Synthesis and antitumor activity of 1,3-benzodioxole derivatives.

Article abstract of DOI:10.1016/S0014-827X(02)01276-4

A series of 1,3-benzodioxoles (5-19) was synthesized and evaluated for their in vitro antitumor activity against human tumor cell lines. Some derivatives exhibited tumor growth inhibition activity. In particular, 6-(4-aminobenzoyl)-1,3-benzodioxole-5-acetic acid methyl ester 8, the most active compound of the series, possesses a significant growth inhibitory activity on 52 cell lines at concentrations ranging from 10(-7) to 10(-5) M.

Full text of DOI:10.1016/S0014-827X(02)01276-4

Il Farmaco 57 (2002) 853ꢀ859
/
www.elsevier.com/locate/farmac
Short Communication
Synthesis and antitumor activity of 1,3-benzodioxole derivatives
Nicola Micale, Maria Zappala`, Silvana Grasso *
Dipartimento farmacoꢀchimico, universita` di Messina, Viale Annunziata, 98168 Messina, Italy
/
Received 25 March 2002; accepted 11 May 2002
Abstract
A series of 1,3-benzodioxoles (5ꢀ
/19) was synthesized and evaluated for their in vitro antitumor activity against human tumor cell
lines. Some derivatives exhibited tumor growth inhibition activity. In particular, 6-(4-aminobenzoyl)-1,3-benzodioxole-5-acetic acid
methyl ester 8, the most active compound of the series, possesses a significant growth inhibitory activity on 52 cell lines at
concentrations ranging from 10ꢁ7 to 10ꢁ5 M.
´
# 2002 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
Keywords: 1,3-Benzodioxoles; Antitumor activity
1. Introduction
the results of a primary antitumor screening performed
by NCI.
The ongoing efforts of research on treatment of
malignancy are focused on the discovery of novel
products that, differently from currently-used drugs,
might have a higher selective cytotoxicity and act with
different mechanisms of action.
Random screening continues to be one of the main
routes to discover new leads in the field of antineoplastic
agents and the National Cancer Institute (NCI,
Bethesda, USA) plays a pivotal role in this area. The
NCI screening program [1] is based on the selection of
compounds characterized by a peculiar chemical struc-
ture.
It has recently been reported that 1,3-benzodioxole
derivatives possess cytotoxic activity against several
human tumor cell lines including human colon carci-
noma cells [2] and multidrug-resistant nasopharyngeal
carcinoma cells [3].
The ability of the NCI selected compounds 5ꢀ17 and
/
19 to inhibit the growth of three human tumor cell lines
in vitro was used as a preliminary indication of their
anticancer activity. Compounds 8, 12, 13, 15, 16 and 19,
which passed criteria for activity in the primary assay,
were evaluated against the full panel of 60 human tumor
cell lines.
2. Chemistry
The synthesis of novel compounds was accomplished
following the reaction sequence reported in Scheme 1.
The methyl ester of 1,3-benzodioxole-5-acetic acid (3)
and 1-(1,3-benzodioxol-5-yl)-2-propanone (4) were pre-
pared by reacting commercially available 1,3-benzodi-
oxole-5-acetonitrile (1) with a methanol solution of
hydrochloric acid and methylmagnesium bromide, res-
pectively. Ketone 4 was also obtained [7] via a pyridi-
nium chlorocromate (PCC) oxidation of alcohol 2. By
condensing alcohol 2 with 4-nitrobenzaldehyde, iso-
chromane derivative 5 was obtained, predominantly as
the trans diastereoisomer, in analogy to related trans-
formations [8]. The reduction of ketoester 7, prepared
On this basis and in pursuing our interest in the study
of new anticancer agents [4ꢀ
our attention on new 1,3-benzodioxole derivatives. We
report here the synthesis, and the physicalꢀchemical
properties of a set of new derivatives (5ꢀ19) along with
/
6], we have now focused
/
/
via Friedelꢀ
/
Crafts acylation of 3 with 4-nitrobenzoic
* Corresponding author
E-mail address: grasso@pharma.unime.it (S. Grasso).
acid in the presence of phosphorous pentoxide, with
´
0014-827X/02/$ - see front matter # 2002 Editions scientifiques et me´dicales Elsevier SAS. All rights reserved.
PII: S 0 0 1 4 - 8 2 7 X ( 0 2 ) 0 1 2 7 6 - 4

Scheme 1. (a) MeOH/HCl, 0 8C, 8 h; (b) MeMgBr, THF, CuBr, D, 30 min; (c) PCC/CH2Cl2, rt, 12 h; (d) O2NC6H4CHO, HCl, (e) O2NC6H4COOH, P2O5, (CH2)Cl2, rt, 12 h; (f) NaBH4, MeOH,
0 8C, 1 h; (g) NH2NH2×H2O, EtOH, rt, 2 h; (h) ethylene glycol, benzene, p-TSOH, D, 2 h; (i) NaBH4, CH2Cl2/MeOH, rt, 2h; (j) MeSO2Cl, CH2Cl2/Et3N, rt, 30 min; (k) NH2NH2×H2O, cat. HCl,
MeOH, rt, 24 h; (m) 10% Pd-C/H2, MeOH, rt, 4 h; (n) SnCl2, MeOH, 70 8C, 1.5 h.
/
/

N. Micale et al. / Il Farmaco 57 (2002) 853ꢀ
/859
855
NaBH4 yielded directly isochromanone derivative 9,
whereas diketoderivative 10, obtained similarly from 4,
gave isoquinoline 12 by treatment with hydrazine
hydrate [7]. The reaction of diketoderivative 10 with 1
equiv. of ethylene glycol primarily involved the aliphatic
ketone group. Nevertheless, monoketal 14 was contami-
nated by minor amounts of diketal 15 (9%) and
trioxepane derivative 16 (12%), which derives from the
involvement of the neighboring carbonyl group as
already observed in related transformations [9]. Com-
pound 14 was also converted by treatment with sodium
borohydride into alcohol 17, which was then trans-
formed into benzopyran derivative 18 and isoquinoline
12. The reduction of the nitro group of derivatives 5, 7,
10 and 12 to yield the corresponding amino derivatives
6, 8, 11 and 13 was carried out under standard
are in full agreement with the proposed structures and
are reported in Section 4.
3. Pharmacological results and conclusions
The growth inhibitory effects of compounds 5ꢀ
/
17 and
M
19 were evaluated in vitro at a concentration of 10ꢁ4
against three human tumor cell lines derived from three
neoplastic diseases, i.e. lung (NCI-H460), CNS (SF-
268), and breast (MCF7) cancers (Table 1).
Compounds 8, 12, 13, 15, 16 and 19 passed the NCI
criteria for activity in the primary assay, i.e. they reduce
the growth of any one of the cell lines to 32% or less,
and, consequently, their cytotoxic and/or growth in-
hibitory effects were evaluated in vitro against approxi-
mately 60 human tumor cell lines derived from nine
neoplastic diseases namely leukemia, melanoma, non-
small cell lung, colon, CNS, ovarian, renal, prostate and
breast cancers.
1
conditions. The elemental analysis (C, H, N) and H
NMR spectral data of all the synthesized compounds
The data reported in Table 2 are average values
referred to all cell lines, obtained by interpolation on
Table 1
Growth percentages at 10ꢁ4 M in three-cell line for compounds 5ꢀ
17
and 19
/
doseꢀresponse curves and which represent concentra-
/
tions that produced 50% cell growth inhibition (GI50),
total cell growth inhibition (TGI, 0% growth) and 50%
Comp. Lung (NCI-H460)
Breast (MCF7)
CNS (SF-268)
cell death (LC50, ꢁ50% growth. The activity against
/
5
6
72
67
68
4
68
53
42
16
39
50
58
26
46
43
26
67
45
12
62
68
44
20
75
51
79
22
52
54
55
20
77
34
specific cell lines is shown in Table 3. Data are reported
only when the value of GI50, TGI or LC50 is higher than 5.
The inhibitory activity of the tested compounds falls
7
8
into the range of 10ꢁ5ꢀ10ꢁ4 M, except 8 which proved
/
9
76
54
76
16
31
56
38
86
74
21
10
11
12
13
14
15
16
17
19
to be the most active derivative within this set of
derivatives; it displays a GI50 mean value of 5.71.
As can be seen in Table 3, compound 8 exhibited high
inhibitory properties on 52 of the tested cell lines at GI50
level and on 15 cell lines at TGI level at 10ꢁ5ꢀ10ꢁ7 M,
/
however, no significant differential cellular or subpanel
sensitivity has been observed. For compounds 12 and
19, only sporadic significant growth inhibition values
are recorded.
The lack of a homogeneous series of derivatives,
together with the very poor activity of the tested
compounds except 8, does not allow a sound SAR
evaluation. In fact, the results obtained with derivatives
7, 8 and 11 should suggest that both the amino and the
ester groups are structural requirements needed for cell
growth inhibition. However, such considerations are not
always true since other nitro and amino derivatives, e.g.
10 versus 11 and 12 versus 13, are equipotent and,
furthermore, a comparison of the activity of derivatives
8 and 19 makes doubts on the role played by the ester
functionality.
In conclusion, this study lead to 6-(4-aminobenzoyl)-
1,3-benzodioxole-5-acetic acid methyl ester (8), the most
active compound of the series, which exhibits good
cytotoxicity; thus the optimization of the substitution is
a future development goal.
Table 2
Mean response parameters of in vitro antitumor activity test for
compounds 8, 12, 13, 15, 16 and 19
Comp.
GI50
TGI
LC50
8
12
13
15
16
19
5.7191.37
4.4890.95
4.4390.68
4.1590.46
4.2390.60
4.5291.94
4.5191.70
4.0590.35
4.0590.35
4.0090.10
4.0490.33
4.1091.92
4.0190.72
4.0090.11
4.0090.00
4.0090.00
4.0090.00
4.0190.38
The response parameters GI50, TGI and LC50 (9SE) are averaged
values referred to all cell lines and represent the molar concentration
(expressed as ꢁlog10) at which the percentage growth is ꢂ50, 0 and
ꢁ50, respectively.

856
N. Micale et al. / Il Farmaco 57 (2002) 853ꢀ859
/
Table 3 (Continued)
Table 3
GI50 and TGI values against different tumor cell lines for compounds
8, 12 and 19
Panel/cell line
8
12
19
GI50
TGI GI50
GI50
TGI
Panel/cell line
8
12
19
MDA-MB231/ATCC
HS 578T
5.30
5.60
6.93
6.88
5.62
GI50
TGI GI50
GI50
TGI
MDA-MB-435
MDA-N
T-47D
6.21
5.78
5.11
Leukemia
CCRF-CEM
HL-60 (TB)
K-562
5.79
5.88
6.13
6.02
5.72
7.08
5.22
5.16
Growth inhibition (GI50) and total growth inhibition (TGI) dose
represent the concentration (expressed as ꢁlog10) at which the
percentage growth is ꢂ50 and 0 as compared with control untreated
cells (ꢂ100), respectively.
MOLT-4
RPMI-8226
SR
5.92
5.29
Non-small cell lung cancer
A549/ATCC
EKVX
5.46
5.53
5.55
4. Experimental
HOP-62
HOP-92
5.42
4.1. Chemistry
NCI-H23
NCI-H322M
NCI-H522
5.21
5.01
6.58
Melting points were determined on a Kofler hot stage
apparatus and are uncorrected. Elemental analyses were
carried out on a C. Erba Model 1106 Elemental
Analyzer for C, H and N) and the results are within
5.71
Colon cancer
COLO 205
HCC-2998
HCT-116
HCT-15
HT29
5.68
6.62
5.97
5.59
6.01
6.50
5.83
6.01
5.46
90.4% of the theoretical values. Merck silica gel 60 F254
/
plates were used for analytical TLC; column chromato-
graphy was performed on Merck silica gel 60 (70-230
1
mesh). H NMR spectra were recorded in CDCl3 by
KM12
SW-620
means a Varian Gemini 300 spectrometer. Chemical
shifts are expressed in d (ppm) relative to TMS as
internal standard and coupling constants (J) in Hz. The
synthesis, physical and analytical properties of com-
pound 10 have been previously described [7].
CNS cancer
SF-268
SF-295
SF-539
SNB-19
SNB-75
U251
5.99
6.13
5.44
5.40
5.92
5.57
5.03
5.46
5.23
4.1.1. Synthesis of 1,3-benzodioxole-5-acetic acid methyl
ester (3)
Melanoma
LOX IMVI
M14
5.41
6.13
6.11
5.29
6.63
5.89
5.89
1,3-Benzodioxole-5-acetonitrile (1) (2 g, 12.4 mmol)
was dissolved in a freshly prepared solution of MeOH/
MeCOCl (1/1, v/v, 60 ml) and the reaction mixture was
stirred at 0 8C for 8 h. The solvent was removed under
vacuum and the residue was treated with water (60 ml)
SK-MEL-2
SK-MEL-28
SK-MEL-5
UACC-257
UACC-62
5.78
and extracted with ethyl ether (2ꢃ80 ml). The pooled
/
Ovarian cancer
IGROV1
organic extracts were dried (Na2SO4) and the solvent
was evaporated at reduced pressure to yield 3 (2.3 g,
95%) as an oil [7].
6.23
6.42
5.21
5.19
5.36
OVCAR3
OVCAR-5
OVCAR-8
SK-OV-3
6.46
6.02
4.1.2. Synthesis of 1-(1,3-benzodioxol-5-yl)-2-propanone
(4)
To a stirred solution of 1 (4 g, 24.8 mmol) and methyl
magnesium bromide (3.0 M solution in THF, 9.5 ml,
28.5 mmol), in THF (80 ml) was added CuBr (64 mg,
0.43 mmol), and the mixture was refluxed under
Renal cancer
786-0
A498
5.24
5.90
5.74
5.81
5.14
5.41
5.12
5.24
CAKI-1
RXF 393
SN12C
TK-10
nitrogen for 30 min. After cooling to 0ꢀ5 8C, 5 ml of
/
H2O was cautiously added, followed by 30 ml of 15%
H2SO4. After stirring for 14 h, 60 ml of ether was added,
the phases were separated, and the aqueous layer was
extracted twice more with 50 ml portions of ether. The
combined organic phase was dried (Na2SO4) and the
Prostate cancer
PC-3
DU-145
5.84
5.59
5.13
Breast cancer
MCF7
NCI/ADR-RES
6.20
6.11

N. Micale et al. / Il Farmaco 57 (2002) 853ꢀ
/859
857
solvent removed under reduced pressure to afford 4
(yield 30%, 1.32 g) as a light brown oil.
(eluant: ethyl ether/light petroleum, 60/40) to afford
pure 7 as yellow solid (2.6 g, 65%).
1
Rf (cyclohexane/EtOAc, 60/40) 0.56; H NMR: 2.14
(s, 3H, CH3), 3.59 (s, 2H, CH2), 5.93 (s, 2H, OCH2O),
Rf (ethyl ether/light petroleum, 50:50) 0.61; m.p. 135ꢀ
/
1
137 8C; H NMR: 3.62 (s, 3H, CH3), 3.87 (s, 2H, CH2),
6.06 (s, 2H, OCH2O), 6.80 and 6.84 (2s, 2H, H-4 and H-
6.61ꢀ
/
6.77 (m, 3H, Ar). Anal. (C10H10O3) C, H, N.
7), 7.92 (d, 2H, Jꢄ
/
8.5, H-2? and H-6?), 8.32 (d, 2H, Jꢄ
/
8.5, H-3? and H-5?). Anal. (C17H13NO7) C, H, N.
4.1.3. Synthesis of trans-7,8-dihydro-7-methyl-5-(4-
nitrophenyl)-5H-[1,3]dioxolo[4,5-g][2]benzopyrane
(5)
4.1.6. Synthesis of 6-(4-aminobenzoyl)-1,3-
benzodioxole-5-acetic acid methyl ester (8)
To a stirred solution of 2 (3 g, 16.6 mmol) in dioxane
(100 ml) was added 4-nitrobenzaldehyde (3 g, 19.8
mmol) and 12N HCl (3 ml). The reaction mixture was
refluxed for 1 h and the solvent was removed under
vacuum. The resulting residue was dissolved in CHCl3
and washed with water (100 ml). The organic layer was
dried (Na2SO4), the solvent was removed at reduced
pressure, and the residue was purified by crystallization
from EtOH to give 4.99 g (94%) of 5.
To a stirred solution of 7 (252 mg, 0.73 mmol) in
anhydrous MeOH (40 ml) tin(II) chloride (825 mg, 3.67
mmol) was added. The reaction mixture was heated at
70 8C for 1.5 h, the solvent was removed under vacuum
and the resulting residue was dissolved in ethyl acetate
and neutralized with 2N NaOH (2ꢃ100 ml). The
/
organic layer was dried (Na2SO4) and the solvent was
removed at reduced pressure. The residue was purified
(202 mg, 88%) by silica gel column chromatography
using EtOAc/light petroleum (50/50) as eluant and
recrystallized from light petroleum to give 8.
Rf (cyclohexane/EtOAc, 70/30) 0.74; m.p. 134ꢀ
/
136 8C; 1H NMR: 1.38 (d, 3H, Jꢄ
/
6.0, CH3), 2.65ꢀ
/
2.89 (m, 2H, CH2), 4.02 (m, 1H, H-7), 5.74 (s, 1H, H-
5), 5.86 (m, 2H, OCH2O), 6.04 and 6.61 (s, 2H, H-4 and
Rf (EtOAc/light petroleum, 50/50) 0.38; m.p. 124ꢀ
/
H-9), 7.52 (d, 2H, Jꢄ
/
8.8, H-2?,6?), 8.22 (d, 2H, Jꢄ
/
8.8,
126 8C; 1H NMR: 3.58 (s, 3H, CH3), 3.70 (s, 2H,
CH2), 4.19 (bs, 2H, NH2), 6.01 (s, 2H, OCH2O), 6.62 (d,
H-3?,5?). Anal. (C17H15NO5) C, H, N.
2H, Jꢄ
and H-7), 7.64 (d, 2H, Jꢄ
(C17H15NO5) C, H, N.
/
8.8, H-3? and H-5?), 6.81 and 6.85 (2s, 2H, H-4
/
8.8, H-2? and H-6?). Anal.
4.1.4. Synthesis of trans-5-(4-aminophenyl)-7,8-dihydro-
7-methyl-5H-[1,3]dioxolo[4,5-g][2]benzopyrane (6)
To a stirred solution of 5 (1.1 g, 3.5 mmol) in MeOH
(50 ml) 10% Pd/C (50 mg) was added. The mixture was
shaken under hydrogen at atmospheric pressure for 4 h
and the Pd/C was filtered out with Celite pad. The
solvent was removed in vacuo and the resulting residue
was purified by column chromatography eluting with
cyclohexane/EtOAc (70/30) as eluant and recrystallized
from EtOH to give 6 (677 mg, 68%).
4.1.7. Synthesis of 5,8-dihydro-5-(4-nitrophenyl)-7H-
[1,3]dioxolo[4,5-g][2]benzopyran-7-one (9)
To a cooled (0ꢀ5 8C) solution of 7 (2.1 g, 6.12 mmol)
/
in MeOH (60 ml) was slowly added NaBH4 (341 mg,
9.18 mmol) and the mixture was stirred for 1 h. The
solvent was removed under reduced pressure and the
resulting residue was poured into water and extracted
Rf (cyclohexane/EtOAc, 70/30) 0.55; m.p. 86ꢀ
/
88 8C;
with chloroform (2ꢃ80 ml). The combined extracts
/
1H NMR: 1.35 (d, 3H, Jꢄ
/
6.0, CH3), 2.60ꢀ2.79 (m, 2H,
/
were dried over Na2SO4. The solvent was removed
under reduced pressure and the resulting residue was
purified by column chromatography using cyclohexane/
EtOAc (50/50) as eluant to give 9 (1.1 g, 58%).
CH2), 3.30 (bs, 2H, NH2), 3.94 (m, 1H, H-7), 5.52 (s,
1H, H-5), 5.84 (m, 2H, OCH2O), 6.16 and 6.56 (s, 2H,
H-4 and H-9), 6.67 (d, 2H, Jꢄ
/
8.5, H-3?,5?), 7.10 (d, 2H,
Jꢄ 8.5, H-2?,6?). Anal. (C17H17NO3) C, H, N.
/
Rf (cyclohexane/EtOAc, 50/50) 0.61; m.p. 204ꢀ
206 8C; 1H NMR: 3.53 and 3.69 (dd, 2H, Jꢄ
18.4,
CH2), 6.00 (s, 2H, OCH2O), 6.36 (s, 1H, H-5), 6.37 (s,
1H, H-9), 6.74 (s, 1H, H-4), 7.53 (d, 2H, Jꢄ 8.5, H-
2?,6?), 8.29 (d, 2H, Jꢄ 8.5, H-3?,5?). Anal. (C16H11NO6)
C, H, N.
/
/
4.1.5. Synthesis of 6-(4-nitrobenzoyl)-1,3-benzodioxole-
5-acetic acid methyl ester (7)
/
/
To a stirred solution of 3 (2.3 g, 11.6 mmol) in 1,2-
dichloroethane (150 ml) 4-nitrobenzoic acid (2.53 g, 15.1
mmol) and phosphorous pentoxide (16.5 g, 117 mmol)
were added. The mixture was stirred at room tempera-
ture overnight, cautiously treated with water (120 ml)
4.1.8. Synthesis of 1-[6-(4-aminobenzoyl)-1,3-
benzodioxol-5-yl]-2-propanone (11)
and extracted with chloroform (2ꢃ
layer was separated and sequentially treated with 10%
NaOH (120 ml), brine (100 ml) and water (3ꢃ70 ml).
The organic phase was dried (Na2SO4) and the solvent
removed under reduced pressure to yield crude 7 which
was purified by a silica gel column chromatography
/
80 ml). The organic
Compound 11 was obtained with a procedure similar
to that reported for compound 6 starting from 10 (219
mg, 0.58 mmol). The product was purified by column
chromatography with EtOAc/cyclohexane/i-PrOH (60/
30/10) as eluant and recrystallized from EtOH to give 11
(159 mg, 80%).
/

858
N. Micale et al. / Il Farmaco 57 (2002) 853ꢀ859
/
Rf (EtOAc/cyclohexane/i-PrOH, 60/30/10) 0.83; m.p.
8H, 2-OCH2CH2O), 5.99 (s, 2H, OCH2O), 7.17 (s, 1H, H-
4), 7.34 (s, 1H, H-7), 7.57 (d, 2H, Jꢄ 9.1, H-2?,6?), 8.05 (d,
2H, Jꢄ 9.1, H-3?,5?). Anal. (C21H21NO8) C, H, N.
16: Rf (CCl4/EtOAc, 85:15) 0.37; m.p. 243ꢀ247 8C
(0.183 g, 12%); H NMR: 1.56 (s, 3H, CH3), 2.92 and
3.09 (dd, 2H, Jꢄ16.0, CH2), 3.58 and 4.27 (m, 4H,
OCH2CH2O), 5.90 (m, 2H, OCH2O), 6.47 (s, 1H, H-10),
6.64 (s, 1H, H-4), 7.64 (d, 2H, Jꢄ 9.1, H-2?,6?), 8.14 (d,
2H, Jꢄ 9.1, H-3?,5?). Anal. (C19H17NO6) C, H, N.
1
83ꢀ
CH2), 4.18 (bs, 2H, NH2), 6.01 (s, 2H, OCH2O), 6.62 (d,
2H, Jꢄ 8.5, H-3?,5?), 6.72 (s, 1H, H-4), 6.88 (s, 1H, H-
7), 7.63 (d, 2H, Jꢄ8.5, H-2?,6?). Anal. (C17H15NO4) C,
H, N.
/
86 8C; H NMR: 2.15 (s, 3H, CH3), 3.77 (s, 2H,
/
/
/
/
1
/
/
4.1.9. Synthesis of 7-methyl-5-(4-nitrophenyl)-1,3-
dioxolo[4,5-g]isoquinoline (12)
/
/
18 (280 mg, 0.88 mmol) was dissolved in MeOH/cat.
HCl and to the solution was added an excess of hydrazine
hydrate (0.15 ml, 3.1 mmol). The mixture reaction was
stirred at room temperature for 24 h. The solvent was
removed under reduced pressure and the resulting residue
was dissolved in chloroform, washed with water,
saturated NaHCO3 and then with NaCl. The organic
phase was dried (Na2SO4), the solvent evaporated and the
residue purified by silica gel column chromatography
(eluant: cyclohexane/EtOAc, 60/40) of the residue gave
4.1.12. Synthesis of [6-(2-methyl-1,3-dioxolan-2yl-
methyl)-1,3-benzodioxol-5-yl]-(4-nitrophenyl)-methanol
(17)
To a solution of 14 (920 mg, 2.46 mmol) in CH2Cl2/
MeOH (100 ml, 40/60) NaBH4 (466 mg, 12.4 mmol) was
added and the mixture was stirred at room temperature
for 2 h. The reaction mixture was poured into water,
extracted with CHCl3 (60 ml) and dried over Na2SO4.
The solvent was removed in vacuo and the crude
product was purified by silica gel column chromatogra-
phy (eluant: light petroleum/EtOAc, 70/30).
198 mg (72%) of 12; m.p. 178ꢀ180 8C [7].
/
4.1.10. Synthesis of 7-methyl-5-(4-aminophenyl)-1,3-
dioxolo[4,5-g]isoquinoline (13)
Rf (eluant: light petroleum/EtOAc, 70/30) 0.35; Oil.
1
(601 mg, 65%); H NMR: 1.46 (s, 3H, CH3), 3.01 and
Compound 13 (343 mg, 85%) was obtained with a
procedure similar to that reported for compound 6
starting from 12 (447 mg, 1.45 mmol).
3.04 (d, 2H, Jꢄ
CH2O), 4.52 (s, 1H, OH), 5.94 (m, 2H, OCH2O), 6.09 (s,
1H, CH), 6.38 (s, 1H, H-7), 6.84 (s, 1H, H-4), 7.60 (d,
/
14.5, CH2), 3.95ꢀ3.97 (m, 4H, OCH2-
/
Rf (EtOAc/cyclohexane/i-PrOH, 60:30:10) 0.53; m.p.
1
2H, Jꢄ
/
8.5, H-2?,6?), 8.23 (d, 2H, Jꢄ 8.5, H-3?,5?).
/
107ꢀ
/
109 8C; H NMR: 2.65 (s, 3H, CH3), 3.70 (bs, 2H,
Anal. (C19H19NO7) C, H, N.
NH2), 6.04 (s, 2H, OCH2O), 7.01 (s, 1H, H-9), 7.27 (s,
1H, H-4), 7.35 (s, 1H, H-8), 6.80 (d, 2H, Jꢄ8.2 Hz, H-
3?,5?), 7.45 (d, 2H, Jꢄ8.2 Hz, H-2?,6?). Anal.
(C17H14N2O2) C, H, N.
/
4.1.13. Synthesis of 7-methyl-5-(4-nitrophenyl)-5H-1,3-
dioxolo[4,5-g][2]benzopyran (18)
/
To a cooled (0ꢀ5 8C) and stirred solution of 17 (475
/
mg, 1.27 mmol) in CH2Cl2 (60 ml) was added triethy-
lamine (264 ml, 1.90 mmol) and methansulfonyl chloride
(114 ml, 1.65 mmol). The ice-bath was removed and the
mixture was stirred at room temperature for 30 min. The
solvent was removed under reduced pressure and the
resulting residue was treated with water, extracted with
4.1.11. Synthesis of [6-(2-methyl-1,3-dioxolan-2yl-
methyl)-1,3-benzodioxol-5-yl]-(4-nitrophenyl)-
methanone (14), 5-(2-methyl-1,3-dioxolan-2yl-methyl)-
6-[2-(4-nitrophenyl)-1,3-dioxolan-2-yl]-1,3-benzo-
dioxole (15) and 4,5-methylenedioxy-9-methyl-1-(4-
nitrophenyl)-10,13,14-trioxa-tricyclo[7.4.1.0*2,7*]-
tetradeca-2,4,6-triene (16)
To a solution of 10 (1.35 g, 4.11 mmol) in anhydrous
benzene (150 ml), ethylene glicol (0.36 ml, 5.34 mmol)
and p-toluensulfonic acid (0.135 g, 0.71 mmol) were
added. The reaction mixture was refluxed for 2 h, was
ethyl acetate (2ꢃ100 ml) and purified by column
/
chromatography using light petroleum/ethyl acetate
(80/20) as eluant.
Rf (light petroleum/ethyl acetate, 70/30) 0.64; m.p.
120ꢀ
5.58 (s, 1H, H-8), 5.93 (s, 2H, OCH2O), 6.07 (s, 1H, H-
5), 6.27 (s, 1H, H-9), 6.52 (s, 1H, H-4), 7.51 (d, 2H, Jꢄ
8.8, H-2?,6?), 8.21 (d, 2H, Jꢄ 8.8, H-3?,5?). Anal.
(C17H13NO5) C, H, N.
/
123 8C (301 mg, 75%); 1H NMR: 1.91 (s, 3H, CH3),
washed with water (2ꢃ80 ml) and then dried over
/
/
anhydrous Na2SO4. The organic phase was concen-
trated in vacuo, and the resulting residue was purified by
column chromatography using CCl4/EtOAc (85/15) as
eluant.
/
14: Rf (CCl4/EtOAc, 85/15) 0.28; oil (1.07 g, 71%);. 1H
4.1.14. Synthesis of (6-methyl-1,3-benzodioxol-5-yl)-(4-
aminophenyl)-methanone (19)
NMR: 1.21 (s, 3H, CH3), 3.13 (s, 2H, CH2), 3.48ꢀ
4H, OCH2CH2O), 6.02 (s, 2H, OCH2O), 6.72 (s, 1H, H-
7), 6.87 (s, 1H, H-4), 7.92 (d, 2H, Jꢄ 8.8, H-2?,6?), 8.28 (d,
2H, Jꢄ 8.8, H-3?,5?). Anal. (C19H17NO7) C, H, N.
15: Rf (CCl4/EtOAc, 85:15) 0.30; oil (0.135 g, 9%); 1H
NMR: 0.98 (s, 3H, CH3), 2.81 (s, 2H, CH2), 3.79ꢀ4.15 (m,
/3.68 (m,
19 (821 mg, 92%) was obtained with a procedure
analogous to that reported for compound 8, starting
from (6-methyl-1,3-benzodioxol-5-yl)-(4-nitrophenyl)-
methanone (1.0 g, 3.5 mmol) obtained in turn as
reported in literature [10].
/
/
/

N. Micale et al. / Il Farmaco 57 (2002) 853ꢀ
/859
859
[2] S. Yokota, M. Kitahara, K. Nagata, Benzylidene lactam com-
pound KNK347, a novel inhibitor of acquisition of thermotoler-
ance and heat shock protein induction in human colon carcinoma
Rf (EtOAc/light petroleum, 50/50) 0.41; m.p. 110ꢀ
112 8C; 1H NMR: 2.23 (s, 3H, CH3), 4.19 (bs, 2H,
NH2), 5.98 (s, 2H, OCH2O), 6.63 (d, 2H, Jꢄ 8.8, H-
3?,5?), 6.72 (s, 1H, H-4), 6.78 (s, 1H, H-7), 7.66 (d, 2H,
Jꢄ8.8, H-2?,6?). Anal. (C17H13NO3) C, H, N.
/
/
cells, Cancer Res. 60 (2000) 2942ꢀ2948.
/
[3] Y. Xia, Z. Yang, P. Xia, K.F. Bastow, Y. Nakanishi, K. Lee,
Antitumor agents. Part 202: Novel 2?-amino chalcones. Design,
synthesis and biological evaluation, Bioorg. Med. Chem. Lett. 10
/
(2000) 699ꢀ
[4] A. Chimirri, S. Grasso, A.M. Monforte, P. Monforte, M.
Zappala`, A. Carotti, Structureꢀactivity relationships of antitu-
mor thiazolo[3,4-a]benzimidazole derivatives, Farmaco 49 (1994)
337ꢀ344.
[5] A. Chimirri, S. Grasso, A.M. Monforte, A. Rao, M. Zappala`,
Synthesis and antitumor activity evaluation of D2-1,2,4-oxadia-
/
701.
4.2. Pharmacology
/
Evaluation of anticancer activity of compounds 5ꢀ
/
17
and 19 was performed at the NCI following the well-
known in vitro disease-oriented antitumor screening
program [11], which is based upon the use of multiple
panels of 60 human tumor cell lines against which our
compounds were tested at 10-fold dilutions of five
concentrations, ranging from 10ꢁ4 to 10ꢁ8 M. The
percentage growth was evaluated spectrophotometri-
cally versus controls not treated with test agents. A 48 h
continuous drug exposure protocol was followed and a
sulforhodamine B (SRB) protein assay was used to
estimate cell viability or growth.
/
zolines, Farmaco 51 (1996) 125ꢀ/129 (and references cited
therein).
[6] S. Grasso, N. Micale, A.M. Monforte, P Monforte, M. Zappala`,
Synthesis and in vitro antitumour activity evaluation of 1-aryl-
1H,3H-thiazolo[4,3-b]quinazolines, Eur. J. Med. Chem. 35
(2000) 1115ꢀ1119.
/
[7] M. Zappala`, S. Grasso, N. Micale, S. Polimeni, C. De Micheli, A
simple and efficient synthesis of GYKI 52466 and GYKI 52895,
Synth. Comm. 32 (2002) 527ꢀ533.
/
[8] T. Lang, J. Koroi, P. Botka, G. Zolyomi, T. Hamori, T. Lang,
Isochromane derivatives, HU 37778 (1986).
[9] K.J. McCullough, A. Masuyama, K.M. Morgan, M. Nojima, Y.
Okada, S. Satake, S. Takeda, Syntheses, structure analyses, and
reactions of 1,3,5-trioxepanes and related compounds, J. Chem.
Acknowledgements
Soc., Perkin Trans. (1998) 2353ꢀ2362.
/
We thank the Antitumor Evaluation Branch of the
National Cancer Institute for performing the biological
evaluation.
[10] S. Grasso, G. De Sarro, A. De Sarro, N. Micale, M. Zappala`,
G. Puja, M.. Baraldi, C. De Micheli, Synthesis and anticon-
vulsant activity of novel and potent 6,7-methylenedioxy-
phthalazin-1(2H)-ones, J. Med. Chem. 43 (2000) 2851ꢀ
/
2860.
References
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(Eds.), Cancer: Principles and Practice of Oncology Updates, vol.
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/