4-Phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione inhibitors of the checkpoint kinase Wee1. structure-activity relationships for chromophore modification and phenyl ring substitution

Article abstract of DOI:10.1021/jm0512591

High-throughput screening has identified a novel class of inhibitors of the checkpoint kinase Wee1, which have potential for use in cancer chemotherapy. These inhibitors are based on a 4-phenylpyrrolo[3,4-c]-carbazole-1,3(2H,6H)- dione template and have been shown by X-ray crystallography to bind at the ATP site of the enzyme. An extensive study of the effects of substitution around this template has been carried out, which has identified substituents which lead to improvements in potency and selectivity for Wee1. While retention of the maleimide ring and pendant 4-phenyl group is necessary for potency, replacement of the carbazole nitrogen by oxygen is well tolerated and results in improved Wee1 selectivity against the related checkpoint kinase Chk1. Wee1 potency and selectivity are also enhanced by the incorporation of lipophilic functionality at the 2′-position of the 4-phenyl ring, and Wee1 selectivity against Chk1 is favored by C3-C5 alkyl substitution of the carbazole nitrogen. These studies provide a basis for the design of active analogues of the pyrrolocarbazole lead with improved physical properties.

Full text of DOI:10.1021/jm0512591

4896
J. Med. Chem. 2006, 49, 4896-4911
4-Phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione Inhibitors of the Checkpoint Kinase Wee1.
Structure-Activity Relationships for Chromophore Modification and Phenyl Ring Substitution
Brian D. Palmer,*,† Andrew M. Thompson,† R. John Booth,‡ Ellen M. Dobrusin,‡ Alan J. Kraker,‡ Ho H. Lee,†
Elizabeth A. Lunney,§ Lorna H. Mitchell,‡ Daniel F. Ortwine,‡ Jeff B. Smaill,† Leesa M. Swan,† and William A. Denny†
Auckland Cancer Society Research Centre, School of Medical Sciences, The UniVersity of Auckland, PriVate Bag 92019,
Auckland 1020, New Zealand, Ann Arbor Laboratories, Pfizer Global Research and DeVelopment, 2800 Plymouth Road,
Ann Arbor, Michigan 48106-1047, and La Jolla Laboratories, Pfizer Global Research and DeVelopment, 10646 Science Center DriVe,
La Jolla, California 92121
ReceiVed December 18, 2005
High-throughput screening has identified a novel class of inhibitors of the checkpoint kinase Wee1, which
have potential for use in cancer chemotherapy. These inhibitors are based on a 4-phenylpyrrolo[3,4-c]-
carbazole-1,3(2H,6H)-dione template and have been shown by X-ray crystallography to bind at the ATP
site of the enzyme. An extensive study of the effects of substitution around this template has been carried
out, which has identified substituents which lead to improvements in potency and selectivity for Wee1.
While retention of the maleimide ring and pendant 4-phenyl group is necessary for potency, replacement of
the carbazole nitrogen by oxygen is well tolerated and results in improved Wee1 selectivity against the
related checkpoint kinase Chk1. Wee1 potency and selectivity are also enhanced by the incorporation of
lipophilic functionality at the 2′-position of the 4-phenyl ring, and Wee1 selectivity against Chk1 is favored
by C3-C5 alkyl substitution of the carbazole nitrogen. These studies provide a basis for the design of
active analogues of the pyrrolocarbazole lead with improved physical properties.
Introduction
The protein tyrosine kinase enzyme Wee1 is involved in
regulation of the DNA damage-sensitive G2/M checkpoint
in the eukaryotic cell cycle, through its inhibitory phosphory-
lation of cdc2 on tyrosine 15.1-3 Many cancer cells lack a
functional p53 signaling pathway, which means that the other
significant DNA damage-sensitive checkpoint, G1/S, is not
controlled. Inhibitors of Wee1 should abrogate the G2/M
checkpoint and should preferentially enhance the cyto-
toxic effects of DNA damaging agents on p53-nega-
tive cells by allowing them to bypass both of the check-
points where damaged cells normally arrest to allow time for
DNA repair. Coadministration of a Wee1 inhibitor with a
conventional DNA-damaging cytotoxic agent in the clinic could
therefore lead to improvements in responses to cancer
therapy.4-6
We recently reported on the development of 6-phenylpyrido-
[2,3-d]pyrimidin-7(8H)-ones (e.g., 1, 2) as inhibitors of Wee1,7
and others have subsequently also noted Wee1 activity for this
template.8,9 Several of these were found to be potent Wee1
inhibitors (IC50 < 100 nM), but this activity could not be
divorced from an even more potent inhibition of the kinase
c-Src. Nevertheless, one of the more effective compounds (1)
from this series was found to sensitize p53 mutant cells to the
DNA damage induced by radiation.10 In a high-throughput
screening (HTS) program, we subsequently found that the
4-phenylpyrrolocarbazole 3 was an equally potent inhibitor of
Wee1 (IC50 ) 97 nM) while proving inactive against c-Src (IC50
> 50 µM).
Although the HTS lead 3 was originally prepared in a
program directed toward the development of inhibitors of the
* To whom correspondence should be addressed. Phone: +64
9 3737 599, ext 86150. Fax: +64 9 3737 502. E-mail b.palmer@
auckland.ac.nz.
protein kinase C family of serine-threonine kinases,11 it had
only low activity against these. The lead 3 was also poorly active
against a series of receptor tyrosine kinases and cyclin-dependent
kinases. However, 3 did display potent inhibition of Chk1
† The University of Auckland.
‡ Ann Arbor Laboratories, Pfizer Global Research and Development.
§ La Jolla Laboratories, Pfizer Global Research and Development.
10.1021/jm0512591 CCC: $33.50 © 2006 American Chemical Society
Published on Web 07/15/2006

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4897
Scheme 2a
Scheme 1a
a Reagents and conditions: (i) [Ph3PCH2Ar]X, LDA, THF, reflux, 5 h;
(ii) NaH, MeI, 20 °C, 15 min; (iii) maleimide, 175 °C, melt, 2-5 h; or
maleimide, toluene, catalyst SnCl2, reflux, 3 h to 6 days; (iv) MnO2, dioxane,
reflux, 5-24 h; or DDQ, toluene-dioxane, reflux, 6-48 h; (v) BBr3,
CH2Cl2, 20 °C, 2-48 h; or PyHCl, 200 °C, 15 min.
a Reagents and conditions: (i) Ph3PdCHCO2CH2Ph, CH2Cl2, 20 °C, 4
h; (ii) maleimide, 175 °C, 3 h; (iii) MnO2, dioxane, reflux, 2 h; (iv) H2/
Pd-C, DMF-MeOH, 60 psi, 2 h; (v) DPPA, Et3N, t-BuOH, reflux 16 h;
then TFA, DCM, 20 °C, 1 h; (vi) aqueous H2SO4, NaNO2, 3 °C, 3 min;
then KI, CuI, 70 °C, 1 h; (vii) PyHCl, 200 °C, 15 min; (viii) ArB(OH)2,
catalyst PdCl2(dppf), 2 N Na2CO3-dioxane, reflux 4 h.
kinase, another enzyme involved in regulation of the G2/M
checkpoint.12 In fact, inhibitors of Chk1 have also been proposed
to sensitize cells to DNA-damaging agents in a manner similar
to Wee1 inhibition.13-15 While the dual Wee1/Chk1 inhibitory
profile of this current series could ultimately prove to be
advantageous, the initial goal of the present work was to prepare
a series of selective Wee1 inhibitors so that their in vitro and
in vivo effects could be evaluated in the absence of other
confounding activities. The best DNA-damaging agent(s) for
use in conjunction with a Wee1 inhibitor may well turn out to
be different from those for use with a Chk1 inhibitor, and the
optimum relative timing of administration of the checkpoint
abrogator and DNA-damaging agent may not be the same for
Wee1 and Chk1 inhibitors.
The synthesis of a series of related compounds lacking the
9-hydroxyl group as potential protein kinase C inhibitors has
been reported,16 but no biological data were provided.
In this paper we report on the synthesis and SAR of
derivatives of 3, defining structural features that are important
for Wee1 activity and identifying substituents in the chro-
mophore that lead to enhanced Wee1 potency and selectivity
against Chk1. In particular, we report results from a compre-
hensive study of the effects of substitutions in the 4-phenyl ring
on the activity of this novel class of Wee1 inhibitor.
carried out using either excess activated MnO2 or 2-5 equiv
of DDQ to give V. This was demethylated to give the VI of
Tables 1 and 2 using either pyridine hydrochloride at 200 °C
or 2-10 equiv of BBr3 in CH2Cl2 at room temperature.
Analogous chemistry starting from a benzofuran-2-carboxalde-
hyde or a benzothiophene-2-carboxaldehyde gave the O- and
S-analogues 8, 9, and 26 of Table 1. Additional examples of
analogues substituted in the 4-phenyl ring, or with this ring
replaced by a heterocycle, were prepared by efficient Suzuki
coupling of the 4-iodide 5 with substituted arylboronic acids
(Scheme 2). This final coupling step was also carried out in a
parallel manner using commercially available boronic acids,
which enabled a large number of compounds, VII, to be
prepared. Wittig reaction of aldehyde 106 with benzyl (triph-
enylphosphoranylidene)acetate gave the E-diene 107 as the sole
product. This reacted in a Diels-Alder reaction with maleimide
at 175 °C to give 108, which was aromatized and converted to
the acid 110 in good yield. Curtius rearrangement of 110 gave
amino compound 111, which was converted into iodide 112 in
a moderate yield of 46% via diazotization. Attempts to introduce
a halogen substituent directly from 110 by halodecarboxylation16
were unsuccessful.
Compounds containing amino functionality in the 4-phenyl
ring were generally prepared by reduction of the corresponding
nitro compounds, usually using nickel boride to preserve any
halogen substituent also in this ring. Compounds containing 2′-
methoxy substitution in the 4-phenyl ring could usually be
selectively demethylated at the 9-methoxy group in the A-ring
with BBr3, leaving the former group intact. Under forcing
conditions both methoxy groups were cleaved, generating
hydroxy-substituted 4-phenyl groups.
Chemistry
The 4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione ring
system was built up as shown in Scheme 1. Substituted
benzyltriphenylphosphonium halide salts were prepared from
known substituted benzyl bromides (or benzyl alcohols follow-
ing bromination) by reaction with triphenylphosphine. These
were reacted in a Wittig condensation with indole-2-carboxal-
dehydes I to give dienes II, usually as an E/Z mixture of alkene
isomers. This diene could be alkylated on the indole nitrogen
to give III. A Diels-Alder reaction of the dienes II or III with
maleimide was best performed as a neat melt at 175 °C or as a
solution in refluxing toluene (with SnCl2 catalyst) to give adduct
IV as a mixture of diastereomers. Aromatization of IV was
The 2′-iodo and 2′-hydroxy compounds 29 and 38 were
obtained via diazotization of the 9-methoxy derivative 139 of
the amino compound 44, followed by reaction with KI and CuI,
which gave both a 2′-iodo and a 2′-hydroxyl derivative (140
and 147, respectively). Demethylation of these gave 29 and 38,
respectively. The primary amide 36 was prepared by partial

4898 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
Scheme 3a
Scheme 4a
a Reagents and conditions: (i) NaBH4, EtOH, 20 °C, 18 h; (ii) TsOH,
MeOH, 20 °C, 30 min; (iii) PhSeH, TsOH, THF, 20 °C, 1 h.
Scheme 5a
a Reagents and conditions: (i) XMgBr, THF, 0-20 °C, 50 min; (ii)
MnO2, CHCl3, reflux, 40 min; (iii) [PhCH2PPh3]Br or [EtPPh3]Br or
[MePPh3]Br, LDA, THF, reflux, 16 h (Y ) Ph or Me or H, respectively);
(iv) maleimide, 180 °C, 40 min, then MnO2, dioxane, reflux, 4 h; (v) BBr3,
CH2Cl2, 20 °C, 3-8 h.
hydrolysis of the 9-methoxy derivative 143 of the nitrile 34 using
H2O2 and K2CO3 in aqueous DMSO, which gave the primary
amide but also converted the imide group to an anhydride 144.
Treatment of this material with an ammonium acetate melt to
reform the imide ring followed by demethylation with BBr3 gave
36. The 2-phenyl derivative 37 was prepared via a Stille
coupling on the 9-methoxy derivative 138 of the bromide 28
with tetraphenyltin, followed by demethylation. The sulfoxide
42 was obtained by oxidation of the thioether 41. Saturated
heterocycles 96 and 99 were obtained by hydrogenations of
pyrrole 95 and pyridine 98, respectively, over Adam’s catalyst
in the presence of hydrochloric acid. The N(2),N(6)-substituted
analogue 20 was prepared by complete methylation of the parent
3 with MeI and NaH to give 133 followed by O-demethylation.
The hydrazide 22 was prepared by reaction of 3 directly with
hydrazine. Finally, the anhydride 21 was obtained as a hydrolysis
byproduct during demethylation of the 9-methoxy derivative
of 311 with a pyridine hydrochloride melt.
Compounds containing functionality in the 5-position of the
pyrrolocarbazole were prepared as shown in Scheme 3. The
required 5-substituent was introduced via Grignard addition to
5-methoxyindole-2-carboxaldehyde 106, followed by MnO2
oxidation of the resulting alcohol VIII to give the ketone IX.
Wittig reaction of IX using benzyl, ethyl, or methylphosphonium
salts was used to introduce the 4-substituent (Ph, Me, or H),
generating dienes X, which were elaborated to final products
XII as described above.
Scheme 4 outlines the routes used to prepare derivatives with
a reduced maleimide ring. Reduction of the lead molecule 3
with NaBH4 gave the 1- and 3-hydroxy analogues 16 and 19 in
a ratio of about 5:1, which were readily separable by chroma-
tography on silica. Only one of the isomers displayed a strong
nuclear Overhauser effect in its 1H NMR spectrum between the
H-10 proton and the hydroxyl doublet, and so this was assigned
as structure 16. Compounds 17 and 18 were prepared from 16
by standard methods as shown. Surprisingly the isomer 19 was
completely unreactive under the same conditions and no
derivatives of it could be prepared.
a Reagents and conditions: (i) maleic anhydride, xylene, reflux, 18 h;
(ii) 2,4-dimethoxybenzylamine, AcOH, reflux, 6 h; (iii) MnO2, dioxane,
reflux, 16 h; (iv) RX, K2CO3, DMF, 90 °C, 3 h; (v) TFA, anisole, 90 °C,
18 h; (vi) BBr3, CH2Cl2, 20 °C, 4 h.
was devised in which the maleimide nitrogen was protected with
a 2,4-dimethoxybenzyl group, thereby enabling alkylation on
the free N6-position to be carried out late in the synthesis
(Scheme 5). Reaction of diene 113 with maleic anhydride gave
anhydride 114, which was reacted with 2,4-dimethoxybenzy-
lamine and aromatized to give 115. Alkylation of 115 on the
N6 nitrogen proceeded in high yield to give XIII. Deprotection
of the maleimide of XIII with trifluoroacetic acid/anisole
followed by demethylation with BBr3 then gave the final
products XIV. This route enabled the introduction of a large
number of substituents at the N6-position using parallel
chemistry techniques.
Binding Mode of Pyrrolocarbazole 3
The mode of binding of the lead compound 3 in the ATP-
binding site of Wee1 was initially predicted using a homology
model of the enzyme, generated using the published crystal
structures of the related kinase Chk1.17,18 The model was
subsequently fully validated when the structures of a cocrystal
of 3 and a construct containing residues 291-575 of the human
For the preparation of the compounds of Table 4 containing
a range of functionality at the N6 position, an alternative route

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4899
Figure 1. (a) Crystal structure of 3 bound in the ATP-binding site of a construct of Wee1 kinase. Coordinates used to generate the figure were
taken from Squire et al.19 and the PDB (code 1X8B). Important hydrogen bonds between the ligand, protein, and ordered water molecules in the
active site are indicated with dotted orange lines. (b) ATP site color-coded by electrostatic potential. Blue areas are negatively charged, red areas
denote positive charge, and neutral areas are in green and cyan. (c) ATP site color-coded by H-bonding regions. Red contains H-bonding donors
in the protein, and blue contains H-bond acceptors.
Wee1 enzyme were solved to 1.8 Å resolution.19 While this
construct contained the substrate and ATP-binding sites of Wee1
and was thus useful for identifying the key features of ligand
binding, it did not retain the full catalytic activity of the larger
construct of Wee1 enzyme that was used in the IC50 enzyme
measurements. On the other hand, this larger construct could
not be induced to crystallize in a form that was suitable for
crystallographic structure determinations.
planar pyrrolocarbazole chromophore. There is room at the
6-position to accommodate moderately sized side chains, while
side chains off the 8-position should protrude beyond the protein
into solvent-accessible space.
Results and Discussion
Compounds were evaluated for their ability to inhibit the
incorporation of radiolabeled phosphate into a synthetic poly-
ornithine-tyrosine peptide by a functional construct of human
Wee1 kinase (aa 215-647). A similar assay was carried out
against Chk1 kinase, using cdc25 as the enzyme substrate. Data
from both assays are reported as the concentration of test
compound required to inhibit phosphate incorporation by 50%
(IC50) in comparison to uninhibited controls and are determina-
tions obtained from isolated enzymes. The final ATP concentra-
tion was 9.5 µM in the Wee1 assays and 4.0 µM in the Chk1
assays.
Initial SAR for Substitutions around the Pyrrolocarbazole
Ring System. The compounds listed in Table 1 explored the
preliminary SAR for potency and selectivity around the pyr-
rolocarbazole ring system. Removal of the 4-phenyl substituent
(4), its replacement by iodine (5), or its movement to the
5-position (15) led to a significant loss in activity against Wee1,
although Chk1 was more tolerant of these modifications, which
therefore resulted in Chk1-selective compounds. Introduction
of an adjacent methyl group at the 5-position (11) was tolerated
by Wee1, but the larger 5-ethyl group (12) was not.
The crystal structure provided additional information con-
cerning the position of ordered water molecules in and around
the active site and also identified a coordinated magnesium atom
in this binding site. While several of these ordered water
molecules were sufficiently close to the ligand to provide useful
targets for drug improvement, the magnesium atom and its
coordination sphere were not. Full details of the magnesium
binding site can be found in the published crystal structure.19
The pyrrolocarbazole binds with the pendant 4-phenyl ring
twisted out of the plane of the chromophore and situated in the
back of the ATP binding pocket (Figure 1). This phenyl binding
pocket is rather hydrophobic, being bounded by the side chains
of Val313, Lys328, and Ile374. Substituents placed on this ring
would be expected to influence the specificity of binding to
the Wee1 enzyme. The 9-hydroxyl group in the A-ring of the
chromophore is placed at the entrance to the pocket and forms
an H-bond contact with Cys379. The 1-carbonyl group accepts
an H-bond from the same Cys379, and the 2-NH imide group
donates an H-bond to Glu377. The 3-carbonyl group accepts
an H-bond from both an ordered water and Asn376. Phe433
forms a π-stacking interaction with the B and C rings of the
The combination of a 4-methyl and a 5-phenyl substituent
(13) resulted in a potent, strongly Chk1-selective compound.

4900 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
Table 1. Inhibitory Activity of Miscellaneous Pyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione Derivatives
IC50 (µM)a
compd
structure
X
Y
Z
Wee1
Chk1
0.047
0.47
0.038
0.30
>10
>10
0.21
ratiob
synthetic routec
3
4
5
6
7
A
A
A
A
A
A
A
A
B
B
B
B
B
C
C
C
D
E
E
E
A
A
A
A
9-OH
9-OH
9-OH
8-OH
10-OH
9-OH
9-OH
9-OH
Me
NH
NH
NH
NH
NH
O
Ph
H
I
Ph
Ph
Ph
Ph
Ph
Ph
Ph
0.097
4.0
2.3
0.31
0.48
0.12
0.017
0.97
1d
3
2
1
1
1
1
1
3
3
3
3
3
4
4
4
>50
8
9
0.43
0.078
0.26
0.13
1.6
9.7
2.3
4.0
>23
2.7
S
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
NMe
0.056
0.24
0.43
0.032
0.37
0.33
0.22
1.8
0.27
0.003
0.16
0.082
Et
Ph
Ph
Ph
Me
Ph
H
OH
OMe
H
OH
NMe
O
N-NH2
9-OH
9-OMe
9-OH
9-OH
>50
20
37
2.8
>50
>50
>10
>10
>10
0.074
>10
0.026
4
NMe
NH
NH
NH
NH
NMe
O
speciale
4.6
5.8
0.44
3.2
0.22
<0.09
1.5
40
5.0
3.9
speciale
3.9
0.011
0.64
0.057
0.033
speciale
2-ClPh
2-ClPh
2-ClPh
2-ClPh
2
1
5A
1
>10
>303
a IC50 values against the isolated enzyme are the average of at least three individual determinations. Replicate values were within 30% of each other.
b Ratio of Chk1 IC50 to Wee1 IC50. c Scheme number used for synthesis. d Reference 11. e See Experimental Section for details.
Methylation of the 9-hydroxyl group (24) or its movement to
the adjacent 10-position (7) significantly reduced activity against
both Wee1 and Chk1, while its movement to the 8-position (6)
led to a 3- to 6-fold loss in potency against both enzymes.
Activity was also significantly reduced when the 1-carbonyl
group was modified (16-18), whereas reduction of the 3-car-
bonyl group (19) was somewhat better tolerated, particularly
by Chk1. A free NH at the 2-position of the maleimide ring
was important for activity against both Wee1 and Chk1 (20-
22), with N-methylation of this group (compare 20 with 10)
leading to a >100-fold loss in potency.
These SARs are largely consistent with the observed binding
mode (see above), where loss of any of the key H-bond contacts
between the ligand and protein leads to a loss in Wee1 activity.
An exception was the relatively modest (3-fold) loss in Wee1
potency upon movement of the 9-hydroxyl group to the adjacent
8-position. The X-ray crystal structure of a Wee1 construct with
inhibitor 3 bound in the active site19 reveals a network of three
ordered water molecules close to the 8-position of the pyrrolo-
carbazole ring system, and it is possible that the 8-hydroxyl
group of 6 forms a stabilizing H-bond with one of these.
A comparison of 3 with 8-10 and 25 showed that the
carbazole N6 nitrogen could be alkylated or replaced by O or
S, with retention of significant Wee1 activity. The oxygen
analogue 8 displayed significantly improved Wee1 selectivity
when compared with the lead 3, with only a 4-fold loss in Wee1
potency. This trend was mirrored in the more potent series
containing a 2-chlorophenyl substituent at the 4-position, with
dibenzofuran 26 displaying an impressive >300-fold selectivity
for Wee1. Comparisons of the pairs 3/23, 10/25, and 8/26
showed that a 2-chloro substituent on the phenyl ring increased
potency against Wee1 by about 10-fold (see later), with
compound 23 in particular displaying excellent Wee1 potency
(IC50 ) 11 nM). A 2′-chloro substituent was similarly observed
to increase Wee1 potency in the 6-phenylpyrido[2,3-d]pyrimi-
din-7(8H)-one series of Wee1 inhibitors7 (e.g., 1, 2), suggesting
that the pendant 6-phenyl ring occupies a similar binding site
in the protein compared to the 4-phenyl ring of the pyrrolocar-
bazoles. As expected from the above data sets, there was no
overall correlation between Wee1 and Chk1 potencies for this
diverse set of analogues.
SAR for Substitution in the 4-Phenyl Ring. Because of the
increase in potency achieved by introduction of a 2′-Cl substit-
uent, we carried out a more general evaluation of the effects of
substitution in the 4-phenyl ring on the activity of the 9-hydroxy-
4-phenylpyrrolocarbazoles as inhibitors of Wee1 and Chk1
kinases. Table 2 lists data for 67 4-phenyl-substituted analogues.
Compounds 23 and 27-44 are 2′-monosubstituted. Lipophilic
2′-substituents such as Cl, Br, I, OMe, and SMe did improve
Wee1 potency, while Me, OH, and NO2 substitution provided
compounds almost equipotent with the unsubstituted compound
3. Other 2-substituents, including the bulky phenyl group (37),
were generally tolerated by Wee1, with only a slight loss in
potency. In contrast, Chk1 was generally much less tolerant of
2′-substitution, with the result that selectivity for Wee1 was
raised from 0.48-fold for 3 to typically 10- to 100-fold.
Compounds 41 and 42 (2′-SMe and 2′-SOMe) displayed
remarkable Wee1 selectivity (>1500-fold and >225-fold,
respectively). Substitution at the 2′-position of the 4-phenyl ring
would have the greatest effect on twisting this ring out of the
plane of the carbazole ring system, leading to more favorable
interactions between this ring and the protein. This 2′-substituent

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4901
Table 2. Inhibitory Activity of Substituted
4-Phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione Derivatives
Table 3. Effect of Heterocyclic Ring Substitution at the 4-Position of
the Pyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione Nucleus
IC50 (µM)a
compd
Z
Wee1
Chk1
ratiob
0.48
synthetic routec
3
27
23
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
77
78
79
80
81
82
83
84
85
86
87
88
89
90
91
H
0.097
0.047
1d
2-F
0.33
0.011
0.023
0.013
0.15
0.51
0.58
0.45
0.19
0.83
0.16
0.57
0.060
0.024
0.26
0.077
0.44
0.15
0.62
0.30
0.23
40
6.5
48
2.0
37
17
1.0
0.17
0.13
15
0.37
20
2Ae
2-Cl
2-Br
2-I
2-Me
2-Et
2-CF3
2
1
specialf
2Ae
19
10
0.45
0.032
0.11
2.4
0.21
1.2
0.50
3.8
2
2
2-CH2OH
2-CN
2-COMe
2-CONH2
2-Ph
2
1
2Ae
specialf
specialf
2-OH
specialf
2-OMe
21
15
2
2-OEt
2-SMe
2
0.033 >50
>1500
>225
85
2
2-SOMe
2-NO2
0.22 >50
from 41
0.047
0.21
4.0
1
2-NH2
0.79
0.073
0.33
0.62
3.8
from 43
3-F
0.22
0.055
0.23
0.33
6
2Ae
3-Cl
2Ae
3-Me
2.7
2Ae
3-CF3
>50
>10
0.97
23
4.2
>10
44
0.26
2.2
2Ae
3-CH2OH
3-CH2NH2
3-CN
3-COMe
3-Ph
0.87
4.4
1.1
5.2
2Ae
a IC50 values against the isolated enzyme are the average of at least three
individual determinations. Replicate values were within 30% of each other.
b Ratio of Chk1 IC50 to Wee1 IC50. c Scheme number used for synthesis.
2
0.18
4.3
40
0.089
0.62
23
2Ae
>2.3
2Ae
1.1
2
3-OH
2.9
from 55
3-OMe
3.5
2Ae
would occupy a moderately lipophilic binding pocket in Wee1
bounded by Val313, Ala326, and Lys328.
3-NO2
0.30 >10
0.070 1.6
16 2.8
0.73 >10
3.3 2.8
>10
0.34
0.12
>33
23
0.18
>14
0.85
1
3-NH2
2
2Ae
Compounds 45-57 are 3′-monosubstituted. The same broad
overall SARs were followed, in that small groups were generally
acceptable, although in this case they provided very little
improvement in Wee1 potency over 3 and smaller increases in
Wee1/Chk1 selectivity (generally 3- to 6-fold). Larger groups
such as phenyl were poorly tolerated. Compounds 58-69 are
4′-monosubstituted. Polar, H-bond donating 4′-substituents (e.g.,
OH, NH2) were well tolerated by both enzymes, resulting in
no overall change in selectivity profile, but most other substitu-
tions resulted in a >10-fold loss in potency.
4-F
4-Cl
2Ae
4-Me
2Ae
4-CF3
>50
2Ae
4-CH2OH
4-CN
4-COMe
4-OH
4-OMe
1.2
0.28
0.07
2
1.8
2Ae
3.6
>10
>2.8
2
0.067
0.021
1.3
0.31
0.11
>0.35
9.1
from 66
12
29
2Ae
4-SMe
>10
10
0.021
2Ae
4-SO2Me
4-NH2
1.1
0.15
0.028
0.012
0.021
0.023
0.024
2Ae
0.14
8.6
81
23
5.6
9.6
2
2-Cl, 3-Cl
2-Cl, 3-OH
2-Cl, 3-NH2
2-Cl, 4-OH
2-Cl, 4-NH2
2-Cl, 5-Cl
2-Cl, 5-OH
2-Cl, 5-NH2
2-Cl, 6-Cl
2-Cl, 6-OH
2-Cl, 6-OMe
2-Br, 4-NH2
2-Br, 6-Br
2-Me, 3-Me
2-Me, 5-Me
2-Me, 6-Me
2-OMe, 4-NH2
2-OMe, 5-NH2
2-OMe, 6-OMe
2-OMe, 6-F
2,6-diCl, 3-OH
2,6-diCl, 4-OH
0.24
0.97
0.48
0.13
0.23
2
1
Compounds 70-80 combine a 2′-Cl group with various 3′-,
4′-, 5′-, or 6′-substituents. Generally, these compounds retained
both the high Wee1 potency (many between 10 and 30 nM)
and the high Wee1/Chk1 selectivity displayed by the 2′-chloro
compound 23. The 2′-Cl, 4′-OH and 2′-Cl, 4′-NH2 compounds
73 and 74 are interesting, showing substantially lower Chk1
activity than the corresponding 4′-OH and 4′-NH2 compounds
65 and 69, respectively. Similar results were seen for the
comparable 2′-Br and 2′-OMe analogues 81 and 86, which show
excellent selectivity due to low Chk1 activity. Addition of a
5′-amino substituent to the 2′-chloro or 2′-methoxyphenyl ring
(77 and 87) also resulted in improved Wee1 selectivity, with
good potency being retained in the former case. The 2′,6′-Cl,
2′,6′-Br, and 2′,6′-diOMe compounds 78, 82, and 88 also
showed excellent Wee1 potency and selectivity, with the 2′,6′-
diMe analogue 85 being slightly less potent but also highly
selective.
1
1
1
0.49 >10
>20
2Ae
0.042
0.020
0.028
0.045
0.015
0.020
0.035
0.27
1.1
26
265
43
1
5.3
1
1.2
1
0.79
0.60
0.22
7.0
18
from 80
40
2A
1
11
200
26
1
7.1
2Ae
2Ae
2
0.96 >10
>10
>133
247
>91
196
24
0.075 >10
0.019
4.7
1
0.11 >10
1
0.027
0.029
0.018
0.049
5.3
2
0.71
2.4
2
133
26
1
1.3
1
a IC50 values against the isolated enzyme are the average of at least three
individual determinations. Replicate values were within 30% of each other.
b Ratio of Chk1 IC50 to Wee1 IC50. c Scheme number used for synthesis.
d Reference 11. e Letter A indicates that array synthesis was used; see
Experimental Section for details. f See Experimental Section for details.
Heterocycles off the 4-Position. A series of compounds were
evaluated in which the 4-phenyl substituent was replaced with
a heterocyclic ring (Table 3). Incorporation of a thienyl or a

4902 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
Table 4. Effect of Steric Bulk at the N6 Position of
4-(2′-Chlorophenyl)pyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione Derivatives
and the SARs seen here are broadly consistent with this binding
mode. Although some of these compounds are potent and
selective inhibitors of the Wee1 enzyme, they have low aqueous
solubilities (typically less than 3 µg/mL) and low permeabilities
across Caco-2 cell monolayers (Papp < 3 × 10-6 cm/s). This
has precluded us from obtaining meaningful data on the effects
of these compounds on abrogation of the G2/M checkpoint in
whole cells. Nevertheless, the results show that 4-phenylpyrrolo-
[3,4-c]carbazole-1,3(2H,6H)-dione is a suitable molecular frame-
work on which to elaborate analogues with solubilizing sub-
stituents at defined positions, especially N6 and C8, guided by
ongoing crystallographic analysis and molecular modeling.
These studies will be reported subsequently.
IC50 (µM)a
compd
R
Wee1
Chk1
ratiob
synthetic routec
25
100
101
102
103
104
105
Me
Et
n-Pr
i-Pr
n-Bu
(CH2)2i-Pr
n-pent
0.057
0.050
0.063
0.053
0.059
0.15
0.22
0.12
0.45
27
35
24
3.9
2.4
7.1
509
593
160
259
5A
5A
5A
5A
5A
5A
5A
Experimental Section
Combustion analyses were performed by the Microchemical
Laboratory, University of Otago, Dunedin, New Zealand. Melting
points were determined using an Electrothermal model 9200 digital
melting point apparatus and are as read. NMR spectra were
measured on a Bruker DRX-400 spectrometer or a Bruker Avance-
400 spectrometer and are referenced to Me4Si. High-resolution mass
spectra were recorded on a Varian VG-70SE spectrometer at
nominal 5000 resolution. Liquid chromatography-mass spectrom-
etry (LC-MS) was performed on an Agilent 1100 LC system
interfaced with an Agilent MSD mass detector. Mass detection was
performed with an APCI source, using simultaneous positive and
negative ion acquisition. Compound purities were determined by
HPLC using simultaneous diode array UV detection. Unless
otherwise indicated, compounds were purified by flash column
chromatography on silica gel 60 support (Scharlau, 230-400 mesh
ASTM), using the indicated eluants.
0.17
44
a IC50 values against the isolated enzyme are the average of at least three
individual determinations. Replicate values were within 30% of each other.
b Ratio of Chk1 IC50 to Wee1 IC50. c Scheme number used for synthesis.
The letter A indicates that array synthesis was used; see Experimental
Section for details.
pyrrole ring gave compounds with comparable Wee1 and Chk1
potencies to the phenyl compound, with the 3′-substituted
derivatives 93 and 95 displaying better activities than their 2′-
substituted analogues (92 and 94). 4-Pyridyl-substituted com-
pounds 97 and 98 were somewhat less active. The reduced
derivatives 96 and 99 of the 3-pyrrole and 3-pyridyl compounds
95 and 98 were much less potent, suggesting either loss of a
favorable π-interaction with the protein or low tolerance for a
more basic nitrogen atom in the active site. If this basic nitrogen
is charged on binding, its presence would likely disfavor ligand
binding in the rather lipophilic Wee1 binding site. None of these
heterocyclic derivatives displayed significantly improved se-
lectivity for Wee1 compared with 4-phenyl-substituted com-
pounds.
SAR for N6 Substitution. In deciding on suitable 4-phenyl
substituents for exploring SAR at other points on the molecule,
Wee1 potency, Wee1/Chk1 selectivity, ease of synthesis and
potential compound metabolic stability were considered. For
these reasons, a 2′-Cl substituent was chosen to explore the SAR
for N6 substitution, as shown in Table 4. Compounds 25 and
100-105 were explored for nonpolar substituents off the N6
position. These analogues showed little change in potency for
Wee1 but a marked increase in selectivity due to a drop-off in
Chk1 inhibition for the i-Pr and longer-chain derivatives,
suggesting less bulk tolerance at this position in the Chk1 ATP
binding site. Indeed, Chk1 has a bulky Glu91 protruding into
this region of the protein, whereas Wee1 has the smaller and
less polar Ser383.
Procedures of Scheme 1. Procedure 1. (2,6-Dichlorobenzyl)-
(triphenyl)phosphonium Bromide (116). A mixture of 2,6-
dichlorotoluene (20.1 g, 0.125 mol), N-bromosuccinimide (24.6 g,
0.138 mol), and 2,2′-azobisisobutyronitrile (0.41 g, 2.50 mmol) in
dry benzene (300 mL) under N2 was stirred at reflux for 6 h with
continuous irradiation from a 100 W lamp. The resulting reaction
mixture was concentrated under vacuum (to 50 mL), cooled, filtered,
and washed with dry benzene. The filtrate (containing the crude
benzyl bromide) was treated directly with triphenylphosphine (49.3
g, 0.188 mol) and was stirred at reflux for 17 h. After the mixture
was cooled, the precipitate was collected by filtration, washed
thoroughly with dry benzene and then pentane, and dried under
vacuum at 50 °C to give 116 (62.1 g, 99%) as a cream powder. An
analytical sample was obtained by recrystallization: mp (CH2Cl2/
1
benzene) 269-272 °C; H NMR (CDCl3) δ 7.76 (m, 9 H), 7.64
(m, 6 H), 7.18 (s, 3 H), 5.50 (d, J ) 14.3 Hz, 2 H). Anal. (C25H20-
BrCl2P) C, H.
Procedure 2. 2-[(E)-2-(2,6-Dichlorophenyl)ethenyl]-5-meth-
oxy-1H-indole (117). Lithium diisopropylamide (52 mL of a 1.5
M solution in cyclohexane, 78 mmol) was added dropwise to a
stirred suspension of 116 (34.6 g, 68.9 mmol) in dry THF (100
mL) under N2 at 0 °C, and then the mixture was stirred at room
temperature for 45 min and then at 50 °C for 30 min, then cooled
to 0 °C. A solution of 5-methoxy-1H-indole-2-carbaldehyde20,21
(106) (10.0 g, 57.1 mmol) in dry THF (60 mL, then 2 × 10 mL to
rinse) was added, and the mixture was stirred at reflux for 5 h.
The cooled solution was added to a mixture of ice and aqueous
sodium bicarbonate (200 mL), the organic layer was separated, and
the aqueous portion was further extracted with CH2Cl2 (5 × 150
mL). The combined organics were washed with water (2 × 200
mL), back-extracted with CH2Cl2 (2 × 100 mL), and evaporated
to dryness. Then the residue was chromatographed on silica gel.
Elution with 0-40% CH2Cl2/light petroleum gave foreruns. Then
further elution with 50% CH2Cl2/light petroleum to 100% CH2Cl2
gave pure E diene 117 (II, 5-OMe, Ar ) 2,6-dichlorophenyl) (17.6
g, 97%) as a yellow solid following recrystallization: mp (CH2-
Conclusions
While the lead 4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione 3 was a potent but nonselective inhibitor of Wee1 versus
Chk1 kinase, good selectivity for Wee1 could be achieved by
the introduction of lipophilic functionality at the 2′-position and
the 2′,6′-positions of the 4-phenyl ring. Enhanced Wee1
selectivity was also observed from substitution of the phenyl
ring with a 2′-lipophilic group together with a 5′-NH2 group
and by incorporation of moderate steric bulk off the N6-position.
Replacement of the N6 atom by oxygen to give the dibenzofuran
analogue of the carbazole also resulted in Wee1-selective
inhibition. Key features of the binding mode for this class of
inhibitor were previously determined by X-ray crystallography
of a cocrystal of an inhibitor bound in a construct of the enzyme,
1
Cl2/hexane) 144-147 °C; H NMR (CDCl3) δ 8.20 (br s, 1 H),
7.36 (d, J ) 8.1 Hz, 1 H), 7.27 (m, 1 H), 7.25 (d, J ) 16.8 Hz, 1

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4903
H), 7.11 (t, J ) 8.0 Hz, 1 H), 7.03 (d, J ) 2.4 Hz, 1 H), 6.93 (d,
J ) 16.8 Hz, 1 H), 6.88 (dd, J ) 8.7, 2.5 Hz, 1 H), 6.61 (d, J )
1.8 Hz, 1 H), 3.86 (s, 3 H). Anal. (C17H13Cl2NO) C, H, N.
(m, 3H), 6.93 (d, J ) 2.1 Hz, 1H), 6.79 (dd, J ) 8.6, 2.1 Hz, 1H).
Anal. (C20H12N2O3‚1/4H2O) C, H, N.
10-Methoxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (126). Reaction of 4-methoxyindole-2-carboxaldehyde23 (123)
(I, 4-OMe) with benzyltriphenylphosphonium bromide using
procedure 2 gave 4-methoxy-2-[2-phenylethenyl]-1H-indole (124)
(II, 4-OMe, Ar ) phenyl) (89%) as a fluorescent oil (mixture of
E/Z isomers), which was used without further purification.
Reaction of the diene 124 with maleimide using procedure 3
gave 10-methoxy-4-phenyl-4,5,6,10c-tetrahydropyrrolo[3,4-c]car-
bazole-1,3(2H,3aH)-dione (125) (IV, 10-OMe, R ) H, Ar )
phenyl) (86%) as a yellow solid, which was used without further
purification.
Aromatization of 125 with DDQ using procedure 4 gave 126
(V, 10-OMe, R ) H, Ar ) phenyl) (78%) as yellow needles: mp
273-276 °C; 1H NMR [(CD3)2SO] δ 12.10 (s, 1H), 10.78 (s, 1H),
7.59 (s, 1H), 7.57 (dd, J ) 8.0, 1.8 Hz, 2H), 7.49 (t, J ) 7.9 Hz,
1H), 7.47-7.39 (m, 3H), 7.16 (d, J ) 7.9 Hz, 1H), 6.80 (d, J )
7.9 Hz, 1H), 3.99 (s, 3H). Anal. (C21H14N2O3) C, H, N.
10-Hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (7). Demethylation of 126 with BBr3 using procedure 5 gave
7 as a yellow-orange solid (92%): mp >300 °C; 1H NMR [(CD3)2-
SO] δ 12.33 (s, 1H), 12.16 (s, 1H), 11.65 (br, 1H), 7.62 (s, 1H),
7.60 (dd, J ) 7.9, 1.8 Hz, 2H), 7.51-7.45 (m, 3H), 7.40 (dd, J )
7.6 Hz, 1H), 7.03 (d, J ) 7.6 Hz, 1H), 6.63 (d, J ) 7.6 Hz, 1H).
Anal. (C20H12N2O3) C, H, N.
Procedure 3. 4-(2,6-Dichlorophenyl)-9-methoxypyrrolo[3,4-
c]carbazole-1,3(2H,6H)-dione (118). A mixture of pure E diene
117 (1.51 g, 4.73 mmol), maleimide (1.90 g, 19.6 mmol), and
powdered SnCl2 (67 mg, 0.35 mmol) in toluene (20 mL) was stirred
in a foil-covered sealed vial at 125 °C for 3 h. The cooled solution
was added to aqueous sodium bicarbonate (100 mL) and extracted
with EtOAc (7 × 100 mL). The combined extracts were evaporated
to dryness to give a crude adduct (IV, 9-OMe, R ) H, Ar ) 2,6-
dichlorophenyl) as a tan powder, which was used directly.
Procedure 4. The crude Diels-Alder adduct above was reacted
directly with DDQ (5.44 g, 24.0 mmol) in toluene (60 mL) and
dioxane (60 mL), stirring at reflux for 2 days. The cooled solution
(and precipitated solids after dissolving these in CHCl3) was
sequentially washed twice with aqueous sodium bicarbonate (300
mL, then 150 mL) and water (2 × 150 mL). Then the aqueous
washings were back-extracted with CHCl3 (8 × 150 mL), washing
these extracts in the same way. The combined organics were
evaporated to dryness, and then the residue was chromatographed
on silica gel. Elution with 0-0.5% MeOH/CH2Cl2 gave foreruns.
Then further elution with 0.5% MeOH/CH2Cl2 gave 118 (V,
9-OMe, R ) H, Ar ) 2,6-dichlorophenyl) (1.52 g, 78%) as a yellow
solid following recrystallization: mp (MeOH/CH2Cl2/hexane) 299-
1
301 °C; H NMR [(CD3)2SO] δ 12.02 (br s, 1 H), 11.15 (br s, 1
5-Methoxy-2-[(E,Z)-2-phenylethenyl]-1-benzofuran (127). A
suspension of benzyltriphenylphosphonium bromide (1.85 g, 4.26
mmol) in THF (30 mL) was treated with a solution of lithium bis-
(trimethylsilyl)amide (4 mL of a 1 M solution in THF, 3.98 mmol).
The reaction mixture was stirred at room temperature for 30 min,
and then a solution of 5-methoxy-1-benzofuran-2-carbaldehyde24
(0.50 g, 2.84 mmol) in THF (10 mL) was added. After 20 min
water was added and the THF was removed at reduced pressure.
The residue was extracted with EtOAc (2 × 50 mL), and the
combined extracts were dried and concentrated. The residue was
purified by column chromatography on silica gel, eluting with DCM
to give 127 as a 1:2 mixture of Z- and E-isomers, which was used
directly (0.63 g, 89%): mp 124-128 °C. Anal. (C17H14O2) C, H.
9-Methoxy-4-phenyl-1H-[1]benzofuro[3,2-e]isoindole-1,3(2H)-
dione (128). The diene mixture 127 was reacted with maleimide
using procedure 3 and then aromatized with MnO2 using procedure
7 to give 128 as a yellow solid (53%): mp 271-275 °C; 1H NMR
[(CD3)2SO] δ 11.41 (br s, 1H), 8.24 (d, J ) 2.7 Hz, 1H), 7.98 (s,
1H), 7.77 (d, J ) 9.0 Hz, 1H), 7.67-7.65 (m, 2H), 7.51-7.45 (m,
3H), 7.30 (dd, J ) 9.0, 2.7 Hz, 1H), 3.91 (s, 3H). Anal. (C21H13-
NO4‚1/3H2O) C, H, N.
H), 8.45 (d, J ) 2.6 Hz, 1 H), 7.61 (d, J ) 7.7 Hz, 2 H), 7.61 (s,
1 H), 7.59 (d, J ) 8.9 Hz, 1 H), 7.50 (dd, J ) 8.8, 7.4 Hz, 1 H),
7.26 (dd, J ) 8.9, 2.7 Hz, 1 H), 3.90 (s, 3 H). Anal. (C21H12Cl2N2O3)
C, H, N.
Procedure 5. 4-(2,6-Dichlorophenyl)-9-hydroxypyrrolo[3,4-
c]carbazole-1,3(2H,6H)-dione (78). Boron tribromide (18.5 mL
of a 1.0 M solution in CH2Cl2, 18.5 mmol) was added to a stirred
solution of carbazole methyl ether 118 (1.52 g, 3.70 mmol) in dry
CH2Cl2 (100 mL) under N2 at 0 °C, and then the mixture was stirred
at room temperature for 2.5 h. The resulting solution was added to
a mixture of ice and aqueous sodium bicarbonate (300 mL) and
extracted with EtOAc (5 × 200 mL). The combined extracts were
washed with water (200 mL) and evaporated to dryness, and then
the residue was chromatographed on silica gel. Elution with 0-2%
MeOH/CH2Cl2 gave foreruns. Then further elution with 2-3%
MeOH/CH2Cl2 gave phenol 78 (1.45 g, 97%) as an orange solid
following recrystallization: mp (MeOH/CH2Cl2/hexane) 205-215
1
°C (dec); H NMR [(CD3)2SO] δ 11.89 (br s, 1 H), 11.08 (br s, 1
H), 9.32 (br s, 1 H), 8.30 (d, J ) 2.4 Hz, 1 H), 7.61 (d, J ) 7.9 Hz,
2 H), 7.55 (s, 1 H), 7.50 (dd, J ) 8.7, 7.4 Hz, 1 H), 7.47 (d, J )
8.7 Hz, 1 H), 7.10 (dd, J ) 8.7, 2.4 Hz, 1 H). Anal. (C20H10Cl2N2O3‚
0.5H2O) C, H, N.
9-Hydroxy-4-phenyl-1H-[1]benzofuro[3,2-e]isoindole-1,3(2H)-
dione (8). Demethylation of 128 with BBr3 using procedure 5 gave
8-Methoxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (122). Reaction of 6-methoxyindole-2-carboxaldehyde22 (119)
(I, 6-OMe) with benzyltriphenylphosphonium bromide using
procedure 2 gave 6-methoxy-2-[2-phenylethenyl]-1H-indole (120)
(II, 6-OMe, Ar ) phenyl) (94%) as a white solid (mixture of E/Z
isomers), which was used without further purification.
Reaction of the diene 120 with maleimide using procedure 3
gave 8-methoxy-4-phenyl-4,5,6,10c-tetrahydropyrrolo[3,4-c]carba-
zole-1,3(2H,3aH)-dione (121) (IV, 8-OMe, R ) H, Ar ) phenyl)
(61%) as a tan solid, which was used without further purification.
1
8 (100%): mp 288-290 °C; H NMR [(CD3)2SO] δ 11.37 (br s,
1H), 9.74 (s, 1H), 8.13 (d, J ) 2.6 Hz, 1H), 7.94 (s, 1H), 7.66-
7.63 (m, 3H), 7.51-7.44 (m, 3H), 7.11 (dd, J ) 8.9, 2.6 Hz, 1H).
Anal. (C20H11NO4‚1/2H2O) C, H, N.
9-Hydroxy-4-phenyl-1H-[1]benzothieno[3,2-e]isoindole-1,3-
(2H)-dione (9). Reaction of 5-methoxy-1-benzothiophene-2-car-
baldehyde25 with benzyltriphenylphosphonium chloride using pro-
cedure 2 gave 5-methoxy-2-[(E,Z)-2-phenylethenyl]-1-benzothiophene
(129) as an E/Z mixture (63%): 1H NMR (CDCl3) minor isomer δ
7.60 (d, 1H), 7.40 (m, 5H), 7.15 (s, 1H), 7.05 (s, 1H), 6.91 (d,
1H), 6.75 (m, 2H), 3.85 (s, 3H); 1H NMR (CDCl3) major isomer δ
7.75 (d, J ) 9 Hz, 1H), 7.55 (m, 2H), 7.25-7.4 (m, 3H), 7.18 (m,
2H), 7.05 (s, 1H), 6.95 (m, 2H), 3.86 (s, 3H).
Aromatization of 121 with DDQ using procedure 4 gave 122
(V, 8-OMe, R ) H, Ar ) phenyl) (65%) as a yellow powder: mp
1
154-156 °C; H NMR [(CD3)2SO] δ 11.87 (br, 1H), 11.02 (br,
1H), 8.75 (d, J ) 8.8 Hz, 1H), 7.61 (dd, J ) 8.0, 1.8 Hz, 2H), 7.60
(s, 1H), 7.49-7.40 (m, 3H), 7.10 (d, J ) 2.2 Hz, 1H), 6.94 (dd, J
) 8.8, 2.2 Hz, 1H), 3.90 (s, 3H). Anal. (C21H14N2O3‚1/4H2O) C, H,
N.
8-Hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (6). Demethylation of 122 with BBr3 using procedure 5 gave
6 as a yellow powder (74%): mp >330 °C; 1H NMR [(CD3)2SO]
δ 11.75 (br, 1H), 10.96 (br, 1H), 10.56 (br, 1H), 8.66 (d, J ) 8.6
Hz, 1H), 7.60 (dd, J ) 8.0, 1.8 Hz, 2H), 7.53 (s, 1H), 7.49-7.39
Reaction of 129 with maleimide for 6 days using procedure 3,
followed by aromatization with MnO2 in CHCl3 for 5 days at 40
°C following procedure 7, gave 9-methoxy-4-phenyl-1H-[1]ben-
zothieno[3,2-e]isoindole-1,3(2H)-dione (130) (44%): 1H NMR
[(CD3)2SO] δ 11.46 (s, 1H), 9.37 (d, J ) 2.5 Hz, 1H), 8.38 (s,
1H), 8.2 (d, J ) 9 Hz, 1H), 7.62 (m, 2H), 7.48 (m, 3H), 7.31 (dd,
J ) 2.5, 9.0 Hz, 1H), 3.94 (s, 3H).
Demethylation of 130 with BBr3 for 48 h using procedure 5 gave
9 (65%): mp 311-313 °C; 1H NMR [(CD3)2SO] δ 11.42 (s, 1H),

4904 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
9.85 (s, 1H), 9.17 (d, J ) 2.5 Hz, 1H), 8.32 (s, 1H), 7.9 (d, J ) 9
Hz, 1H), 7.62 (m, 2H), 7.5 (m, 3H), 7.17 (dd, J ) 2.5, 9 Hz, 1H).
Anal. (C20H11NO3S‚0.2H2O) C, H, N, S.
hydrochloride (50 g) at 200 °C under nitrogen. After 25 min the
reaction mixture was cooled, diluted with water, and extracted with
ethyl acetate (3×). The combined organic extracts were washed
with saturated sodium bicarbonate and then brine before being dried
over anhydrous sodium sulfate and concentrated in vacuo. Chro-
matography on silica, eluting with ethyl acetate/dichloromethane
(1:9 to 1:3), gave at highest Rf 21 (108 mg, 17%) as an orange
9-Methoxy-6-methyl-4-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (132). To a stirred solution of 2-[2-phenylethenyl]-
5-methoxy-1H-indole11 (131) (II, 5-OMe, Ar ) phenyl) (0.20 g,
0.80 mmol) in DMF (5 mL) under nitrogen was added sodium
hydride (91 mg of a 50% dispersion in oil, 1.9 mmol) followed by
methyl iodide (0.041 mL, 0.71 mmol). After 15 min the reaction
mixture was diluted with water and extracted with ethyl acetate
(3×). The combined organic extracts were washed with brine, dried
over anhydrous sodium sulfate, and concentrated in vacuo. The
residue was then dissolved in toluene (20 mL), and maleimide (92
mg, 0.95 mmol) was added before the solution was heated at reflux
for 24 h and the reaction mixture was concentrated to dryness. The
product was redissolved in dioxane (100 mL) before activated MnO2
(1.24 g, 14.25 mmol) was added, and the resulting suspension was
heated at reflux for 18 h. Filtration through Celite, concentration
under reduced pressure, and chromatography on silica, eluting with
ethyl acetate/dichloromethane (1:3), followed by trituration from
methanol, gave 132 (V, 9-OMe, R ) Me, Ar ) phenyl) (0.20 g,
70%) as a yellow powder: mp 286-291 °C; 1H NMR [(CD3)2SO]
δ 11.11 (br s, 1H), 8.53 (d, J ) 2.6 Hz, 1H), 7.80 (s, 1H), 7.66 (m,
3H), 7.47 (m, 3H), 7.30 (dd, J ) 9.0, 2.6 Hz, 1H), 3.96 (s, 3H),
3.90 (s, 3H). Anal. (C22H16N2O3) C, H, N.
1
powder: mp >300 °C; H NMR [(CD3)2SO] δ 12.13 (br s, 1H),
9.43 (br s, 1H), 8.20 (d, J ) 2.4 Hz, 1H), 7.82 (s, 1H), 7.67 (m,
2H), 7.54-7.46 (m, 4H), 7.14 (dd, J ) 8.7, 2.4 Hz, 1H). EIMS
found 329.0691; C20H11NO4 requires 329.0688. Further elution gave
at lowest Rf 9-hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3-(2H,6H)-
dione11 (3) (510 mg, 78%). 1H NMR data are identical to those for
an authentic sample.
2-Amino-9-hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (22). To a solution of 311 (80 mg, 0.24 mmol) in
ethanol (10 mL) was added hydrazine hydrate (0.12 mL, 2.4 mmol).
The reaction mixture was heated at reflux for 30 min, and then
concentrated sulfuric acid (4 drops) was added. Reflux was
continued overnight, and then the mixture was poured onto ice.
The orange precipitate was collected by filtration to give 22
1
(67 mg, 80%): mp 280-286 °C; H NMR [(CD3)2SO] δ 11.75
(br s, 1H), 9.28 (br s, 1H), 8.37 (d, J ) 2.4 Hz, 1H), 7.61-7.59
(m, 2H), 7.56 (s, 1H), 7.50-7.43 (m, 4H), 7.07 (dd, J ) 8.7, 2.4
Hz, 1H), 4.85 (s, 2H). EIMS found 343.0961; C20H13N3O3 requires
343.0957.
Procedure 6. 9-Hydroxy-6-methyl-4-phenylpyrrolo[3,4-c]car-
bazole-1,3(2H,6H)-dione (10). Compound 132 (180 mg, 0.50
mmol) was heated in a melt with pyridine hydrochloride (10 g) at
200 °C under nitrogen. After 15 min the reaction mixture was
cooled and diluted with water to precipitate the product, which was
collected by filtration and washed with water before being dried in
vacuo. Chromatography on silica, eluting with ethyl acetate/
dichloromethane (1:3), followed by trituration from dichloromethane/
hexane, gave 10 (155 mg, 90%) as an orange powder: mp 297-
2-[2-(2-Chlorophenyl)ethenyl]-5-methoxy-1H-indole (113).
Lithium diisopropylamide (34.4 mL of a 1.5 N solution, 0.052 mol)
was added dropwise under nitrogen to a suspension of (2-
chlorobenzyl)(triphenyl)phosphonium chloride (20.17 g, 0.048 mol)
in dry THF (200 mL), and the solution was stirred at room
temperature for 15 min. A solution of 5-methoxy-1H-indole-2-
carbaldehyde (106) (6.99 g, 0.040 mol) in THF (30 mL) was added,
and stirring was continued at room temperature for 15 min. Then
the reaction mixture was heated at reflux for 6 h. The cooled
solution was diluted with water and extracted with EtOAc, and the
organic phase was dried. The drying agent was removed, and the
solution was concentrated to dryness to give an oil, which was
adsorbed onto silica and chromatographed on silica. Elution with
ethyl acetate/petroleum ether (1:1) gave 113 (II, 5-OMe, Ar )
2-chlorophenyl) as a mixture of E/Z isomers (9.76 g, 87%).
Crystallization of a small sample from methanol afforded pure
1
300 °C; H NMR [(CD3)2SO] δ 11.05 (br s, 1H), 9.34 (s, 1H),
8.39 (d, J ) 2.5 Hz, 1H), 7.76 (s, 1H), 7.65 (m, 2H), 7.54 (d, J )
8.8 Hz, 1H), 7.46 (m, 3H), 7.13 (dd, J ) 8.8, 2.5 Hz, 1H), 3.93 (s,
3H). Anal. (C21H14N2O3‚H2O) C, H, N.
2,6-Dimethyl-9-methoxy-4-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (133). To a stirred solution of 9-hydroxy-4-
phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione11 (3) (0.15 g, 0.46
mmol) in THF/DMF (4:1, 25 mL) under nitrogen was added sodium
hydride (110 mg of a 50% dispersion in oil, 2.3 mmol). After 20
min, excess methyl iodide (1.0 mL) was added and the mixture
was stirred for a further 30 min before being diluted with water
and extracted with ethyl acetate (3×). The combined organic
extracts were washed with brine, dried over anhydrous sodium
sulfate, and concentrated in vacuo. Chromatography on silica,
eluting with ethyl acetate/hexane (1:1), followed by trituration from
methanol, gave 133 (141 mg, 83%) as a yellow powder: mp 237-
238 °C; 1H NMR [(CD3)2SO] δ 8.49 (d, J ) 2.6 Hz, 1H), 7.76 (s,
1H), 7.68-7.61 (m, 3H), 7.51-7.44 (m, 3H), 7.27 (dd, J ) 8.9,
2.6 Hz, 1H), 3.92 (s, 3H), 3.90 (s, 3H), 3.05 (s, 3H). Anal.
(C23H18N2O3) C, H, N.
2,6-Dimethyl-9-hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (20). To a stirred solution of 133 (85 mg, 0.21
mmol) in CH2Cl2 (20 mL) under nitrogen was added boron
tribromide (2.1 mL of a 1 M solution in CH2Cl2, 2.1 mmol). After
2 h, the reaction mixture was diluted with water and extracted with
ethyl acetate (3×). The combined organic extracts were washed
with saturated sodium bicarbonate and then brine before being dried
over anhydrous sodium sulfate and concentrated in vacuo. Chro-
matography on silica, eluting with ethyl acetate/hexane (3:2),
followed by trituration from methanol, gave 20 (56 mg, 69%) as
an orange powder: mp 278-281 °C; 1H NMR [(CD3)2SO] δ 9.35
(s, 1H), 8.40 (d, J ) 2.4 Hz, 1H), 7.74 (s, 1H), 7.65 (m, 2H), 7.52
(d, J ) 8.8 Hz, 1H), 7.50-7.43 (m, 3H), 7.13 (dd, J ) 8.8, 2.4
Hz, 1H), 3.91 (s, 3H), 3.06 (s, 3H). Anal. (C22H16N2O3) C, H, N.
9-Hydroxy-4-phenyl-1H-furo[3,4-c]carbazole-1,3-(6H)-di-
one (21). 9-Methoxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione11 (677 mg, 1.98 mmol) was heated in a melt with pyridine
1
E-isomer: mp 135-137 °C; H NMR (CDCl3) δ 11.39 (s, 1H),
7.86 (dd, J ) 7.8, 1.5 Hz, 1H), 7.49 (dd, J ) 8.0, 1.2 Hz, 1H),
7.43 (d, J ) 16.4 Hz, 1H), 7.37 (m, 1H), 7.31-7.23 (m, 3H), 7.01
(d, J ) 2.4 Hz, 1H), 6.77 (dd, J ) 8.7, 2.4 Hz, 1H), 6.56 (s, 1H),
3.75 (s, 3H). Anal. (C17H14ClNO) C, H, N.
4-(2-Chlorophenyl)-9-methoxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (24). A solution of 113 (1.5 g, 5.29 mmol),
maleimide (0.61 g, 6.34 mmol), and SnCl2 (0.20 g, 1.05 mmol) in
toluene (25 mL) was refluxed for 6 h. After dilution with ethyl
acetate, the solution was washed with water and the organic phase
was dried. The drying agent was removed, and the solution was
concentrated to dryness to give 4-(2-chlorophenyl)-9-methoxy-
4,5,6,10c-tetrahydropyrrolo[3,4-c]carbazole-1,3(2H,3aH)-dione (134)
(IV, 9-OMe, R ) H, Ar ) 2-chlorophenyl) (1.98 g, 98%) as a
yellow solid (a mixture of diastereomers), which was used without
further purification.
Procedure 7. Manganese dioxide (12.0 g) was added to a
solution of 134 (2.1 g, 5.51 mmol) in dioxane (100 mL), and the
mixture was refluxed with stirring for 16 h. The mixture was filtered
hot through a plug of Celite, washing through with more dioxane
and then 10% methanol/dioxane. The combined filtrate and
washings were concentrated to dryness, and the residue was
adsorbed onto silica and chromatographed. Elution with ethyl
acetate/petroleum ether (3:7) gave 24 (1.66 g, 79%), which
crystallized from THF/petroleum ether as a yellow powder: mp
170-175 °C (dec); 1H NMR [(CD3)2SO] δ 11.96 (br s, 1H), 11.08
(br s, 1H), 8.46 (d, J ) 2.6 Hz, 1H), 7.60-7.55 (m, 3H), 7.51-
7.42 (m, 3H), 7.24 (dd, J ) 8.9, 2.6 Hz, 1H), 3.89 (s, 3H). Anal.
(C21H13ClN2O3‚1/2H2O) C, H, N.

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4905
4-(2-Chlorophenyl)-9-methoxy-1H-[1]benzofuro[3,2-e]isoin-
dole-1,3(2H)-dione (136). Reaction of 5-methoxy-1-benzofuran-
2-carbaldehyde24 with (2-chlorobenzyl)(triphenyl)phosphonium bro-
mide using procedure 2 gave 5-methoxy-2-[(E,Z)-2-(2-chlorophenyl)-
ethenyl]-1-benzofuran (135) as a 2:1 mixture of E/Z isomers (24%),
which was used without further purification.
Reaction of 135 with maleimide using procedure 3 and aroma-
tization with MnO2 using procedure 7 gave 136 (49%): mp 246-
248 °C; 1H NMR [(CD3)2SO] δ 11.46 (br s, 1H), 8.22 (d, J ) 2.7
Hz, 1H), 7.99 (s, 1H), 7.80 (d, J ) 9.0 Hz, 1H), 7.61-7.58 (m,
1H), 7.53-7.44 (m, 3H), 7.33 (dd, J ) 9.0, 2.7 Hz, 1H), 3.92 (s,
3H). Anal. (C21H12ClNO4) C, H, N.
treated dropwise with a solution of NaNO2 (13.6 mg, 0.197 mmol)
in cold water (2 × 0.5 mL) and stirred at 7 °C for 8 min. A solution
of urea (5 mg, 0.083 mmol) in cold water (2 × 0.4 mL) was added,
and the mixture was stirred at 7 °C for 4 min. Finally, a suspension
of KI (140 mg, 0.843 mmol) and CuI (140 mg, 0.735 mmol) in
cold water (2 × 1 mL) was added. Then the cooling bath was
removed, and the mixture was stirred for 10 min and then at 60-
65 °C for 2 h. An aqueous solution of sodium metabisulfite (100
mL of 0.5%) was added, and then the mixture was extracted with
20% THF/EtOAc (5 × 100 mL). The extracts were washed with
water (200 mL), the water was extracted with EtOAc, and the
combined organic extracts were concentrated to dryness. The residue
was chromatographed twice on silica gel, eluting with 0-0.5%
MeOH/DCM and 0.5% MeOH/DCM, and then on the second
column with 1% EtOH/CHCl3 to give 140 (V, 9-OMe, R ) H, Ar
) 2-iodophenyl) (18 mg, 30%): mp (MeOH/THF/DCM/hexane)
311-315 °C; 1H NMR [(CD3)2SO] δ 11.95 (br s, 1H), 11.07 (br s,
1H), 8.46 (d, J ) 2.6 Hz, 1H), 7.97 (dd, J ) 7.7, 0.9 Hz, 1H), 7.58
(d, J ) 8.8 Hz, 1H), 7.50 (td, J ) 7.6, 0.9 Hz, 1H), 7.48 (s, 1H),
7.44 (dd, J ) 7.6, 1.8 Hz, 1H), 7.25 (dd, J ) 8.9, 2.7 Hz, 1H),
7.19 (td, J ) 7.6, 1.8 Hz, 1H), 3.89 (s, 3H). Anal. (C21H13IN2O3)
C, H, N.
4-(2-Iodophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione (29). Demethylation of 140 with BBr3 using procedure 5
gave 29 (88%): mp (MeOH/DCM/hexane) 217-223 °C; 1H NMR
[(CD3)2SO] δ 11.82 (br s, 1H), 11.01 (br s, 1H), 9.28 (br s, 1H),
8.32 (d, J ) 2.4 Hz, 1H), 7.96 (br d, J ) 7.8 Hz, 1H), 7.49 (br t,
J ) 7.7 Hz, 1H), 7.46 (d, J ) 8.9 Hz, 1H), 7.42 (s, 1H), 7.42 (dd,
J ) 7.4, 1.7 Hz, 1H), 7.19 (td, J ) 7.6, 1.7 Hz, 1H), 7.08 (dd, J )
8.7, 2.5 Hz, 1H). Anal. (C20H11IN2O3) C, H, N.
(2-Cyanobenzyl)(triphenyl)phosphonium Bromide (141). Bro-
mination of o-tolunitrile with NBS/AIBN, followed by reaction of
the crude bromide with triphenylphosphine (1.2 equiv.), using
procedure 1, except that the reaction time for the bromination was
2 h, gave 141 (70%) as a light-brown powder: mp (CH2Cl2/
benzene) 237-241 °C; 1H NMR (CDCl3) δ 7.90 (dd, J ) 7.8, 2.3
Hz, 1H), 7.85-7.63 (m, 15H), 7.52 (br t, J ) 7.7 Hz, 1H), 7.44
(br d, J ) 7.5 Hz, 1H), 7.38 (tdd, J ) 7.6, 2.1, 1.0 Hz, 1H), 5.86
(d, J ) 14.7 Hz, 2H). Anal. (C26H21BrNP) C, H, N.
4-(2-Chlorophenyl)-9-hydroxy-1H-[1]benzofuro[3,2-e]isoindole-
1,3(2H)-dione (26). Demethylation of 136 with BBr3 using
1
procedure 5 gave 26 (89%): mp 140-145 °C; H NMR [(CD3)2-
SO] δ 11.39 (br s, 1H), 9.78 (s, 1H), 8.12 (d, J ) 2.6 Hz, 1H),
7.93 (s, 1H), 7.66 (d, J ) 8.9 Hz, 1H), 7.60-7.58 (m, 1H), 7.52-
7.43 (m, 3H), 7.13 (dd, J ) 8.9, 2.6 Hz, 1H). EIMS found M+,
363.0294, 365.0269; C20H10ClNO4 requires 363.0298, 365.0289.
2-[(E)-2-(2-Bromophenyl)ethenyl]-5-methoxy-1H-indole (137).
Reaction of 106 and (2-bromobenzyl)(triphenyl)phosphonium bro-
mide using procedure 2 gave 137 (II, 5-OMe, Ar ) 2-bromophenyl)
(88%): mp (DCM/hexane) 120-123 °C; 1H NMR (CDCl3) δ 8.21
(br s, 1H), 7.66 (dd, J ) 7.9, 1.5 Hz, 1H), 7.59 (dd, J ) 8.1, 1.0
Hz, 1H), 7.32 (br t, J ) 7.1 Hz, 1H), 7.27 (d, J ) 8.3 Hz, 1H),
7.22 (d, J ) 16.4 Hz, 1H), 7.12 (td, J ) 7.6, 1.5 Hz, 1H), 7.05 (d,
J ) 16.4 Hz, 1H), 7.04 (d, J ) 2.4 Hz, 1H), 6.88 (dd, J ) 8.8, 2.4
Hz, 1H), 6.59 (d, J ) 1.7 Hz, 1H), 3.86 (s, 3H). Anal. (C17H14-
BrNO) C, H, N.
4-(2-Bromophenyl)-9-methoxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (138). Maleimide (1.48 g, 15.2 mmol) and 137 (1.00
g, 3.05 mmol) were reacted in dry toluene (10 mL) in a sealed vial
at reflux for 24 h, and the product was treated with DDQ (3.49 g,
15.4 mmol) in toluene (30 mL) and dioxane (40 mL), stirring under
reflux for 24 h. The resulting mixture was partitioned between
saturated aqueous NaHCO3 (250 mL) and 15% MeOH/DCM. The
organic extracts were washed with aqueous NaHCO3 solution and
water, the aqueous layers were back-extracted with 15% MeOH/
DCM, and the combined organic extracts were concentrated to
dryness. The residue was chromatographed on silica gel, eluting
with 0-0.75% MeOH/DCM and then with 0.75% MeOH/DCM,
to give 138 (V, 9-OMe, R ) H, Ar ) 2-bromophenyl) (1.23 g,
96%): mp (MeOH/DCM/hexane) 273-275 °C; 1H NMR [(CD3)2-
SO] δ 11.96 (br s, 1H), 11.08 (br s, 1H), 8.46 (d, J ) 2.6 Hz, 1H),
7.74 (d, J ) 7.8 Hz, 1H), 7.58 (d, J ) 8.9 Hz, 1H), 7.55 (s, 1H),
7.48 (m, 2H), 7.39 (m, 1H), 7.25 (dd, J ) 8.9, 2.7 Hz, 1H), 3.89
(s, 3H). Anal. (C21H13BrN2O3) C, H, N.
2-[(E)-2-(5-Methoxy-1H-indol-2-yl)ethenyl]benzonitrile (142).
5-Methoxy-1H-indole-2-carbaldehyde (106) was reacted with 141
as described in procedure 2, except that the LDA and aldehyde
were (sequentially) added at 0 °C and the ratio LDA/aldehyde was
1.55:1, to give (after crystallization from CH2Cl2/hexane) the diene
142 (II, 5-OMe, Ar ) 2-cyanophenyl) as a yellow solid (the pure
1
E isomer) (60%): mp 192-196 °C; H NMR (CDCl3) δ 8.40 (br
s, 1H), 7.79 (d, J ) 8.1 Hz, 1H), 7.66 (dd, J ) 8.1, 0.9 Hz, 1H),
7.58 (td, J ) 7.9, 1.3 Hz, 1H), 7.32 (td, J ) 8.0, 0.9 Hz, 1H), 7.29
(d, J ) 8.5 Hz, 1H), 7.28 (d, J ) 16.9 Hz, 1H), 7.18 (d, J ) 16.4
Hz, 1H), 7.04 (d, J ) 2.4 Hz, 1H), 6.91 (dd, J ) 8.9, 2.4 Hz, 1H),
6.64 (d, J ) 1.8 Hz, 1H), 3.86 (s, 3H). Anal. (C18H14N2O) C, H,
N.
4-(2-Bromophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (28). Demethylation of 138 with BBr3 for 4 h using
procedure 5 gave 28 (89%): mp (MeOH/DCM/hexane) 244-246
°C; 1H NMR [(CD3)2SO] δ 11.83 (br s, 1H), 11.01 (br s, 1H), 9.29
(br s, 1H), 8.31 (d, J ) 2.4 Hz, 1H), 7.73 (br d, J ) 7.9 Hz, 1H),
7.47 (m, 4H), 7.38 (m, 1H), 7.08 (dd, J ) 8.7, 2.5 Hz, 1H). Anal.
(C20H11BrN2O3‚1/2H2O) C, H, N.
2-(9-Methoxy-1,3-dioxo-1,2,3,6-tetrahydropyrrolo[3,4-c]car-
bazol-4-yl)benzonitrile (143). The E diene 142 was subjected to
successive reactions with maleimide and then DDQ as described
in procedures 3 and 4 except that the poorly soluble final product
was obtained by removal of the reaction solvents, washing of the
solid residue thoroughly with aqueous NaHCO3 solution and water,
and then trituration in 15% MeOH/CH2Cl2 and crystallization of
the liquors from THF/CH2Cl2/hexane to give 143 (V, 9-OMe, R )
H, Ar ) 2-cyanophenyl) (96%) as a yellow solid: mp 348-350
°C; 1H NMR [(CD3)2SO] δ 12.06 (br s, 1H), 11.19 (br s, 1H), 8.48
(d, J ) 2.6 Hz, 1H), 7.96 (dd, J ) 7.7, 1.0 Hz, 1H), 7.82 (td, J )
7.7, 1.4 Hz, 1H), 7.71 (br d, J ) 7.4 Hz, 1H), 7.71 (s, 1H), 7.66
(td, J ) 7.6, 1.1 Hz, 1H), 7.60 (d, J ) 8.9 Hz, 1H), 7.27 (dd, J )
8.8, 2.6 Hz, 1H), 3.90 (s, 3H). EIMS found M+, 367.0956;
C22H13N3O3 requires 367.0957.
4-(2-Aminophenyl)-9-methoxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (139). A solution of 149 (see later for preparation)
(71 mg, 0.183 mmol) in 2:1 THF/MeOH (45 mL) containing wet
10% Pd-C (71 mg) was hydrogenated at 60 psi for 7 h. The
solution was filtered through Celite and washed thoroughly with
MeOH and THF. Then the filtrate was concentrated to dryness to
give 139 (V, 9-OMe, R ) H, Ar ) 2-aminophenyl) (67 mg, 97%)
1
as a yellow solid: mp (MeOH/DCM/hexane) >320 °C; H NMR
[(CD3)2SO] δ 11.80 (br s, 1H), 10.94 (br s, 1H), 8.47 (d, J ) 2.6
Hz, 1H), 7.54 (d, J ) 8.8 Hz, 1H), 7.51 (s, 1H), 7.21 (dd, J ) 8.8,
2.6 Hz, 1H), 7.08 (td, J ) 7.7, 1.5 Hz, 1H), 7.01 (dd, J ) 7.6, 1.5
Hz, 1H), 6.73 (dd, J ) 8.1, 0.6 Hz, 1H), 6.61 (td, J ) 7.4, 0.9 Hz,
1H), 4.70 (s, 2H), 3.89 (s, 3H). Anal. (C21H15N3O3‚H2O) C, H, N.
4-(2-Iodophenyl)-9-methoxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione (140). A solution of 139 (46 mg, 0.129 mmol) in 90% H2-
SO4 (2.5 mL) was cooled to -5 °C, diluted with ice-water (2.5
mL), then allowed to warm to 7 °C. The resulting suspension was
2-(9-Hydroxy-1,3-dioxo-1,2,3,6-tetrahydropyrrolo[3,4-c]car-
bazol-4-yl)benzonitrile (34). The methyl ether 143 was demethy-
lated with BBr3 using procedure 5, except that the reaction time
was 5 h with 5 equiv of BBr3 and then a further 4 h with an extra

4906 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
5 equiv of BBr3, to give (after crystallization from THF/CH2Cl2/
(d, J ) 2.6 Hz, 1H), 7.50 (m, 5H), 7.38 (s, 1H), 7.20 (dd, J ) 8.9,
2.6 Hz, 1H), 7.10 (m, 5H), 3.86 (s, 3H). Anal. (C27H18N2O3) C, H,
N.
hexane) the phenol 34 (83%) as a yellow solid: mp 199-204 °C
1
(dec); H NMR [(CD3)2SO] δ 11.93 (br s, 1H), 11.13 (br s, 1H),
9.31 (br s, 1H), 8.34 (d, J ) 2.4 Hz, 1H), 7.96 (dd, J ) 7.6, 0.9
Hz, 1H), 7.82 (td, J ) 7.7, 1.4 Hz, 1H), 7.70 (br d, J ) 7.5 Hz,
1H), 7.65 (s, 1H), 7.65 (td, J ) 7.7, 1.0 Hz, 1H), 7.48 (d, J ) 8.7
Hz, 1H), 7.10 (dd, J ) 8.7, 2.5 Hz, 1H). Anal. (C21H11N3O3‚1/
4H2O) C, H, N.
4-[1,1′-Biphenyl]-2-yl-9-hydroxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (37). Demethylation of 146 with BBr3 for 4 h using
procedure 5 gave 37 (98%): mp (MeOH/DCM/hexane) 198-203
1
°C (dec). H NMR [(CD3)2SO] δ 11.61 (br s, 1H), 10.82 (br s,
1H), 9.22 (br s, 1H), 8.24 (d, J ) 2.4 Hz, 1H), 7.52 (m, 1H), 7.45
(m, 3H), 7.37 (d, J ) 8.7 Hz, 1H), 7.31 (s, 1H), 7.11 (m, 5H), 7.03
(dd, J ) 8.7, 2.5 Hz, 1H). Anal. (C26H16N2O3‚1/4H2O) C, H, N.
4-(2-Hydroxyphenyl)-9-methoxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (147). Diazotization of 139 as described above for
the preparation of 140 and elution of the first chromatography
column with 1% MeOH/DCM gave 147 (V, 9-OMe, R ) H, Ar )
2-hydroxyphenyl) (24%): mp (THF/DCM/pentane) 264-267 °C;
1H NMR [(CD3)2SO] δ 11.82 (br s, 1H), 10.95 (br s, 1H), 9.41 (br
s, 1H), 8.47 (d, J ) 2.6 Hz, 1H), 7.56 (s, 1H), 7.54 (d, J ) 8.9 Hz,
1H), 7.25 (dd, J ) 7.4, 1.6 Hz, 1H), 7.23 (td, J ) 7.9, 1.7 Hz, 1H),
7.22 (dd, J ) 8.7, 2.6 Hz, 1H), 6.91 (br d, J ) 7.5 Hz, 1H), 6.87
(td, J ) 7.4, 0.9 Hz, 1H), 3.89 (s, 3H). Anal. (C21H14N2O4) C, H,
N.
9-Hydroxy-4-(2-hydroxyphenyl)pyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (38). Demethylation of 147 with BBr3 for 4 h using
procedure 5 gave 38 (73%): mp (MeOH/THF/CH2Cl2/hexane)
291-296 °C; 1H NMR [(CD3)2SO] δ 11.67 (br s, 1H), 10.88 (br s,
1H), 9.37 (br s, 1H), 9.22 (br s, 1H), 8.31 (d, J ) 2.4 Hz, 1H),
7.51 (s, 1H), 7.42 (d, J ) 8.7 Hz, 1H), 7.24 (dd, J ) 7.5, 1.5 Hz,
1H), 7.22 (td, J ) 7.7, 1.8 Hz, 1H), 7.05 (dd, J ) 8.7, 2.5 Hz, 1H),
6.91 (br d, J ) 8.0 Hz, 1H), 6.86 (td, J ) 7.4, 0.9 Hz, 1H). Anal.
(C20H12N2O4‚H2O) C, H, N.
2-(9-Methoxy-1,3-dioxo-1,2,3,6-tetrahydropyrrolo[3,4-c]car-
bazol-4-yl)benzamide (145). Hydrogen peroxide (0.5 mL of a 35%
aqueous solution, 5.8 mmol) was added dropwise to a stirred
mixture of carbazole methyl ether 143 (61 mg, 0.166 mmol) and
potassium carbonate (229 mg, 1.66 mmol) in DMSO (3 mL), and
then the mixture was stirred at room temperature for 26 h. Further
hydrogen peroxide (0.5 mL) in DMSO (3 mL) was added, and the
mixture was stirred at 50 °C for 4 days. The cooled solution was
diluted with water (100 mL) and extracted with EtOAc (4 × 100
mL). The aqueous portion was then acidified (to pH 2) with
concentrated HCl and extracted with EtOAc (4 × 100 mL). These
extracts were washed with water (100 mL), back-extracted with
EtOAc (100 mL), and evaporated to dryness to give crude 2-(9-
methoxy-1,3-dioxo-3,6-dihydro-1H-furo[3,4-c]carbazol-4-yl)benza-
mide (144) (75 mg) as a solid, which was used directly: 1H NMR
[(CD3)2SO] δ 12.26 (br s, 1 H), 8.27 (d, J ) 2.5 Hz, 1 H), 7.74 (s,
1 H), 7.71-7.48 (m, 6 H), 7.30 (dd, J ) 8.9, 2.6 Hz, 1 H), 7.21
(br s, 1 H), 3.91 (s, 3 H). FABMS found [M]+, 386.0905;
C22H14N2O5 requires 386.0903.
A mixture of crude anhydride 144 (73 mg) and ammonium
acetate (10 g) was stirred at 141 °C for 7 h. The cooled solution
was added to a mixture of ice and aqueous sodium bicarbonate
(100 mL) and extracted with EtOAc (5 × 100 mL). The combined
extracts were washed with water (200 mL) and evaporated to
dryness, and then the residue was recrystallized to give 145 (V,
9-OMe, R ) H, Ar ) 2-benzamide) (55 mg, 76% for 2 steps) as
a yellow solid: mp (MeOH/THF/EtOAc/pentane) 345-348 °C; 1H
NMR [(CD3)2SO] δ 11.87 (br s, 1 H), 10.97 (br s, 1 H), 8.46 (d,
J ) 2.6 Hz, 1 H), 7.62 (m, 1 H), 7.56-7.38 (m, 6 H), 7.22 (dd, J
) 8.8, 2.5 Hz, 1 H), 7.14 (br s, 1 H), 3.89 (s, 3 H). Anal.
(C22H15N3O4) C, H, N.
5-Methoxy-2-[(E)-2-(2-nitrophenyl)ethenyl]-1H-indole (148).
Reaction of 106 and (2-nitrobenzyl)(triphenyl)phosphonium chloride
using procedure 2 gave 148 (II, 5-OMe, Ar ) 2-nitrophenyl)
(47%): mp (DCM/hexane) 136-138 °C; 1H NMR (CDCl3) δ 8.25
(br s, 1H), 7.98 (dd, J ) 8.2, 1.2 Hz, 1H), 7.78 (dd, J ) 7.9, 1.0
Hz, 1H), 7.61 (td, J ) 7.6, 0.9 Hz, 1H), 7.41 (d, J ) 16.3 Hz, 1H),
7.39 (td, J ) 7.8, 1.3 Hz, 1H), 7.27 (m, 1H), 7.13 (d, J ) 16.3 Hz,
1H), 7.04 (d, J ) 2.4 Hz, 1H), 6.90 (dd, J ) 8.8, 2.5 Hz, 1H), 6.62
(d, J ) 1.8 Hz, 1H), 3.86 (s, 3H). Anal. (C17H14N2O3) C, H, N.
9-Methoxy-4-(2-nitrophenyl)pyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (149). Reaction of 148 successively with maleimide
using procedure 3 and then DDQ using procedure 4 gave 149 (V,
9-OMe, R ) H, Ar ) 2-nitrophenyl) (97%): mp (MeOH/EtOAc/
2-(9-Hydroxy-1,3-dioxo-1,2,3,6-tetrahydropyrrolo[3,4-c]car-
bazol-4-yl)benzamide (36). Boron tribromide (2.0 mL of a 1.0 M
solution in CH2Cl2, 2.0 mmol) was added to a stirred solution of
carbazole methyl ether 145 (44.6 mg, 0.103 mmol) in dry CH2Cl2
(3 mL) under N2, and then the mixture was stirred at room
temperature for 5 h. The resulting solution was added to a mixture
of ice and aqueous sodium bicarbonate (100 mL) and extracted
with EtOAc (5 × 100 mL). The combined extracts were washed
with water (200 mL) and evaporated to dryness, and then the residue
was chromatographed on silica gel. Elution with 50-80% EtOAc/
light petroleum gave foreruns. Then further elution with 80%
EtOAc/light petroleum and EtOAc gave 36 (34 mg, 89%) as an
orange solid following recrystallization: mp (THF/hexane) 330-
1
DCM/hexane) 294-298 °C; H NMR [(CD3)2SO] δ 12.01 (br s,
1H), 11.10 (br s, 1H), 8.44 (d, J ) 2.5 Hz, 1H), 8.21 (dd, J ) 8.1,
1.0 Hz, 1H), 7.86 (td, J ) 7.5, 1.4 Hz, 1H), 7.74 (td, J ) 7.8, 1.3
Hz, 1H), 7.67 (s, 1H), 7.65 (dd, J ) 7.6, 1.4 Hz, 1H), 7.59 (d, J )
8.9 Hz, 1H), 7.25 (dd, J ) 8.7, 2.6 Hz, 1H), 3.90 (s, 3H). Anal.
(C21H13N3O5) C, H, N.
9-Hydroxy-4-(2-nitrophenyl)pyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (43). Demethylation of 149 using procedure 5 for
5 h with 5 equiv of BBr3 and then for a further 21 h with an extra
5 equiv of BBr3 gave 43 (53%): mp (MeOH/DCM/hexane) 322-
330 °C; 1H NMR [(CD3)2SO] δ 11.88 (br s, 1H), 11.05 (br s, 1H),
9.29 (br s, 1H), 8.30 (d, J ) 2.4 Hz, 1H), 8.20 (dd, J ) 8.2, 1.1
Hz, 1H), 7.85 (td, J ) 7.5, 1.2 Hz, 1H), 7.73 (td, J ) 7.9, 1.4 Hz,
1H), 7.63 (dd, J ) 7.6, 1.4 Hz, 1H), 7.61 (s, 1H), 7.47 (d, J ) 8.7
Hz, 1H), 7.09 (dd, J ) 8.7, 2.5 Hz, 1H). Anal. (C20H11N3O5‚1/2-
MeOH) C, H, N.
4-(2-Aminophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (44). A solution of 43 (81 mg, 0.217 mmol) in 2:1
THF/MeOH (60 mL) containing wet 10% Pd-C (81 mg) was
hydrogenated at 60 psi for 12 h. The solution was filtered through
Celite, the Celite and catalyst were washed thoroughly with MeOH
and THF, and the combined filtrates were evaporated to give 44
(60 mg, 81%): mp (MeOH/DCM/hexane) >330 °C; 1H NMR
[(CD3)2SO] δ 11.67 (br s, 1H), 10.88 (br s, 1H), 9.21 (s, 1H), 8.32
(d, J ) 2.4 Hz, 1H), 7.46 (s, 1H), 7.42 (d, J ) 8.6 Hz, 1H), 7.08
(td, J ) 7.9, 1.5 Hz, 1H), 7.05 (dd, J ) 8.7, 2.4 Hz, 1H), 7.00 (dd,
J ) 7.6, 1.4 Hz, 1H), 6.72 (br d, J ) 8.1 Hz, 1H), 6.60 (td, J )
7.4, 0.9 Hz, 1H), 4.69 (s, 2H). Anal. (C20H13N3O3‚H2O) C, H, N.
1
334 °C; H NMR [(CD3)2SO] δ 11.73 (br s, 1 H), 10.91 (br s, 1
H), 9.23 (br s, 1 H), 8.31 (d, J ) 2.4 Hz, 1 H), 7.61 (m, 1 H),
7.54-7.37 (m, 6 H), 7.13 (br s, 1 H), 7.05 (dd, J ) 8.7, 2.5 Hz, 1
H). Anal. (C21H13N3O4) C, H, N.
4-[1,1′-Biphenyl]-2-yl-9-methoxypyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (146). A mixture of bromide 138 (85.5 mg, 0.203
mmol), [1,1′-bis(diphenylphosphino)ferrocene]dichloropalladium-
(II)‚DCM (85 mg, 0.104 mmol), and tetraphenyltin (0.451, 1.06
mmol) in dry DMF (2.5 mL) was stirred in a sealed vial at 130 °C
for 4 days. The mixture was then partitioned between aqueous
NaHCO3 (100 mL) and 2:1 CH2Cl2/EtOAc (5 × 80 mL), and the
combined organic extracts were concentrated to dryness. The residue
was chromatographed several times on silica gel, eluting with DCM,
to give the crude product. This was further purified by preparative
reversed phase C-18 HPLC, using a gradient of 65-99% MeCN/
aqueous HCO2NH4 buffer, pH 3.45, to give 146 (V, 9-OMe, R )
H, Ar ) 2-biphenyl) (21%): mp (MeOH/DCM/hexane) 220-223
°C; 1H NMR [(CD3)2SO] δ 11.76 (br s, 1H), 10.87 (br s, 1H), 8.39

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4907
2-[(E)-2-(2-Chloro-5-methoxyphenyl)ethenyl]-5-methoxy-1H-
indole (150). Reaction of 106 and (2-chloro-5-methoxybenzyl)-
(triphenyl)phosphonium bromide26 using procedure 2 gave 150 (II,
5-OMe, Ar ) 2-chloro-5-methoxyphenyl) (85%): mp (DCM/
The extracts were washed with water, concentrated, and chromato-
graphed on silica gel. Elution with 0-2% MeOH/CH2Cl2 and then
3% MeOH/CH2Cl2 gave 77 (97%): mp (THF/DCM/pentane) 301-
1
306 °C (dec); H NMR [(CD3)2SO] δ 11.78 (br s, 1H), 10.97 (br
1
hexane) 119-121 °C; H NMR (CDCl3) δ 8.22 (br s, 1H), 7.29
s, 1H), 9.26 (br s, 1H), 8.31 (d, J ) 2.4 Hz, 1H), 7.45 (s, 1H), 7.45
(d, J ) 8.7 Hz, 1H), 7.13 (d, J ) 8.3 Hz, 1H), 7.07 (dd, J ) 8.7,
2.5 Hz, 1H), 6.62 (dd, J ) 8.2, 2.8 Hz, 1H), 6.60 (d, J ) 2.3 Hz,
1H), 5.29 (s, 2H). Anal. (C20H12ClN3O3‚1/4H2O) C, H, N.
(d, J ) 8.9 Hz, 1H), 7.26 (br d, J ) 8.7 Hz, 1H), 7.22 (d, J ) 16.5
Hz, 1H), 7.18 (d, J ) 3.0 Hz, 1H), 7.06 (d, J ) 16.7 Hz, 1H), 7.04
(br s, 1 H), 6.88 (dd, J ) 8.7, 2.5 Hz, 1H), 6.77 (dd, J ) 8.8, 3.0
Hz, 1H), 6.59 (d, J ) 1.8 Hz, 1H), 3.86 (s, 3H), 3.85 (s, 3H). Anal.
(C18H16ClNO2) C, H, N.
4-(2-Chloro-5-methoxyphenyl)-9-methoxypyrrolo[3,4-c]car-
bazole-1,3(2H,6H)-dione (151). Successive reaction of 150 with
maleimide using procedure 3 and then with DDQ using procedure
4 gave 151 (V, 9-OMe, R ) H, Ar ) 2-chloro-5-methoxyphenyl)
(84%): mp (MeOH/DCM/hexane) 284-286 °C; 1H NMR [(CD3)2-
SO] δ 11.96 (br s, 1H), 11.08 (br s, 1H), 8.46 (d, J ) 2.6 Hz, 1H),
7.58 (s, 1H), 7.57 (m, 1H), 7.46 (d, J ) 8.7 Hz, 1H), 7.25 (dd, J
) 8.8, 2.6 Hz, 1H), 7.07 (d, J ) 2.9 Hz, 1H), 7.04 (dd, J ) 8.7,
3.1 Hz, 1H), 3.89 (s, 3H), 3.80 (s, 3H). Anal. (C22H15ClN2O4‚H2O)
C, H, N.
Procedures of Scheme 2. Benzyl (2E)-3-(5-Methoxy-1H-indol-
2-yl)-2-propenoate (107). Benzyl (triphenylphosphoranylidene)-
acetate (49.2 g, 0.120 mol) was added to a stirred solution of
5-methoxy-1H-indole-2-carbaldehyde (106) (20.0 g, 0.114 mol) in
DCM (500 mL), and the solution was stirred at room temperature
for 4 h. The solvent was evaporated, and the residue was slurried
with MeOH (200 mL), filtered, and washed with cold MeOH to
1
give 107 (30.46 g, 87%): mp 155-157 °C; H NMR (CDCl3) δ
8.28 (s, 1H), 7.69 (d, J ) 16.1 Hz, 1H), 7.43-7.32 (m, 5H), 7.23
(d, J ) 9.1 Hz, 1H), 7.03 (d, J ) 2.3 Hz, 1H), 6.92 (dd, J ) 9.1,
2.3 Hz, 1H), 6.74 (d, J ) 1.8 Hz, 1H), 6.24 (d, J ) 16.1 Hz, 1H),
5.26 (s, 2H), 3.84 (s, 3H). Anal. (C19H17NO3) C, H, N.
4-(2-Chloro-5-hydroxyphenyl)-9-hydroxypyrrolo[3,4-c]carba-
zole-1,3(2H,6H)-dione (76). Bis-demethylation of 151 for 6.5 h
with 10 equiv of BBr3 using procedure 5 gave 76 (90%): mp
(MeOH/DCM/hexane) 335-340 °C; 1H NMR [(CD3)2SO] δ 11.80
(br s, 1H), 11.00 (br s, 1H), 9.78 (br s, 1H), 9.27 (br s, 1H), 8.31
(d, J ) 2.4 Hz, 1H), 7.49 (s, 1H), 7.45 (d, J ) 8.7 Hz, 1H), 7.32
(d, J ) 8.5 Hz, 1H), 7.07 (dd, J ) 8.7, 2.5 Hz, 1H), 6.84 (dd, J )
8.5, 2.9 Hz, 1H), 6.82 (d, J ) 2.8 Hz, 1H). Anal. (C20H11ClN2O4‚
1/2H2O‚1/4hexane) C, H, N.
Procedure 8. Benzyl 9-Methoxy-1,3-dioxo-1,2,3,6-tetrahydro-
pyrrolo[3,4-c]carbazole-4-carboxylate (109). Maleimide (4.82 g,
0.050 mol) was added to a solution of 107 (12.71 g, 0.041 mol) in
THF (150 mL), and the mixture was stirred until it was homoge-
neous. The THF was evaporated, and the residue was dried under
high vacuum for 30 min and then stirred at 175 °C for 3 h. The
solid melt was cooled and stirred vigorously with EtOAc (100 mL)
overnight. The precipitate was filtered and washed with Et2O to
give benzyl 9-methoxy-1,3-dioxo-1,2,3,3a,4,5,6,10c-octahydropy-
rrolo[3,4-c]carbazole-4-carboxylate (108) (13.76 g, 83%), which
was used directly: mp 179-181 °C; 1H NMR [(CD3)2SO] δ 10.98
(br s, 1H), 10.87 (s, 1H), 7.45-7.32 (m, 5H), 7.20 (d, J ) 2.4 Hz,
1H), 7.17 (d, J ) 8.7 Hz, 1H), 6.69 (dd, J ) 8.7, 2.4 Hz, 1H), 5.21
(s, 1H), 4.22 (br d, J ) 7.8 Hz, 1H), 4.14 (dd, J ) 7.8, 4.2 Hz,
1H), 3.75 (s, 3H), 3.19-3.13 (m, 1H), 3.00 (dd, J ) 16.5, 4.8 Hz,
1H), 2.79-2.70 (m, 1H).
Activated MnO2 (69 g) was added to a solution of 108 (13.76 g,
0.034 mol) in dioxane (300 mL), and the mixture was refluxed
with vigorous stirring for 2 h. The mixture was filtered while hot
through a plug of Celite, which was washed exhaustively with a
MeOH/p-dioxane (1:1) mixture until the washings were colorless.
The combined washings and filtrate were concentrated to dryness,
and the residue was triturated several times with Et2O to give 109
(12.47 g, 91%): mp 245 °C. A sample was crystallized from
EtOAc: mp 289-292 °C; 1H NMR [(CD3)2SO] δ 12.11 (br s, 1H),
11.27 (br s, 1H), 8.45 (d, J ) 2.6 Hz, 1H), 7.98 (s, 1H), 7.59 (d,
J ) 8.9 Hz, 1H), 7.55-7.51 (m, 2H), 7.44-7.34 (m, 3H), 7.27
(dd, J ) 8.9, 2.6 Hz, 1H), 5.41 (s, 2H), 3.88 (s, 3H). Anal.
(C23H16N2O5) C, H, N.
9-Methoxy-1,3-dioxo-1,2,3,6-tetrahydropyrrolo[3,4-c]carba-
zole-4-carboxylic Acid (110). A solution of 109 (2.00 g) in DMF/
MeOH (4:1) (50 mL) containing 5% Pd-C (0.50 g) was hydro-
genated at 60 psi for 2 h (Parr apparatus). The solution was filtered
through a plug of Celite, which was washed six times with neat
DMF followed by MeOH (several cycles). The combined filtrate
and washings were concentrated to dryness, and the residue was
slurried with Et2O and filtered. The products from five identical
mixtures were combined to give crude 110 (8.1 g, 84%). A sample
was purified by chromatography on silica gel, eluting with EtOAc,
followed by trituration with Et2O: mp >300 °C; 1H NMR [(CD3)2-
SO] δ 12.15 (s, 1H), 8.42 (d, J ) 2.5 Hz, 1H), 7.97 (s, 1H), 7.57
(d, J ) 8.8 Hz, 1H), 7.25 (dd, J ) 8.8, 2.5 Hz, 1H), 3.82 (s, 3H),
3.40 (br, 2H). Anal. (C16H10N2O5‚H2O) C, H, N.
4-Amino-9-methoxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (111). Diphenylphosphorylazide (1.81 mL, 8.38 mmol) was
added to a mixture of 110 (2.55 g, 8.22 mmol) and Et3N (1.17 mL,
8.38 mmol) in anhydrous t-BuOH (300 mL), and the mixture was
refluxed under nitrogen for 16 h. The solution was evaporated, and
the residue was partitioned between EtOAc and saturated aqueous
NaHCO3. Insoluble material was removed by filtration of the two
layers through Celite, washing with more EtOAc. The organic phase
(2-Chloro-5-nitrobenzyl)(triphenyl)phosphonium Bromide
(152). Bromination of (2-chloro-5-nitrophenyl)methanol with 30%
HBr in acetic acid, followed by reaction of the crude bromide with
triphenylphosphine using procedure 1, gave 152 (63%): mp (CH2-
1
Cl2/benzene) 239-243 °C; H NMR (CDCl3) δ 8.22 (br s, 1 H),
8.07 (br d, J ) 8.7 Hz, 1 H), 7.87-7.65 (m, 15 H), 7.41 (d, J )
8.9 Hz, 1 H), 5.80 (d, J ) 14.8 Hz, 2 H). Anal. (C25H20BrClNO2P)
C, H, N.
2-[(E)-2-(2-Chloro-5-nitrophenyl)ethenyl]-5-methoxy-1H-in-
dole (153). Reaction of 106 and 152 using procedure 2 gave 153
(II, 5-OMe, Ar ) 2-chloro-5-nitrophenyl) (57%): mp (DCM/
1
pentane) 191-193 °C; H NMR (CDCl3) δ 8.54 (d, J ) 2.6 Hz,
1H), 8.25 (br s, 1H), 8.02 (dd, J ) 8.8, 2.6 Hz, 1H), 7.56 (d, J )
8.9 Hz, 1H), 7.28 (d, J ) 8.7 Hz, 1H), 7.26 (d, J ) 16.4 Hz, 1H),
7.20 (d, J ) 16.5 Hz, 1H), 7.06 (d, J ) 2.4 Hz, 1H), 6.91 (dd, J )
8.8, 2.5 Hz, 1H), 6.70 (br s, 1H), 3.86 (s, 3H). Anal. (C17H13ClN2O3‚
1/4H2O) C, H, N.
4-(2-Chloro-5-nitrophenyl)-9-methoxypyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (154). Successive reaction of 153 with maleimide
using procedure 3 and then DDQ using procedure 4 gave 154 (V,
9-OMe, R ) H, Ar ) 2-chloro-5-nitrophenyl) (95%): mp THF/
1
DCM/pentane) 285-287 °C; H NMR [(CD3)2SO] δ 12.06 (br s,
1H), 11.16 (br s, 1H), 8.46 (d, J ) 2.6 Hz, 1H), 8.36 (d, J ) 2.8
Hz, 1H), 8.33 (dd, J ) 8.7, 2.7 Hz, 1H), 7.91 (d, J ) 8.7 Hz, 1H),
7.71 (s, 1H), 7.60 (d, J ) 8.9 Hz, 1H), 7.27 (dd, J ) 8.9, 2.6 Hz,
1H), 3.90 (s, 3H). Anal. (C21H12ClN3O5‚1/4CH2Cl2) C, H, N.
4-(2-Chloro-5-nitrophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (155). Demethylation of 154 using procedure 5
for 6 h with 10 equiv of BBr3 and then for a further 4 h with an
extra 10 equiv of BBr3 gave 155 (VI, 9-OH, R ) H, Ar ) 2-chloro-
5-nitrophenyl) (88%): mp (THF/CH2Cl2/pentane) 268 °C (dec);
1H NMR [(CD3)2SO] δ 11.94 (br s, 1H), 11.11 (br s, 1H), 9.31 (br
s, 1H), 8.35 (d, J ) 2.6 Hz, 1H), 8.32 (dd, J ) 8.6, 2.9 Hz, 1H),
8.32 (d, J ) 2.4 Hz, 1H), 7.91 (d, J ) 8.7 Hz, 1H), 7.66 (s, 1H),
7.48 (d, J ) 8.7 Hz, 1H), 7.10 (dd, J ) 8.7, 2.5 Hz, 1H). Anal.
(C20H10ClN3O5‚1/4H2O) C, H, N.
4-(5-Amino-2-chlorophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (77). A mixture of 155 (70 mg, 0.172 mmol)
and freshly prepared (wet) nickel boride (266 mg) in MeOH (5.6
mL) and 1 M HCl (1.4 mL) was stirred at reflux for 3 h.
Concentrated aqueous ammonia and aqueous NaHCO3 (100 mL)
were added, and the mixture extracted with EtOAc (5 × 70 mL).

4908 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
was dried and evaporated, and the residue was dissolved in DCM/
TFA (1:1) (200 mL) and kept at room temperature for 1 h. The
solvents were evaporated, and the residue was partitioned between
EtOAc and saturated aqueous NaHCO3 solution. The EtOAc
solution was dried and evaporated, and the residue was chromato-
graphed on silica gel, eluting with EtOAc/petroleum ether (1:1)
followed by EtOAc and then MeOH/EtOAc (1:9), to give 111 (2.16
g, 93%): mp 342-345 °C; 1H NMR [(CD3)2SO] δ 11.18 (s, 1H),
10.78 (br s, 1H), 8.18 (d, J ) 2.5 Hz, 1H), 7.29 (d, J ) 8.7 Hz,
1H), 7.01 (dd, J ) 8.7, 2.5 Hz, 1H), 6.83 (s, 1H), 6.28 (br s, 2H),
3.82 (s, 3H). Anal. (C15H11N3O3‚1/2H2O) C, H, N.
SO] δ 11.58 (br s, 1H), 7.34 (d, J ) 9.0 Hz, 1H), 7.23 (s, 1H),
7.12 (d, J ) 2.4 Hz, 1H), 6.94 (dd, J ) 9.0, 2.4 Hz, 1H), 3.77 (s,
3H), 2.52 (s, 3H). Anal. (C11H11NO2) C, H, N.
Procedure 11. 9-Hydroxy-5-methyl-4-phenylpyrrolo[3,4-c]-
carbazole-1,3(2H,6H)-dione (11). A suspension of benzyl(triph-
enyl)phosphonium bromide (3.4 g, 7.9 mmol) in THF (30 mL) was
treated with a solution of LDA (4.9 mL of a 1.5 M solution in
cyclohexane, 7.4 mmol). The red-orange reaction mixture was
stirred for 10 min, and then a solution of 156 (1.0 g, 5.3 mmol) in
THF (15 mL) was added. The mixture was heated at reflux
overnight. Then water was added and the solvent was removed
under reduced pressure. The residue was extracted with EtOAc (3
× 50 mL), and the combined extracts were dried and evaporated.
The residue was chromatographed on silica gel, eluting with DCM,
to give methyl 2-[(E,Z)-1-methyl-2-phenylethenyl]-1H-indol-5-yl
ether (157) (X, X ) Me, Y ) phenyl) (0.26 g, 19%) as a mixture
of E and Z isomers, which was used without further purification.
Reaction of 157 with maleimide at 180 °C for 40 min using
procedure 8, followed by aromatization with MnO2 using procedure
7, gave 9-methoxy-5-methyl-4-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (158) (XI, X ) Me, Y ) phenyl), which was used
directly.
4-Iodo-9-methoxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione
(112). Concentrated H2SO4 (10 mL) was added at room temperature
to powdered 111 (0.50 g, 1.78 mmol), and the mixture was stirred
for 5 min and then cooled in an ice bath. An ice-water slurry (∼40
mL) was added in one portion with vigorous stirring, and the
mixture was stirred for a further 15 min. When the internal
temperature had reached 3 °C, a solution of NaNO2 (0.18 g, 2.65
mmol) in cold water (1 mL) was added dropwise over 30 s, and
the mixture was stirred for an additional 3 min. Powdered urea (74
mg, 1.23 mmol) was added, and the mixture was stirred for another
3 min. Finally a suspension of KI (1.46 g, 8.79 mmol) and CuI
(1.46 g, 7.66 mmol) in cold water (10 mL) was added, and the
mixture was stirred vigorously for 5 min and then warmed slowly
to 70 °C and held at this temperature for 1 h. EtOAc was added,
and the two-phase mixture was filtered through a plug of Celite
and washed with more EtOAc. The combined organic portions were
washed with 0.5 N aqueous Na2SO3 and evaporated, and the residue
was chromatographed on silica gel. Elution with EtOAc gave 112
(0.32 g, 46%): mp (THF/petroleum ether) 322-325 °C; 1H NMR
[(CD3)2SO] δ 11.89 (s, 1H), 11.29 (s, 1H), 8.40 (d, J ) 2.6 Hz,
1H), 8.18 (s, 1H), 7.54 (d, J ) 8.9 Hz, 1H), 7.24 (dd, J ) 8.9, 2.6
Hz, 1H), 3.87 (s, 3H). Anal. (C15H9IN2O3‚1/4H2O) C, H, N.
9-Hydroxy-4-iodopyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione (5).
Powdered 112 (0.50 g, 1.27 mmol) was added in one portion to a
dry, freshly prepared pyridine hydrochloride melt at 200 °C under
a CaCl2 drying tube, and the mixture was stirred at this temperature
for 15 min. The mixture was partitioned between EtOAc and water,
and the EtOAc extracts were dried and evaporated. The residue
was chromatographed on silica gel, eluting with EtOAc, to give 5
Demethylation of 158 with BBr3 using procedure 5 gave 11 (40%
1
overall): mp 270-280 °C; H NMR [(CD3)2SO] δ 11.60 (s, 1H),
10.82 (s, 1H), 9.22 (s, 1H), 8.31 (d, J ) 2.0 Hz, 1H), 7.47-7.42
(m, 4H), 7.36-7.23 (m, 2H), 7.06 (dd, J ) 8.7, 2.0 Hz, 1H), 2.32
(s, 3H). EIMS found M+, 342.1005; C21H14N2O3 requires 342.1004.
5-Methoxy-2-vinyl-1H-indole (159). Reaction of 106 with
methyl(triphenyl)phosphonium bromide and LDA using procedure
1
11 gave 159 (X, X ) Y ) H) (87%): mp 80-81 °C; H NMR
[(CD3)2SO] δ 11.09 (s, 1H), 7.21 (d, J ) 8.7 Hz, 1H), 6.97 (d, J
) 2.3 Hz, 1H), 6.75-6.66 (m, 2H), 6.37 (s, 1H), 5.76 (d, J ) 17.3
Hz, 1H), 5.21 (d, J ) 11.6, 1H), 3.73 (s, 3H). Anal. (C11H11NO)
C, H, N.
9-Methoxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione (160). Re-
action of 159 with maleimide at 180 °C using procedure 8, followed
by aromatization of the adduct with MnO2 using procedure 7, gave
160 (XI, X ) Y ) H) (76%): mp 260-270 °C; 1H NMR [(CD3)2-
SO] δ 11.91 (s, 1H), 11.10 (s, 1H), 8.38 (d, J ) 2.6 Hz, 1H), 7.81
(d, J ) 8.2 Hz, 1H), 7.76 (d, J ) 8.2 Hz, 1H), 7.54 (d, J ) 8.8 Hz,
1H), 7.22 (dd, J ) 8.8, 2.6 Hz, 1H), 3.88 (s, 3H). Anal.
(C15H10N2O3‚1/2H2O) C, H, N.
1
(0.43 g, 89%): mp >350 °C; H NMR [(CD3)2SO] δ 11.76 (br s,
1H), 11.24 (br s, 1H), 9.29 (br s, 1H), 8.27 (d, J ) 2.4 Hz, 1H),
8.13 (s, 1H), 7.44 (d, J ) 8.7 Hz, 1H), 7.08 (dd, J ) 8.7, 2.4 Hz,
1H). Anal. (C14H7IN2O3) C, H, N.
9-Hydroxypyrrolo[3,4-c]carbazole-1,3(2H,6H)-dione (4). De-
methylation of 160 with BBr3 using procedure 5 gave 4 (79%):
1
mp 335-345 °C; H NMR [(CD3)2SO] δ 11.77 (s, 1H), 11.04 (s,
Procedure 9. 4-(2-Chlorophenyl)-9-hydroxypyrrolo[3,4-c]car-
bazole-1,3(2H,6H)-dione (23). A mixture of 5 (41.8 mg, 0.110
mmol) and 2-chlorobenzeneboronic acid (52 mg, 0.332 mmol) in
p-dioxane (3 mL) and 2 N Na2CO3 (0.5 mL) was purged with
nitrogen. Pd(dppf)Cl2 (35 mg, 0.011 mmol) was added, and the
mixture was refluxed under N2 for 4 h and then partitioned between
EtOAc and water. The organic layer was dried and evaporated,
and the residue was chromatographed on silica gel, eluting with
EtOAc/petroleum ether (1:4) followed by EtOAc/petroleum ether
(2:3), to give 23 (33.1 mg, 83%): mp (EtOAc/petroleum ether)
215-220 °C (dec); 1H NMR [(CD3)2SO] δ 11.83 (br s, 1H), 11.01
(br s, 1H), 9.27 (br s, 1H), 8.32 (d, J ) 2.4 Hz, 1H), 7.65-7.40
(m, 5H), 7.08 (dd, J ) 8.7, 2.4 Hz, 1H). Anal. (C20H11ClN2O3‚1/
4EtOAc) C, H, N.
1H), 9.23 (s, 1H), 8.24 (d, J ) 2.3 Hz, 1H), 7.75 (2d, J ) 8.2 Hz,
2H), 7.43 (d, J ) 8.7 Hz, 1H), 7.05 (dd, J ) 8.7, 2.3 Hz, 1H).
Anal. (C14H8N2O3‚1/2H2O) C, H, N.
1-(5-Methoxy-1H-indol-2-yl)-1-propanone (161). Reaction of
5-methoxy-1H-indole-2-carbaldehyde (106) with ethylmagnesium
bromide and oxidation of the resulting alcohol with MnO2 using
procedure 10 gave 161 (IX, X ) Et) (82%): mp 170-171.5 °C;
1H NMR [(CD3)2SO] δ 11.56 (s, 1H), 7.34 (d, J ) 9.1 Hz, 1H),
7.22 (d, J ) 1.7 Hz, 1H), 7.11 (d, J ) 2.4 Hz, 1H), 6.93 (d, J )
9.1, 2.4 Hz, 1H), 3.77 (s, 3H), 2.96 (q, J ) 7.3 Hz, 2H), 1.13 (t, J
) 7.3 Hz, 3H). Anal. (C12H13NO2) C, H, N.
5-Ethyl-9-hydroxy-4-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione (12). Reaction of 161 with benzyltriphenylphosphonium
bromide using procedure 11 gave 2-[(E,Z)-1-ethyl-2-phenylethenyl]-
5-methoxy-1H-indole (162) (X, X ) Et, Y ) phenyl) (38%): mp
95-97 °C. Anal. (C19H19NO) C, H, N.
Procedures of Scheme 3. Procedure 10. 1-(5-Methoxy-1H-
indol-2-yl)ethanone (156). A solution of 5-methoxy-1H-indole-
2-carbaldehyde (106)20,21 (2.0 g, 11.0 mmol) in THF (30 mL) at 0
°C was treated dropwise with a solution of methylmagnesium
bromide (11 mL of a 3 M solution in Et2O, 34.0 mmol). The
solution was allowed to warm to room temperature over 50 min,
saturated NH4Cl was then added, solvent was removed under
reduced pressure, and the residue was extracted with EtOAc (2 ×
60 mL). The combined extracts were washed, dried, and concen-
trated, and the product was dissolved in CHCl3 (40 mL) and treated
with MnO2 (15 g, 0.171 mol) at reflux for 40 min. The mixture
was filtered through Celite and concentrated to give 156 (IX, X )
Reaction of 162 with maleimide at 180 °C, followed by
aromatization of the crude adduct with MnO2 using procedures 8
and 7, gave 5-ethyl-9-methoxy-4-phenylpyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (163) (XI, X ) Et, Y ) phenyl) (68%): mp
301-303 °C; 1H NMR [(CD3)2SO] δ 11.78 (s, 1H), 10.87 (s, 1H),
8.47 (d, J ) 2.6 Hz, 1H), 7.56 (d, J ) 8.8 Hz, 1H), 7.48-7.41 (m,
3H), 7.32-7.30 (m, 2H), 7.23 (dd, J ) 8.8, 2.6 Hz, 1H), 3.89 (s,
3H), 2.76 (q, J ) 7.4 Hz, 2H), 1.07 (t, J ) 7.4 Hz, 3H).
Demethylation of 163 with BBr3 using procedure 5 gave 12
1
1
Me) (1.80 g, 83%): mp (DCM) 170-172 °C; H NMR [(CD3)2-
(97%): mp 190-196 °C; H NMR [(CD3)2SO] δ 11.63 (s, 1H),

Pyrrolocarbazole Inhibitors of Wee1 Kinase
Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16 4909
10.81 (s, 1H), 9.21 (s, 1H), 8.31 (d, J ) 2.4 Hz, 1H), 7.48-7.40
(m, 4H), 7.31-7.29 (m, 2H), 7.06 (dd, J ) 8.7, 2.4 Hz, 1H), 2.74
(q, J ) 7.4 Hz, 2H), 1.06 (t, J ) 7.4 Hz, 3H). Anal. (C22H16N2O3‚
1/2H2O) C, H, N.
9-Hydroxy-5-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (15). Demethylation of 170 with BBr3 using procedure 5 gave
15 (89%): mp 335-345 °C; 1H NMR [(CD3)2SO] δ 11.50 (s, 1H),
11.10 (s, 1H), 9.26 (d, J ) 2.4 Hz, 1H), 8.30 (d, J ) 2.4 Hz, 1H),
7.77-7.75 (m, 2H), 7.65-7.61 (m, 3H), 7.56-7.52 (m, 1H), 7.46
(d, J ) 8.7 Hz, 1H), 7.06 (dd, J ) 8.7, 2.4 Hz, 1H). Anal.
(C20H12N2O3) H, N. C: calcd, 73.16; found, 72.71.
Procedures of Scheme 4. 1,9-Dihydroxy-4-phenyl-1,6-dihy-
dropyrrolo[3,4-c]carbazol-3(2H)-one (16) and 3,9-Dihydroxy-
4-phenyl-3,6-dihydropyrrolo[3,4-c]carbazol-1(2H)-one (19). NaBH4
(four portions of 0.24 g, 0.025 mol total) was added over 5 h to a
solution of 311 (0.50 g, 1.50 mmol) in EtOH (80 mL), and the
solution was left overnight. An additional four portions of NaBH4
were added at 1 h intervals, and the solution was then diluted with
water and extracted with EtOAc. The organic phase was dried and
evaporated, and the residue was chromatographed on silica gel.
Elution with EtOAc/petroleum ether (2:1) gave starting material
(12 mg), followed by 19 (0.07 g, 14%): mp 300-310 °C (dec);
1H NMR [(CD3)2SO] δ 11.26 (br s, 1H), 8.98 (s, 1H), 8.83 (s, 1H),
8.50 (d, J ) 2.3 Hz, 1H), 7.70 (br d, J ) 7.0 Hz, 2H), 7.53 (s, 1H),
7.49-7.31 (m, 4H), 6.96 (dd, J ) 8.7, 2.3 Hz, 1H), 6.25 (d, J )
9.7 Hz, 1H), 5.87 (d, J ) 9.7 Hz, 1H). Anal. (C20H13N2O3‚1/4H2O)
C, H, N.
5-Methoxy-2-[(1E)-1-phenyl-1-propenyl]-1H-indole (164). Re-
action of 166 (see later) with ethyl(triphenyl)phosphonium bromide
and LDA using procedure 11 gave 164 (X, X ) phenyl, Y ) Me)
1
(72%): mp 128-130 °C; H NMR [(CD3)2SO] δ 10.97 (s, 1H),
7.47-7.39 (m, 3H), 7.28-7.19 (m, 3H), 6.88 (d, J ) 2.4 Hz, 1H),
6.69 (dd, J ) 8.7, 2.4 Hz, 1H), 6.38 (q, J ) 6.9 Hz, 1H), 5.69 (s,
1H), 3.69 (s, 3H), 1.67 (d, J ) 6.9 Hz, 3H). Anal. (C18H17NO) C,
H, N.
9-Methoxy-4-methyl-5-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (165). Reaction of 164 with maleimide using
procedure 8, followed by aromatization with MnO2 using procedure
7, gave 165 (XI, X ) phenyl, Y ) Me) (74%): mp 284-286 °C;
1H NMR [(CD3)2SO] δ 11.06 (br s, 1H), 10.93 (s, 1H), 8.43 (d, J
) 2.6 Hz, 1H), 7.64-7.54 (m, 3H), 7.45-7.42 (m, 3H), 7.14 (dd,
J ) 8.9, 2.6 Hz, 1H), 3.32 (s, 3H), 2.48 (s, 3H). Anal. (C22H16N2O3‚
1/3H2O) C, H, N.
9-Hydroxy-4-methyl-5-phenylpyrrolo[3,4-c]carbazole-1,3-
(2H,6H)-dione (13). Demethylation of 165 with BBr3 using
procedure 5 gave 13 (85%): mp >300 °C; 1H NMR [(CD3)2SO] δ
11.01 (s, 1H), 10.79 (s, 1H), 9.17 (s, 1H), 8.28 (d, J ) 2.4 Hz,
1H), 7.34-7.60 (m, 2H), 7.57-7.52 (m, 1H), 7.43-7.41 (m, 2H),
7.34 (d, J ) 8.7 Hz, 1H), 6.98 (dd, J ) 8.7, 2.4 Hz, 1H), 2.26 (s,
3H). EIMS found M+, 342.1003; C21H14N2O3 requires 342.1004.
(5-Methoxy-1H-indol-2-yl)(phenyl)methanone (166). Reaction
of 106 with phenylmagnesium bromide and oxidation of the
resulting alcohol with MnO2 using procedure 10 gave 166 (IX, X
) phenyl) (91%): mp 159-161 °C; 1H NMR [(CD3)2SO] δ 11.86
(s, 1H), 7.94-7.91 (m, 2H), 7.71-7.66 (m, 1H), 7.61-7.57 (m,
2H), 7.42 (d, J ) 8.9 Hz, 1H), 7.16 (d, J ) 2.4 Hz, 1H), 7.03 (s,
1H), 6.99 (dd, J ) 8.9, 2.4 Hz, 1H), 3.77 (s, 3H). Anal. (C16H13-
NO2) C, H, N.
Elution with EtOAc gave a mixed fraction (0.014 g), followed
1
by 16 (0.32 g, 64%): mp 300-310 °C (dec); H NMR [(CD3)2-
SO] δ 11.38 (br s, 1H), 9.06 (s, 1H), 8.55 (br s, 1H), 7.72 (d, J )
2.2 Hz, 1H), 7.55 (dd, J ) 7.8, 2.1 Hz, 2H), 7.47-7.35 (m, 4H),
7.32 (s, 1H), 6.98 (dd, J ) 8.6, 2.2 Hz, 1H), 6.38 (d, J ) 10.3 Hz,
1H), 6.20 (d, J ) 10.3 Hz, 1H). Anal. (C20H13N2O3‚H2O) C, H, N.
9-Hydroxy-1-methoxy-4-phenyl-1,6-dihydropyrrolo[3,4-c]car-
bazol-3(2H)-one (17). A solution of 16 (0.050 g, 0.16 mmol) and
p-TsOH (15 mg) in MeOH (5 mL) was stirred at room temperature
for 30 min and then poured into saturated aqueous NaHCO3
solution. The mixture was extracted with EtOAc, and the organic
phase was dried and concentrated to dryness to give 17 (0.041 g,
1
2-[(E)-1,2-Diphenylethenyl]-5-methoxy-1H-indole (167). Reac-
tion of 166 with benzyl(triphenyl)phosphonium bromide and LDA
using procedure 11 gave 167 (X, X ) Y ) phenyl) (0.95 g, 73%):
74%): mp (EtOAc/petroleum ether) 290-300 °C (dec); H NMR
[(CD3)2SO] δ 11.47 (br s, 1H), 9.15 (br s, 1H), 8.75 (s, 1H), 7.57-
7.53 (m, 3H), 7.44-7.37 (m, 4H), 7.36 (s, 1H), 7.09 (dd, J ) 8.7,
2.4 Hz, 1H), 6.26 (s, 1H), 3.25 (s, 3H). Anal. (C21H15N2O3‚1/2H2O)
C, H, N.
1
mp (DCM) 144-147 °C; H NMR [(CD3)2SO] δ 11.24 (s, 1H),
7.49-7.42 (m, 3H), 7.29-7.26 (m, 4H), 7.17-7.08 (m, 3H), 6.97-
6.95 (m, 2H), 6.92 (d, J ) 2.4 Hz, 1H), 6.75 (dd, J ) 8.7, 2.4 Hz,
1H), 5.81 (s, 1H), 3.70 (s, 3H). Anal. (C23H19NO‚1/10H2O) C, H,
N.
9-Hydroxy-4-phenyl-1,6-dihydropyrrolo[3,4-c]carbazol-3(2H)-
one (18). A solution of 16 (0.20 g, 0.605 mmol) in THF (30 mL)
was treated sequentially with p-TsOH (23 mg, 0.121 mmol) and
PhSeH (1.48 mL, 4.24 mmol), and the solution was stirred at room
temperature for 1 h. The mixture was partitioned between water
and EtOAc, and the organic layer was washed with aqueous
NaHCO3 solution. The organic phase was dried and evaporated,
and the residue was chromatographed on silica gel. Elution with
EtOAc/petroleum ether (1:1) and then EtOAc followed by EtOAc/
MeOH (95:5) gave 18 (0.174 g, 91%): mp 270-280 °C (dec); 1H
NMR [(CD3)2SO] δ 11.41 (br s, 1H), 9.13 (br s, 1H), 8.27 (br s,
1H), 7.60-7.52 (m, 2H), 7.47-7.36 (m, 4H), 7.34-7.31 (m, 1H),
7.30 (s, 1H), 6.99 (dd, J ) 8.6, 2,2 Hz, 1H), 4.78 (br s, 2H). Anal.
(C20H14N2O2‚1/4H2O) C, H. N: calcd, 8.78; found, 8.37.
9-Hydroxy-4,5-diphenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-
dione (14). Reaction of 167 with maleimide at 180 °C, followed
by aromatization with MnO2 using procedures 8 and 7, gave
9-methoxy-4,5-diphenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (168) (XI, X ) Y ) phenyl) (77%): 1H NMR [(CD3)2SO] δ
11.08 (s, 1H), 11.02 (br s, 1H), 8.52 (d, J ) 2.6 Hz, 1H), 7.50 (d,
J ) 8.9 Hz, 1H), 7.36-7.28 (m, 3H), 7.21-7.11 (m, 8H), 3.89 (s,
3H).
Demethylation of 168 with BBr3 using procedure 5 gave 14
(87%): mp >300 °C; 1H NMR [(CD3)2SO] δ 10.96 (s, 1H), 10.94
(s, 1H), 9.23 (s, 1H), 8.37 (d, J ) 2.4 Hz, 1H), 7.39 (d, J ) 8.7
Hz, 1H), 7.35-7.29 (m, 3H), 7.22-7.10 (m, 7H), 7.03 (dd, J )
8.7, 2.4 Hz, 1H). Anal. (C26H16N2O3) C, H, N.
Methyl 2-(1-Phenylvinyl)-1H-indol-5-yl Ether (169). Reaction
of 166 with methyl(triphenyl)phosphonium bromide and LDA using
procedure 11 gave 169 (X, X ) Ph, Y ) H) (95%): mp 119-121
°C; 1H NMR [(CD3)2SO] δ 11.13 (s, 1H), 7.47-7.39 (m, 5H), 7.26
(d, J ) 8.8 Hz, 1H), 6.97 (d, J ) 2.4 Hz, 1H), 6.76 (dd, J ) 8.8,
2.4 Hz, 1H), 6.12 (s, 1H), 5.77 (s, 1H), 5.30 (s, 1H), 3.72 (s, 3H).
Anal. (C17H15NO) C, H, N.
9-Methoxy-5-phenylpyrrolo[3,4-c]carbazole-1,3(2H,6H)-di-
one (170). Reaction of 169 with maleimide at 180 °C using
procedure 8, followed by aromatization of the adduct with MnO2
using procedure 7, gave 170 (XI, X ) Ph, Y ) H) (73%): mp
281-285 °C; 1H NMR [(CD3)2SO] δ 11.63 (br s, 1H), 11.15 (br s,
1H), 8.45 (d, J ) 2.5 Hz, 1H), 7.78-7.76 (m, 2H), 7.68 (s, 1H),
7.65-7.62 (m, 2H), 7.57-7.53 (m, 2H), 7.22 (dd, J ) 8.8, 2.5 Hz,
1H), 3.89 (s, 3H). Anal. (C21H14N2O3‚1/3H2O) C, H, N.
Procedures of Scheme 5. 4-(2-Chlorophenyl)-9-methoxy-
4,5,6,10c-tetrahydro-1H-furo[3,4-c]carbazole-1,3(3aH)-dione (114).
A solution of 113 (0.30 g, 1.06 mmol) and maleic anhydride (0.16
g, 1.59 mmol) in xylene (30 mL) was heated at reflux for 18 h,
then concentrated under reduced pressure and chromatographed on
silica gel. Elution with EtOAc/hexane (1:2) gave 114 (0.29 g,
1
72%): mp (EtOAc/hexane) 189-191 °C; H NMR [(CD3)2SO] δ
11.16 (br s, 1H), 7.69 (dd, J ) 7.7, 1.4 Hz, 1H), 7.51 (dd, J ) 7.7,
1.4 Hz, 1H), 7.43 (ddd, J ) 7.7, 7.7, 1.4 Hz, 1H), 7.36 (ddd, J )
7.7, 7.7, 1.4 Hz, 1H), 7.25 (d, J ) 8.6 Hz, 1H), 7.15 (d, J ) 2.4
Hz, 1H), 6.77 (dd, J ) 8.6, 2.4 Hz, 1H), 4.70 (d, J ) 7.7 Hz, 1H),
4.47 (dd, J ) 7.7, 3.5 Hz, 1H), 3.78 (s, 3H), 3.71-3.66 (m, 1H),
3.36 (dd, J ) 15.9, 13.0 Hz, 1H), 2.99 (dd, J ) 15.9, 4.1 Hz, 1H).
Anal. (C21H16ClNO4) C, H, N.
4-(2-Chlorophenyl)-2-(2,4-dimethoxybenzyl)-9-methoxypyr-
rolo[3,4-c]carbazole-1,3(2H,6H)-dione (115). A solution of an-
hydride 114 (2.80 g, 7.42 mmol) in acetic acid (70 mL) was treated

4910 Journal of Medicinal Chemistry, 2006, Vol. 49, No. 16
Palmer et al.
with 2,4-dimethoxybenzylamine (1.67 mL, 11.1 mmol), and the
resulting solution was heated at reflux for 6 h, then partially
concentrated under reduced pressure and diluted with water. The
precipitate was collected by filtration, washed with water and dried,
then aromatized in dioxane with MnO2 using procedure 7. The
product was chromatographed on silica gel, eluting with EtOAc/
hexane (1:1), to give 115 (1.46 g, 37%): mp (MeOH) 224-226
°C; 1H NMR [(CD3)2SO] δ 12.02 (br s, 1H), 8.45 (d, J ) 2.6 Hz,
1H), 7.62 (s, 1H), 7.58 (m, 2H), 7.53-7.42 (m, 3H), 7.25 (dd, J )
8.8, 2.6 Hz, 1H), 6.95 (d, J ) 8.4 Hz, 1H), 6.57 (d, J ) 2.3 Hz,
1H), 6.45 (dd, J ) 8.4, 2.3 Hz, 1H), 4.68 (s, 2H), 3.88 (s, 3H),
3.80 (s, 3H), 3.72 (s, 3H). Anal. (C30H23ClN2O5) C, H, N.
Array Synthesis of N6-Substituted Analogues of Table 4 from
115. To a solution of carbazole 115 (50 mg, 0.09 mmol per reaction)
in dimethylformamide (5 mL) under nitrogen were added potassium
carbonate (0.13 g, 0.93 mmol) and alkyl halide (0.28 mmol). The
resulting suspension was warmed to 90 °C with stirring for 3 h
before being diluted with water and extracted with ethyl acetate.
The organic phase was dried, the drying agent was removed, and
the solution was concentrated to dryness to give alkylated carbazole
(XIII), which was used directly.
A solution of the crude carbazole XIII from the above step (0.09
mmol) in anisole (0.5 mL) and TFA (2.0 mL) was heated in a sealed
vessel at 90 °C for 18 h, and then solvent was removed under
reduced pressure. The residue was diluted with water and extracted
with Et2O (3×). The combined organic extracts were washed with
1 N KOH and brine, then dried, and evaporated. The aqueous layer
was acidified with concentrated HCl and extracted with Et2O, and
the organic extract was evaporated. The combined crude material
was triturated with Et2O/hexane and then dissolved in DCM (20
mL) and demethylated with BBr3 as described in procedure 5.
Chromatography of the product on silica gel, eluting with EtOAc/
hexane (1:1 to 1:0), gave the compounds of Table 4 as yellow solids.
The products were characterized by HPLC-MS analysis.
The following compounds were prepared by the above-described
general method.
0.026 µCi/pL γ-32P-ATP (ICN Biomedicals, Inc.). The plates were
mixed at room temperature for 20 min. The reaction was stopped
by adding 50 µL of ice-cold 20% TCA with 0.1 M tetrasodium
pyrophosphate. Plates were incubated on ice or refrigerated at 4
°C for 1 h. The liquid reaction mixture was removed on a vacuum
manifold, and the precipitated phosphorylated substrate was rinsed
five times with 200 µL of ice-cold 10% TCA with 0.1 M
tetrasodium pyrophosphate. An amount of 25 µL of liquid scintil-
lation cocktail was added to the membrane bound substrate, and
the plate was read in a Microbeta plate reader (Perkin-Elmer).
Activity of compounds was calculated in comparison to uninhibited
control determinations in each assay.
Chk1 Inhibition. The assay was carried out in round-bottom
polypropylene 96-well plates (Costar). Compounds were tested in
serial dilutions beginning with a high concentration of 50 µM
followed by up to nine 3-fold dilutions. Compounds were dissolved
and diluted in DMSO. An amount of 2 µL of drug was spotted on
the bottom of the assay plates, then diluted with 58 µL of Chk1
buffer (20 mM Tris, pH 8.0, 50 mM NaCl, 10% glycerol, 10 mM
MgCl2, 5 mM dithiothreitol), and mixed at room temperature for 1
min. An amount of 20 µL of buffer containing 250 ng/well Chk1
enzyme (Onyx Pharmaceuticals) and 1 µg/well GST-Cdc25 sub-
strate (Onyx Pharmaceuticals) was added. Contents of the wells
were mixed for 1 min and incubated at room temperature for 10
min. Then 20 µL of buffer containing 20 µM ATP and 0.4 µCi
ATP [γ-33P] was added. The contents were mixed for 1 min and
incubated at 30 °C for 30 min. The reaction was stopped by the
addition of 50 µL of 120 mM EDTA to each well except to the
control wells already containing EDTA. Then 140 µL of the
contents of the wells was transferred to the wells of Reacti-Bind
glutathione coated 96-well plates (Pierce). Contents were mixed
for 1 min and incubated at room temperature for 1 h. All of the
wells were rinsed three times each with 300 µL of PBS and air-
dried. Then 200 µL of MicroScint (Packard) was added to the wells.
Plates were sealed with an adhesive cover and counted in a
TopCount microplate scintillation counter (Packard). Activity of
the compounds was calculated by comparison to uninhibited control
determinations in each assay.
4-(2-Chlorophenyl)-9-hydroxy-6-methylpyrrolo[3,4-c]carba-
zole-1,3(2H,6H)-dione (25). 25 was prepared from 115 and MeI.
Found: M + H ) 377. Purity 100%.
4-(2-Chlorophenyl)-6-ethyl-9-hydroxypyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (100). 100 was prepared from 115 and EtI.
Found: M + H ) 391. Purity 97%.
4-(2-Chlorophenyl)-9-hydroxy-6-propylpyrrolo[3,4-c]carba-
zole-1,3(2H,6H)-dione (101). 101 was prepared from 115 and
n-PrBr. Found: M + H ) 405. Purity 99%.
4-(2-Chlorophenyl)-9-hydroxy-6-isopropylpyrrolo[3,4-c]car-
bazole-1,3(2H,6H)-dione (102). 102 was prepared from 115 and
i-PrBr. Found: M + H ) 405. Purity 99%.
Acknowledgment. We thank Chuck Moore for performing
the enzyme assays and the Auckland Division of the Cancer
Society of New Zealand for continued support.
Supporting Information Available: Additional experimental
procedures and characterizations; combustion analytical data. This
material is available free of charge via the Internet at http://
pubs.acs.org.
6-Butyl-4-(2-chlorophenyl)-9-hydroxypyrrolo[3,4-c]carbazole-
1,3(2H,6H)-dione (103). 103 was prepared from 115 and n-BuBr.
Found: M + H ) 419. Purity 98%.
4-(2-Chlorophenyl)-9-hydroxy-6-isopentylpyrrolo[3,4-c]car-
bazole-1,3(2H,6H)-dione (104). 104 was prepared from 115 and
isopentyl bromide. Found: M + H ) 433. Purity 98%.
4-(2-Chlorophenyl)-9-hydroxy-6-pentylpyrrolo[3,4-c]carba-
zole-1,3(2H,6H)-dione (105). was 105 prepared from 115 and
pentyl bromide. Found: M + H ) 433. Purity 98%.
Determinations of Enzyme Inhibition. Wee1 Inhibition. The
assay was carried out in 96-well filter microtiter plates (Millipore;
MADP NOB 10). Compounds were dissolved and diluted in
DMSO. Then 10 µL of 3× EDB buffer (150 mM Tris, pH 8.0, 30
mM NaCl, 30 mM MgC12, 3 mM DTT), 18 µL of water, and 2 µL
of drug dilution were added to the test wells and mixed thoroughly.
An amount of 10 µL of enzyme-substrate mixture was added to
the wells. The Wee1 enzyme (human Wee1 kinase aa 215-647,
Onyx Pharmaceuticals, expressed in and purified from a baculovirus
protein expression system) concentration was 0.01 µg/µL, and the
substrate (polyornithine/tyrosine (4: 1), Sigma Chemical Co.) was
0.6 µg/µL in 1× EDB buffer. The plates were mixed thoroughly
for 5 min at room temperature. The reaction was started by adding
10 µL of 1× EDB buffer containing 47.5 µM ATP (Sigma) and
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