s, 1 H, H-8),
7.43 (s, 1 H, H-5), 7.44 (m, 2 H, H-3′, H-6′), 7.62, 7.69 (both t,
J ) 7.5 Hz, 1 H each, H-4′, H-5′). Anal. (C16H10FNO3) C, H, N.
2′-F lu or o-6,7-(m eth ylen ed ioxy)-2-p h en yl-4-(O-eth yl a c-
eta te)qu in olin e (2). Compound 1 (283 mg, 1 mmol) was
dissolved in dry DMF (12 mL), and NaH (60% in oil, 110 mg,
2.8 mmol) was added portionwise with stirring at 40 °C. Ethyl
chloroacetate (500 mg, 4.08 mmol) was added, and the reaction
mixture was stirred for 2 h at 60 °C. The reaction mixture
was poured into ice-water and filtered. The precipitate
obtained was washed with water and recrystallized from CH2-
Cl2-MeOH to afford 260 mg of 2; yield 71.2%, mp 119-120
°C. 1H NMR (CDCl3): δ 1.31 (t, J ) 3.7 Hz, 3 H, CH3), 4.32 (q,
J ) 7.2 Hz, 2 H, CH2CH3), 4.88 (s, 2 H, OCH2COO), 6.13 (s, 2
H, OCH2O), 7.04 (s, 1 H, H-3), 7.17 (m, 1 H, H-3′), 7.30 (m, 1
H, H-5′), 7.40 (m, 1 H, H-4′), 7.42 (s, 1 H, H-8), 7.60 (s, 1 H,
H-5), 8.06 (m, 1 H, H-6′). Anal. (C20H16FNO5) C, H, N.
Among the new compounds, 13 (a fluorinated qui-
nolone with a heterocyclic ring at the 6-position) was
the most potent in all assays. It was more cytotoxic than
1 in virtually all cases, especially against RXF-393 renal
and SK-Mel5 melanoma cancer cells with log GI50
values of less than -8.00. Over all 60 cell lines, 13 was
about 6-fold more active than 1, as shown by the mean
log GI50 values. In keeping with its greater cytotoxicity
13 was more potent than 6 as an inhibitor of tubulin
assembly, but the greater affinity of 13 relative to 6 was
best demonstrated by its substantially greater activity
as an inhibitor of the binding of [3H]colchicine to
tubulin. In the latter assay, 13 was nearly as active as
the highly potent combretastatin A-4.14
2′-F lu or o-6,7-(m et h ylen ed ioxy)-2-p h en yl-4-(O-a cet ic
a cid )qu in olin e (3). Compound 2 (160 mg, 0.44 mmol) was
treated with aqueous NaOH (10%, 15 mL). The reaction
mixture was refluxed for 2 h and cooled to room temperature.
Aqueous HCl (10%) was added until the pH was 1-2. The
resulting precipitate was harvested by filtration and recrystal-
lized from MeOH-CHCl3 to give a light-yellow solid, 3; 130
Previously, 2-phenyl-4-quinolones were found to in-
hibit tubulin polymerization and the binding of radio-
labeled colchicine to tubulin.5-7 Numerous compounds
with various substitutions on both A and C rings were
studied. The “biaryl system”, composed of rings A and
C, is probably15 analogous to the similar biaryl system
occurring in many antimitotic natural products such as
colchicine, podophyllotoxin, and combretastatin A-44
(Figure 1). However, the pharmacophores in the B ring
of phenylquinolones have been relatively unexplored,
and we show here that the ketone functional moiety is
essential for a strong interaction with tubulin, providing
additional insight into the mechanism of ligand binding
at the colchicine site. The ketone oxygen and, most
1
mg, yield 87.0%, mp >300 °C. H NMR (DMSO-d6): δ 5.18 (s,
2 H, OCH2COO), 6.33 (s, 2 H, OCH2O), 7.42 (s, 1 H, H-3), 7.45
(s, 1 H, H-8), 7.48 (m, 2 H, H-3′ and H-5′), 7.54 (s, 1 H, H-5),
7.65 (m, 1 H, H-4′), 7.91 (m, 1 H, H-6′). Anal. (C18H12FNO5‚
0.25 H2O) C, H, N.
2′-F lu or o-6,7-(m eth ylen ed ioxy)-2-p h en yl-4-(O-eth yl 4′-
bu tyr a te)qu in olin e (4). 4 was obtained from 1 and ethyl
4-chlorobutyrate; yield 77.6%, mp 93-94 °C. 1H NMR (CDCl3):
δ 1.27 (t, 3 H, CH3, J ) 7.0 Hz), 2.29 (m, 2 H, CH2CH2CH3),
2.62 (t, J ) 7.29 Hz, 2 H, H-3′), 4.18 (q, J ) 7.0 Hz, 2 H, OCH2-
CH3), 4.28 (q, J ) 6.0 Hz, 2 H, OCH2CH2,), 6.12 (s, 2 H,
OCH2O), 7.13 (s, 1 H, H-3), 7.15-7.30 (m, 3 H, H-3′, H-4′, H-5′),
7.41 (s, 1 H, H-8), 7.46 (s, 1 H, H-5), 8.02 (m, 1 H, H-6′). Anal.
(C22H20FNO5) C, H, N.

3936 J ournal of Medicinal Chemistry, 2001, Vol. 44, No. 23
Xia et al.
2′-F lu or o-6,7-(m eth ylen ed ioxy)-2-p h en yl-4-(O-eth yl 4′-
bu tyla cetic a cid )qu in olin e (5). 5 was obtained by hydrolysis
of 4 with aqueous NaOH, using the same synthetic procedure
as for 3; yield 82.5%, mp >300 °C. 1H NMR (DMSO-d6): δ 2.10
(m, 2 H, CH2CH2CH2), 2.49 (t, 2 H, CH2COO), 4.29 (t, 2 H,
OCH2), 6.22 (s, 2 H, OCH2O), 7.23 (s, 1 H, H-3), 7.33 (m, 1 H,
H-3′), 7.35 (m, 1 H, H-5′), 7.38 (s, 1 H, H-8), 7.44 (s, 1 H, H-5),
7.52 (m, 1 H, H-4′), 7.95 (m, 1 H, H-6′). Anal. (C20H16FNO5) C,
H, N.
Ack n ow led gm en t . This investigation was sup-
ported by a grant from the National Cancer Institute
(Grant CA-17625) awarded to K. H. Lee.
Refer en ces
(1) For part 210, see: Shi, Q.; Wang, H. K.; Bastow, K. F.;
Tachibana, Y.; Chen, K.; Lee, F. Y.; Lee, K. H. Bioorg. Med.
Chem., submitted.
2′-F lu or o-6,7-(m et h ylen ed ioxy)-2-p h en ylq u in o-4-t h i-
on e (6). Compound 1 (500 mg, 1.77 mmol) in 30 mL of dry
toluene was stirred for a few minutes at room temperature,
and Lawessen reagent (1.07 g, 2.65 mmol) was added with
continued stirring. The mixture was stirred at 110-120 °C
for 24 h and became clear with a deep-orange color. The
mixture was cooled to room temperature, poured into water,
and extracted with CH2Cl2. The organic layer was dried over
sodium sulfate and concentrated. Chromatography using CH2-
Cl2/CH3OH as eluant afforded 430.6 mg of 6; yield 81.5%, mp
226-228 °C. 1H NMR (DMSO-d6): δ 6.24 (s, 2 H, OCH2O),
7.18 (s, 1 H, H-3), 7.33 (s, 1 H, H-8), 7.50 (m, 2 H, H-3′, H-5′),
7.72 (m, 1 H, H-4′), 7.77 (m, 1 H, H-6′), 8.08 (s, 1 H, H-5), 12.93
(s, 1 H, NH). Anal. (C16H10FNO2S‚1.05 H2O) C, H, N.
N-Boc-2′-flu or o-6,7-(m eth ylen ed ioxy)-2-p h en yl-4-qu i-
n olon e (7). To a solution of 1 (283 mg, 1 mmol) in 6 mL of
methylene chloride were added triethylamine (0.15 mL, 1
mmol), di-tert-butyl dicarbonate (436 mg, 2 mmol), and 4-(dim-
ethylamino)pyridine (61.25 mg, 1 mmol). The solution was
stirred for 24 h at room temperature under N2. The mixture
was poured into water, extracted with CH2Cl2, and washed
with water. The organic layer was dried over sodium sulfate
and concentrated. Chromatography using EtOAc-hexane as
eluant afforded 7; yield 86.8%, mp 118-120 °C. 1H NMR
(CDCl3): δ 1.61 (s, 9 H, 3 × CH3), 6.14 (s, 2 H, OCH2O), 7.14
(m, 1 H, H-3′), 7.20 (s, 1 H, H-3), 7.29 (s, 1 H, H-5), 7.40 (m, 1
H, H-5′), 7.46 (s, 1 H, H-8), 7.71 (m, 1 H, H-4′), 8.07 (m, 1 H,
H-6′). Anal. (C21H18FNO5) C, H, N.
(2) Rowinsky, E. K.; Donehower, R. C. The Clinical Pharmacology
and Use of Antimicrotubule Agents in Cancer Chemotherapeu-
tics. Pharmacol. Ther. 1992, 52, 35-84.
(3) Verweij, J .; Clavel, M.; Chevalier, B. Paclitaxel (Taxol) and
Docetaxel (Taxotere): Not Simply Two of a Kind. Ann. Oncol.
1994, 5, 495-505.
(4) Hastie, S. B. Interactions of Colchicine with Tubulin. Pharmacol.
Ther. 1991, 51, 377-401.
(5) Brossi, A.; Yeh, H. J .; Chrzanowska, M.; Wolff, J .; Hamel, E.;
Lin, C. M.; Quinn, F.; Suffness, M.; Silverton, J . Colchicine and
Its Analogues: Recent Findings. Med. Res. Rev. 1988, 8, 77-
94.
(6) Kuo, S. C.; Lee, H. Z.; J uang, J . P.; Lin, Y. T.; Wu, T. S.; Chang,
J . J .; Lednicer, D.; Paull, K. D.; Lin, C. M.; Hamel, E.; Lee, K.
H. Synthesis and Cytotoxicity of 1,6,7,8-Substituted 2-(4′-
Substituted phenyl)-4-quinolones and Related Compounds: Iden-
tification as Antimitotic Agents Interacting with Tubulin. J .
Med. Chem. 1993, 36, 1146-1156.
(7) Li, L.; Wang, H. K.; Kuo, S. C.; Wu, T. S.; Lednicer, D.; Lin, C.
M.; Hamel, E.; Lee, K. H. Antitumor Agents. 150. 2′,3′,4′,5′,5,6,7-
Substituted 2-Phenyl-4-quinolones and Related Compounds:
Their Synthesis, Cytotoxicity, and Inhibition of Tubulin Polym-
erization. J . Med. Chem. 1994, 37, 1126-1135.
(8) Li, L.; Wang, H. K.; Kuo, S. C.; Wu, T. S.; Lednicer, D.; Lin, C.
M.; Hamel, E.; Lee, K. H. Antitumor Agents. 155. Synthesis and
Biological Evaluation of 3′,6,7-Substituted 2-Phenyl-4-quinolones
as Antimicrotubule Agents. J . Med. Chem. 1994, 37, 3400-3407.
(9) Bundgaard, H. In Bioreversible Carriers in Drug Design Theory
and Application; Pergamon Press: New York, 1987; pp 13-94.
(10) Burger, A. In Medicinal Chemistry; Wiley-Interscience: New
York, 1997; Vol. 1, pp 72-80.
(11) Grever, M. R.; Schepartz, S. A.; Chabner, B. A. The National
Cancer Institute: Cancer Drug Discovery and Development
Program. Semin. Oncol. 1992, 19, 622-638.
(12) Monks, A.; Scudiero, D.; Skehan, P.; Shoemaker, R.; Paull, K.;
Vistica, D.; Hose, C.; Langley, J .; Cronise, P.; Vaigro-Wolff, A.;
Gray-Goodrich, M.; Campbell, H.; Mayo, J .; Boyd, M. Feasibility
of a High-flux Anticancer Drug Screen Using a Diverse Panel
of Cultured Human Tumor Cell Lines. J . Nat. Cancer Inst. 1991,
83, 757-766.
(13) Paull, K. D.; Lin, C. M.; Malspeis, L.; Hamel, E. Identification
of Novel Antimitotic Agents Acting at the Tubulin Level by
Computer-Assisted Evaluation of Differential Cytotoxicity Data.
Cancer Res. 1992, 52, 3892-3900.
(14) Lin, C. M.; Ho, H. H.; Pettit, G. R.; Hamel, E. Antimitotic Natural
Products Combretastatin A-4 and Combretastatin A-2: Studies
on the Mechanism of Their Inhibition of the Binding of Colchi-
cine to Tubulin. Biochemistry 1989, 28, 6984-6991.
(15) Xia, Y.; Yang, Z. Y.; Xia, P.; Bastow, K.; Tachibana, Y.; Kuo, S.
C.; Hamel, E.; Hackl, T.; Lee, K. H. Antitumor Agents. 181.
Synthesis and Biological Evaluation of 6,7,2′,3′,4′-Substituted-
2′-Flu or o-6-pyr r oyl-2-ph en yl-4-qu in olon e (13). 2-Amino-
5-pyrroylacetophenone (11, 1 g, 4.9 mmol), prepared from
commercially available 3′-chloroacetophenone according to
literature procedures,8,17 was dissolved in 10 mL of THF and
2 mL of triethylamine. The mixture was cooled in an ice bath.
A solution of 2-fluorobenzoyl chloride (855 mg, 5.39 mmol) was
added dropwise. After 30 min at 0 °C, the mixture was stirred
at room temperature overnight and poured onto 50 mL of ice-
water. The precipitate was collected and washed successively
with water and MeOH. The solid (12) was dried under vacuum
and suspended in 20 mL of tert-butyl alcohol. Potassium tert-
butoxide (1.65 g, 14.7 mmol) was added, and the mixture was
heated under N2 at 70 °C for 16 h. The mixture was cooled
and poured into 30 mL of ice-water. Aqueous 10% HCl was
added to attain pH ) 6. The solid was collected and washed
several times with water. The crude product was recrystallized
from a mixture of CH2Cl2 and MeOH to afford 13; yield 59.3%.
1H NMR (DMSO-d6): δ 2.01 (m, 4 H, CH2CH2CH2CH2), 3.33
(m, 4 H, CH2CH2CH2CH2), 6.04 (s, 1 H, H-3), 7.04 (d, J ) 2.5
Hz, 1 H, H-8), 7.10 (dd, J ) 2.5, 9.1 Hz, 1 H, H-7), 7.39 (d, J
) 9.0 Hz, 1 H, H-5), 7.43-7.71 (m, 4 H, H-3′, H-4′, H-5′, H-6′).
Anal. (C19H17FN2O‚0.25 H2O) C, H, N.
1,2,3,4-tetrahydro-2-phenyl-4-quinolones as
a New Class of
Antimitotic Antitumor Agents. J . Med. Chem. 1998, 41, 1155-
1162.
(16) Hamel, E.; Lin, C. M.; Plowman, J .; Wang, H. K.; Lee, K.H.;
Paull, K. D. Antitumor 2,3-Dihydro-2-(aryl)-4(1H)-quinazolinone
Derivatives. Interactions with Tubulin. Biochem. Pharmacol.
1996, 51, 53-59.
(17) Simpson, J . C. E.; Atkinson, C. M.; Schofield, K.; Stephenson,
O. o-Amino-ketones of the Acetophenone and Benzophenone
Types. J . Chem. Soc. 1945, 646-657.
Biologica l Assa ys. The tubulin polymerization and [3H]-
colchicine binding assays were performed as described previ-
ously.7 In the polymerization assay, reaction mixtures con-
tained 10 µM tubulin, and in the colchicine binding assay, the
reaction mixtures contained 1.0 µM tubulin and 5.0 µM [3H]-
colchicine.
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