Synthesis and antitumour activity of new derivatives of flavone-8-acetic acid (FAA). Part 4: Variation of the basic structure

Article abstract of DOI:10.1002/1521-4184(20006)333:6<181::AID-ARDP181>3.0.CO;2-O

A range of 11 derivatives of flavone-8-acetic acid (FAA) in which the structure has been substantially altered in different ways have been prepared and their anti-tumour activity evaluated in vitro against a panel of human and murine tumour cell lines and in vivo against MAC 15A. The generally poor activity observed shows that the basic structure cannot be altered much without destroying the activity.

Full text of DOI:10.1002/1521-4184(20006)333:6<181::AID-ARDP181>3.0.CO;2-O

New Derivatives of Flavone-8-acetic Acid
181
Synthesis and Antitumour Activity of New Derivatives of
Flavone-8-acetic Acid (FAA). Part 41): Variation of the Basic Structure✩
a)
b)
b)
b)
b)
R. Alan Aitken *, Michael C. Bibby *, Patricia A. Cooper , John A. Double *, Andrea L. Laws ,
a)
a)
(the late) Robert B. Ritchie , and David W. J. Wilson
a) School of Chemistry, University of St. Andrews, North Haugh, St. Andrews, Fife, KY16 9ST, UK
b) Clinical Oncology Unit, University of Bradford, Bradford, West Yorkshire, BD7 1DP, UK
Key Words: Flavone acetic acid; antitumour activity; structure-activity relationships
treatment of solid tumours, its good activity against murine
Summary
[2,3]
tumour models
did not translate into useful clinical activ-
[4]
ity. In an attempt to clarify the mechanism of action of 1 a
number of groups have reported synthesis and anti-tumour
evaluation of a variety of structurally related compounds.
These include analogues of 1 in which the ring oxygen is
replaced by S or NH , xanthenone-4-acetic acids such as2
and flavone-8-carboxylic acids such as 3 . Further examples
A range of 11 derivatives of flavone-8-acetic acid (FAA) in which
the structure has been substantially altered in different ways have
been prepared and their anti-tumour activity evaluated in vitro
against a panel of human and murine tumour cell lines and in vivo
against MAC 15A. The generally poor activity observed shows
that the basic structure cannot be altered much without destroying
the activity.
[5]
[6]
[7]
investigated more recently include the coumarin-, flavonol-
[8]
and flavanone-acetic acids 4, 5 and 6 , flavone-2′-carbox-
[9]
[10]
[11]
ylic acids 7 , the nitro and amino compounds 8 and 9
and
conformationally restricted compounds such as 10 . In
previous papers in this series, we have described the synthesis
Introduction
[12]
and activity of the 6-methyl compounds 11 , and a wide
[13]
[1]
range of examples 12 with both subtituted phenyl groups
and substituted heteroaromatic groups
Although flavone-8-acetic acid 1 (FAA, NSC347512,
LM975) initially seemed a promising new agent for the
[14]
at the 2-position.
In this paper we describe the preparation and in vitro and in
vivo activity of a range of analogues of 1 in which the basic
structure has been altered more fundamentally.
O
O
O
Me
Me
O
Me
O
CO2H
1
Ph
O
Ar
Results and Discussion
Me
CO2H
CO2H
3
Synthesis
2
O
O
The first series of analogues were obtained fortuitously
during the previously reported synthesis of compounds 12.
Treatment of 3-allyl-2-hydroxyacetophenone 14 obtained by
thermal rearrangement of 13 with the appropriate methyl
ester 15 and base followed by in situ cyclisation of the
condensation product 16 gave the 8-allylflavones 17. When
Ar
O
OH
Ar
O
O
CO2H
6
Ar
O
CO2H
CO2H
4
O
5
O
these were treated with 3 equivalents of KMnO rather than
4
O
O
NO2
Ph
the large excess used to obtain 12, the oxidation proceeded
NH2
Ar
O
O
OH
R2
O
O
O
R1
Me
OH
Ar
2 NaH
heat
Me
HO2C
CO2H
9
ArCO2Me
OH
O
15
7
8
13
16
O
O
O
O
14
OMe
OMe
Me
H2SO4/MeOH
Ar =
O
Ar
O
Ar
O
O
O
a
b
OMe
KMnO4
CO2H
12
CO2H
Me
OMe
OMe
CO2H
Me
O
Ar
O
Ar
OH
S
10
11
OMe
MeO
OMe
—–—
1) Part 3: Ref. [14]
17
O
e
d
18
c
.
Arch. Pharm. Pharm. Med. Chem.
© WILEY-VCH Verlag GmbH, D-69451 Weinheim, 2000
0365-6233/00/0505/0181 $17.50 +.50/0

182
Aitken and co-workers
only as far as the stage of the hydroxyketones 18. As com-
pared to the structure of 12, these show a considerable simi-
larity, with C=O and acidic OH groups being present in
approximately the same location, but the absence of an actual
carboxylic acid function is significant and whether or not
these compounds are still active will thus be of interest.
In an attempt to examine the required spacing between the
8-acetic acid and 2-arylpyranone groups of 1 for activity, we
designed an extended analogue 33 in which this spacing is
increased. As shown, this was prepared by a variation of the
usual synthesis employing the acetylenic methyl ester 30.
Rather than cyclising to give the 2-(phenylethynyl)ben-
zopyranone, the intermediate instead cyclised to give the
pyran product 31. This behaviour of acetylenic 1,3-diketones
is well known, having first been reported by Ruhemann in
O
O
O
KMnO4
[18]
[19]
1908
and subsequently applied in a variety of cases
.
O
Ph
O
Ph
O
Ph
Ph
Methylation of the free OH group in 31 followed by perman-
ganate oxidation gave the extended analogue 33 which was
tested as its water-soluble sodium salt 34.
CO2H
CO2Me
1
17a
21
H
O
N
N
O
O
O
O
O
Ph
OH
2 NaH
Me
CONHNH2
Ph
O
OH
O
Ph
PhC≡C–CO2Me
OH
23
O
O
Ph
Br
30
OH
OH
O
20
14
O
O
Ph
31
19
O
Ph
N
N
Me2SO4
CONH2
O
O
Me
O
O
22
24
Ph
O
Ph
NaHCO3
KMnO4
O
Ph
Conventional reactions of both 8-allylflavone 17a and FAA
1 were used to obtain a further series of analogues: the
epoxide 19, the allylic bromide 20, the methyl ester 21 and
OMe
OMe
OMe
CO2H
33
CO2– Na+
32
34
the amide 22. The last two compounds have been re-
[1,15]
ported
before but with no anti-tumour activity results.
An attempt to prepare the hydrazide by reaction of 21 with
hydrazine hydrate led not only to the desired functional group
transformation but also to cleavage of the pyran ring to afford
the pyrazole 23. The 1,2,4-oxadiazole 24 was also prepared
The final analogue prepared was the dimeric bis(FAA)
ether 37 and the idea for this came from the work of Breslow
[20]
and co-workers
where a series of bis(amides) gave much
greater anti-tumour activity than monoamides, supposedly
due to cooperative binding at two proximate receptor sites.
When the standard synthetic method was applied to the
diphenyl ether 4,4′-diester 35 the target compound was ob-
tained albeit in poor yield.
since this group has been reported to be an effective bio-isos-
[16,17]
tere for a carboxylic ester in various situations
.
O
OH
O
O
O
Me
R
2 NaH
O
O
O
O
OH
OH
RCO2Me
R
4 NaH
Me
OH
H2SO4/MeOH
25
2
MeO2C
CO2Me
O
26
14
27
O
O
14
KMnO4
36
35
O
O
O
KMnO4
O
O
O
O
NaHCO3
R = CH2Ph CHPh2
NaHCO3
O
O
O
O
O
R
R
CO2H
O
HO2C
b
a
CO2– Na+
O
Na+ –O2C
CO2H
CO2– Na+
37
38
28
29
Anti-Tumour Activity in Vitro
We were also interested to prepare the 2-benzyl compound
29a since this was reported in the original paper to show
good anti-tumour activity against colon adenocarcinoma C38
in vivo but the structure differs considerably from all other
active analogues reported to date. The diphenylmethyl ana-
logue 29b was similarly prepared to examine the effect of
Activity of 7 of the compounds was assessed against MAC
15A derived from an ascitic murine adenocarcinoma of the
colon. In addition, 18a, 23 and 29a were evaluated against a
[1]
[3]
panel of human tumour cell lines. These were the human
[21]
colon adenocarcinoma DLD-1 , the human rectal adeno-
[22]
carcinoma HRT-18
and the human chronic myelogenous
[23]
placing a more bulky and non-planar group at the 2-position. leukaemia with erythroid characteristics K562 . In all stud-
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)

New Derivatives of Flavone-8-acetic Acid
183
Table 1. Activity against tumour cell lines in vitro.
Table 2. Activity against MAC 15A in vivo.
——————————————————————————————
——————————————————————————————
Com-
pound
Dose
Growth
delay (d)
Significance (r)
Com-
pound
IC50 value (mM) against
———————————————————
(mg kg–1
)
MAC 15A DLD-1
HRT-18
K562
——————————————————————————————
——————————————————————————————
1
200
300
28
2.2
NS
1
23±5.0
123±34
31±5.8
79±13
12±4.1
>1280
209±50
60±11
31±4.3
9.4±2.1
146±15
40±12
81±6.5
68±24
23±4.0
>260
7.25
< 0.01
< 0.01
NS
18d
23
2
13.3
18d
60
0
52±9.7
100
200
240
212
38
0.2
0
NS
24
19
20
21
22
23
24
NS
29a
29b
34
22±2.2
73±21
54±3.2
0
NS
0
NS
1.6
0
NS
38
30
NS
——————————————————————————————
60
0
NS
ies the compounds were reconstituted in saline and che-
mosensitivity was assessed using an MTT assay following
100
34
2.5
0
< 0.01
NS
[24]
29a
the continuous (96 hours) exposure of cell lines to each
compound. All results were expressed in terms of % survival,
taking the control absorbance values to represent 100% sur-
40
0
NS
60
0
NS
29b
34
500
500
0
NS
vival. From the dose response curves constructed, IC values
50
1.0
NS
were estimated. The results given in Table 1 show a broad
spectrum of activity with IC values for MAC 15A ranging
——————————————————————————————
50
from 12 to over 1200 µM. Although it is clear that all the
compounds here have relatively low activity and high con-
centrations are required for any significant effect, some of the
results are interesting. Thus it appears that both the 2-benzyl
compound 29a and the compound 23 with a distinctly non-
flavonoid structure have activity comparable to 1, while the
hydroxyketone 18d, the oxadiazole 24 and the dimeric com-
pound 38 are less active, and both the extended analogue 34
and the diphenylmethyl compound 29b are essentially inac-
tive.
NS = not significant (r > 0.05)
contribute to our understanding of the mechanism of action
of FAA.
Acknowledgment
We thank the Association for International Cancer Research for their
generous support of this work and Bradford’s War on Cancer (M.C.B. and
J.A.D.)
The results in a broader panel comprising two lines derived
from human large bowel tumours, DLD-1 and HRT-18 and a
human leukaemia, K562 are also interesting. As shown in
Table 1, the hydroxyketone 18d was much more active than
1 against DLD-1 but essentially inactive against K562. Both
23 and 29a showed roughly comparable activity to 1 against
these three cell lines.
Experimental
Melting points were determined on a Reichert hot-stage microscope and
are uncorrected. NMR spectra were recorded at 300 MHz for 1H and at
75 MHz for 13C on a Bruker AM300 instrument using solutions in CDCl3 or
CD3SOCD3 and are reported in ppm relative to Me4Si as internal standard
with coupling constants in Hz. Infrared spectra were obtained using a
Perkin-Elmer SP-1200 spectrophotometer on thin films for liquids and on
Nujol mulls for solids. Mass spectra were obtained, unless otherwise indi-
cated, on an A.E.I./Kratos MS–50 spectrometer using electron impact (EI)
at 70 eV. Fast atom bombardment (FAB) spectra were obtained on a VG
Autospec spectrometer using glycerol as the matrix. Dry THF was freshly
distilled from potassium benzophenone ketyl under N2. Solutions of products
were dried over anhydrous MgSO4 and evaporated under reduced pressure.
Anti-Tumour Activity in Vivo
The results of evaluation of the compounds against the
MAC 15A subcutaneous tumour are shown in Table 2 and
these proved to be almost uniformly negative. Only for the
oxadiazole compound 24 is there any hint of significant
activity. In the case of 29a which showed good activity in
Preparation of 8-(3-Hydroxy-2-oxopropyl)flavone Derivatives 18a–e
To a stirred solution of the appropriate 8-allylflavone 17 prepared as
described previously[13,14] (11 mmol) in a mixture of acetic acid (100 ml),
water (100 ml) and acetone (100 ml) was added potassium permanganate (2.6
g, 16.5 mmol) over a period of 1 h. After a further 1 h the mixture was
decolourised by the addition of a saturated solution of sodium metabisulfite
(approx. 50 ml) and then reduced in volume and extracted with CH2Cl2 (400
ml). The extract was washed with water (2 × 100 ml), dried and evaporated.
The resulting yellow solid was triturated with dry ether at 0 °C and filtered
to yield the product as shown in Table 3.–1H NMR see Table 4.–13C NMR
see Table 5.
vitro this is not entirely unexpected since the benzylic CH
2
group is susceptible to biological oxidation and it may be
destroyed by metabolism before it can reach the site of action.
It seems clear from these results that by introducing the more
fundamental alterations to the FAA structure we have re-
moved some of the elements required for activity. However
these results do serve to further define the requirements for
an active compound of this type and, taken together with the
results for compounds 1–12 mentioned earlier, they may
The compounds 19–24 were prepared as follows and had properties as
listed in Table 3.–1H NMR see Table 4.–13C NMR see Table 5.
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)

184
Aitken and co-workers
Table 3. Preparation and properties of flavone derivatives.
—————————————————————————————————————————————————————————————–
Com-
pound
Yield
[%]
Mp
Formula or lit. mp
(C, H, N)a
IR
MS
[ûC]
[cm–1
]
[m/z (%)]
—————————————————————————————————————————————————————————————–
18a
18b
18c
18d
18e
19
66
66
57
45
52
62
75
64
90
32
12
49
12
88
42
9
200–202
155–157
182–184
181–183
214–216
117–118
170–172
169–170
235–240
236–239
189–190
59–60
C18H14O4 + 0.5 H2O
20H18O6
3300, 1723, 1641, 1591
3306, 1730, 1624, 1599
3440, 1720, 1630, 1581
3250, 1710, 1565
3322, 3020, 1695, 1550
1705, 1600, 1560
1620, 1580, 1560
1720, 1638, 1600
—
294 (15) [M+] , 279 (100), 235 (53)
C
354 (73) [M+], 324 (21), 295 (63), 133 (100)
354 (95) [M+], 324 (14), 311 (6), 133 (100)
384 (0.5) [M+], 352 (5), 346 (8), 268 (100)
330 (55) [M+], 314 (3), 271 (59), 133 (100)
278 (92) [M+], 262 (48), 235 (40), 156 (100)
342/340 (4) [81/79Br–M+], 261 (100) [M+–Br]
294 (90) [M+], 266 (3), 235 (74), 133 (100)
279 (1) [M+], 262 (1), 247 (5), 105 (100)
308 (3) [M+], 276 (100), 248 (68), 220 (16)
318 (100) [M+], 290 (9), 279 (7), 261 (11)
276 (100) [M+], 185 (6), 161 (22), 132 (23)
294 (100) [M+], 277 (30), 249 (29), 179 (26)
339 (39) [M+Na]+, 317 (20) [M+H]+, 176 (100)b
352 (100%) [M+], 274 (12), 191 (16), 161 (84)
370 (23) [M+] , 325 (20), 283 (18), 167 (100)
C20H18O6
C21H20O7 + 0.3 H2O
C17H14O5S
C18H14O3
20
C18H13BrO2
ref.[1] 168–169
ref.[14] 232–258
21
22
23
C17H16N4O2 + 0.5 H2O 3345, 1620, 1575
24
C19H14N2O3
C19H16O2
ref.[1] 143–145
—
1615, 1570
27a
28a
29a
27b
28b
29b
31
1643, 1599
140–142
350
1716, 1650, 1603, 1590
1610, 1560
135–137
171–172
350
C25H20O2
C24H18O4
—
1647, 1598
3400–2800, 1733, 1635
79
74
45
38
89
29
13
94
1610
—
178–180
bp 194
C20H16O3
2520, 1652, 1585, 1568
1600
304 (100) [M+], 276 (83), 157 (52), 129 (44)
318 (100) [M+], 290 (83), 275 (9), 258 (14)
336 (100) [M+], 308 (75), 292 (13), 265 (44)
—
c
32
33
212–214
267–268
189–191
267–269
350
C20H16O5
—
3060–2940, 1720, 1621
1713, 1612, 1555
1733, 1634, 1582
3400, 1701, 1640, 1595
1571, 1530
34
36
C36H26O5 + 0.5 H2O
538 (6) [M+], 298 (3), 256 (5), 91 (100)
574 (10) [M+] , 530 (17) [M+–CO2], 486 (100)
—
d
37
38
—
—————————————————————————————————————————————————————————————–
a Satisfactory elemental analysis obtained
b FAB
c HRMS (C21H18O3): calcd, 318.1254; found, 318.1256.
d HRMS (C33H22O3 [M+–CO2]): calcd, 530.1366; found, 530.1355.
8-(2,3-Epoxypropyl)-flavone (19)
evaporated. The residue was recrystallised from ethyl acetate to give the title
compound as a yellow powder.
A solution of 8-allylflavone (17a) (262 mg, 1 mmol) in dichloromethane
(10 ml) was stirred at 0–5 °C while m-chloroperbenzoic acid (240 mg, 1.1
mmol) was added. The solution was stirred at this temperature for 3 h and
for a further 1 h at room temperature before being shaken with 10% aqueous
sodium bicarbonate (20 ml). The organic layer was separated, dried and
evaporated to give an orange semi-solid. This was triturated with ethanol and
the resulting solid recrystallised from ethanol gave the title compound as
colourless crystals.
8-(Methoxycarbonylmethyl)flavone (21)
A mixture of flavone-8-acetic acid (1) (150 mg, 5.3 mmol) and methanol
(15 ml) containing conc. sulfuric acid (0.5 ml) was heated under reflux for
5 h. The reaction mixture was cooled and poured into water (20 ml). The
colourless precipitate was filtered off, washed with 10% sodium bicarbonate
solution and water, dried and recrystallised from methanol to give the title
compound as colourless crystals.
8-(3-Bromoprop-1-enyl)flavone (20)
8-(Carboxamidomethyl)flavone (22)
8-Allylflavone (17a) (3.0 g, 11.5 mmol) was added to a suspension of
1,3-dibromo-5,5-dimethylhydantoin (1.66g, 5.8 mmol) and AIBN (0.18 g,
1.0 mmol) in carbon tetrachloride (75 ml) and the mixture was heated under
reflux for 8 h. The resulting suspension was cooled and filtered and the filtrate
A solution of flavone-8-acetic acid (1) (0.5 g, 1.79 mmol) in thionyl
chloride (10 ml) was heated under reflux for 30 min and then evaporated.
The residue was stirred with conc. aqueous ammonia (5 ml) for 3 h and then
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)

New Derivatives of Flavone-8-acetic Acid
185
Table 4. 1H NMR data for substituted benzopyran-4-ones, δH.
—————————————————————————————————————————————————————————————–
Com-
pound
Benzopyranone
H-3
H-5a
H-6b
H-7a
8-Substituent
2-Substituent
—————————————————————————————————————————————————————————————–
18a
18b
18c
6.79 (s)
6.84 (s)
7.07 (s)
8.16 (dd)
8.00 (dd)
8.19 (dd)
7.38 (t)
7.47 (t)
7.39 (t)
7.56 (dd)
7.70 (dd)
7.57 (dd)
4.41 (s, 2 H), 4.09 (s, 2 H),
3.15 (br s, 1 H)
7.77 (m, 2 H), 7.52–7.39
(m, 3 H)
5.40 (br s, 1 H), 4.25 (s, 2 H),
4.18 (s, 2 H) 7
31 (m, 3 H), 3.90 (s, 3 H),
3.79 (s, 3 H)
4.37 (s, 2 H), 4.05 (s, 2 H),
3.20 (br s, 1 H)
7.27 (m, 1 H), 7.00 (m, 2 H),
3.87 (s, 3 H), 3.85 (s, 3 H)
18d
18e
7.14 (s)
6.80 (s)
7.98 (dd)
7.95 (m)
7.46 (t)
7.40 (t)
7.72 (dd)
7.62 (dd)
5.34 (t, J = 5, 1 H), 4.24 (m, 4 H)
7.29 (s, 2 H), 3.91 (s, 6 H),
5.42 (t, J = 6, 1 H), 4.38
(d, J = 6, 2 H)c
3.75 (s, 3 H)
7.95 (m, 1 H), 7.26 (d, J = 6, 1 H),
19
20
6.88 (s)
6.79 (s)
8.17 (dd)
8.14 (dd)
7.41 (t)
7.35 (t)
7.65 (dd)
7.80 (dd)
3.4–3.25 (m, 3 H), 2.88
(m, 1 H)c
4.06 (s, 3 H)
7.95 (m, 2 H), 7.58 (m, 3 H)
7.18 (d, J = 12, 1 H), 4.22
(d, 2 H, J = 7)c
7.92–7.85 (m, 2 H), 7.6–7.5
(m, 3 H)
21
6.85 (s)
7.30 (s)
6.82 (s)
6.13 (s)
8.18 (dd)
7.67 (dd)
8.19 (m)
8.03 (dd)
7.40 (t)
6.90 (t)
7.39 (t)
7.38 (t)
7.62 (dd)
7.14 (dd)
7.78 (m)
7.47 (dd)
4.02 (s, 2 H), 3.75 (s, 3 H)
9.2 (br s, 2H), 4.40 (br s, 2 H)
4.55 (s, 2 H), 2.35 (s, 3 H)
7.95–7.90 (m, 2 H), 7.6–7.5 (m, 3 H)
7.87 (m, 2H), 7.55–7.38 (m, 3 H)
7.88 (m, 2 H), 7.57 (m, 3 H)
22
24
27a
5.97–5.83 (m, 1 H), 5.06
(m, 2 H)c
7.38–7.25 (m, 5 H), 3.93 (s, 2 H)
28a
6.26 (s)
7.90 (dd)
7.27 (t)
7.67 (dd)
12.5–11.5 (br s, 1 H), 3.83
(s, 2 H)
7.40–7.33 (m, 5 H), 3.99 (s, 2 H)
29ad
27b
7.15 (s)
6.17 (s)
7.67 (dd)
8.05 (dd)
7.50–7.20 (m, 2 H)
3.53 (s, 2 H)
7.50–7.20 (m, 5 H), 3.55 (s, 2 H)
7.40–7.20 (m, 10 H), 5.37 (s, 1 H)
7.30 (m)
7.40 (m)
7.07 (t)
7.37 (t)
7.45 (dd)
5.85–5.64 (m, 1 H),
4.95–4.81 (m, 2 H)c
28b
29bd
36
6.32 (s)
6.25 (s)
6.81 (s)
8.12 (dd)
7.58 (dd)
8.12 (dd)
7.52 (dd)
7.40 (dd)
7.56 (dd)
3.59 (s, 2 H) (acid
proton not apparent)
7.36–7.19 (m, 10 H), 5.38 (s, 1 H)
3.52 (s, 2 H)
7.84 (m, 2 H), 7.41–7.22 (m, 8 H),
6.18 (s, 1 H)
6.16–6.03 (m, 2 H),
5.20–5.13 (m, 4 H),
3.76 (d, J = 6, 4 H)
7.96 and 7.21 (AB pattern,
J = 9, 8 H)
37
7.14 (s)
6.71 (s)
8.15 (dd)
7.73 (dd)
7.47 (t)
7.01 (t)
7.77 (dd)
7.52 (dd)
4.04 (s, 4 H) (acid
protons not apparent)
8.01 (m, 4 H), 7.20 (m, 4 H)
38c
3.70 (s, 4 H)
7.87 and 7.11 (AB pattern, J = 9, 8 H)
—————————————————————————————————————————————————————————————–
a J = 8, 2 b J = 8 c Additional signals: 18e 4.16 (s, 2 H); 19 2.63 (m, 1 H); 20 6.61 (dt, J = 12, 7, 1 H); 27a 3.54 (d, 2 H, J = 8);
27b , 3.42 (d, J = 8, 2 H) d in D2O
the mixture evaporated. The residual solid was recrystallised from ethanol to
give the product as colourless crystals.
H), 9.25 (br s, 31 H), 7.90 (m, 2 H), 7.70 (d, J = 8, 1 H), 7.59–7.42 (m, 3 H),
7.18 (s, 1 H), 7.16 (d, J = 8, 1 H), 6.93 (t, J = 8, 1 H), 4.34 (br s, 2 H).–13
C
NMR: δ = 170.3 (CO), 153.7 (4ry), 130.4 (CH), 129.3 (2 CH), 129.1 (4ry),
129.0 (4ry), 128.9 (CH), 128.8 (CH), 125.7 (2 CH, 4ry), 123.8 (CH), 119.2
(4ry), 116.8 (4ry), 100.1 (CH), 35.2 (CH2).
3-(3-Hydrazidocarbonylmethyl-2-hydroxyphenyl)-5-phenylpyrazole (23)
A solution of 8-(methoxycarbonylmethyl)flavone (21) (294 mg, 1.0 mmol)
in methanol (10 ml) was stirred at room temperature while hydrazine hydrate
(200 mg, 4.0 mmol) in methanol (5 ml) was added. After 8 h, the mixture
was poured into water and the resulting precipitate was filtered off and
recrystallised from ethanol to give the product.–1H NMR: δ = 11.40 (br s, 31
8-((3-Methyl-1,2,4-oxadiazol-5-yl)methyl)flavone (24)
A solution of acetamide oxime[25] (0.33 g, 4.4 mmol) in dry tetrahydro-
furan (15 ml) was heated to reflux in the presence of sodium hydride (0.21 g,
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)

186
Aitken and co-workers
Table 5. 13C NMR data for substituted benzopyran-4-ones, δC.
—————————————————————————————————————————————————————————————–
Com- Benzopyranone
pound C–2 C–3
8-substituent
C–8a C–1 C–2
2-substituent
Other
C–4
C–4a C–5
C–6
C–7
C–8
C–3
C–1′ C–2′ C–3′ C–4’
—————————————————————————————————————————————————————————————–
18a
18b
18c
18d
18e
19
163.4 107.9 178.3 122.7 125.6 125.2 135.5 124.2 154.3 40.2 205.9 68.0 131.7 126.2 129.3 131.8
160.5 111.2 177.1 123.0 124.9 123.4 135.9 124.6 154.2 40.5 207.6 67.4 125.4 152.9 147.2 115.8
160.8 112.8 178.6 122.6 125.5 124.9 135.3 124.1 154.4 40.2 206.1 67.9 121.2 153.5 113.1 117.8
162.3 107.0 177.5 123.5 126.7 125.1 136.2 123.9 154.3 39.9 207.5 67.6 103.9 104.0 153.5 140.7
159.9 105.5 176.6 123.4 125.0 124.7 135.9 123.8 153.7 39.6 207.5 67.9
—
109.8 157.9 117.9
163.1 107.5 178.6 124.0 125.0 124.5 134.7 126.7 154.5 32.7 51.2 46.9 131.8 126.2 129.2 131.6
163.2 107.6 178.1 124.3 125.7 125.0 131.1 126.0 153.2 129.1 126.6 32.9 131.8 126.3 129.2 131.7
20
21
163.0 107.5 178.4 123.9 125.2 125.0 135.3 124.1 154.4 35.8 170.9
163.1 107.7 178.0 123.2 125.8 125.2 134.9 124.4 154.1 27.6
—
—
131.7 126.2 129.1 131.7 52.4 (OMe)
131.6 126.3 129.2 131.8
24
—
27a
28a
29a
27b
28b
29b
36
167.5 110.3 178.6 123.6 127.4 123.7 135.3 124.7 154.4 33.8 133.8 116.7 134.9 128.9 129.3 129.3 40.8 (CH2)
167.8 109.5 176.9 123.0 124.7 123.6 135.4 125.1 154.2 34.6 171.8
161.8 111.4 183.9 124.5 126.4 120.2 139.9 130.8 157.4 41.4 181.5
—
—
135.6 128.6 129.1 127.0 39.5 (CH2)
138.8 131.4 132.5 134.7 47.2 (CH2)
169.2 112.1 178.6 123.8 124.8 123.7 135.1 129.6 154.4 33.9 133.9 116.6 138.9 128.8 129.0 127.5 56.1 (CH)
169.4 112.0 178.7 123.6 125.0 123.6 135.3 125.3 154.7 34.9 174.8
172.8 111.5 183.4 125.0 125.4 124.7 136.1 129.6 157.3 41.1 181.4
—
—
138.8 128.8 129.0 127.5 56.1 (CH)
137.9 131.7 132.2 138.6 57.1 (CH)
162.3 107.0 178.6 124.0 125.0 124.0 135.3 127.7 154.2 34.0 134.2 117.0 129.4 128.3 119.6 159.2
37
161.9 106.8 177.2 123.5 125.4 124.0 136.0 126.6 154.4 35.8 172.3
161.5 109.4 177.7 121.3 128.3 121.7 135.0 128.2 156.7 41.7 181.9
—
—
125.2 132.1 119.0 159.1
139.1 133.8 121.1 156.7
38
—————————————————————————————————————————————————————————————–
Additional signals: 18b 124.5 (C–5′), 120.2 (C–6′), 60.4 (OMe), 55.9 (OMe); 18c 152.2 (C–5′), 114.3 (C–6′), 56.1 (OMe), 55.9 (OMe); 18d 60.4 (OMe),
56.3 (2 C, OMe); 18e 131.1 (C–5′), 59.6 (OMe); 24 176.7 (C-5′), 167.6 (C-2′), 11.6 (2′-Me)
8.8 mmol). After 0.5 h 8-(methoxycarbonylmethyl)flavone (20) (1.29 g, 4.4
mmol) was added and heating continued for a further 4 h. Upon cooling, the
mixture was filtered and evaporated. The residue was extracted with dichlo-
romethane which was evaporated to give crude product. This was recrystal-
lised from methanol to give the title compound as colourless needles.
For synthesis of the remaining compounds the required carboxylic acids
were commercially available. They were converted into the corresponding
methyl esters by treatment with boiling thionyl chloride for 15 min, evapo-
ration, heating the residue with methanol for 1 h, followed by evaporation
and distillation.
Methyl Phenylpropiolate (30)
Phenylpropiolic acid was prepared by treatment of a solution of phenylace-
tylene in dry tetrahydrofuran with butyllithium in hexane followed by CO2
gas. The acid was taken up in the calculated quantity of aqueous 2M sodium
hydroxide which was filtered and evaporated to dryness. A suspension of the
residual sodium salt in ether and excess thionyl chloride was heated under
reflux for 6 h and then evaporated. Distillation of the residue gave the acid
chloride which was heated with methanol for 1 h and then evaporated and
the residue distilled to give the product as a colourless liquid, bp (oven temp.)
107 °C/0.13 Torr (ref.[29] 132–133 °C/16 Torr).–1H NMR: δ = 7.95 (s, 5 H),
3.80 (s, 3 H).
The compounds 29a,b and 38 were prepared by the previously reported
method[13] involving reaction of 3-allyl-2-hydroxyacetophenone (14), so-
dium hydride and the appropriate methyl ester followed by in situ acid-in-
duced cyclisation, permanganate oxidation and conversion into the sodium
salts using NaHCO3. The properties of the intermediates and products are
listed in Table 3.–1H NMR see Table 4.–13C NMR see Table 5.
The preparation of 34 followed the same route save for the addition of the
methylation step described below before the permanganate oxidation. The
properties of the intermediates and products are listed in Table 3 and the 1H
and 13C NMR spectra are given below.
Methyl Phenylacetate (25a)
From phenylacetic acid (84%) as a colourless liquid bp (oven temp.)
81 °C/0.3 Torr (ref.[26] 220 °C).–1H NMR: δ = 7.18 (m, 5 H), 3.67 (s, 3 H),
3.59 (s, 2 H).
Methyl Diphenylacetate (25b)
From diphenylacetic acid (91%) as cream plates, mp 58–60 °C (ref.[27]
59–62 °C).–1H NMR: δ = 7.31 (m, 10 H), 5.05 (s, 1 H), 3.76 (s, 3 H).
2-(3-Allyl-2-hydroxyphenyl)-6-phenylpyran-4-one (31)
1H NMR: δ = 9.3 (br s, 1 H, OH), 7.85 (d, J = 2, 1 H), 7.83 (d, J = 4, 1 H),
7.71 (dd, J = 8, 2, 1 H), 7.59 (d, J = 2, 1 H), 7.53–7.47 (m, 3 H), 7.29 (dd, J
= 8, 2, 1 H), 6.98 (t, J = 8, 1 H), 6.83 (d, J = 6, 1 H), 6.13–6.02 (m, 1 H),
5.27–5.11 (m, 2 H), 3.70 (d, J = 6, 2 H).–13C NMR: δ = 181.8, 164.1, 162.7,
154.5, 136.2, 133.4, 131.7, 131.5, 129.2, 128.4, 126.7, 126.1, 120.1, 118.9,
116.4, 115.1, 110.8, 34.7.
Dimethyl 4,4′-Oxydibenzoate (35)
From 4,4′-oxydibenzoic acid (60%) as colourless needles, mp 154–155 °C
(ref.[28] 156–158 °C).–1H NMR: δ = 8.05 and 7.05 (AB pattern, J = 8, 8 H),
3.90 (s, 3 H).
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)

New Derivatives of Flavone-8-acetic Acid
187
2-(3-Allyl-2-methoxyphenyl)-6-phenylpyran-4-one (32)
activity was tested up to the maximum tolerated dose. At the dose levels used
there was no significant body weight loss.
Sodium hydride (0.24 g, 9.8 mmol) was washed with light petroleum (40
ml) which was decanted off and replaced by THF (200 ml). The suspension
was then heated under reflux for 10 min and 31 (3.0 g, 9.8 mmol) was added
portionwise over 10 min followed by dimethyl sulfate (3.7 g, 39 mmol). After
heating for a further 2 h, the mixture was allowed to cool and concentrated
ammonia (40 ml) was added. After stirring for 12 h, water (200 ml) was added
and the mixture extracted with methylene chloride which was dried and
evaporated to yield a yellow oil. Column chromatography (silica, ethyl
acetate/petroleum 2:1) gave the product.–1H NMR: δ = 7.88–7.81 (m, 2 H),
7.63 (dd, J = 8, 2, 1 H), 7.52–7.49 (m, 3 H), 7.37 (dd, J = 8, 2, 1 H), 7.20 (t,
J = 8, 1 H), 7.00 (d, J = 2, 1 H), 6.83 (d, J = 2, 1 H), 6.10–5.90 (m, 1 H),
5.20–5.03 (m, 2 H), 3.69 (s, 3 H), 3.50 (d, J = 7, 2 H).–13C NMR: δ = 180.5,
163.7, 162.0, 156.8, 136.6, 134.8, 133.5, 131.4, 129.1, 129.0, 127.7, 125.9,
125.3, 124.4, 116.5, 115.6, 111.1, 61.3, 33.8.
Chemotherapy was administered on day 5 after tumour implantation to
allow for vascularization to occur, as determined histologically. Tumour
growth was followed by serial calliper measurements and anti-tumour activ-
ity assessed by tumour volume. This was calculated by the formula a2 × b/2
where a and b were the smaller and larger tumour diameters respectively[30]
.
Growth delay was determined as the difference in time taken for the median
tumours of the analogue treated and solvent control treated mice to reach a
relative tumour volume of two. The significance of the growth delay was
determined using a Mann-Whitney statistical analysis.
References
✩
Dedicated to Professor Richard Neidlein, Heidelberg on the occasion
of his 70th birthday.
2-(3-Carboxymethyl-2-methoxyphenyl)-6-phenylpyran-4-one (33)
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harvested from subconfluent stocks using 0.25% trypsin, counted on a
haemocytometer (Improved Neubauer chamber, Weber UK) and diluted in
complete RPMI 1640 for use at a concentration of 1 × 104 ml–1 MAC15A.
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RPMI tissue culture medium immediately prior to use and serially diluted.
100 µl per well of cell suspension was plated in 96 well plates (U bottomed,
tissue culture treated, Costar Cat No3799) and incubated for 3–4 hours before
addition of the test solution. To one row of eight wells, 100 µl complete
RPMI 1640 was added to serve as the control. Subsequent rows of eight wells
received a concentration of test solution over the range 1 mg ml–1 to 0.01 µg
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at 37 °C in an atmosphere of 5% CO2, 95% air for four days before being
assessed.
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used medium and compound solution was removed from each of the wells
following the 4 day exposure and replaced with 150 µl fresh medium and
20 µl of 5 mg ml–1 MTT solution. After 4 hours all of the medium and MTT
was removed from all of the wells and replaced with 150 µl DMSO. The
formazan crystals produced during the assay were dissolved and mixed by
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Received: January 17, 2000 [FP450]
Arch. Pharm. Pharm. Med. Chem. 333, 181–188 (2000)