1,5-dimethyl-6H-pyridazino[4,5-b]carbazole, a 3-aza bioisoster of the antitumor alkaloid olivacine.

Article abstract of DOI:10.1248/cpb.50.1479

A series of b-fused carbazoles structurally related to pyrido[4,3-b]carbazole-type alkaloids was prepared, utilizing the Diels-Alder reaction of 1-methylpyrano[3,4-b]indol-3(9H)-one with electron-deficient acetylenic dienophiles as the key step. The title

Full text of DOI:10.1248/cpb.50.1479

November 2002
Chem. Pharm. Bull. 50(11) 1479—1483 (2002)
1479
1,5-Dimethyl-6H-pyridazino[4,5-b]carbazole, a 3-Aza Bioisoster of the
Antitumor Alkaloid Olivacine
1)
Norbert HAIDER* and Eddy SOTELO
Institute of Pharmaceutical Chemistry, University of Vienna; Althanstraße 14, A-1090 Vienna, Austria.
Received July 1, 2002; accepted August 5, 2002
A series of b-fused carbazoles structurally related to pyrido[4,3-b]carbazole-type alkaloids was prepared,
utilizing the Diels–Alder reaction of 1-methylpyrano[3,4-b]indol-3(9H)-one with electron-deficient acetylenic
dienophiles as the key step. The title compound (14) thus obtained in only four steps represents a new 3-aza ana-
log of the antitumor natural product, olivacine.
Key words 3-aza-olivacine; pyridazino[4,5-b]carbazole; antitumor agent
Olivacine (1,5-dimethyl-6H-pyrido[4,3-b]carbazole) is a dazino[4,5-b]carbazoles and thus cannot be adopted to the
member of the pyridocarbazole family of alkaloids. It was synthesis of our target structure. Here, we wish to report on a
first isolated from the bark and stem of Aspidosperma oli- short synthesis of 3-aza-olivacine and on the results of an
vaceum2) and it has been found to possess similar antitumor initial in-vitro testing for antitumor activity.
activity as its 5,11-dimethyl isomer, ellipticine, and related
In a first approach to the 1,5-disubstituted pyridazino[4,5-
compounds like 9-methoxyellipticine.3,4) As the main mecha- b]carbazole skeleton, we investigated the selective addition
nism of these agents’ antineoplastic action, a stabilization of methyllithium to an appropriate 1-methylcarbazole-2,3-di-
of the “cleavable complex” which is formed between DNA carboxylic acid derivative. After some unsuccessful attempts
and the enzyme, topoisomerase II,5) is generally accepted, with the corresponding anhydride, we employed the pro-
although additional mechanisms have been discussed for tected N-phenylimide (5) as the electrophile. This compound
some pyridocarbazoles, e.g. reactivation of the lost function- can be easily prepared starting from commercially available
ality of the p53 protein.6) Based on the naturally occurring 1-methylpyrano[3,4-b]indol-3(9H)-one20) (1) by Diels–Alder
lead compounds, numerous modifications of this system have reaction with dimethyl acetylenedicarboxylate which gives
been studied,7) leading to the development of drugs like the diester (2),20,21) followed by ester hydrolysis and subse-
elliptinium,8) datelliptium,9) retelliptine,10) or pazelliptine.11) quent formation of the cyclic anhydride (3).22) Heating of the
More recently, the highly potent olivacine analog, S16020- latter with excess aniline, followed by protection of the car-
2,12,13) has attracted considerable attention due to its apparent bazole nitrogen with a benzenesulfonyl group gives 5 in
ability to circumvent P-glycoprotein-mediated multidrug re- good overall yield. For steric reasons, attack of a carbanionic
sistance.14)
reagent could be expected to take place preferentially at the
less hindered CO group of the cyclic imide structure. Indeed,
this was found to be the case: treatment of 5 with methyl-
lithium in tetrahydrofuran at ꢀ10 °C afforded the two acetyl-
carbazolecarboxamides 6a and 6b in a ratio of 1.6 : 1 in favor
of the desired isomer 6a; structural assignment is based on
nuclear Overhauser enhancement (NOE) difference spec-
troscopy. Whereas the moderate regioselectivity of this trans-
formation is compensated to some extent by the high com-
bined yield (90%), the separation of the two isomers turned
Chart 1
Among the structural variations which have been de- out to be very difficult, requiring multiple runs of medium-
scribed so far, there are several examples for incorporation of pressure liquid chromatography. Refluxing of the main prod-
an additional nitrogen atom into the tetracyclic skeleton, like uct (6a) in hydrazine hydrate for 24 h smoothly effected the
in the case of 9-aza-ellipticine15) or pazelliptine.11,16) Based desired pyridazinone ring closure as well as deprotection of
on previous investigations in the 3-aza-ellipticine series by the carbazole nitrogen to afford the key intermediate (7a) in
Landelle et al.,17) we recently reported the synthesis of sev- high yield. However, in view of the cumbersome 6a/6b sepa-
eral pyridazino[4,5-b]carbazoles, some of which exhibited ration, we decided to investigate alternative routes to this
significant antitumor activity.18) In the course of our ongoing fused pyridazinone in such a manner that all of the required
research in this field, we now became interested in the syn- carbon atoms should be introduced already at the stage of the
thesis of the hitherto unknown title molecule, 1,5-dimethyl- initial Diels–Alder reaction with the pyrone (1).
6H-pyridazino[4,5-b]carbazole, as a 3-aza isoster of oli-
Based on findings previously reported by Moody23) who
vacine, which was required as a reference compound. Until had used similar reagents, we succeeded in transforming the
now, only one method for the preparation of 1,5-dialkyl-6H- diene 1 regiospecifically into the lactone 8 in high yield
pyridazino[4,5-b]carbazoles has been described, which is (92%), using ethyl 4-hydroxy-2-pentynoate as the dienophile
based on a ring transformation of 5-acyl-11H-pyrido[4,3- in refluxing bromobenzene solution. In order to introduce the
a]carbazoles under Wolff-Kishner conditions.19) However, required ketone functionality, an attempt was made to selec-
this sequence inherently leads to 5-ethyl-substituted pyri- tively brominate the C1 atom of the fused system in analogy
∗ To whom correspondence should be addressed. e-mail: norbert.haider@univie.ac.at
© 2002 Pharmaceutical Society of Japan

1480
Vol. 50, No. 11
Chart 2
Chart 3
to syntheses of other benzo-fused bromo lactones,24,25) by
treatment of 8 with N-bromosuccinimide (NBS) and diben-
zoyl peroxide in refluxing chloroform. However, we were not
able to obtain the desired bromo lactone: employment of one
equivalent of NBS resulted in the formation of a 1 : 1 mixture
of unreacted starting material and a compound which must
be a dibrominated derivative. The yield of this dibromo com-
pound for which structure 10 is proposed (cf. Chart 3) could
be raised to about 80% by increasing the NBS amount to 2.2
eq. Although the unstable dibromo lactone could not be iso-
lated and purified, it could be easily transformed into a stable
crystalline product by prolonged refluxing in hydrazine hy-
drate. Under these conditions, the protecting group is split
off and the pyridazinone ring is formed. Interestingly, the lat-
ter was found to bear a hydroxymethyl substituent at position
1.
ment of ethyl 4-oxo-2-pentynoate as the dienophile in the
Diels–Alder reaction with 1, however at the expense of an al-
most complete loss of regioselectivity, which is not surpris-
ing in view of previous observations with similar reagents.23)
The ratio of the isomeric keto esters (12a, b) thus formed
was determined by 1H-NMR as 1.2 : 1 in favor of the desired
isomer (12a). Whereas the combined yield is very high and
the cycloaddition proceeds smoothly within only 2 h of re-
fluxing in bromobenzene solution, the main problem turned
out to be the complete failure to separate the isomers by any
means. Moreover, hydrazinolysis of the ester mixture by re-
fluxing in ethanolic hydrazine hydrate for 18 h expectedly
gave the fused pyridazinones 7a and 7b in excellent com-
bined yield, but again all attempts failed to separate 7a from
its isomer at this stage. However, careful reaction monitoring
1
of this cyclocondensation step by H-NMR revealed that the
desired pyridazinone (7a) is formed from its precursor (12a)
much faster than the isomer (7b) from the corresponding
keto ester (12b). Taking advantage of this kinetic aspect and
of the low solubility of the tetracyclic product in the reaction
medium, a convenient method for the preparation of pure 7a
Although reductive transformation of the CH2OH group in
11 into the required methyl substituent could be envisaged, a
more direct route to the target structure was sought by a
modified pathway which should avoid the need to oxidize
any carbon atom. This could be accomplished by employ-

November 2002
1481
Chart 4
Preliminary evaluation of the cytotoxic activity of 3-aza-
olivacine (14) towards four human tumor cell lines, using the
XTT26) assay (Table 1), showed significant effects only at a
relatively high concentration of 3.16 mg/ml (approx. 10ꢀ5 M),
whereas at lower concentrations (10ꢀ6 to 10ꢀ9 M) no dose-de-
pendent activity was observed. For all cell lines, growth in-
hibitory activity of 14 was found to exceed that of the 5-
monomethyl congener (15), the synthesis of which we had
reported recently.18)
In conclusion, a series of b-fused carbazoles could be
made available, utilizing the Diels–Alder reaction of 1-
methylpyrano[3,4-b]indol-3(9H)-one (1) with electron-defi-
cient acetylenic dienophiles as the key step. Employment of a
4-oxo-2-pentynoic acid ester in this cycloaddition reaction
thus permits the synthesis of the title compound (14) as a
new 3-aza bioisoster of the natural product, olivacine, in only
four steps from commercially available material. Further
structural modifications of this compound type will be car-
ried out, aiming at an enhancement of antineoplastic activity.
Table 1. Cytotoxic Activity of 3-aza-olivacine (14) and 5-Methyl-6H-pyri-
dazino[4,5-b]carbazole18) (15) (% Inhibition of Cell Growth) at a Fixed Con-
centration of 3.16 mg/ml
Cell linea)
Compound
KB
L-1210
SK-OV-3
MCF-7
14 (RꢁMe)
15 (RꢁH)
71.2
51.0
88.2
73.0
69.5
43.3
41.9
n.d.b)
a) Cell lines: KB, cervical carcinoma; L-1210, lymphatic leukemia; SK-OV-3, ovar-
ian carcinoma; MCF-7, mamma carcinoma. b) Not detectable.
was developed, simply by interrupting the hydrazinolysis
process after only 10 min of refluxing, followed by isolation
of the precipitate thus formed (see Experimental). The one-
pot sequence 1→12a, b→7a thus affords the key intermedi-
Experimental
Melting points (uncorrected) were determined on a Kofler hot-stage mi-
1
ate in 30% yield based on the commercially available starting croscope (Reichert). H-NMR spectra were recorded on a Varian UnityPlus
300 spectrometer (300 MHz). IR spectra were recorded on a Perkin–Elmer
material. After isolation of 7a, refluxing of the remaining re-
action mixture was continued for 18 h to give another precip-
itate which was found to be a mixture of 7a and 7b in a ratio
1605 FTIR or on a Perkin–Elmer Spectrum 1000 FTIR instrument. Mass
spectra were recorded on a Shimadzu QP5050A spectrometer, high-resolu-
tion mass spectra were obtained on a Finnigan MAT 8230 instrument at the
of 0.6 : 1 from which a pure sample of the latter pyridazinone
could be isolated by fractional crystallization.
Institute of Organic Chemistry, University of Vienna, and on a Micromass
Autospec instrument at the Department of Organic Chemistry, University of
Santiago de Compostela. Elemental analyses were carried out at the Institute
of Physical Chemistry (Microanalytical Laboratory), University of Vienna.
Column chromatography was performed on Merck Kieselgel 60 (0.063—
0.200 mm). Medium-pressure liquid chromatography (MPLC) was done on
Merck Lobar columns filled with LiChroprep Si 60 (0.040—0.063 mm),
with UV detection at 280 nm. For TLC, Merck aluminium sheets pre-coated
with Kieselgel 60 F254 were used.
The last two steps to the target compound are straightfor-
ward: first, the pyridazinone (7a) was converted into the
chloro compound (13) in very good yield by refluxing in
phosphorus oxychloride, and finally the chloro substituent
was reductively removed by catalytic transfer hydrogenation,
using ammonium formate as the hydrogen source. Work-up
by continuous extraction with ethyl acetate of an aqueous
suspension of the crude product gave analytically pure 14 in
satisfactory yield.
Compounds 1,20) 2,20,21) and 322) were prepared by known procedures. Sol-
vent names are abbreviated as follows: tetrahydrofuran (THF), N,N-di-
methylformamide (DMF), dimethylsulfoxide (DMSO).
3,5-Dihydro-4-methyl-2-phenylpyrrolo[3,4-b]carbazole-1(2H),3-

1482
Vol. 50, No. 11
dione27) (4) A mixture of 1-methylcarbazole-2,3-dicarboxylic acid anhy-
dride (3) (1.0 g, 3.98 mmol) and aniline (20 ml) was refluxed for 2 h. After
cooling, it was poured into ice-water (50 ml). The pH was adjusted to 1—2
by addition of concd hydrochloric acid and the product was collected by
filtration, washed with water, and dried. Recrystallization from methanol
afforded the imide 4 (1.17 g, 90%) as colorless crystals, mp 312—313 °C
(lit.27) mp 297—298 °C).
5-Benzenesulfonyl-3,5-dihydro-4-methyl-2-phenylpyrrolo[3,4-b]car-
bazole-1(2H),3-dione (5) To a stirred, ice-cooled solution of the imide 4
(2.0 g, 6.13 mmol) in a 1 : 1 mixture of toluene and DMF (50 ml) was added
sodium hydride (80% suspension; 0.28 g, 9.3 mmol) in small portions. After
10 min, benzenesulfonyl chloride (1.42 g, 8.04 mmol) was added, and the
mixture was stirred at room temperature for 4 h. Excess sodium hydride was
then decomposed with 7% sulfuric acid, and the product was extracted with
toluene. The extract was washed with water, dried, and evaporated. Recrys-
tallization of the residue from acetonitrile gave compound 5 (2.63 g, 92%) as
colorless crystals, mp 257—259 °C. 1H-NMR (DMSO-d6) d: 3.04 (3H, s, 4-
CH3), 7.20—7.31 (4H, m, phenyl-H), 7.40—7.63 (8H, m, 7-H, 8-H, phenyl-
H), 8.11 (1H, d, Jꢁ8.1 Hz, 6-H), 8.16 (1H, d, Jꢁ7.2 Hz, 9-H; shows positive
NOE on irraditiation at 8.53 ppm), 8.53 (1H, s, 10-H). IR (KBr) cmꢀ1: 1762,
1712, 1502, 1446, 1374, 1362, 1224, 1168, 1104, 750, 692, 588. MS m/z
(rel. int.): 466 (Mꢂ, 10%), 402 (4), 326 (29), 325 (82), 297 (12), 242 (20),
178 (19), 123 (38), 95 (20), 77 (100), 51 (34). Anal. Calcd for
C27H18N2O4S·0.15H2O: C, 69.11; H, 3.93; N, 5.97. Found: C, 69.04; H,
4.15; N, 6.27.
Reaction of the Imide 5 with Methyllithium A 1.4 M solution of
methyllithium in diethyl ether (1.1 ml, 1.54 mmol) was added dropwise at
ꢀ10 °C to a stirred suspension of the imide 5 (620 mg, 1.33 mmol) in dry
THF (40 ml) under an atmosphere of nitrogen. The mixture was stirred at
ꢀ10 °C for 30 min and then at room temperature for 1 h. The volatile com-
ponents were removed under reduced pressure and the solid residue was pre-
purified by short-column chromatography (eluent: ethyl acetate/light petro-
leum, 1 : 1). The mixture of the two isomeric products 6a and 6b was sepa-
rated by repeated MPLC with recycling of mixed fractions (eluent: ethyl ac-
etate/light petroleum, 1 : 1).
Evaporation of the first fraction afforded N-phenyl-3-acetyl-1-methyl-9-
benzenesulfonyl-9H-carbazole-2-carboxamide (6a) as colorless crystals
(360 mg, 55%), mp 130—131 °C (ethyl acetate/light petroleum). 1H-NMR
(DMSO-d6) d: 1.57 (3H, s, COCH3), 3.03 (3H, s, 1-CH3), 6.72 (1H, s, NH),
7.11—7.15 (2H, m, phenyl-H), 7.22—7.28 (2H, m, phenyl-H), 7.36—7.58
(8H, m, 6-H, 7-H, phenyl-H), 8.00 (1H, d, Jꢁ7.8 Hz, 5-H), 8.07 (1H, d,
Jꢁ8.1 Hz, 8-H), 8.16 (1H, s, 4-H). IR (KBr) cmꢀ1: 1700, 1676, 1496, 1368,
1186, 1172, 770, 756, 576. MS m/z (rel. int.): 482 (Mꢂ, 1%), 464 (9), 400
(6), 341 (7), 326 (10), 324 (24), 323 (100), 308 (30), 293 (11), 191 (7), 77
(34), 51 (15). HR-MS m/z: 482.1301 (Calcd for C28H22N2O4S: 482.1300).
Anal. Calcd for C28H22N2O4S·0.5H2O: C, 68.42; H, 4.72; N, 5.70. Found: C,
68.34; H, 4.93; N, 5.41.
Calcd for C16H13N3O: C, 72.99; H, 4.98; N, 15.96. Found: C, 72.65; H, 4.90;
N, 15.95.
1,4-Dimethyl-1,5-dihydro-3H-furo[3,4-b]carbazol-3-one (8) A solu-
tion of 1-methylpyrano[3,4-b]indol-3(9H)-one (1) (1.0 g, 5 mmol) and ethyl
4-hydroxy-2-pentynoate28) (1.42 g, 10 mmol) in bromobenzene (50 ml) was
refluxed under argon for 72 h. After cooling, the mixture was diluted with
ethyl acetate (50 ml) and filtered through a short column of silica (complete
elution with ethyl acetate). The residue obtained on evaporation of the volatile
components was recrystallized from toluene to give the lactone 8 (1.16 g,
92%) as almost colorless crystals, mp 233—234 °C. 1H-NMR (DMSO-d6)
d: 1.61 (3H, d, Jꢁ6.6 Hz, 1-CH3), 2.86 (3H, s, 4-CH3), 5.68 (1H, q,
Jꢁ6.6 Hz, 1-H), 7.19—7.25 (1H, m, 8-H), 7.46—7.52 (1H, m, 7-H), 7.54—
7.59 (1H, m, 6-H), 8.16 (1H, s, 10-H; shows positive NOE on irradiation at
1.61 ppm), 8.21 (1H, d, Jꢁ7.8 Hz, 9-H), 11.59 (1H, s, NH). IR (KBr) cmꢀ1
:
3289, 3058, 2973, 2926, 1729, 1628, 1511, 1349, 1325, 1306, 1249, 1033,
747. MS m/z (rel. int.): 252 (12%), 251 (Mꢂ, 69), 237 (13), 236 (80), 209
(15), 208 (100), 180 (42), 152 (15), 118 (11), 90 (15), 77 (18), 76 (22). Anal.
Calcd for C16H13NO2: C, 76.48; H, 5.21; N, 5.57. Found: C, 76.09; H, 5.14;
N, 5.48.
5-Benzenesulfonyl-1,4-dimethyl-1,5-dihydro-3H-furo[3,4-b]carbazol-
3-one (9) To a stirred, ice-cooled solution of the lactone 8 (200 mg,
0.8 mmol) in a 1 : 1 mixture of toluene and DMF (10 ml) was added sodium
hydride (80% suspension; 45 mg, 1.5 mmol) in small portions. After 10 min,
benzenesulfonyl chloride (176 mg, 1 mmol) was added and the mixture was
stirred at room temperature for 4 h. Excess sodium hydride was then decom-
posed with 7% sulfuric acid, and the product was extracted with toluene.
The extract was washed with water, dried, and evaporated. Recrystallization
of the residue from acetonitrile gave compound 9 (260 mg, 83%) as color-
less crystals, mp 235—239 °C. 1H-NMR (DMSO-d6) d: 1.59 (3H, d,
Jꢁ6.9 Hz, 1-CH3), 2.97 (3H, s, 4-CH3), 5.68 (1H, q, Jꢁ6.9 Hz, 1-H), 7.07—
7.12 (2H, m, phenyl 2-H, 6-H; shows positive NOE on irradiation at
2.97 ppm), 7.18—7.24 (2H, m, phenyl 3-H, 5-H), 7.39—7.48 (2H, 8-H,
phenyl 4-H), 7.55—7.61 (1H, m, 7-H), 7.93 (1H, d, Jꢁ7.5 Hz, 9-H), 8.04
(1H, s, 10-H), 8.08 (1H, d, Jꢁ8.4 Hz, 6-H; shows positive NOE on irradia-
tion at 7.07—7.12 ppm). IR (KBr) cmꢀ1: 3079, 2975, 2930, 1753, 1448,
1363, 1339, 1183, 1029, 772, 762, 690, 576, 559. MS m/z (rel. int.): 391
(13%), 327 (9), 251 (18), 250 (100), 208 (79), 207 (20), 180 (31), 179 (24),
178 (16), 152 (11), 77 (29), 51 (14). Anal. Calcd for C22H17NO4S: C, 67.50;
H, 4.38; N, 3.58. Found: C, 67.70; H, 4.47; N, 3.50.
3,6-Dihydro-1-hydroxymethyl-5-methyl-4H-pyridazino[4,5-b]car-
bazol-4-one (11) A solution of the lactone 9 (196 mg, 0.5 mmol) and N-
bromosuccinimide (196 mg, 1.1 mmol) in dry chloroform (10 ml) was heated
to reflux. Dibenzoyl peroxide (70%; 10 mg, 0.03 mmol) was added, and re-
fluxing was continued for 3 h. The solvent was evaporated under reduced
pressure, the residue of crude dibromo compound 10 was taken up in hy-
drazine hydrate (5 ml) and the mixture was refluxed for 18 h. It was then di-
luted with water (20 ml) and the precipitate was collected by filtration,
washed with water, and dried. The material was washed several times with
boiling methanol to afford the pure product (130 mg, 93%) as pale yellow
crystals, mp ꢃ320 °C (decomp). 1H-NMR (DMSO-d6) d: 3.16 (3H, s, 5-
CH3), 4.80 (2H, d, Jꢁ5.7 Hz, CH2OH), 5.43 (1H, t, Jꢁ5.7 Hz, CH2OH),
7.25—7.31 (1H, m, 9-H), 7.52—7.58 (1H, m, 8-H), 7.62 (1H, d, Jꢁ8.1 Hz,
7-H), 8.29 (1H, d, Jꢁ7.8 Hz, 10-H), 8.73 (1H, s, 11-H; shows positive NOE
on irradiation at 4.80 ppm), 11.72 (1H, s, carbazole NH; shows positive
NOE on irradiation at 3.16 ppm), 12.00 (1H, s, pyridazine NH). IR (KBr)
cmꢀ1: 3415, 3335, 3251, 3062, 2926, 1627, 1495, 1453, 1417, 1376, 1337,
1239, 1026, 822, 768, 754. MS m/z (rel. int.): 280 (18%), 279 (Mꢂ, 100),
278 (21), 263 (15), 250 (40), 233 (24), 205 (20), 192 (11), 164 (8), 140 (12),
102 (19), 98 (18), 88 (12), 81 (28). Anal. Calcd for C16H13N3O2: C, 68.81;
H, 4.69; N, 15.04. Found: C, 68.53; H, 4.77; N, 15.21.
Evaporation of the second fraction gave N-phenyl-2-acetyl-1-methyl-9-
benzenesulfonyl-9H-carbazole-3-carboxamide (6b) as colorless crystals
(225 mg, 35%), mp 186—188 °C (ethyl acetate/light petroleum). 1H-NMR
(DMSO-d6) d: 1.71 (3H, s, COCH3), 2.95 (3H, s, 1-CH3), 6.90 (1H, s, NH),
7.09—7.13 (2H, m, phenyl-H), 7.22—7.28 (2H, m, phenyl-H), 7.34—7.59
(8H, m, 6-H, 7-H, phenyl-H), 8.04 (1H, d, Jꢁ7.8 Hz, 5-H; shows positive
NOE on irraditiation at 8.23 ppm), 8.07 (1H, d, Jꢁ8.9 Hz, 8-H), 8.23 (1H, s,
4-H). IR (KBr) cmꢀ1: 1684, 1676, 1496, 1448, 1370, 1178, 722, 688, 576.
MS m/z (rel. int.): 482 (Mꢂ, 6%), 467 (8), 464 (10), 400 (16), 399 (13), 326
(20), 324 (26), 323 (100), 308 (29), 293 (35), 249 (16), 147 (14), 77 (64), 51
(25). HR-MS m/z: 482.1310 (Calcd for C28H22N2O4S: 482.1300). Anal.
Calcd for C28H22N2O4S: C, 69.69; H, 4.60; N, 5.81. Found: C, 69.35; H,
4.71; N, 5.59.
One-Pot Synthesis of the Pyridazinones 7a and 7b from the Pyranoin-
dole 1 A solution of 1-methylpyrano[3,4-b]indol-3(9H)-one (1) (398 mg,
2 mmol) and ethyl 4-oxo-2-pentynoate29) (560 mg, 4 mmol) in bromobenzene
(30 ml) was refluxed under argon for 2 h. The volatile components were re-
moved under reduced pressure and the oily residue consisting of the two iso-
meric esters 12a and 12b was dissolved in ethanol (8 ml). After addition of
hydrazine hydrate (300 mg, 6 mmol), the solution was refluxed for 10 min,
then it was immediately chilled. The precipitate was collected by filtration,
washed with ethanol, and dried to afford pure 7a (160 mg, 30%). The filtrate
was refluxed for another 18 h. The precipitate thus obtained was filtered off,
washed with ethanol, and dried to afford a mixture of 7a and 7b (225 mg,
43%) in a ratio of 0.6 : 1. Fractional crystallization from ethanol gave pure
2,6-dihydro-4,5-dimethyl-1H-pyridazino[4,5-b]carbazol-1-one (7b) as color-
3,6-Dihydro-1,5-dimethyl-4H-pyridazino[4,5-b]carbazol-4-one (7a)
A mixture of compound 6a (241 mg, 0.5 mmol) and hydrazine hydrate (5 ml)
was refluxed for 24 h. The excess reagent was removed under reduced pres-
sure and the residue was triturated with 1 M hydrochloric acid (10 ml). The
solid was collected by filtration, washed with water, and dried. Recrystalliza-
tion from ethanol afforded the product (116 mg, 88%) as colorless crystals,
mp 312—314 °C (decomp). 1H-NMR (DMSO-d6) d: 2.62 (3H, s, 1-CH3),
3.16 (3H, s, 5-CH3), 7.25—7.31 (1H, m, 9-H), 7.52—7.58 (1H, m, 8-H),
7.61 (1H, d, Jꢁ8.1 Hz, 7-H), 8.38 (1H, d, Jꢁ7.8 Hz, 10-H), 8.58 (1H, s, 11-
H; shows positive NOE on irradiation at 2.62 ppm), 11.71 (1H, s, carbazole-
NH; shows positive NOE on irradiation at 3.16 ppm), 11.88 (1H, s, pyri-
dazine-NH). IR (KBr) cmꢀ1: 3415, 3239, 3052, 1642, 1498, 1387, 1364,
1267, 1247, 789, 745, 726. MS m/z (rel. int.): 264 (15%), 263 (Mꢂ 100), 218
(5), 204 (4), 163 (4), 132 (3), 91 (12), 84 (7), 72 (7), 71 (13), 55 (11). Anal.
1
less crystals, mp ꢃ350 °C (decomp). H-NMR (DMSO-d6) d: 2.79 (3H, s,

November 2002
1483
4-CH3; shows positive NOE on irradiation at 2.99 ppm), 2.99 (3H, s, 5-CH3),
7.23—7.29 (1H, m, 9-H), 7.50—7.56 (1H, m, 8-H), 7.60 (1H, d, Jꢁ8.1 Hz,
7-H), 8.34 (1H, d, Jꢁ7.8 Hz, 10-H), 8.99 (1H, s, 11-H), 11.69 (1H, s, car-
bazole NH; shows positive NOE on irradiation at 2.99 ppm), 12.13 (1H, s,
pyridazine NH). IR (KBr) cmꢀ1: 3284, 3064, 2987, 1642, 1625, 1603, 1551,
1455, 1386, 1353, 1293, 1234, 1176, 795, 742, 731, 557. MS m/z (rel. int.):
264 (18%), 263 (100), 234 (19), 219 (11), 218 (26), 206 (20), 205 (12), 204
(22), 191 (15), 102 (11), 88 (8). HR-MS m/z: 263.1071 (Calcd for
C16H13N3O: 263.1059). Anal. Calcd for C16H13N3O·0.1H2O: C, 72.49; H,
5.02; N, 15.85. Found: C, 72.46; H, 4.81; N, 15.88.
Santiago de Compostela, Spain.
2) Schmutz J., Hunziker F., Pharm. Acta Helv., 33, 341—347 (1958).
3) Mosher C. W., Crews O. P., Action E. M., Goodman L., J. Med. Chem.,
9, 237—241 (1966).
4) Gribble G. W., “The Alkaloids,” Vol. 39, ed. by Brossi A., Academic
Press, New York, 1990, pp. 239—352.
5) Tewey K. M., Chen G. L., Nelson E. M., Liu L. F., J. Biol. Chem., 259,
9182—9187 (1984).
6) Ohashi M., Sugikawa E., Nakanishi N., Jpn. J. Cancer Res., 86, 819—
827 (1995).
4-Chloro-1,5-dimethyl-6H-pyridazino[4,5-b]carbazole (13)
A
mix-
7) Ohashi M., Oki T., Expert Opin. Ther. Pat., 6, 1285—1294 (1996).
8) Le Pecq J. B., Paoletti C., Nguyen Dat X., DE 2618223 (1976) [Chem.
Abstr., 86, 55620 (1977)].
9) Le Pecq J. B., Paoletti C., US 4310667 (1982) [Chem. Abstr., 96,
181489 (1982)].
ture of the pyridazinone 7a (263 mg, 1 mmol) and POCl3 (25 ml) was re-
fluxed for 3 h. The excess reagent was distilled off under reduced pressure
and the residue was treated with crushed ice (20 g). The mixture was basi-
fied by addition of concd ammonia and the precipitate was collected by fil-
tration, washed with water, and dried to afford the product (260 mg, 92%) as 10) Bisagni E., Ducrocq C., Rivalle C., Tambourin P., Wendling F., Civier
a yellow solid, mp ꢃ300 °C (decomp). 1H-NMR (DMSO-d6) d: 3.01 (3H, s,
1-CH3), 3.20 (3H, s, 5-CH3), 7.31—7.37 (1H, m, 9-H), 7.59—7.67 (2H, m,
7-H, 8-H), 8.49 (1H, d, Jꢁ8.1 Hz, 10-H), 9.02 (1H, s, 11-H; shows positive
NOE on irradiation at 3.01 ppm). IR (KBr) cmꢀ1: 3236, 3063, 2922, 1627,
1609, 1496, 1467, 1400, 1360, 1310, 1260, 745, 591. MS m/z (rel. int.): 283
A., Montagnier L., Chermann J. C., Gruest J., Lidereau R., EP 10029
(1980) [Chem. Abstr., 94, 30988 (1981)].
11) Bisagni E., Ducrocq C., Rivalle C., Tambourin P., Wendling F., Cher-
mann J. C., Montagnier L., DE 2815724 (1978) [Chem. Abstr., 90,
87427 (1979)].
(Mꢂ, 4%), 281 (Mꢂ, 10), 264 (18), 263 (100), 246 (7), 234 (9), 218 (15), 12) Guillonneau C., Pierré A., Charton Y., Guilbaud N., Kraus-Berthier L.,
204 (13), 191 (10), 132 (11), 121 (16), 109 (14), 102 (22), 95 (22), 91 (15),
Léonce S., Michel A., Bisagni E., Atassi G., J. Med. Chem., 42,
83 (16), 57 (29), 55 (29). HR-MS m/z: 281.0712 (Calcd for C16H12ClN3:
2191—2203 (1999).
281.0720). Anal. Calcd for C16H12ClN3·0.6H2O: C, 65.69; H, 4.55; N, 13) Le Mée S., Pierré A., Markovits J., Atassi G., Jacquemin-Sablon A.,
14.36. Found: C, 65.56; H, 4.29; N, 14.50.
1,5-Dimethyl-6H-pyridazino[4,5-b]carbazole (14) A mixture of the
chloro compound 13 (281 mg, 1 mmol), ammonium formate (630 mg,
Saucier J.-M., Mol. Pharmacol., 53, 213—220 (1998).
14) Pierré A., Léonce S., Pérez V., Atassi G., Cancer Chemother. Pharma-
col., 42, 454—460 (1998).
10 mmol), and 10% Pd/C catalyst (60 mg) in methanol (60 ml) was refluxed 15) Rivalle C., Ducrocq C., Bisagni E., J. Chem. Soc., Perkin Trans. 1,
under argon for 1 h. After addition of another portion of ammonium formate
(315 mg, 5 mmol), refluxing was continued for 2 h. The catalyst was filtered
off and the filtrate was evaporated under reduced pressure. The residue was
1979, 138—141 (1979).
16) Ducrocq C., Bisagni E., Rivalle C., Lhoste J. M., J. Chem. Soc., Perkin
Trans. 1, 1979, 142—145 (1979).
taken up in water (200 ml) and the mixture was basified by addition of concd 17) Landelle H., Laduree D., Cugnon de Sevricourt M., Robba M., Chem.
ammonia (2 ml). The suspension was continuously extracted with ethyl ac- Pharm. Bull., 37, 2679—2682 (1989).
etate (400 ml) for 24 h, using a rotary perforator. The extract was kept in the 18) Haider N., Jbara R., Khadami F., Wanko R., Heterocycles, 48, 1609—
refrigerator for 16 h, then the separated solid was collected by filtration, 1622 (1998).
washed with cold ethyl acetate, and dried to afford the pure product (150 mg, 19) Baradarani M. M., Joule J. A., J. Chem. Soc., Chem. Commun., 1978,
1
61%) as pale yellow crystals, mp ꢃ340 °C (decomp). H-NMR (DMSO-d6)
d: 2.95 (3H, s, 5-CH3; shows positive NOE on irradiation at 9.73 ppm), 3.03
(3H, s, 1-CH3; shows positive NOE on irradiation at 8.92 ppm), 7.27—7.33
(1H, m, 9-H), 7.55—7.64 (2H, m, 7-H, 8-H), 8.45 (1H, d, Jꢁ7.8 Hz, 10-H),
309—310 (1978).
20) Plieninger H., Müller W., Weinerth K., Chem. Ber., 97, 667—681
(1964).
21) Moody C. J., J. Chem. Soc., Perkin Trans. 1, 1985, 2505—2508
8.92 (1H, s, 11-H), 9.73 (1H, s, 4-H), 11.75 (1H, s, NH). IR (KBr) cmꢀ1
:
(1985).
3059, 2924, 2874, 2765, 1611, 1498, 1467, 1416, 1401, 1379, 1315, 1253,
22) Lancelot J. C., Landelle H., Robba M., Chem. Pharm. Bull., 32, 902—
1228, 865, 745, 590, 554. MS m/z (rel. int.): 248 (20%), 247 (Mꢂ, 100), 246
908 (1984).
(17), 219 (14), 205 (13), 189 (51), 178 (10), 151 (13), 134 (17), 124 (11), 23) Moody C. J., Shah P., J. Chem. Soc., Perkin Trans. 1, 1988, 1407—
109 (20), 102 (24), 96 (21), 94 (23), 88 (14), 82 (16), 70 (20). Anal. Calcd 1415 (1988).
for C16H13N3: C, 77.71; H, 5.30; N, 16.99. Found: C, 77.57; H, 5.43; N, 24) Sugimoto A., Tanaka H., Eguchi Y., Ito S., Takashima Y., Ishikawa M.,
17.02.
J. Med. Chem., 27, 1300—1305 (1984).
25) Neudeck H. K., Monatsh. Chem., 127, 201—217 (1996).
Acknowledgements The authors are grateful to Zentaris AG, Frank- 26) Scudiero D. A., Shoemaker R. H., Paull K. D., Monks A., Tierney S.,
furt/Germany, for performing the in-vitro evaluation of antitumor activity. E.
S. wishes to thank the University of Santiago de Compostela and the Aus-
trian Academic Exchange Service for support of his postdoctoral stay at the
University of Vienna. Experimental contributions by Mr. J. A. Maasai are
gratefully acknowledged.
Nofziger T. H., Currens M. J., Seniff D., Boyd M. R., Cancer Res., 48,
4827—4833 (1988).
27) Pindur U., Haber M., Erfanian-Abdoust H., Heterocycles, 34, 781—
790 (1992).
28) Herrmann J. L., Berger M. H., Schlessinger R. H., J. Am. Chem. Soc.,
101, 1544—1549 (1979).
References and Notes
1) Present address: Department of Organic Chemistry, Laboratory of
Medicinal Chemistry, University of Santiago de Compostela, 15782
29) Yerino L. V., Osborn M. E., Mariano P. S., Tetrahedron, 38, 1579—
1591 (1982).