Discovery of 4-aryl-4H-chromenes as a new series of apoptosis inducers using a cell- and caspase-based high-throughput screening assay. 3. Structure-activity relationships of fused rings at the 7,8-positions

Article abstract of DOI:10.1021/jm070216c

As a continuation of our efforts to discover and develop the apoptosis-inducing 4-aryl-4H-chromenes as novel anticancer agents, we explored the SAR of fused rings at the 7,8-positions. It was found that a five-member aromatic ring, such as pyrrolo with nitrogen at either the 7- or 9-position, is preferred. A six-member aromatic ring, such as benzo or pyrido, also led to potent compounds. The SAR of the 4-aryl group was found to be similar for chromenes with a fused ring at the 7,8-positions. These compounds were found to inhibit tubulin polymerization, indicating that cyclization of the 7,8-positions into a ring does not change the mechanism of action. Compound 2h was identified to be a highly potent apoptosis inducer with an EC₅₀ of 5 nM and a highly potent inhibitor of cell proliferation with a GI₅₀ of 8 nM in T47D cells.

Full text of DOI:10.1021/jm070216c

2858
J. Med. Chem. 2007, 50, 2858-2864
Discovery of 4-Aryl-4H-chromenes as a New Series of Apoptosis Inducers Using a Cell- and
Caspase-Based High-Throughput Screening Assay. 3. Structure-Activity Relationships of Fused
Rings at the 7,8-Positions
William Kemnitzer,† John Drewe,† Songchun Jiang,† Hong Zhang,† Jianghong Zhao,† Candace Crogan-Grundy,† Lifen Xu,†
Serge Lamothe,# Henriette Gourdeau,# Re´al Denis,# Ben Tseng,† Shailaja Kasibhatla,† and Sui Xiong Cai*,†
Epicept Corporation, 6650 Nancy Ridge DriVe, San Diego, California 92121, and Shire-BioChem Inc., 275 Armand-Frappier BouleVard,
LaVal, Que´bec H7V 4A7, Canada
ReceiVed February 23, 2007
As a continuation of our efforts to discover and develop the apoptosis-inducing 4-aryl-4H-chromenes as
novel anticancer agents, we explored the SAR of fused rings at the 7,8-positions. It was found that a five-
member aromatic ring, such as pyrrolo with nitrogen at either the 7- or 9-position, is preferred. A six-
member aromatic ring, such as benzo or pyrido, also led to potent compounds. The SAR of the 4-aryl group
was found to be similar for chromenes with a fused ring at the 7,8-positions. These compounds were found
to inhibit tubulin polymerization, indicating that cyclization of the 7,8-positions into a ring does not change
the mechanism of action. Compound 2h was identified to be a highly potent apoptosis inducer with an EC50
of 5 nM and a highly potent inhibitor of cell proliferation with a GI50 of 8 nM in T47D cells.
Introduction
limited the use of tubulin targeting agents. The colchicine
binding site located on the monomeric unpolymerized R/â-
tubulin represents another potential tubulin target for the
development of apoptosis inducing chemotherapeutic agents.
Combretastatin A-4 phosphate prodrug 1a (CA4P) and amide
prodrug 1b (AVE-8062) (Chart 1) both inhibit tubulin polym-
erization by binding at the colchicine site and are currently in
clinical trials.8 These compounds have additionally been identi-
fied as vascular-disrupting agents (VDA) for their ability to
disrupt the tumor vasculature by targeting endothelial cells while
sparing the normal vasculature.8,9 Vascular-disrupting agents that
induce apoptosis in cancer cells by targeting the clinically
validated tubulin/microtubule system offer the attractive pos-
sibility of treating all tumor types and improving the effective-
ness of conventional clinical chemotherapeutic agents.10
Apoptosis, or programmed cell death, is the normal cellular
process of eliminating unneeded cells that may threaten tissue
homeostasis and organ morphogenesis.1 Apoptosis proceeds
with characteristic biochemical and cytological features, in-
cluding nuclear condensation and DNA fragmentation. Two
main pathways of apoptosis have been established.2 The
binding of death activators to receptors such as TNF-Ra at the
cell surface initiates apoptosis through an extrinsic pathway.
On the other hand, chemotherapeutic agents, genotoxic stress,
and other death stimuli will initiate apoptosis through an
intrinsic or mitochondrial pathway. Both pathways involve a
cascade of initiator and effector caspases that are activated
sequentially. Caspases are a family of cysteine proteolytic
enzymes that are normally present inside cells as inactive
zymogens.3 Within the caspase family, caspase-3, -6, and
-7 have been identified as the key effector caspases that
cleave multiple protein substrates inside cells, leading to
irreversible cell death.4 The acquisition of resistance to apoptosis
is one of the hallmarks of cancer.5 In addition, it has been
reported that apoptosis may be involved in the regulation of
metastasis.6
Apoptosis is known to play a pivotal role in the antitumor
efficacy of several clinically useful cytotoxic agents.7 The
proapoptotic chemotherapeutic agents that target tubulin, includ-
ing paclitaxel and vinblastine, are among the most successful
and commonly prescribed anticancer therapies. Paclitaxel is a
microtubule-stabilizing agent that binds to the fully formed
microtubules and prevents depolymerization of the tubulin
subunits. Vinblastine functions by binding to the tubulin
monomers and inhibiting their polymerization into microtubules.
The emergence of drug-resistant tumor cells, along with dose-
limiting neurologic and bone marrow toxicity, however, has
We have been interested in the discovery and development
of apoptosis inducers as potential anticancer agents and have
developed a cell- and caspase-based high-throughput screening
(HTS) system for the discovery of apoptosis inducers.11 We have
recently reported the discovery of 4-aryl-4H-chromenes as a
new series of potent apoptosis-inducing agents possessing
vascular-disrupting activity.12 The 4-aryl-4H-chromenes inhibit
tubulin polymerization and bind at or close to the binding site
of colchicine. They are also active in the multidrug resistant
MES-SA/DX5 tumor cells and are highly active as single agents
and in combination with other anticancer agents in several tumor
animal models.13 Starting from our screening hit 2-amino-3-
cyano-7-dimethylamino-4-(3-methoxy-4,5-methylenedioxyphe-
nyl)-4H-chromene (1c) (Chart 1), we have reported the SAR
of the 4-aryl group and the identification of 2-amino-3-cyano-
7-dimethylamino-4-(3-bromo-4,5-dimethoxyphenyl)-4H-
chromene (1d) as a lead compound.14 More recently, we have
reported SAR of the 7- and 5-, 6-, 8-positions of 4-aryl-4H-
chromenes and identified several potent 7-substituted and 7,8-
disubstituted analogues, including 2,7,8-triamino-3-cyano-4-(3-
bromo-4,5-dimethoxyphenyl)-4H-chromene (1e).15 As a continua-
tion of our SAR study, we have explored the synthesis of novel
chromenes via cyclization of the 7,8-positions into a five- or
six-member ring. Herein, we report the SAR of 7,8-fused-4-
* To whom correspondence should be addressed. Phone: 858-202-4006.
Fax: 858-202-4000. E-mail: scai@epicept.com.
† EpiCept Corporation.
# Shire-BioChem Inc.
a Abbreviations: TNF-R, tumor necrosis factor-R; VDA, vascular-
disrupting agents.
10.1021/jm070216c CCC: $37.00 © 2007 American Chemical Society
Published on Web 05/11/2007

4-Aryl-4H-chromenes as Apoptosis Inducers
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12 2859
Table 1. SAR of 4-Aryl-2-amino-3-cyano-4H-chromenes with a Fused
Ring at the 7,8-Positions in the Caspase Activation Assay
Chart 1
Scheme 1a
a Conditions: (a) EtOH, piperidine, room temp.
Scheme 2a
a Data are the mean of three or more experiments and are reported as
the mean ( standard error of the mean (SEM). b Data from ref 14. c Data
from ref 15. d NA ) not applicable.
a Conditions: (a) 1,4-dioxane-2,3-diol, EtOH, room temp, 24 h; (b)
butane-2,3-dione, EtOH, reflux, 1 h.
fused at the 7,8-positions, was only 2-fold less potent than the
7,8-diamino analogue 1e in T47D cells, suggesting that a ring
structure is tolerated at the 7,8-positions. The 2-pyrido (2b) and
3-pyrido (2c) analogues had activity similar to that of 2a,
suggesting that the aromatic nitrogen at those sites is tolerated.
The 5-pyrido (2d) analogue was >2-fold more potent than 2a,
suggesting that an aromatic nitrogen at that position might
contribute to activity. Replacing the benzo ring in 2a with a
nonaromatic tetrahydrobenzo ring (2e) resulted in >2-fold
reduction in potency, suggesting that a planar structure is
preferred. Interestingly, the pyrazino analogue (2f) was >6-
fold less potent than 2a and almost 16-fold less potent than 2d.
The dimethyl substituted pyrazino analogue (2g) was >4-fold
less potent than 2f, suggesting a size-limiting pocket around
the 8,9-positions.
aryl-4H-chromenes as inducers of apoptosis and as potential
anticancer agents.
Results and Discussion
Chemistry. Compounds 2a-e,h-j, 3a-h, 4b,d-e, 5a,b, and
6b,c were prepared by a one-pot reaction of the commercially
available substituted arylaldehydes and aryl alcohols with
malononitrile in good yields according to methods previously
described.14 As an illustration, Scheme 1 depicts the one-pot
reaction for the synthesis of pyrrolochromenes. Substituted
arylaldehydes for the synthesis of compounds 3i-j and 6d-e
are not commercially available and were synthesized according
to published procedures.14,15 For compounds 4a, 4c, and 6a the
benzylidene intermediate was synthesized first and then reacted
with the appropriate aryl alcohol according to published
procedures.14 Compounds 2f and 2g were synthesized in good
yields from cyclization of 1e15 with 1,4-dioxane-2,3-diol and
butane 2,3-dione, respectively (Scheme 2).
Structure-Activity Relationship (SAR) Studies. The apo-
ptosis-inducing activities of 4-aryl-4H-chromenes were mea-
sured by our proprietary cell- and caspase-based HTS assay16
in human breast cancer cells T47D and human non-small-cell
lung cancer cells H1299 (Table 1). By use of the 2-amino,
3-cyano, and the preferred 4-(3-bromo-4,5-dimethoxyphenyl)
groups of 1e, the SAR of chromenes with a fused ring at the
7,8-positions was explored. Compound 2a, with a benzo ring
We next explored chromenes with a five-member ring fused
at the 7,8-positions. The pyrrolo analogue (2h) is highly potent
with an EC50 of 5 nM, which is >6-fold more potent than 1e
and almost 15-fold more potent than 2a, suggesting that a five-
member aromatic ring fits well into a pocket at the 7,8-positions.
The 8-methylpyrrolo analogue (2i) was 6-fold less potent than
2h, which is similar to the observed decrease in potency from
2f to 2g, confirming that there is a size-limiting pocket off the
8-position of the fused ring. The reverse pyrrolo analogue (2j)
was found to be highly potent with an EC50 of 16 nM and was
about 3-fold less potent than 2h.

2860 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12
Kemnitzer et al.
Table 3. SAR of 4-Aryl-2-amino-3-cyanobenzo-4H-chromenes and
Table 2. SAR of 4-Aryl-2-amino-3-cyanopyrrolo-4H-chromenes in the
Caspase Activation Assay
4-Aryl-2-amino-3-cyanopyrido-4H-chromenes in the Caspase Activation
Assay
a Data are the mean of three or more experiments and are reported as
the mean ( standard error of the mean (SEM).
The SAR of the 4-aryl group of chromenes with a six-member
ring fused at the 7,8-positions was also found to be similar to
the reported SAR of 7-substituted and 7,8-disubstituted com-
pounds14,15 (Table 3). For the analogues with a benzo ring at
the 7,8-positions, the 3,4,5-trisubstituted phenyl analogues 5a,b
were found to have potencies similar to or slightly more potent
than that of 2a. For the analogues with a pyrido group at the
7,8-positions, the 3,4,5-trisubstituted phenyl analogues 6a-c
were found to have potencies similar to or slightly less potent
than that of 2b and 2c, and the 5-substituted-3-pyridyl analogues
6d and 6e were found to be slightly less potent than 2c.
The activities of these compounds toward the human non-
small-cell lung cancer cell line H1299 were roughly parallel to
their activity toward T47D cells. In general, H1299 cells were
slightly less sensitive (about 2-fold less sensitive as indicated
by the EC50 value) to the compounds than T47D cells in this
assay.
Selected compounds were also tested by the traditional growth
inhibition assay to confirm that the active compounds can inhibit
tumor cell growth. The growth inhibition assays in T47D and
H1299 cells were run in a 96-well microtiter plate as described
previously.16 In brief, T47D and H1299 cells were exposed
continuously to test compound for 48 h at 37 °C. CellTiter-Glo
reagent (Promega) was added, and the samples were mixed by
agitation, incubated at room temperature for 10-15 min, and
then read using a luminescent plate reader (Tecan Spectrafluor
Plus instrument). The GI50 is defined as 50% inhibition of cell
proliferation. GI50 values, in comparison with their EC50 values,
are summarized in Table 4.
a Data are the mean of three or more experiments and are reported as
the mean ( standard error of the mean (SEM).
To determine whether a fused ring at the 7,8-positions
changes the SAR of the 4-aryl group of the chromene, a group
of 7,8-fused pyrrolo and 7,8-fused reverse pyrrolo analogues
with different aryls at the 4-position were synthesized (Table
2). The SAR of the 4-aryl groups of 7,8-fused pyrrolochromenes
was found to be similar to the reported SAR of 7-substituted
and 7,8-disubstituted compounds.14,15 A 3,4,5-trisubstituted
phenyl group, such as 3-bromo-4-hydroxy-5-methoxyphenyl
(3a) and 3,4,5-trimethoxyphenyl (3b), was found to result in
potent analogues with potency 2- to 6-fold less than that of 2h.
The 3,5-dimethoxyphenyl analogue (3c) and 3-substituted
phenyl analogues, including 3-methoxy (3d), 3-nitro (3f), and
3-cyano (3g), all had good activity ranging from 30 to 68 nM.
The nonsubstituted phenyl analogue (3e) was much less active,
confirming the importance of substituents on the phenyl ring,
especially at the 3-position. Similar to the reported SAR for
7-NMe2-chromenes,14 the 3-pyridyl analogue (3h) was >2-fold
more potent than the phenyl analogue 3e, and 5-substituted
3-pyridyl analogues 3i and 3j were highly potent with potency
approaching that of 2h.
Compounds 2h and 2j were found to be the most potent
inhibitors of tumor cell growth among the compounds tested.
Compound 2h has a GI50 of 8 and 19 nM, while 2j has a GI50
of 9 and 6 nM, in T47D and H1299 cells, respectively.
Compounds 2h and 2j are significantly more potent in the
growth inhibition assay in T47D cells than the initial lead
compound 1d. Compound 2h is >11-fold more potent than 1d,
while 2j is 10-fold more potent than 1d in the T47D cell line.
In general, the compound that is more active in the apoptosis
induction assay, as measured by caspase activation, also is more
potent in the growth inhibition assay.
The SAR of the 4-aryl group of reverse pyrrolochromene
followed that of the pyrrolochromene. The 3-bromo-4-hydroxy-
5-methoxyphenyl analogue (4a), 3,4,5-trimethoxyphenyl ana-
logue (4b), and 3,5-dimethoyphenyl analogue (4c) were 2- to
4-fold less potent than 2j. The 3-substituted phenyl analogues,
including the 3-methoxy (4d) and 3-cyano (4e), had good
activity ranging from 88 to 120 nM.

4-Aryl-4H-chromenes as Apoptosis Inducers
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12 2861
Table 4. Comparison of Caspase Activation Activity and Inhibition of
Cell Proliferation Activity of 4-Aryl-2-amino-3-cyano-4H-chromenes
Solvents were removed by rotary evaporation under reduced
pressure; where appropriate, the compound was further dried using
a vacuum pump. The 1H NMR spectra were recorded at 300 MHz.
All samples were prepared as dilute solutions in either deuterio-
chloroform (CDCl3) with 0.05% v/v tetramethylsilane (TMS),
acetone-d6 (CD3COCD3, 99.9%) with 1.0% v/v tetramethylsilane
(TMS), or dimethyl-d6-sulfoxide (CD3SOCD3) with 0.05% v/v
TMS. Chemical shifts are reported in parts per million (ppm)
downfield from TMS (0.00 ppm) and J coupling constants are
reported in hertz. Elemental analyses were performed by Numega
Resonance Labs, Inc. (San Diego, CA). Melting points were
determined on a glass capillary melting point apparatus. Human
breast T47D and human small-cell lung H1299 tumor cell lines
were purchased from the American Type Culture Collection
(Manasas, VA). Tubulin was obtained from Cytoskeleton (Boulder,
CO).
T47D
EC50 a (µM)
1d 0.019 ( 0.004 0.092 ( 0.013 0.043 ( 0.001 0.016 ( 0.007
1e 0.034 ( 0.005 0.15 ( 0.02 0.081 ( 0.014 0.031 ( 0.001
2c 0.061 ( 0.003 0.057 ( 0.003 0.150 ( 0.017 0.071 ( 0.006
2f 0.46 ( 0.018 0.45 ( 0.018 0.86 ( 0.24 0.55 ( 0.003
H1299
GI50 b (µM)
EC50 a (µM) GI50 b (µM)
2h 0.005 ( 0.003 0.008 ( 0.001 0.013 ( 0.003 0.019 ( 0.009
2i
2j
0.030 ( 0.004 0.089 ( 0.026 0.057 ( 0.007 0.140 ( 0.051
0.016 ( 0.001 0.009 ( 0.001 0.053 ( 0.008 0.006 ( 0.001
3a 0.013 ( 0.002 0.057 ( 0.003 0.025 ( 0.001 0.071 ( 0.006
3i
4a 0.035 ( 0.001 0.070 ( 0.006 0.080 ( 0.013 0.070 ( 0.009
0.013 ( 0.002 0.073 ( 0.005 0.065 ( 0.022 0.100 ( 0.007
a From Tables 1 and 2. b Data are the mean of three experiments and
are reported as the mean ( standard error of the mean (SEM).
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-naph-
tho[1,2-b]pyran (2a). To a mixture of 5-bromoveratraldehyde
(0.100 g, 0.408 mmol), naphthalen-1-ol (0.059 g, 0.41 mmol), and
malononitrile (0.027 g, 0.41 mmol) in ethanol (2.0 mL) was added
piperidine (0.081 mL, 0.82 mmol). The mixture was stirred for 12
h, and the resulting white solid was collected by filtration, washed
with methanol, and dried in vacuo to yield 0.035 g (20%) of 2a as
a white solid: mp 190-192 °C (dec); 1H NMR (DMSO-d6) δ 8.23
(d, J ) 8.1, 1H), 7.91 (d, J ) 8.1, 1H), 7.67-7.59 (m, 3H), 7.23
(br s, 2H), 7.20 (d, J ) 8.4, 1H), 7.06 (m, 1H), 6.95 (m, 1H), 4.93
(s, 1H), 3.80 (s, 3H), 3.70 (s, 3H). Anal. (C22H17BrN2O3) C, H, N.
The following compounds were prepared from the corresponding
aryl aldehyde, aryl alcohol, and malononitrile with piperidine in
ethanol by a procedure similar to that described for the preparation
of compound 2a.
Similar to the hit 1c and initial lead compound 1d, compounds
2h (MX-76747) and 2j were found to be tubulin inhibitors
that bind at or close to the colchicine site of â-tubulin.12
Compounds 2h and 2j were found to inhibit tubulin polymer-
ization with IC50 values of 400 and 900 nM, respectively. In
addition, compound 2h was found to arrest MCF-7 breast
carcinoma cells in the G2/M phase of the cell cycle followed
by apoptosis as measured by cell cycle analysis,12 confirming
that 2h and related compounds induce apoptosis similar to other
chromenes. Therefore, cyclization of the 7,8-positions into a
ring system does not change the mechanism of action of these
chromenes.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4,7-dihy-
dro-8-methylpyrano[2,3-e]indole (2i). Compound 2i was prepared
from 5-bromoveratraldehyde, 4-hydroxy-2-methyl-1H-indole, and
malononitrile and was isolated as a white solid (48%): mp 224-
Conclusion
In conclusion, we have explored the SAR of the apoptosis-
inducing 4-aryl-4H-chromenes by fusing the 7,8-positions into
a five- or six-member ring. It was found that six-member
aromatic rings, including benzo and pyrido, were generally well
tolerated while a nonaromatic tetrahydrobenzo ring resulted in
significant reduction of apoptotic activity. Interestingly, a
pyrazino ring with two heteroatoms also led to a dramatic
decrease in apoptotic activity. Significantly, a pyrrolo ring with
nitrogen at either the 7- or 9-position was found to result
in highly potent analogues. The SAR of the 4-aryl group was
found to be similar for chromenes with a fused ring at the
7,8-positions and similar to the previously reported 7-NMe2,
7-OMe, 7-NHEt, and 7,8-disubstituted analogues. The 7,8-
fused chromenes were found to be potent in the cell growth
inhibition assay and to inhibit tubulin polymerization, similar
to chromenes without a fused ring at the 7,8-positions. Several
analogues, such as 2h and 2j, were identified that have low
nanomolar potency in both the caspase assay and the growth
inhibition assay, and compound 2h is significantly more
potent than the initial lead compounds 1d and 1e. Through
SAR studies of the 4-aryl-4H-chromenes, an anticancer drug
candidate (MX-116407, EPC2407) with potent vascular dis-
rupting activity and in vivo efficacy has been identified13 and
is currently in phase I clinical trial. Additional SAR and in
vivo studies of 4-aryl-chromenes will be reported in future
publications.
1
226 °C (dec); H NMR (DMSO-d6) δ 11.15 (s, 1H), 7.01-6.97
(m, 2H), 6.94 (br s, 2H), 6.86 (d, J ) 1.8, 1H), 6.64 (d, J ) 8.4,
1H), 6.16 (br s, 1H), 4.77 (s, 1H), 3.79 (s, 3H), 3.70 (s, 3H), 2.38
(s, 3H). Anal. (C21H18BrN3O3) C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-(3,4,5-trimethoxyphenyl)-
pyrano[2,3-e]indole (3b). Compound 3b was prepared from 3,4,5-
trimethoxybenzaldehyde, 4-hydroxy-1H-indole, and malononitrile
and was isolated as a white solid (43%): mp 220-222 °C (dec);
1H NMR (DMSO-d6) δ 11.29 (s, 1H), 7.35 (t, J ) 2.7, 1H), 7.10
(d, J ) 8.7, 1H), 6.90 (br s, 2H), 6.76 (d, J ) 8.4, 1H), 6.52 (s,
2H), 6.46-6.45 (m, 1H), 4.75 (s, 1H), 3.70 (s, 6H), 3.62 (s, 3H).
Anal. (C21H19N3O4) C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-(3-pyridyl)pyrano[2,3-e]in-
dole (3h). Compound 3h was prepared from pyridine-3-carboxal-
dehyde, 4-hydroxy-1H-indole, and malononitrile and was isolated
as a yellow solid (11%): mp 226-229 °C (dec); 1H NMR (DMSO-
d6) δ 11.32 (s, 1H), 8.49-8.41 (m, 2H), 7.53 (d, J ) 8.1, 1H),
7.37-7.30 (m, 2H), 7.11 (d, J ) 8.4, 1H), 7.01 (br s, 2H), 6.67
(d, J ) 8.4, 1H), 6.47 (s, 1H), 4.88 (s, 1H). Anal. (C17H12N4O) C,
H, N.
2-Amino-3-cyano-4,7-dihydro-4-(5-methyl-3-pyridyl)pyrano-
[2,3-e]indole (3i). Compound 3i was prepared from 5-methylpy-
ridine-3-carboxaldehyde, 4-hydroxy-1H-indole, and malononitrile
and was isolated as a yellow solid (52%): mp 230-233 °C (dec);
1H NMR (DMSO-d6) δ 11.32 (br s, 1H), 8.31-8.26 (m, 2H), 7.37
(t, J ) 2.9, 1H), 7.32 (br s, 1H), 7.11 (dd, J ) 8.4 and 0.6, 1H),
6.98 (br s, 2H), 6.65 (d, J ) 8.4, 1H), 6.47 (t, J ) 2.1, 1H), 4.82
(s, 1H), 2.23 (s, 3H). Anal. (C18H14N4O) C, H, N.
2-Amino-3-cyano-4,9-dihydro-4-(3,4,5-trimethoxyphenyl)-
pyrano[3,2-g]indole (4b). Compound 4b was prepared from 3,4,5-
trimethoxybenzaldehyde, 7-hydroxy-1H-indole, and malononitrile
and was isolated as a black solid (28%): mp 222-226 °C (dec);
1H NMR (acetone-d6) δ 10.42 (br s, 1H), 7.35-7.34 (m, 1H), 7.27
(d, J ) 8.2, 1H), 6.75 (d, J ) 8.2, 1H), 6.61 (s, 2H), 6.48-6.47
(m, 1H), 5.97 (br s, 2H), 4.81 (s, 1H), 3.76 (s, 6H), 3.68 (s, 3H).
Anal. (C21H19N3O4‚1.0H2O) C, H, N.
Experimental Sections
General Methods and Materials. Commercial-grade reagents
and solvents obtained from Acros, Aldrich, Apin Lancaster, TCI,
or VWR were used without further purification unless otherwise
indicated. All mixtures were stirred magnetically; moisture-sensitive
reactions were performed under argon in oven-dried glassware.
Thin-layer chromatography (TLC), usually using ethyl acetate/
hexane as the solvent system, was used to monitor reactions.

2862 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12
Kemnitzer et al.
2-Amino-3-cyano-4-(5-methyl-3-pyridyl)-4H-pyrano[2,3-f]iso-
quinoline (6e). Compound 6e was prepared from 5-methylpyridine-
3-carbaldehyde, 5-hydroxyisoquinoline, and malononitrile and was
pyridine-3-carboxaldehyde, 4-hydroxy-1H-indole, and malononitrile
and was isolated as a yellow solid (8%): mp 210-214 °C (dec);
1H NMR (acetone-d6) δ 10.44 (br 1H), 8.17 (d, J ) 1.8, 1H), 8.15
(d, J ) 2.7, 1H), 7.36 (t, J ) 2.7, 1H), 7.19-7.16 (m, 2H), 6.76
(d, J ) 8.1, 1H), 6.58-6.56 (m, 1H), 6.29 (s, 2H), 4.92 (s, 1H),
3.82 (s, 3H). Anal. (C18H14N4O2‚0.3H2O) C, H, N.
2-Amino-3-cyano-4,9-dihydro-4-(3-methoxyphenyl)pyrano-
[3,2-g]indole (4d). Compound 4d was prepared from 3-methoxy-
benzaldehyde, 7-hydroxy-1H-indole, and malononitrile and was
isolated as a yellow solid (25%): mp 204-206 °C; 1H NMR
(CDCl3) δ 8.38 (br s, 1H), 7.31-7.19 (m, 3H), 6.84-6.68 (m, 4H),
6.53 (q, J ) 2.1, 1H), 4.83 (s, 1H), 4.62 (br s, 2H), 3.76 (s, 3H).
Anal. (C19H15N3O2) C, H, N.
1
isolated as a white solid (38%): mp 252-256 °C (dec); H NMR
(acetone-d6) δ 9.27 (d, J ) 0.9, 1H), 8.63 (d, J ) 5.7, 1H), 8.45
(d, J ) 2.1, 1H), 8.33 (d, J ) 1.5, 1H), 8.06 (dt, J ) 6.3 and 1.2,
1H), 7.83 (dd, J ) 8.4 and 0.6, 1H), 7.49 (s, 1H), 7.33 (d, J ) 8.4,
1H), 6.60 (br s, 2H), 5.06 (s, 1H), 2.28 (s, 3H). Anal. (C19H14N4O)
C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4,7-dihy-
dropyrano[2,3-e]indole (2h). To a mixture of 5-bromoveratralde-
hyde (0.100 g, 0.408 mmol), 4-hydroxy-1H-indole (0.054 g, 0.41
mmol), and malononitrile (0.027 g, 0.41 mmol) in ethanol (2.0 mL)
was added piperidine (0.081 mL, 0.82 mmol). The mixture was
stirred for 12 h, and then the solvent was evaporated. The re-
sulting yellow oil was diluted with dichloromethane, and the
precipitate was collected by filtration. Recrystallization (EtOAc/
hexanes) yielded 0.017 g (10%) of 2h as a yellow solid: mp 235-
237 °C (dec); 1H NMR (DMSO-d6) δ 11.33 (br s, 1H), 7.36
(t, J ) 2.4, 1H), 7.12 (d, J ) 8.4, 1H), 7.00 (br s, 2H), 6.99 (s,
1H), 6.88 (d, J ) 1.8, 1H), 6.74 (d, J ) 8.7, 1H), 6.46 (br s, 1H),
4.80 (s, 1H), 3.80 (s, 3H), 3.69 (s, 3H). Anal. (C20H16BrN3O3) C,
H, N.
2-Amino-3-cyano-4-(3-cyanophenyl)-4,9-dihydropyrano[3,2-
g]indole (4e). Compound 4e was prepared from 3-cyanobenzalde-
hyde, 7-hydroxy-1H-indole, and malononitrile and was isolated as
1
a white solid (34%): mp 225-227 °C (dec); H NMR (CDCl3) δ
8.43 (br s, 1H), 7.55-7.51 (m, 2H), 7.47-7.40 (m, 2H), 7.33 (dd,
J ) 8.1 and 0.9, 1H), 7.28-7.25 (m, 1H), 6.61-6.55 (m, 2H),
4.92 (s, 1H), 4.73 (br s, 2H). Anal. (C19H12N4O) C, H, N.
2-Amino-3-cyano-4-(3-iodo-4,5-dimethoxyphenyl)-4H-naph-
tho[1,2-b]pyran (5b). Compound 5b was prepared from 3-iodo-
4,5-dimethoxybenzaldehyde, naphthalen-1-ol, and malononitrile and
1
2-Amino-3-cyano-4-(3,4,5-trimethoxyphenyl)-4H-naphtho-
[1,2-b]pyran (5a). Compound 5a was prepared from 3,4,5-
trimethoxybenzaldehyde, naphthalen-1-ol, and malononitrile by a
procedure similar to that described for the preparation of 2h and
was isolated as a white solid (20%): mp 221-222 °C; H NMR
(DMSO-d6) δ 8.24 (d, J ) 7.5, 1H), 7.91 (d, J ) 7.5, 1H), 7.67-
7.56 (m , 3H), 7.22 (br s, 2H), 7.19 (d, J ) 8.7, 1H), 7.13 (d, J )
1.2, 1H), 7.05 (d, J ) 1.5, 1H), 4.90 (s, 1H), 3.78 (s, 3H), 3.67 (s,
3H). Anal. (C22H17IN2O3) C, H, N.
1
was isolated as a white solid (8%): mp 191-193 °C; H NMR
(DMSO-d6) δ 8.23 (d, J ) 8.1, 1H), 7.90 (d, J ) 8.4, 1H), 7.66-
7.55 (m, 3H), 7.20 (d, J ) 8.7, 1H), 7.15 (s, 2H), 6.58 (s, 2H),
4.87 (s, 1H), 3.71 (s, 6H), 3.63 (s, 3H). Anal. (C23H20N2O4‚1.5H2O)
C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-(3,5-dimethoxyphenyl)pyrano-
[2,3-e]indole (3c). To a solution of 3,5-dimethoxybenzaldehyde
(0.039 g, 0.21 mmol), 4-hydroxy-1H-indole (0.028 g, 0.21 mmol),
and malononitrile (0.014 g, 0.21 mmol) in ethanol (1.0 mL) was
added piperidine (0.041 mL, 0.41 mmol). The resulting solution
was stirred for 12 h, the solvent was evaporated, and the residue
was purified by flash column chromatography (elution with EtOAc/
hexanes, 1:1), yielding a crude yellow solid. It was recrystallized
(dichloromethane:hexanes) to yield 0.009 g (12%) of 3c as a yellow
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4,9-dihy-
dropyrano[3,2-g]indole (2j). To 5-bromoveratraldehyde (0.245 g,
1.00 mmol) and malononitrile (0.066 g, 1.0 mmol) in ethanol (4.0
mL) was added piperidine (0.050 mL, 0.50 mmol) and 7-hydroxy-
1H-indole (0.133 g, 1.00 mmol). The mixture was stirred at room
temperature overnight. The solvent was evaporated, and the residue
was purified by flash column chromatography (elution with EtOAc/
hexanes, 1:2) to yield 0.056 g (13%) of 2j as a white solid: mp
1
solid: mp 218-220 °C (dec); H NMR (DMSO-d6) δ 11.30 (s,
1H), 7.35 (t, J ) 2.7, 1H), 7.09 (d, J ) 9.7, 1H), 6.91 (s, 2H), 6.73
(d, J ) 8.4, 1H), 6.45 (s, 1H), 6.34 (s, 3H), 4.70 (s, 1H), 3.69 (s,
6H). Anal. (C20H17N3O3) C, H, N.
1
191-192 °C; H NMR (CDCl3) δ 8.39 (br s, 1H), 7.34-7.25 (m,
2H), 6.91 (d, J ) 2.1, 1H), 6.76 (d, J ) 2.1, 1H), 6.67 (d, J ) 8.1,
1H), 6.56 (d, J ) 2.1, 1H), 4.80 (s, 1H), 4.67 (br s, 2H), 3.84 (s,
3H), 3.83 (s, 3H). Anal. (C20H16BrN3O3) C, H, N.
The following compounds were prepared from the corresponding
arylaldehyde, aryl alcohol, and malononitrile with piperidine in
ethanol by a procedure similar to that described for the preparation
of compound 2j.
2-Amino-3-cyano-4,7-dihydro-4-(3-methoxyphenyl)pyrano-
[2,3-e]indole (3d). Compound 3d was prepared from 3-methoxy-
benzaldehyde, 4-hydroxy-1H-indole, and malononitrile and was
isolated as a white solid (25%): mp 195-198 °C; 1H NMR (CDCl3)
δ 8.26 (br s, 1H), 7.26-7.18 (m, 2H), 7.09-7.06 (m, 1H), 6.84-
6.76 (m, 4H), 6.65-6.63 (m, 1H), 4.80 (s, 1H), 4.65 (br s, 2H),
3.76 (s, 3H). Anal. (C19H15N3O2) C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-phenylpyrano[2,3-e]indole (3e).
Compound 3e was prepared from benzaldehyde, 4-hydroxy-1H-
indole, and malononitrile and was isolated as a yellow solid
(42%): mp 209-211 °C; 1H NMR (DMSO-d6) δ 11.30 (br s, 1H),
7.37-7.08 (m, 7H), 6.90 (br s, 2H), 6.67 (d, J ) 8.1, 1H), 6.46 (s,
1H), 4.77 (s, 1H). Anal. (C18H13N3O) C, H, N.
2-Amino-3-cyano-4-(3-cyanophenyl)-4,7-dihydropyrano[2,3-
e]indole (3g). Compound 3g was prepared from 3-cyanobenzal-
dehyde, 4-hydroxy-1H-indole, and malononitrile by a procedure
similar to that described for the preparation of compound 3c and
1
was isolated as a yellow solid (33%): mp 220-222 °C (dec); H
NMR (DMSO-d6) δ 11.34 (s, 1H), 7.71-7.68 (m, 2H), 7.54-7.52
(m, 2H), 7.37 (t, J ) 3.9, 1H), 7.11 (d, J ) 8.4, 1H), 7.04 (s, 2H),
6.67 (d, J ) 8.7, 1H), 6.47 (s, 1H), 4.93 (s, 1H). Anal. (C19H12N4O)
C, H, N.
2-Amino-3-cyano-4-(3-iodo-4,5-dimethoxyphenyl)-4H-pyrano-
[2,3-f]isoquinoline (6c). To a solution of 3-iodo-4,5-dimethoxy-
benzaldehyde (0.292 g, 1.00 mmol) and malononitrile (0.066 g,
1.0 mmol) in ethanol (2.0 mL) were added 5-hydroxyisoquinoline
(0.145 g, 1.00 mmol) and piperidine (0.050 mL, 0.50 mmol). The
mixture was refluxed overnight and then cooled to room temper-
ature. The resulting orange precipitate was collected by filtration,
washed with ethanol and hexanes, and then dried in vacuo to yield
1
0.274 g (57%) of 6c as a white solid: mp 214-216 °C; H NMR
(CDCl3) δ 9.22 (d, J ) 0.9, 1H), 8.65 (d, J ) 6.0, 1H), 7.95 (d, J
) 6.3, 1H), 7.68 (d, J ) 8.5, 1H), 7.17 (d, J ) 8.7, 1H), 7.13 (d,
J ) 1.8, 1H), 6.77 (d, J ) 1.8, 1H), 4.83 (br s, 2H), 4.82 (s, 1H),
3.82 (s, 6H). Anal. (C21H16IN3O3) C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-(3-nitrophenyl)pyrano[2,3-e]-
indole (3f). Compound 3f was prepared from 3-nitrobenzaldehyde,
4-hydroxy-1H-indole, and malononitrile and was isolated as a
2-Amino-4-(3-bromo-4-hydroxy-5-methoxyphenyl)-3-cyano-
4,7-dihydropyrano[2,3-e]indole (3a). To a solution of 4-acetoxy-
3-bromo-5-methoxybenzaldehyde (1.02 g, 3.74 mmol), malononi-
trile (0.247 g, 3.74 mmol), and 4-hydroxy-1H-indole (0.497 g, 3.74
mmol) in ethanol (10.0 mL) was added piperidine (0.19 mL, 1.9
mmol). The mixture was refluxed for 4 h and cooled to room
temperature, and then the solvent was evaporated to yield the crude
product. It was purified by flash column chromatography (elution
1
yellow solid (17%): mp 202-204 °C; H NMR (CDCl3) δ 8.30
(br s, 1H), 8.12-8.08 (m, 1H), 8.05 (t, J ) 2.1, 1H), 7.63 (td, J )
4.5 and 2.4, 1H), 7.49 (t, J ) 8.1, 1H), 7.25-7.24 (m, 1H), 7.11
(dd, J ) 8.4 and 0.9, 1H), 6.69-6.66 (m, 2H), 4.99 (s, 1H), 4.76
(br s, 2H). Anal. (C18H12N4O3) C, H, N.
2-Amino-3-cyano-4,7-dihydro-4-(5-methoxy-3-pyridyl)pyrano-
[2,3-e]indole (3j). Compound 3j was prepared from 5-methoxy-

4-Aryl-4H-chromenes as Apoptosis Inducers
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12 2863
with EtOAc/hexanes, 1:1), yielding 0.10 g (7%) of 3a as a yellow
solid: mp 238-242 °C; 1H NMR (CDCl3) δ 8.27 (br s, 1H), 7.31-
7.27 (m, 1H), 7.12-7.10 (m, 1H), 6.91 (d, J ) 2.1, 1H), 6.75-
6.70 (m, 2H), 6.65 (s, 1H), 5.82 (s, 1H), 4.76 (s, 1H), 4.68 (br s,
2H), 3.86 (s, 3H). Anal. (C19H14BrN3O3‚0.2H2O) C, H, N.
for 2 h and then cooled to room temperature. The solvent was
evaporated, and the residue was diluted with ethyl acetate (50 mL),
washed with brine (10 mL), dried over Na2SO4, and concentrated
in vacuo to yield a crude product. It was purified by flash column
chromatography (elution with EtOAc/hexanes, 1:1), yielding 0.011
1
g (3%) of 4a as a yellow solid: mp 197-201 °C (dec); H NMR
2-Amino-3-cyano-4-(3,4,5-trimethoxyphenyl)-4H-pyrano[2,3-
f]isoquinoline (6b). Compound 6b was prepared from 3,4,5-
trimethoxybenzaldehyde, 5-hydroxyisoquinoline, and malononitrile
by a procedure similar to that described for the preparation of 3a
and was isolated as a white solid (4%): mp 215-219 °C (dec); 1H
NMR (CDCl3) δ 9.22 (s, 1H), 8.65 (d, J ) 5.8, 1H), 7.95 (d, J )
6.6, 1H), 7.67 (d, J ) 8.8, 1H), 7.21 (d, J ) 8.5, 1H), 6.42 (s, 2H),
4.85 (s, 1H), 4.82 (br s, 2H), 3.83 (s, 3H), 3.80 (s, 6H). Anal.
(C22H19N3O4‚0.4H2O) C, H, N.
(CDCl3) δ 8.39 (br s, 1H), 7.31 (d, J ) 8.7, 1H), 7.26-7.25 (m,
1H), 6.90 (d, J ) 2.1, 1H), 6.71-6.65 (m, 2H), 6.54-6.52 (m,
1H), 5.84 (s, 1H), 4.78 (s, 1H), 4.66 (br s, 2H), 3.87 (s, 3H). Anal.
(C19H14BrN3O3‚0.2H2O) C, H, N.
The following compounds were prepared by a procedure similar
to that described for the preparation of compound 4a.
2-Amino-3-cyano-4,9-dihydro-4-(3,5-dimethoxyphenyl)pyrano-
[3,2-g]indole (4c). Compound 4c was prepared from 7-hydroxy-
1H-indole and 2-(3,5-dimethoxybenzylidene)malononitrile and was
isolated as a white solid (26%): mp 185-186 °C; 1H NMR (CDCl3)
δ 8.39 (br s, 1H), 7.30 (d, J ) 8.1, 1H), 7.23-7.21 (m, 1H), 6.71
(d, J ) 8.4, 1H), 6.54-6.52 (m, 1H), 6.38 (d, J ) 2.1, 2H), 6.34-
6.32 (m, 1H), 4.78 (s, 1H), 4.63 (br s, 2H), 3.75 (s, 6H). Anal.
(C20H17N3O3) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-pyr-
ano[2,3-f]quinoline (2b). To a solution of 5-bromoveratraldehyde
(0.245 g, 1.00 mmol) and malononitrile (0.066 g, 1.0 mmol) in
ethanol (2.0 mL) was added piperidine (0.10 mL, 1.0 mmol). The
mixture was stirred at room temperature for 15 min, then refluxed
for 1 h, and cooled to room temperature. To the mixture were added
piperidine (0.10 mL, 1.0 mmol) and 5-hydroxyquinoline (0.145 g,
1.00 mmol). The mixture was refluxed for 1 h, and the solvent
was evaporated. The residue was purified by flash column chro-
matography (elution with EtOAc/hexanes, 1:2), yielding a white
solid. It was recrystallized by EtOH to yield 0.050 g (11%) of 2b
as a white solid: mp 235-237 °C (dec); 1H NMR (CDCl3) δ 8.95
(s, 1H), 8.50 (d, J ) 9.0, 1H), 7.82 (d, J ) 9.0, 1H), 7.50 (br s,
1H), 7.30 (s, 1H), 6.92 (s, 1H), 6.76 (s, 1H), 4.82 (br s, 3H), 3.84
(s, 6H). Anal. (C21H16BrN3O3) C, H, N.
2-Amino-3-cyano-4-(3,4,5-trimethoxyphenyl)-4H-pyrano[2,3-
f]quinoline (6a). Compound 6a was prepared from 2-(3,4,5-
trimethoxybenzylidene)malononitrile and 5-hydroxyquinoline and
1
was isolated as a white solid (14%): mp 229-230 °C (dec); H
NMR (CDCl3) δ 8.96 (m, 1H), 8.52 (d, J ) 8.7, 1H), 7.82 (d, J )
9.0, 1H), 7.52 (m, 1H), 7.33 (d, J ) 8.7, 1H), 6.43 (s, 2H), 4.83 (s,
3H), 3.83 (s, 3H), 3.80 (s, 6H). Anal. (C22H19N3O4) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-pyr-
ano[2,3-f]quinoxaline (2f). To a mixture of 2,7,8-triamino-4-(3-
bromo-4,5-dimethoxyphenyl)-3-cyano-4H-chromene (0.300 g, 0.719
mmol) and ethanol (3.6 mL) was added glyoxal (0.095 mL, 40%
in H2O, 0.79 mmol). The mixture was stirred at room temperature
overnight. The resulting precipitate was collected and washed with
ethanol and hexanes and then dried in vacuo to yield 0.20 g (62%)
of 2f as a brown solid: mp 157-159 °C (dec); 1H NMR (CDCl3)
δ 8.94 (s, 2H), 7.85 (d, J ) 9.1, 1H), 7.39 (d, J ) 9.1, 1H), 6.97
(d, J ) 1.9, 1H), 6.77 (d, J ) 1.6, 1H), 5.02 (br s, 2H), 4.88 (s,
1H), 3.84 (s, 6H). Anal. (C20H15BrN4O3‚1.0H2O) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-pyr-
ano[2,3-f]isoquinoline (2c). A mixture of 5-bromoveratraldehyde
(0.245 g, 1.00 mmol) and malononitrile (0.066 g, 1.0 mmol) in
ethanol (2.0 mL) and piperidine (0.10 mL, 1.0 mmol) was stirred
at room temperature for 15 min, then refluxed for 1 h, and cooled
to room temperature. To the mixture was added piperidine (0.10
mL, 1.0 mmol) and 5-hydroxyisoquinoline (0.145 g, 1.00 mmol).
The mixture was refluxed for 1 h, and the resulting white solid
was collected by filtration, washed with ethanol and ether, and then
dried in vacuo to yield 0.070 g (16%) of 2c as a white solid: mp
218-220 °C (dec); 1H NMR (CDCl3) δ 9.23 (s, 1H), 8.66 (d, J )
6.0, 1H), 7.95 (d, J ) 5.7, 1H), 7.69 (d, J ) 8.4, 1H), 7.18 (d, J )
8.7, 1H), 6.92 (d, J ) 1.8, 1H), 6.75 (d, J ) 1.8, 1H), 4.85 (m,
3H), 3.84 (s, 6H). Anal. (C21H16BrN3O3) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-8,9-di-
methyl-4H-pyrano[2,3-f]quinoxaline (2g). To a mixture of 2,7,8-
triamino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-
chromene (0.050 g, 0.12 mmol) and ethanol (0.60 mL) was added
butane-2,3-dione (0.010 mL, 0.12 mmol). The mixture was refluxed
for 1 h and then cooled to room temperature. The resulting
precipitate was collected and washed with ethanol and then dried
in vacuo to yield 0.051 g (91%) of 2g as a brown solid: mp 245-
The following compounds were prepared by a procedure similar
to that described for the preparation of compound 2c.
1
247 °C (dec); H NMR (CDCl3) δ 7.70 (d, J ) 8.7, 1H), 7.25-
7.24 (m, 1H), 6.95 (d, J ) 2.1, 1H), 6.73 (d, J ) 2.1, 1H), 4.98 (br
s, 2H), 4.83 (s, 1H), 3.83 (s, 3H), 3.81 (s, 3H), 2.80 (s, 3H), 2.75
(s, 3H). Anal. (C22H19BrN4O3‚0.5H2O) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-4H-pyr-
ano[3,2-h]quinoline (2d). Compound 2d was prepared from
5-bromoveratraldehyde, malononitrile, and 8-hydroxyquinoline and
2-Amino-4-(5-bromo-3-pyridyl)-3-cyano-4H-pyrano[2,3-f]iso-
quinoline (6d). To 5-bromopyridine-3-carbaldehyde (0.093 g, 0.50
mmol) and malononitrile (0.033 g, 0.50 mmol) in ethanol (5.0 mL)
at 0 °C was added piperidine (0.100 mL, 1.00 mmol). The mixture
became a yellow solution upon stirring overnight, and then
5-hydroxyisoquinoline was added (0.074 mL, 0.51 mmol). The
resulting mixture was heated at 80 °C for 6 h, and the resulting
white solid was collected by filtration, washed with ether, and then
dried in vacuo to yield 0.031 g (16%) of 6d as a white solid: mp
238-240 °C (dec); 1H NMR (acetone-d6) δ 9.29 (d, J ) 2.5, 1H),
8.64-8.63 (m, 2H), 8.60 (d, J ) 2.5, 1H), 8.07 (d, J ) 5.8, 1H),
7.95 (t, J ) 2.2, 1H), 7.86 (d, J ) 8.5, 1H), 7.38 (d, J ) 8.5, 1H),
6.71 (br s, 2H), 5.18 (s, 1H). Anal. (C18H11BrN4O) C, H, N.
1
was isolated as a white solid (41%): mp 126-128 °C (dec); H
NMR (CDCl3) δ 9.00 (dd, J ) 4.3 and 1.5, 1H), 8.16 (dd, J ) 8.4
and 1.5, 1H), 7.55-7.50 (m, 2H), 7.15 (d, J ) 8.5, 1H), 6.98 (dd,
J ) 1.9 and 0.8, 1H), 6.76 (d, J ) 2.2, 1H), 5.11 (br s, 2H), 4.85
(s, 1H), 3.83 (s, 6H). Anal. (C21H16BrN3O3‚1.5H2O) C, H, N.
2-Amino-4-(3-bromo-4,5-dimethoxyphenyl)-3-cyano-7,8,9,10-
tetrahydro-4H-naphtho[1,2-b]pyran (2e). Compound 2e was
prepared from 5-bromoveratraldehyde, malononitrile, and 5,6,7,8-
tetrahydro-1-hydroxynaphthalene and was isolated as a white solid
1
(10%): mp 200-202 °C; H NMR (CDCl3) δ 6.89 (d, J ) 1.9,
1H), 6.80-6.72 (m, 3H), 4.64-4.63 (m, 3H), 3.85 (s, 3H), 3.83
(s, 3H), 2.73-2.72 (m, 4H), 1.81-1.79 (m, 4H). Anal. (C22H21-
BrN2O3‚0.2H2O) C, H, N.
Supporting Information Available: Details of the caspase
activation assay (EC50), cell growth inhibition assays (GI50), and
tubulin inhibition assay and table of elemental analysis data for
the targeted compounds 2a-j, 3a-j, 4a-e, 5a,b, and 6a-e. This
material is available free of charge via the Internet at http://
pubs.acs.org.
2-Amino-4-(3-bromo-4-hydroxy-5-methoxyphenyl)-3-cyano-
4,9-dihydropyrano[3,2-g]indole (4a). To 2-(3-bromo-4-hydroxy-
5-methoxybenzylidene)malononitrile (0.279 g, 1.00 mmol) and
7-hydroxy-1H-indole (0.133 g, 1.00 mmol) in ethanol (5.0 mL) was
added piperidine (0.050 mL, 0.50 mmol). The mixture was refluxed

2864 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 12
Kemnitzer et al.
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