Synthesis and cytotoxic activities of pyrrole[2,3-d]pyridazin-4-one derivatives

Article abstract of DOI:10.1248/cpb.50.754

1-Methyl-2-phenyl (1) and 1,3-dimethyl-2-phenyl (2) -substituted pyrrole[2,3-d]pyridazinones, as well as their tetracyclic analogues 3-6, were synthesized and evaluated in vitro by the National Cancer Institute against 60 human tumor cell lines derived fr

Full text of DOI:10.1248/cpb.50.754

754
Chem. Pharm. Bull. 50(6) 754—759 (2002)
Vol. 50, No. 6
Synthesis and Cytotoxic Activities of Pyrrole[2,3-d]pyridazin-4-one
Derivatives
Gabriele MURINEDDU,a Giorgio CIGNARELLA,b Giorgio CHELUCCI,c Giovanni LORIGA,a and
,a
*
Gérard Aimè PINNA
a Dipartimento Farmaco Chimico Tossicologico, Università di Sassari; Via F. Muroni 23, 07100 Sassari, Italy: b Istituto di
Chimica Farmaceutica e Tossicologica, Università di Milano; Viale Abruzzi 42, 20131 Milano, Italy: and c Dipartimento di
Chimica, Università di Sassari; Via Vienna 2, 07100 Sassari, Italy.
Received December 3, 2001; accepted February 28, 2002
1-Methyl-2-phenyl (1) and 1,3-dimethyl-2-phenyl (2) -substituted pyrrole[2,3-d]pyridazinones, as well as
their tetracyclic analogues 3—6, were synthesized and evaluated in vitro by the National Cancer Institute against
60 human tumor cell lines derived from nine cancer cell types. Biological results showed that the antitumor ac-
tivities of these compounds were related to the planarity of their ring systems with potency increasing in the
order 2Ͻ4Ӎ5Ͻ6Ͻ3. Among them, the most potent compound 3 showed significant cell line cytotoxicity, particu-
larly against the renal cancer subpanel [GI50 (mM) 5.07] and displayed significant potency [GI50 (mM) 3.04—4.32]
against MOLT-4, SR (leukemia), NCI-H460 (non-small cell lung), HCT-116 (colon), and SF-295 (CNS) cancer
cells, respectively.
Key words pyrrole[2,3-d]pyridazin-4-one derivative; cytotoxic activity; structure–activity relationship
1-acetyl-2-phenyl-3-methylpyrrole-4-carboxylate 13,8) fol-
The pyrrole[2,3-d]pyridazine (I) system is rarely reported
on the literature. Although the synthesis of I derivatives was
first described by Fisher more than 70 years ago,1—3) to our
knowledge only two reports have appeared since then dealing
with pyrrole[2,3-d]pyridazin-4-ones (II) as inhibitors of
human cancer cell proliferation4) and with pyrrole[2,3-d]-
pyridazin-7-ones (III) as carbohydrate-modified nucleosides
active as antiviral and antiproliferative agents.5)
Our continuing interest in the chemistry and pharmacol-
ogy of pyridazines and polycyclic congeners6) led us to incor-
porate II into tetracyclic derivatives 3—6 provided with an
almost planar structure potentially intercalating7) with DNA,
and to test them as inhibitors of human cancer cell prolifera-
tion. In addition, 1-methyl-2-phenylpyrrole[2,3-d]pyridazin-
4-one (1) and substituted 1,3-dimethyl-2-phenylpyrrole[2,3-
d]pyridazin-4-one (2) derived from 3 by removal or cleavage
of the carbon bridge between the phenyl and pyrrole moieties
were synthesized to compare the variation in biological ef-
fects due to the loss of planarity of the ring system.
lowed by N-methylation with Me2SO4 and KOH in EtOH
(Chart 5).
The synthesis of the benzocycloheptapyrrole ester 7e was
carried out by a sigmatropic rearrangement9,10) of the adduct
(16) of benzosuberone oxime 15 with methyl propiolate to
give 17, which was finally N-methylated as reported above to
7e (Chart 6). Finally, the benzo[g]indole derivative 7f was
obtained by dehydrogenation of 7d with DDQ in CH2Cl2
(Chart 7).
Results and Discussion
The tetracyclic compounds 3—6 as well as their frame-
works 1 and 2 have been submitted to a preliminary screen-
ing by the National Cancer Institute (NCI) for evaluation in
an in vitro preclinical antitumor screening program11—13)
against 60 human tumor cell lines derived from leukemia,
non-small cell lung cancer, colon cancer, central nervous sys-
tem (CNS) cancer, melanoma, ovarian cancer, renal cancer,
prostate cancer and breast cancer. The dose-response curves
for each cell line were measured in all compounds with five
different drug concentrations (10Ϫ4—10Ϫ8 M) and the concen-
A survey of the literature revealed that not only the tetra-
cyclic compounds 3—6 but also the pyrrolepyridazinones 1
and 2 were unknown.
Chemistry
The synthesis of 1 and 2 as well as of 3—6 was performed
following a common strategy. That is, formylation of the ap-
propriate esters 7a—f with Vilsmeier’s reagent in acetonitrile
afforded the corresponding 2-formyl derivatives 8a—f which
were finally condensed with hydrazine to give the desired
compounds 1—6 (Chart 3).
Chart 1
The starting esters 7 were obtained by the following proce-
dures. The synthesis of 7a, c, d was performed by condensing
the appropriate bromoketone 9a, c, d with ethyl cyanoacetate
to give the cyanoketoesters 10, which after reaction with
gaseous HCl in Et2O cyclized to the pyrrole derivatives 11.
N-Methylation of 11 with Me2SO4 and KOH in EtOH, fol-
lowed by dehalogenation of 12 with HCOONH4 and 10%
Pd–C, gave 7a, c, d (Chart 4). The 3-methyl derivative 7b
was in turn obtained by N-deacetylation of the known ethyl
Chart 2
To whom correspondence should be addressed. e-mail: pinger@uniss.it
© 2002 Pharmaceutical Society of Japan

June 2002
755
Reagents: i) (CH3)2NϩϭCHCl ClϪ, CH3CN; ii) H2NNH2·H2O 98%.
Chart 3
Reagents: i) NCCH2COOEt, K2CO3, Me2CO; ii) gaseous HCl, Et2O; iii) KOH, EtOH, Me2SO4, Me2CO; iv) HCOONH4, 10% Pd–C, MeOH.
Chart 4
Reagents: i) 2.5 M NaOH; ii) KOH, EtOH, Me2SO4, Me2CO.
Chart 5
tration causing 50% cell growth inhibition (GI50) as com- series seem to be related to the planarity of the ring system,
pared to the control was calculated (Table 1). Among the with compounds 4 (XϭCH2CH2) and 5 (XϭCH2CH2CH2)
tetracyclic structures 3—6, a significant activity was found being less active (GI50 of 26.9 and 24.5 mM, respectively).
for the indenopyrrole[2,3-d]pyridazinone 3 (XϭCH2) (GI50ϭ
However, this hypothesis is not consistent with the results
11.7 mM) and for the benzo[g]pyridazin[4,5-b]indol-7-one 6 for 1-methyl-2-phenylpyrrole[2,3-d]pyridazinone 1, which
(XϭCHϭCH) (GI50ϭ15.5 mM). In particular, compound 3 was inactive, and 1,3-dimethyl-2-phenylpyrrole[2,3-d]pyri-
exhibited significant cell line-selective cytotoxicity against dazinone 2, which possessed only weak activity (GI50ϭ36.3
the renal cancer subpanel (GI50ϭ3.46 mM) and against mM), thus revealing that planarity does not greatly influence
MOLT-4, SR (leukemia), NCI-H460 (non-small cell lung), anticellular activity.
HCT-116 (colon) and SF-295 (CNS) cancer cells, respec-
In conclusion, the synthesis and preliminary evaluation of
tively (Fig. 1). Interestingly, the inhibition properties in this the tetracyclic structures 3—6 derived from indenopyridazi-

756
Vol. 50, No. 6
Experimental
none suggest that the observed cytotoxic activity could de-
rived from intercalation of the planar structures with DNA.
On the basis of this hypothesis compounds 3 and 6 will be
selected as lead compounds for a structure–activity study in-
Chemistry, General Unless otherwise noted, all materials were ob-
tained from commercial suppliers and used without purification. Flash chro-
matography was performed using Merck Silica Gel 60 (230—400 mesh
ASTM). Thin-layer chromatography (TLC) was performed with Polygram®
1
volving the replacement of the amidic 4-carbonyl with SIL N-HR-/HV254 precoated plastic sheets (0.2 mm). H-NMR spectra were
measured in CDCl3 with superconducting FT-NMR using a XL-200 Varian
aminic side chains to increase interaction with nucleic acids.
Further research on this is in progress.
apparatus at 200 MHz.
Chemical shifts are expressed in d (ppm) downfield from internal tetra-
methylsilane (TMS) and coupling constants in Hz. Significant 1H-NMR data
are reported in the following order: multiplicity (s, singlet; d, doublet; dd,
doublet of doublets; t, triplet; q, quartet; m, multiplet), number of protons,
and coupling constants in Hz. IR spectra were recorded as thin films or
Nujol mulls on NaCl plates with a Perkin-Elmer 781 IR spectrophotometer
and are expressed in n (cmϪ1). UV–VIS spectra were recorded as ethanolic
solutions with Perkin-Elmer Lambda 5 and Hitachi U-2001 spectrophotome-
ters and the absorption wavelengths are expressed in nm followed by (log e).
Melting points were determined on a Thomas Hoover capillary melting
point apparatus and are uncorrected.
Starting benzosuberone oxime and a-bromotetral-1-ones were prepared
as described in the literature,14,15) a-bromoacetophenone, a-bromoindan-1-
one were commercially available (Aldrich Chemical Co.).
The physico-chemical data of the compounds 1—17 are reported in Ta-
bles 2 and 3.
General Alkylation Procedure for Compounds 10a, c, d A solution of
the bromo-derivative (5 mmol) in acetone (11 ml) was added dropwise to a
suspension of NCCH2CO2Et (40 mmol) and K2CO3 (10 mmol) at 40—45 °C.
The reaction mixture was stirred at 40—45 °C for 1 h and then cooled at
room temperature. After the addition of AcOEt (10 ml) and water (10 ml)
under stirring, the organic layer was separated, washed with a solution of
10% KH2PO4 (7.5 ml) and brine (5 ml), dried (Na2SO4), concentrated under
reduced pressure to give a crude oil. The oil was distilled (150 °C/1 mmHg)
to remove excess NCCH2CO2Et and the residue was purified by flash chro-
Reagents: i) Methylpropiolate, DMSO, TEA: ii) D; iii) KOH, EtOH, Me2SO4,
Me2CO.
Chart 6
matography (Petroleum ether/AcOEt, 8/2) to give the desired products
10a, c, d.
Ethyl 2-Cyano-4-oxo-4-phenylbutanoate (10a): mp: 51—52 °C [lit16)
:
53—55 °C].
Ethyl 2-Cyano-2-(1-oxo-2,3-dihydro-1H-inden-2-yl)acetate (10c): IR:
1715 (CϭO), 1745 (CϭO), 2250 (CN); UV: 247.0 (4.14), 289.5 (3.56),
294.5 (3.57); 1H-NMR [CDCl3]: 1.21 (t, 3H, CH3), 2.58—2.59 (dd, 2H,
CH2), 2.93—3.03 (dd, 2H, CH), 3.40—3.52 (dd, 1H, CH), 4.11 (q, 2H,
CH2), 7.38 (t, 1H, CH), 7.46 (d, 1H, CH), 7.60 (t, 1H, CH), 7.77 (d, 1H,
CH). Anal. Calcd for C14H13NO3: C, 69.12; H, 5.38; N, 5.75; Found: C,
68.96; H, 5.36; N, 5.71.
Reagents: i) DDQ, CH2Cl2.
Chart 7
Ethyl 2-Cyano-2-(1-oxo-1,2,3,4-tetrahydronaphthalen-2-yl)acetate (10d):
a)
Table 1. Inhibition of in Vitro Cancer Cell Lines by Pyrrolpyridazinone Derivatives 1—6 (GI50
)
RϭH (1), CH3 (2) Xϭ–CH2 (3), –(CH2)2 (4), –(CH2)3 (5), –CHϭCH (6)
1
2
3
4
5
6
Cell line
Leukemia
Non-small cell lung cancer
Colon cancer
CNS cancer
GI50 (mM)
GI50 (mM)
GI50 (mM)
GI50 (mM)
GI50 (mM)
GI50 (mM)
N.A.b)
N.A.
N.A.
N.A.
N.A.
N.A.
N.A.
N.A.
N.A.
N.A.
36.3
46.7
31.6
40.7
31.6
38.0
38.0
36.3
30.2
36.3
9.1
13.8
8.9
12.5
14.7
12.8
7.7
15.1
16.2
11.7
23.9
30.9
26.9
34.6
21.8
39.8
23.4
31.6
22.9
26.9
21.5
28.5
24.5
32.2
19.4
37.4
21.0
29.2
20.5
24.5
14.6
14.0
13.6
15.5
14.9
17.1
19.0
17.3
15.8
15.5
Melanoma
Ovarian cancer
Renal cancer
Prostate cancer
Breast cancer
Mean
a) GI50 These values are the concentrations [mM] needed to achieve 50% growth inhibition of the given human cancer cell line subpanels. Data obtained from NCI in vitro dis-
ease-oriented tumor cells screening. b) N.A.ϭNot active compound in primary anticancer assays on a three cell line panel consisting of MCF7 (breast), NCI-H460 (lung) and
SF-268 (CNS).

June 2002
757
Table 2. Physicochemical Data of the Compounds 1—6
mp (°C)a)
Yield (%)
1
2
3
4
5
6
321—322
286—287
287—288
254—256
210—212
300—301
52.9
27.3
45.5
33.3
37.0
35.5
a) All compounds were crystallized from EtOH.
Table 3. Physicochemical Data of the Compounds 7—17
mp (°C or bp/mmHg)
Yield (%)
7b
7c
7d
7e
7f
8a
8b
8c
8d
59—60/0.5
94—96
59—60/0.5
79—80/0.5
116—118
60/0.5
40/0.5
118—120
74—76
43
52
58—60
54—56
206—208
185—186
59—60
102—104
53—55
108—110
Colorless oil
124
85.6
81.9
70.6
95.6
87.9
51.6
95.5
70.0
66.7
67.6
77.5
72.3
96.5
69.8
43.0
95.6
95.2
95.3
60.0
95.0
25.0
8e
8f
10c
10d
11c
11d
12a
12c
12d
14
16
17
IR: 1680 (CϭO), 1740 (CϭO), 2250 (CN); UV: 208.7 (3.80), 238.7 (3.96);
1H-NMR [CDCl3]: 1.37 (t, 3H, CH3), 2.36 (q, 2H, CH2), 3.12 (t, 2H, CH2),
4.33 (q, 2H, CH2), 4.36 (q, 1H, CH), 4.45 (d, 1H, CH), 7.30 (d, 1H, CH),
7.34 (t, 1H, CH), 7.54 (t, 1H, CH), 8.06 (d, 1H, CH). Anal. Calcd for
C15H15NO3: C, 70.02; H, 5.87; N, 5.44; Found: C, 70.20; H, 5.89; N, 5.47.
General Procedure for Compounds 11a, c, d A solution of the
cyanoketo esters (10a, c, d) (6.7 mmol) in Et2O (30 ml for 10a, d, 15 ml for
10c), cooled at a 0—5 °C in an ice-water bath, was bubbled with gaseous
HCl (1.86 g, 5.1 mmol). The solution was stirred at room temperature for
24 h. Then excess HCl and Et2O were removed by flashing with N2. The
solid residue was triturated in MeOH to give 11a, c, d as cream-colored crys-
tals.
Ethyl 2-Chloro-5-phenyl-1H-pyrrole-3-carboxylate (11a): mp: 119 °C
[lit16): 118—120 °C].
Fig. 1. In Vitro Cytotoxic Activities of 3
Ethyl 2-Chloro-1H-indeno[1,2-b]pyrrole-3-carboxylate (11c): IR: 1670
(CϭO), 3240 (NH); UV: 226.5 (4.34), 232.0 (4.31), 285.5 (4.51), 294.5
(4.43); 1H-NMR [CDCl3]: 1.40 (t, 3H, CH3), 3.66 (s, 2H, CH2), 4.35 (q, 2H,
CH2), 7.11 (t, 1H, CH), 7.25 (t, 1H, CH), 7.42 (d, 1H, CH), 7.47 (d, 1H,
CH), 11.58 (br s, 1H, NH exchanged with D2O). Anal. Calcd for
C14H12ClNO2: C, 64.25; H, 4.62; Cl, 13.54; N, 5.35; Found: C, 64.08; H,
4.60; Cl, 13.40; N, 5.32.
Ethyl 2-Chloro-4,5-dihydro-1H-benzo[g]indole-3-carboxylate (11d): IR:
1660 (CϭO), 3210 (NH); UV: 222.5 (4.55), 233.5 (4.36), 291.5 (4.45),
296.0 (4.47), 309.0 (4.36); 1H-NMR [CDCl3]: 1.38 (t, 3H, CH3), 2.88—3.05
(m, 4H, CH2ϫ2), 4.35 (q, 2H, CH2), 7.10—7.26 (m, 4H, Ph), 8.78 (br s 1H,
NH exchanged with D2O). Anal. Calcd for C15H14ClNO2: C, 65.34; H, 5.11;
Cl, 12.85; N, 5.07; Found: C, 65.51; H, 5.14; Cl, 12.90; N, 5.09.
The numerical values listed are GI50 values, which are the molar concentrations caus-
ing 50% growth inhibition.
The solid precipitate was filtered off and the solution concentrated under re-
duced pressure to give an oil that solidified on standing.
Ethyl 1-Methyl-2-chloro-5-phenyl-1H-pyrrole-3-carboxylate (12a): IR:
1
1700 (CϭO): UV: 225.0 (4.36), 270.8 (4.32); H-NMR [CDCl3]: 1.36 (t,
3H, CH3), 3.58 (s, 3H, CH3), 4.33 (q, 2H, CH2), 6.63 (s, 1H, CH), 7.40 (m,
5H, CHϫ5). Anal. Calcd for C14H14ClNO2: C, 63.76; H, 5.35; Cl, 13.44; N,
5.31; Found: C, 63.56; H, 5.31; Cl, 13.41; N, 5.28.
Ethyl 1-Methyl-2-chloro-1,4-dihydro-indeno[1,2-b]pyrrole-3-carboxylate
1
(12c): IR: 1700 (CϭO); UV: 238.0 (3.45), 277.0 (3.84); H-NMR [CDCl3]:
1.39 (t, 3H, CH3), 3.65 (s, 2H, CH2), 3.87 (s, 3H, CH3), 4.35 (q, 2H, CH2),
7.15 (t, 1H, CH), 7.29 (t, 1H, CH), 7.44 (t, 1H, CH), 7.50 (d, 1H, CH). Anal.
Calcd for C15H14ClNO2: C, 65.34; H, 5.11; Cl, 12.87; N, 5.08; Found: C,
65.52; H, 5.12; Cl, 12.91; N, 5.10.
Ethyl 1-Methyl-2-chloro-4,5-dihydro-1H-benzo[g]indole-3-carboxylate
(12d): IR: 1700 (CϭO); UV: 223.5 (4.48), 234.0 (4.35), 296.5 (4.38), 308.0
General Procedure for Compounds 12a, c, d and 7b, e KOH 0.28 g
(4.99 mmol) was dissolved in a solution of chloroesters 11a, c, d or esters 14
and 17 (4.14 mmol) in EtOH (23 ml). The solvent was then removed under
reduced pressure and to the residue dissolved in acetone (18.6 ml) and
Me2SO4 0.78 ml (8.28 mmol) was added. The mixture was stirred at room
temperature (30 min for 11c, 1 h for 11a and 14, 3 h for 17, 4.5 h for 11d).

758
Vol. 50, No. 6
(4.35); 1H-NMR [CDCl3]: 1.40 (t, 3H, CH3), 2.16 (s, 3H, CH3), 3.77 (s, 3H,
CH3), 4.40 (q, 2H, CH2), 7.25—7.30 (m, 2H, CHϫ2), 7.45—7.52 (m, 3H,
CHϫ3), 10.37 (s, 1H, CH). Anal. Calcd for C16H17NO3: C, 70.83; H, 6.31;
N, 5.16; Found: C, 70.48; H, 6.30; N, 5.14.
1
(4.29); H-NMR [CDCl3]: 1.38 (t, 3H, CH3), 2.82—2.98 (m, 4H, CH2ϫ2),
3.88 (s, 3H, CH3), 4.34 (q, 2H, CH2), 7.13—7.41 (m, 4H, CHϫ4). Anal.
Calcd for C16H16ClNO2: C, 66.32; H, 5.56; Cl, 12.23; N, 4.83; Found: C,
66.05; H, 5.52; Cl, 12.10; N, 4.80.
Ethyl 1-Methyl-2-formyl-1,4-dihydro-indeno[1,2-b]pyrrole-3-carboxylate
(8c): IR: 1640 (CϭO), 1700 (CϭO); UV: 268.0 (4.08), 281.0 (3.99), 291.0
(3.87), 355.5 (4.35); 1H-NMR [CDCl3]: 1.43 (t, 3H, CH3), 3.76 (s, 2H, CH2),
4.33 (s, 3H, CH3), 4.41 (q, 2H, CH2), 7.30—7.40 (m, 2H, CHϫ2), 7.54 (d,
1H, CH), 7.64 (d, 1H, CH), 10.41 (s, 1H, CH). Anal. Calcd for C16H15NO3:
C, 71.36; H, 5.61; N, 5.20; Found: C, 71.18; H, 5.59; N, 5.18.
Ethyl 1-Methyl-2-formyl-4,5-dihydro-1H-benzo[g]indole-3-carboxylate
(8d): IR: 1655 (CϭO), 1700 (CϭO), UV: 226.0 (4.78), 234.0 (4.65), 253.0
(4.24), 294.0 (4.61), 307.0 (4.47); 1H-NMR [CDCl3]: 1.40 (t, 3H, CH3),
2.87—3.00 (m, 4H, CH2ϫ2), 4.25 (s, 3H, CH3), 7.25—7.35 (m, 3H,
CHϫ3), 7.60 (d, 1H, CH), 10.38 (s, 1H, CH). Anal. Calcd for C17H17NO3: C,
72.07; H, 6.04; N, 4.94; Found: C, 72.35; H, 6.07; N, 4.96.
Ethyl 1,4-Dimethyl-5-phenyl-1H-pyrrole-3-carboxylate (7b): IR: 1705
(CϭO); UV: 235.0 (4.39), 271.0 (4.36); 1H-NMR [CDCl3]: 1.34 (t, 3H,
CH3), 2.21 (s, 3H, CH3), 3.49 (s, 3H, CH3), 4.30 (q, 2H, CH2), 7.29—7.31
(m, 3H, CHϫ3), 7.36—7.50 (m, 3H, CHϫ3). Anal. Calcd for C15H17NO2:
C, 74.05; H, 7.04; N, 5.75; Found: C, 73.85; H, 7.01; N, 5.72.
Methyl 1-Methyl-1,4,5,6-tetrahydrobenzo[6,7]cyclohepta[1,2-b]pyrrole-3-
carboxylate (7e): IR: 1695 (CϭO); UV: 240.0 (3.77), 245.0 (3.75), 306.5
(4.55); 1H-NMR [CDCl3]: 1.37 (t, 3H, CH3), 2.13 (q, 2H, CH2), 2.35 (t, 2H,
CH2), 2.54 (t, 2H, CH2), 3.96 (s, 3H, CH3), 4.32 (q, 2H, CH2), 6.90 (s, 1H,
CH), 7.25—7.35 (m, 4H, CHϫ4). Anal. Calcd for C16H17NO2: C, 75.25; H,
6.71; N, 5.48; Found: C, 75.10; H, 6.65; N, 5.45.
General Procedure for Compounds 7a, c, d To
a solution of
Methyl 1-Methyl-2-formyl-1,4,5,6-tetrahydrobenzo[6,7]cyclohepta[1,2-
b]pyrrole-3-carboxylate (8e): IR: 1650 (CϭO), 1700 (CϭO); UV: 226.0
(4.74), 230.0 (4.45), 251.5 (4.29), 297.0 (4.50); H-NMR [CDCl3]: 1.35 (t,
3H, CH3), 2.15 (q, 2H, CH2), 2.37 (t, 2H, CH2), 2.58 (t, 2H, CH2), 3.97 (s,
3H, CH3), 4.33 (q, 2H, CH2), 7.26—7.34 (m, 4H, CHϫ4), 10.30 (s, 1H,
CH). Anal. Calcd for C18H19NO3: C, 72.70; H, 6.44; N, 4.71; Found: C,
72.63; H, 6.42; N, 4.70.
chloroesters 12a, c, d (2.72 mmol) and HCOONH4 0.86 g (13.6 mmol) in
MeOH (22 ml) 0.18 g of 10% Pd–C was added. The mixture was stirred
under nitrogen (2 h for 12a, d and 4 h for 12c) and then filtered on Celite and
the solution concentrated. The oily residue was taken up with water and ex-
tracted with ethyl ether. The organic layer was separated, dried (Na2SO4),
concentrated under reduced pressure to give an oil (7a, d) or a solid (7c).
Ethyl 1-Methyl-5-phenyl-1H-pyrrole-3-carboxylate (7a): bp: 145 °C/
0.5 mmHg [lit17): 148 °C/0.5 mmHg].
1
Ethyl 1-Methyl-2-formyl-1H-benzo[g]indole-3-carboxylate (8f): IR: 1660
(CϭO), 1700 (CϭO); UV: 226.0 (4.70), 243.0 (4.65), 253.0 (4.24), 292.5
Ethyl 1-Methyl-1,4-dihydro-indeno[1,2-b]pyrrole-3-carboxylate (7c): IR:
1
1
(4.7), 307.3 (4.46); H-NMR [CDCl3]: 1.43 (t, 3H, CH3), 2.88—2.98 (m,
1695 (CϭO); UV: 234.5 (4.27), 282.5 (4.39); H-NMR [CDCl3]: 1.37 (t,
4H, CH2ϫ2), 4.24 (s, 3H, CH3), 7.45—7.54 (m, 2H, CHϫ2), 7.64 (d, 1H,
CH, JABϭ8.2 Hz), 7.96 (d, 1H, CH), 8.30 (d, 1H, CH, JABϭ8.2 Hz), 8.40 (d,
1H, CH), 10.35 (s, 1H, CH). Anal. Calcd for C17H15NO3: C, 72.58; H, 5.37;
N, 4.98; Found: C, 72.62; H, 5.38; N, 5.00.
3H, CH3), 3.66 (s, 2H, CH2), 3.93 (s, 3H, CH3), 4.32 (q, 2H, CH2), 7.16 (d,
1H, CH), 7.27 (s, 1H, CH), 7.30 (t, 1H, CH), 7.44 (d, 1H, CH), 7.50 (d, 1H,
CH). Anal. Calcd for C15H15NO2: C, 74.66; H, 6.26; N, 5.80; Found: C,
74.80; H, 6.29; N, 5.82.
Methyl (E,E)/(Z,E)-3-[(6,7,8,9-Tetrahydro-5H-benzo-[a]cyclohepten-
5-ylidenamido)oxy]-2-propenoate (16) To a solution of the benzo-
suberone oxime 1514) (6.2 mmol) and a few drops of triethylamine in anhy-
drous dimethyl sulfoxide (DMSO) (6 ml), a solution of methylpropiolate
(12.4 mmol) in anhydrous DMSO (2.5 ml) was added dropwise at room tem-
perature and in the presence of drops of triethylamine. The reaction mixture
was heated to 65—70 °C for 24 h. The cooled reaction solution was poured
into crushed ice and the aqueous layer was extracted with CH2Cl2. The or-
ganic phase was collected, washed with water, dried (Na2SO4) and concen-
trated to yield a brown residue. Purification by flash chromatography (petro-
leum ether/AcOEt, 8/2) afforded the desired O-vinyl oxime ether 16 as a
mixture of (E,E) and (Z,E) isomers.
IR: 1740 (CϭO), 1644 (CϭN), 1602 (CϭC); UV: 257.8 (4.50), 206.4
(4.38); 1H-NMR [CDCl3]: 1.57—1.87 (m, 4H, CH2ϫ2), 2.68—2.84 (m, 4H,
CH2ϫ2), 3.72 and 3.74 (sϫ2, 3H, CH3, of (Z,E) and (E,E) isomers), 6.23
and 6.90 (q ABϫ2, 2H, JABϭ7.4, 12.80 Hz, C2H and C3H of (Z,E) and
(E,E) isomers), 7.14—7.44 (m, 4H, CHϫ4). Anal. Calcd for C15H17NO3: C,
69.48; H, 6.60; N, 5.40; Found: C, 69.32; H, 6.58; N, 5.38.
Ethyl 1-Methyl-4,5-dihydro-1H-benzo[g]indole-3-carboxylate (7d): IR:
1700 (CϭO); UV: 226.0 (4.78), 234.0 (4.65), 253.0 (4.24), 294.0 (4.61),
307.0 (4.47); H-NMR [CDCl3]: 1.35 (t, 3H, CH3), 2.86 (t, 2H, CH2), 2.98
(t, 2H, CH2), 3.93 (s, 3H, CH3), 4.34 (q, 2H, CH2), 7.10—7.30 (m, 4H,
CHϫ4), 7.43 (d, 1H, CH). Anal. Calcd for C16H17NO2: C, 75.25; H, 6.71; N,
5.48; Found: C, 75.07; H, 6.68; N, 5.46.
1
Ethyl 4-Methyl-5-phenyl-1H-pyrrole-3-carboxylate (14) A suspen-
sion of N-acetyl-pyrrolester 138) (1.10 mmol) in 2.5 M NaOH (3 ml) was re-
fluxed for 30 min. The solution was cooled at room temperature and the pre-
cipitate was filtered off, washed with water, dried to give the ester 14. IR:
1675 (CϭO), 3270 (NH); UV: 235.0 (4.39), 271.0 (4.36); 1H-NMR
[CDCl3]: 1.36 (t, 3H, CH3), 2.43 (s, 3H, CH3), 4.32 (q, 2H, CH2), 7.41—
7.47 (m, 6H, CHϫ6), 8.42 (br s, 1H, NH exchanged with D2O). Anal. Calcd
for C14H15NO2: C, 73.34; H, 6.59; N, 6.10; Found: C, 73.07; H, 6.57; N,
6.08.
Ethyl 1-Methyl-1H-benzo[g]indole-3-carboxylate (7f) To a solution
of the N-methyl-ester 7d (2.66 mmol) in CH2Cl2 (10 ml) 1.81 g (7.98 mmol)
of 2,3-dichloro-5,6-dicyanobenzoquinone (DDQ) was added and the mixture
was stirred at room temperature for 5 min. The solvent was evaporated and
the residue was purified by flash chromatography on Al2O3 (EtPet/AcOEt
8/2) to give 7f as a solid, mp 116—118 °C. IR: 1690 (CϭO); UV: 226.3
(4.75), 236.0 (4.70), 255.0 (4.47); 1H-NMR [CDCl3]: 1.43 (t, 3H, CH3), 4.24
(s, 3H, CH3), 4.41 (q, 2H, CH2), 7.45—7.54 (m, 2H, CHϫ2), 7.64 (d, 1H,
CH, JABϭ8.2 Hz), 7.70 (s, 1H, CH), 7.96 (d, 1H, CH), 8.30 (d, 1H, CH,
Methyl 1,4,5,6-Tetrahydrobenzo[6,7]cyclohepta[b]pyrrole-3-carboxy-
late (17) A neat mixture of the (E,E) and (Z,E) isomers 16 (3.6 mmol) was
heated to 130 °C for 15 h to give a brown residue. Purification by flash chro-
matography (petroleum ether/AcOEt 8/2) afforded the desired product 17.
IR: 3300 (NH), 1680 (CϭO), 1600 (CϭC); UV: 309.8 (4.15), 235.2
1
(3.81), 231.0 (3.79), 204.8 (4.01); H-NMR [CDCl3]: 1.97—2.15 (m, 2H,
CH2), 2.76—2.86 (m, 4H, CH2ϫ2), 3.86 (s, 3H, CH3), 6.81 (d, 1H,
Jϭ2.2 Hz, C2H), 7.16—7.50 (m, 4H, CHϫ4), 9.11 (br s, 1H, NH exchanged
with D2O). Anal. Calcd. for C15H15NO2: C, 74.66; H, 6.26; N, 5.80; Found:
C, 74.80; H, 6.43; N, 5.40.
J
ABϭ8.2 Hz), 8.40 (d, 1H, CH). Anal. Calcd for C16H16NO2: C, 75.86; H,
6.36; N, 5.53; Found: C, 76.01; H, 6.39; N, 5.55.
General Procedure for Compounds 8a—f To a solution of pyrrole
ester 7a—f (4.71 mmol) in CH3CN (11.5 ml), N-NЈ-dimethylchloromethyl-
ene ammonium chloride (Vilsmeier’s reagent) (6.11 mmol) was added and
the suspension was stirred at room temperature (1.5 h for 7b, c, d, 2 h for 7a,
3 h for 7e, f). The solvent was concentrated under reduced pressure and the
oily residue was taken up with saturated aqueous NaHCO3. The aqueous
layer was extracted with AcOEt, the organic layer was washed (H2O), dried
(Na2SO4) and evaporated under reduced pressure to yield alternatively an oil
(8a, b) purified by bulb-to-bulb distillation under a vacuum, or a solid (8c—
f) purified by flash chromatography.
Ethyl 1-Methyl-2-formyl-5-phenyl-1H-pyrrole-3-carboxylate (8a): IR:
1660 (CϭO), 1715 (CϭO); UV: 247.3 (4.28), 269.1 (4.14), 247.2 (4.11),
325.1 (4.38), 340.5 (4.28); 1H-NMR [CDCl3]: 1.38 (t, 3H, CH3), 3.93 (s, 3H,
CH3), 4.38 (q, 2H, CH2), 6.74 (s, 1H, CH), 7.37—7.51 (m, 5H, CHϫ5),
10.47 (s, 1H, CH). Anal. Calcd for C15H15NO3: C, 70.02; H, 5.87; N, 5.44;
Found: C, 70.22; H, 5.89; N, 5.48.
General Ring Closure Procedure with Hydrazine Hydrate for Com-
pounds 1—6
A mixture of formyl ester 8a—f (1.85 mmol) in
H2NNH2·H2O (98%, 1.80 ml, 37.1 mmol) was refluxed for 0.25—4 h (0.25 h
for 8c, 1 h for 8a, b, d, 4 h for 8e, f) and then poured onto ice. The solid pre-
cipitate was filtered and washed with H2O to give a crude product that was
purified by trituration with ethyl ether to yield the expected products 1—6.
1-Methyl-2-phenyl-1H-pyrrol[2,3-d]pyridazin-4(5H)-one (1): IR: 1630
(CϭN), 1660 (CϭO), 3210 (NH); UV: 232.3 (4.336), 304.1 (4.295); 1H-
NMR [CDCl3]: 3.84 (s, 3H, CH3), 6.81 (s, 1H, CH), 7.47—7.58 (m, 5H,
CHϫ5), 8.20 (s, 1H, CH), 12.08 (br s, 1H, NH exchanged with D2O). Anal.
Calcd for C13H11N3O: C, 69.37; H, 4.91; N, 18.65; Found: C, 69.20; H, 4.89;
N, 18.50.
1,3-Dimethyl-2-phenyl-1H-pyrrol[2,3-d]pyridazin-4(5H)-one (2): IR:
1640 (CϭO), 3280 (NH); UV: 278.0 (4.043), 310.0 (4.398); 1H-NMR
[CDCl3]: 2.42 (s, 3H, CH3), 3.68 (s, 3H, CH3), 7.33—7.54 (m, 5H, CHϫ5),
8.07 (s, 1H, CH), 10.25 (br s, 1H, NH exchanged with D2O). Anal. Calcd for
Ethyl 1,4-Dimethyl-2-formyl-5-phenyl-1H-pyrrole-3-carboxylate (8b):
IR: 1665 (CϭO), 1700 (CϭO); UV: 249.0 (4.08), 278.0 (3.95), 331.0

June 2002
759
C14H13N3O: C, 70.27; H, 5.47; N, 17.56; Found: C, 70.57; H, 5.50; N, 17.59.
5-Methyl-5,10-dihydroindenopyrrol[2,3-d]pyridazin-1(2H)-one (3): IR:
1650 (CϭO); UV: 235.5 (4.631), 243.5 (4.085), 313.0 (4.329), 327.0
(1932).
4) Marquet J. P., Bisagni E., Andre-Louisfert J., Chim. Ther., 3, 348—355
(1968).
5) Meade E. A., Wotring L. L., Drach J. C., Townsend L. B., J. Med.
Chem., 40, 794—801 (1997).
6) Cignarella G., Barlocco D., Pinna G. A., Murineddu G., 7th Interna-
tional Symposium on the Chemistry and Pharmacology of Pyri-
dazines, IL-8, Santiago de Compostela, September 13—16, 2000.
7) Denny W. A., “Cancer Chemotherapeutic Agents,” ed. by Foye W. O.,
American Chemical Society, Washington, DC, 1995, pp. 218—239.
8) Gabbutt C. D., Hepwoth J. D., Heron B. M., Elsegood M. R. J., Clegg
W., Chem. Commun., 1999, 289—290 (1999).
1
(4.653), 340.0 (4.779); H-NMR [CDCl3]: 2.80 (s, 2H, CH2), 4.15 (s, 3H,
CH3), 7.29 (t, 1H, CH), 7.39 (t, 1H, CH), 7.58 (d, 2H, CHϫ2), 8.23 (s, 1H,
CH), 12.10 (br s, 1H, NH exchanged with D2O). Anal. Calcd for C14H11N3O:
C, 70.87; H, 4.67; N, 17.71; Found: C, 71.10; H, 4.71; N, 17.73.
11-Methyl-5,6-dihydrobenzo[g]pyridazin[4,5-b]indol-7(8H)-one (4): IR:
1650 (CϭO), 3170 (NH); UV: 240.5 (4.600), 276.5 (5.084), 318.0 (4.609),
331.0 (4.705); 1H-NMR [CDCl3]: 2.94 (t, 2H, CH2), 3.16 (t, 2H, CH2), 4.09
(s, 3H, CH3), 7.26—7.39 (m, 3H, CHϫ3), 7.62 (d, 1H, CH), 8.09 (s, 1H,
CH), 12.27 (br s, 1H, NH exchanged with D2O). Anal. Calcd for C15H13N3O:
C, 71.70; H, 5.21; N, 16.72; Found: C, 71.98; H, 5.23; N, 16.73.
9) Pinna G. A., Pirisi M. A., Paglietti G., J. Chem. Res. (S), 1990, 360—
361 (1990).
12-Methyl-5,6,7,12-tetrahydrobenzo[3Ј,4Ј]cyclohepta[1Ј,2Ј-4,5]pyrrol-
[2,3-d]pyridazin-8(9H)-one (5): IR: 1645 (CϭO), 3160 (NH); UV: 227.5 10) Pinna G. A., Pirisi M. A., Paglietti G., J. Chem. Res. (M), 1990,
(4.950), 236.5 (4.924), 301.0 (4.609), 331.0 (5.005); 1H-NMR [CDCl3]: 2.18
(q, 2H, CH2), 2.39 (t, 2H, CH2), 2.58 (t, 2H, CH2), 4.02 (s, 3H, CH3), 7.28—
7.38 (m, 5H, CHϫ5), 8.60 (s, 1H, CH), 12.20 (br s, 1H, NH exchanged with
D2O). Anal. Calcd for C16H15N3O: C, 72.43; H, 5.69; N, 15.83; Found: C,
72.10; H, 5.67; N, 15.80.
2777—2795 (1990).
11) 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., J. Natl. Cancer Inst., 83, 757—766
(1991).
11-Methylbenzo[g]pyridazin[4,5-b]indol-7(8H)-one (6): IR: 1645 (CϭO),
3160 (NH); UV: 257.5 (4.658), 275.5 (5.084), 318.0 (4.589), 331.0 (4.705);
1H-NMR [CDCl3]: 4.28 (s, 3H, CH3), 7.47—7.56 (m, 2H, CHϫ2), 7.66 (d,
12) Paull K. D., Shoemaker R. H., Hods L., Monks A., Scudiero D. A.,
Rubinstein L., Plowman J., Boyd M. R., J. Natl. Cancer Inst., 81,
1088—1092 (1989).
1H, CH, JABϭ8.2 Hz), 7.97 (d, 2H, CHϫ2), 8.31 (d, 1H, CH, JABϭ8.2 Hz), 13) Boyd M. R., Paull K. D., Rubinstein L. R., “Cytotoxic Anticancer
8.35 (d, 1H, CH), 8.40 (s, 1H, CH), 12.30 (br s, 1H, NH exchanged with
D2O). Anal. Calcd for C15H11N3O: C, 72.27; H, 4.45; N, 16.85; Found: C,
71.97; H, 4.41; N, 16.83.
Drugs: Models and Concept for Drug Discovery,” ed. by Valeriote F.
A., Corbett T., Baker L., Kluwer Academic Publishers, Amsterdam,
1992, pp. 11—25.
14) Sinha A. K., Rastogi S. N., Das S. R., Indian J. Chem., Sect. B, 30B,
1041—1046 (1991).
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