Design, synthesis, and biological evaluation of novel 7-azaindolyl- heteroaryl-maleimides as potent and selective glycogen synthase kinase-3β (GSK-3β) inhibitors

Article abstract of DOI:10.1016/j.bmc.2004.04.010

Two approaches were developed to synthesize the novel 7-azaindolyl- heteroarylmaleimides. The first approach was based upon the palladium-catalyzed Suzuki cross-coupling or Stille cross-coupling of 2-chloro-maleimide 5 with various arylboronic acids or ar

Full text of DOI:10.1016/j.bmc.2004.04.010

Bioorganic & Medicinal Chemistry 12 (2004) 3167–3185
Design, synthesis, and biological evaluation of novel 7-azaindolyl-
heteroaryl-maleimides as potent and selective glycogen synthase
kinase-3b (GSK-3b) inhibitors
David J. O’Neill,a Lan Shen,a Catherine Prouty,a Bruce R. Conway,a Lori Westover,a
Jun Z. Xu,a Han-Cheng Zhang,b Bruce E. Maryanoff,b William V. Murray,a
Keith T. Demaresta and Gee-Hong Kuoa,*
aDrug Discovery Division, Johnson and Johnson Pharmaceutical Research and Development,
L.L.C., Raritan, NJ 08869-0602, USA
bDrug Discovery Division, Johnson and Johnson Pharmaceutical Research and Development,
L.L.C., Spring House, PA 19477-0776, USA
Received 23 February 2004; accepted 6 April 2004
Available online 12 May 2004
Abstract—Two approaches were developed to synthesize the novel 7-azaindolyl-heteroarylmaleimides. The first approach was based
upon the palladium-catalyzed Suzuki cross-coupling or Stille cross-coupling of 2-chloro-maleimide 5 with various arylboronic acids
or arylstannanes. The second approach was based upon the condensation of ethyl 7-azaindolyl-3-glyoxylate 12 with various
acetamides. The hydroxypropyl-substituted 7-azaindolylmaleimide template was first used to screen different heteroaryls attached to
the maleimide. Replacement of hydroxypropyl with different chain lengths and different functional groups were studied next. Many
compounds synthesized were demonstrated to have high potency at GSK-3b, good GS activity in HEK293 cells and good to
excellent metabolic stability in human liver microsomes. Three representative compounds (21, 33, and 34) were demonstrated to
have good selectivity against a panel of 80 kinase assays. Among them, compound 33 exhibited very weak inhibitions at the other 79
kinase assays, and behaved as a highly selective GSK-3b inhibitor.
ꢀ 2004 Elsevier Ltd. All rights reserved.
1. Introduction
receptor substrate-1 (IRS-1) and GS, the two key targets
in the insulin signaling pathway.6 Suppression of these
targets may limit most insulin-mediated biological re-
sponses. In addition, elevated GSK-3 activity was found
in diabetic tissues, reinforcing GSK-3 as a promising
therapeutic target for insulin resistance and Type 2
diabetes. Another important substrate of GSK-3 is Tau,
a protein that is critical in microtubule function in cer-
tain neurons. Tau hyperphosphorylation has been pos-
tulated to promote microtubule disassembly, an early
event in the progression of Alzheimer’s disease.7 Inhi-
bition of GSK-3 has also been shown to attenuate
apoptotic signals.8 Therefore, GSK-3 inhibitors may be
useful for the treatment of Alzheimer’s disease and
protection against cell death.9 Lithium ions and valproic
acid have been used as mood stabilizers for the chronic
treatment of patients with bipolar disorder. Recently,
these compounds have been shown to be GSK-3 inhib-
itors.10 Finally, studies on fibroblasts from the GSK-3b
knockout mice indicate that inhibition of GSK-3 may be
Glycogen synthase kinase-3 (GSK-3) is a serine/threo-
nine protein kinase that was first identified over 20 years
ago because of its ability to phosphorylate and inhibit
glycogen synthase (GS),1 the rate-limiting enzyme of
glycogen biosynthesis.2 Mammalian GSK-3 exists as
two isoforms, GSK-3a and GSK-3b, sharing 98%
homology in their catalytic domain.3 Both isoforms are
ubiquitously expressed in cells and tissues, and have
similar biochemical properties.3 Today, it is known that
GSK-3 is involved in diverse cellular processes and
might have multiple substrates.4;5 For example, GSK-3
phosphorylates and inhibits the functioning of insulin
Keywords: Highly selective inhibitor; Kinase; Azaindolemaleimide;
Cross-coupling.
* Corresponding author. Tel.: +1-908-704-4330; fax: +1-908-203-8109;
e-mail: gkuo@prdus.jnj.com
0968-0896/$ - see front matter ꢀ 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2004.04.010

3168
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
Cl
N
H
H
R
N
H3C
N
O
O
N
O
O
Cl
N
HN
N
N
N
H
HN
H
N
R
R1
N
R1
N
N
H
N
R
N
R2
N
R1
CN
Purines
Bisindolylmaleimides
Anilinomaleimides
H
CHIR 99021
H
N
H
N
O
N
O
O
N
O
O
O
R
N
N
tero-
Hetero-
O
N
N
O
aryl
aryl
N
N
N
N
N
N
N
S
R1
(CH2)nR
OH
O
O
2
Thiadiazolidinones
Macrocyclic polyoxygenated
bis-7-azaindolylmaleimide,
Compound A
Macrocyclic bis-7-azaindolyl-
maleimide, Compound B
7-Azaindolyl-heteroaryl
maleimides, Targets
Scheme 1.
useful in treating inflammatory disorders or diseases
through the negative regulation of NFjB activity.11
2. Chemistry
Two approaches were used to synthesize these novel
heteroaryl-7-azaindolylmaleimides. The first approach
was based upon the palladium-catalyzed Suzuki cross-
coupling20 or Stille cross-coupling21 of 2-chloro-
maleimide 5 with various arylboronic acids (method A)
or arylstannanes (method B) (Scheme 2). The chloride 5
was prepared from a readily available 7-azaindole-3-
acetonitrile 122 in four steps. Oxidation of 1 gave acet-
amide 2, and N-1 alkylation of 2 with the bromide gave
the three-carbon side chain attached acetamide 3. Using
a procedure for the synthesis of aryl substituted hy-
droxymaleimide derivatives,23 acetamide 3 was reacted
with diethyl oxalate in the presence of potassium tert-
butoxide to give 2-hydroxy-maleimide 4 in 80% yield.
Reaction of 4 with oxalyl chloride gave the key common
intermediate 5. It is known in the literature that the
electron-rich/steric bulky P(t-Bu)3 is one of the most
desirable ligands for oxidative addition of palladium(0)
into the C–Cl bond.20;21 Indeed, treatment of the com-
mercially available Pd(P(t-Bu)3)2 and the chloride 5 with
either arylboronic acids or arylstannanes gave moderate
to good yields of the desired coupling products 6–9, 21,
23, 29–40 after deprotection with n-Bu4NF in THF
(Scheme 2).
Since previous report showed that purines12 (Scheme 1)
are capable of exhibiting GSK-3 inhibitory activi-
ties, many chemical series13–15 (pyrazines, pyrimidines,
heterocyclic-pyrimidones, amine pyrazoles, bisindolylma-
leimides, hymenialdisine, paullones, indirubines, anil-
inomaleimides, [1,2,3]triazole-furazan, and CHIR99021)
have been developed as ATP competitive GSK-3
inhibitors while thiadiazolidinones16 were reported to
be the first ATP noncompetitive GSK-3 inhibitors.
However, with the exception of [1,2,3]triazole-fura-
zan,13c CHIR99021,14 and anilinomaleimides,15 which
were reported as selective GSK-3 inhibitors versus 20–40
protein kinases, the majority of ATP competitive GSK-3
inhibitors all showed equivalent activities at other pro-
tein kinases. In general, it is always desirable to have
a therapeutic agent to specifically inhibit the enzyme
target to minimize the potential side effects.
In an attempt to identify potent and selective protein
kinase C gamma (PKCc) inhibitors for the treatment of
chronic pain, we unexpectedly discovered that macro-
cyclic polyoxygenated bis-7-azaindolylmaleimide (com-
pound A, Ki ¼ 0:017 lM, Scheme 1)17a and macrocyclic
bis-7-azaindolylmaleimide
(compound
B,
Ki ¼
0:011 lM)17b are potent and highly selective GSK-3b
inhibitors. However, these compounds were found to be
metabolically liable (decreased metabolic stability in
human liver microsomes (HLM)), partially due to the
metabolism of the bottom cyclic linkers.18 This article
describes our continued efforts toward the identification
of several novel 7-azaindolyl-heteroarylmaleimides as
potent and selective GSK-3b inhibitors with improved
metabolic stability in HLM.19
The second approach was based upon the condensation
of ethyl 7-azaindolyl-3-glyoxylate 12 with various
acetamides.24 The common intermediate 12 was pre-
pared in two steps (Scheme 3). Reaction of diethyl
oxalate with the magnesium salt of 10 gave 11. Alkyl-
ation of 11 with the bromide gave glyoxylate 12. The
acetamide 15 was prepared from the known chloride
1325 in two steps (Scheme 4). Displacement of the
chloride with KCN gave 14, followed by oxidation gave

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3169
H
N
O
O
CONH2
CONH2
CN
OH
i
ii
iii
N
N
N
H
N
H
N
N
N
N
(CH2)3R
(CH2)3R
1
2
3, R = OTBDPS
45, R = Ph
4, R = OTBDPS
46, R = Ph
57, R = OPh
61, R = CN
58, R = OPh
62, R = CN
H
H
N
N
O
O
O
O
Cl
Ar
iv
v
vi
(method A or
method B)
N
N
N
N
(CH2)3R
(CH2)3R
5, R = OTBDPS
47, R = Ph
6-9, 21, 23, 29-40
48-49, 60, 64-66
59, R = OPh
63, R = CN
Scheme 2. Reagents and conditions: (i) H2O2, K2CO3, DMSO; (ii) Br(CH2)3R, Cs2CO3, DMF; (iii) (CO2Et)2, KOt-Bu, THF; (iv) (COCl)2, DMF/
CH2Cl2; (v) method A: Pd2(dba)3, Pd(P(t-Bu)3)2, ArB(OH)2; method B: Pd(P(t-Bu)3)2, ArSnBu3; (vi) TBAF, THF (for 5, R ¼ OTBDPS series only).
O
OC2H5
O
O
OC2H5
O
i
ii
N
N
N
H
N
H
N
N
(CH2)3OTBDMS
10
11
12
Scheme 3. Reagents and conditions: (i) EtMgBr, (C2H5OCO)2, THF; (ii) Br(CH2)3OTBDMS, Cs2CO3, DMF.
H
H
N
N
O
O
O
O
SEM
N
N
N
N
NH
ii
iii
iv
i
CONH2
Cl
CN
N
N
N
N
N
N
N
N
N
SEM
SEM
SEM
(CH2)3OTBDMS
(CH2)3OH
15
13
14
16
17
Scheme 4. Reagents and conditions: (i) KCN, EtOH; (ii) H2O2, K2CO3, DMSO; (iii) 12, KOt-Bu, THF; (iv) TFA, CH2Cl2.
H
N
H
N
H
N
H
N
O
O
O
O
O
O
O
N
O
N
CONH2
N
N
iii
i, ii
N
+
+
N
N
N
N
N
H
N
N
N
N
N
N
N
N
(CH2)3OTBDMS
(CH2)3OH
(CH2)3OTBDMS
(CH2)3OH
18
19
20
21
22
Scheme 5. Reagents and conditions: (i) CH3I, Cs2CO3, DMF; (ii) 12, KOt-Bu, THF; (iii) TBAF, THF.
15. Condensation of 12 with 15 in the presence of KOt-
Bu gave 16, removal of the silyl-protecting groups with
TFA gave the product 17 (Scheme 4).
2-(2-methyl-2H-pyrazol-3-yl)-acetamide. Condensation
of 12 with these mixtures in the presence of base gave 19
and 20 (Scheme 5). Removal of the silyl-protecting
group from 19 (or 20) with TBAF gave product 21
(or 22). The structure differentiation of 21 and 22
was confirmed by an independent synthesis of 21 via
Methylation of pyrazole-acetamide 1826 gave a 2:1
mixture of 2-(1-methyl-1H-pyrazole-3-yl)-acetamide and

3170
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
palladium-catalyzed Stille coupling reaction of 5 with 2-
1-methyl-3-tributylstannanyl-1H-pyrazole27 (Scheme 2).
catalyzed cross-coupling reactions of the chlorides gave
desired products 48–49, 60, and 64–66. The N-methyl
analogue of the key intermediate 5, that is chloride 50,
could also be prepared in a direct manner30 (Scheme 8).
Condensation of the Grignard reagent of 10 with 2,3-
dichloro-N-methylmaleimide gave 50 in 21% yield.
Alkylation, followed by cross-coupling reaction with
organo-tin 43 in microwave gave 53 (or 54). Hydrolysis
and ammonolysis31 gave the desired product 55 (or 56).
Alkylation of acetamide 2428 with the bromides gave 25
(or 26) (Scheme 6). Condensation of 12 with 25 (or 26),
followed by TBAF treatment gave product 27 (or 28).
During the palladium-catalyzed Stille reaction of 5 with
organo-tin 43, we also isolated 21% yield of the n-butyl
coupling product 44 (Scheme 7) in addition to the ex-
pected product 34 (36% yield). The butyl- (vs phenyl-)
transferred from phenyltributyltin was also observed
previously by Farina et al.29
3. Results and discussion
Again in Scheme 2, alkylation of acetamide 2 with
various bromides gave 45, 57, and 61, condensation with
diethyl oxalate gave 46, 58, and 62, followed by chlori-
nation gave the chlorides 47, 59, and 63. Palladium-
It has been demonstrated that the acyclic bis-
indolylmaleimides
(GF
109,203
where
R ¼
Me2N(CH2)3, R1 ¼ H; Ro 31-8220 where R ¼
NH2CS(@NH)(CH2)3, R1 ¼ CH3, Scheme 1), widely
H
N
O
O
H2NOC
H2NOC
N
N
N
i
ii, iii
N
(CH2)nOTBDMS
N
N
N
N
H
(CH2)nOH
(CH2)3OH
25, n = 3
26, n = 2
24
27, n = 3
28, n = 2
Scheme 6. Reagents and conditions: (i) Br(CH2)nOTBDMS, Cs2CO3, DMF, 80 ꢁC; (ii) 12, KOt-Bu, THF; (iii) TBAF, THF.
H
N
H
N
H
N
O
O
O
O
O
O
H3CO
Bu3Sn
OCH3
N
N
Cl
i, ii
+
+
OCH3
N
N
N
N
N
N
N
N
OCH3
43
(CH2)3OTBDPS
(CH2)3OH
(CH2)3OH
5
44
34
Scheme 7. Reagents and conditions: (i) Pd(P(t-Bu)3)2, THF; (ii) TBAF, THF.
N
N
N
O
O
O
O
O
O
OCH3
N
iv, v
i
ii
Cl
Cl
iii
N
H
N
N
H
N
N
N
N
N
N
OCH3
(CH2)nPh
(CH2)nPh
10
50
51, n = 2
52, n = 1
53, n = 2
54, n = 1
H
N
O
O
N
OCH3
N
N
N
OCH3
(CH2)nPh
55, n = 2
56, n = 1
Scheme 8. Reagents and conditions: (i) EtMgBr, 2,3-dichloro-N-methylmaleimide, toluene; (ii) Ph(CH2)nBr, NaH, DMF; (iii) 43, Pd2(dba)3,
P(t-Bu)3, THF/DMF, microwave; (iv) KOH, EtOH, 10% citric acid; (v) HMDS, MeOH/DMF, 80 ꢁC.

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3171
used as protein kinase C (PKC) inhibitors are also po-
tent GSK-3 inhibitors.32 We thus decided to focus our
initial chemistry efforts on the preparation of three-
carbon chain molecules. Since the hydroxyl analog
(compound B, Ki ¼ 0:011 lM, Scheme 1) was the most
potent GSK-3b inhibitor in our previous studies,17b we
felt the combination of a hydroxypropyl side chain and
7-azaindolylmaleimide may provide a good template for
searching for novel molecules with high potency and
selectivity at GSK-3b.
1 lM), pyrazole 23 (Ki ¼ 0:098 lM), and thiazole 30
(8.5% inhibition @ 1 lM) also displayed a wide range of
potency, it appears that the basicity of the heteroaryls
did not have any impact on the GSK-3b inhibitory
potency. Attachment of a heterocycle instead of the
heteroaryl to the maleimide resulted in compound 35
with lower potency (Ki ¼ 0:084 lM) as compared to
compound 33 (Ki ¼ 0:025 lM).
We continued to examine the impact of fused-biaryl
rings (36–39) on the GSK-3b inhibitory potency. Similar
to the observation of methoxy-substituted pyrimidine
34, the extra ring fused at either 2,3-position (36,
Ki ¼ 0:057 lM) or 3,4-position (Ki ¼ 0:029 lM for 37;
Ki ¼ 0:081 lM for 38, and Ki ¼ 0:098 lM for 39) did not
seem to be detrimental to the potency. However, the
potency was reduced significantly if the fused-triaryl ring
was installed (40, 55% inhibition at 1 lM). While the
replacement of the heteroaryl with small hydroxyl group
or nitrile group totally abolished the kinase inhibition
(4% inhibition at 1 lM for 41 and 1% inhibition at 1 lM
for 42, Table 2), the n-butyl group replacement retained
moderate potency (44, Ki ¼ 0:129 lM).
Four pyrrole or furan attached 7-azaindolylmaleimides
were first synthesized (6–9, Table 1). We were pleased to
see that all four acyclic maleimides showed comparable
high potency (Ki ¼ 0:006–0:089 lM) to the cyclic bis-7-
azaindolylmaleimide (compound B, Ki ¼ 0:011 lM) al-
though the furan 9 displayed slightly lower potency
(Ki ¼ 0:089 lM). We then explored the influence of bis-
nitrogen containing five-member-heteroaryl maleimides
(17, 21–23) on the GSK-3b inhibitory potency. It was
very surprised to find out that while all three pyrazole
analogs (21–23) exhibited good to moderate potency
(Ki ¼ 0:048–0:285 lM), imidazole 17 was poorly active
(29.9% inhibition at 1 lM). Among the three pyrazole
analogs (21–23), pyrazole 22 with the N-CH3 substitu-
tion at the 2-position relative to the maleimide displayed
the lowest potency (Ki ¼ 0:285 lM).
After exploring the structure–activity relationship at
the heteroaryl site, we turned our attention to optimize
the hydroxypropyl side chain portion. Replacing the
hydroxyl group of compound 34 (Ki ¼ 0:006 lM) and 31
(Ki ¼ 0:020 lM) with phenyl group gave 48
(Ki ¼ 0:006 lM, Table 3) and 49 (Ki ¼ 0:178 lM). It
seems the more lipophilic phenyl group did not offer
improvement in potency. Shortening the chain length to
two-carbon or one-carbon linker afforded 55
(Ki ¼ 0:063 lM) and 56 (61.4% inhibition at 1 lM). As
compared to the first phenyl analog 48 (Ki ¼ 0:006 lM),
the three-carbon chain is still more desirable from the
potency point of view. Insertion of one oxygen atom
into the side chain of 48, which potentially might
serve as the hydrogen-bond acceptor, gave 60
(Ki ¼ 0:026 lM). No improvement was observed
when compared to 48. Replacing the hydroxyl group
of 34 (Ki ¼ 0:006 lM), 33 (Ki ¼ 0:025 lM), and 21
(Ki ¼ 0:048 lM) with a nitrile group gave the corre-
sponding 64 (Ki ¼ 0:018 lM), 65 (Ki ¼ 0:045 lM), and
66 (Ki ¼ 0:051 lM). Once more, the hydroxyl group
proved to be highly favorable for GSK-3b inhibitory
potency in the 7-azaindolylmaleimide series, although
for a reason not clearly understood yet.
We next examined the impact of introduction of the
second hydroxyalkyl side chain to the molecule. Both
compound 27 and compound 28 with additional hy-
droxypropyl or hydroxyethyl side chains gave much
lower potency (Ki ¼ 5:54 lM for 27 and Ki ¼ 1:9 lM for
28) unfortunately. It was decided to re-visit the mono-
hydroxypropyl-substituted series. Both bismethyl-
substituted isoxazole 29 and thiazole 30 gave poor
activity (28.5% inhibition at 1 lM for 29 and 8.5%
inhibition at 1 lM for 30). Up to this stage, it seems
consistent that the steric hindrance at the 2-position of
the heteroaryls (such as 22 and 29) or the installation of
heteroatoms at both of the 2-position of the five-mem-
ber rings (such as 17 and 30) are unfavorable to the
kinase inhibition at GSK-3b.
Replacement of the five-member heteroaryls with six-
member heteroaryls gave compounds 31–34 with high
potency (Ki ¼ 0:006–0:025 lM) except compound 32. It
is interesting to note that while the installation of
nitrogen atoms at both of the ortho-position of the six-
member ring gave, consistently, poor activity (21.5%
inhibition at 1 lM for 32); the increased steric hindrance
of the OCH3 substituent at the 2-position of the
pyrimidine gave even better potency (Ki ¼ 0:006 lM for
34 vs Ki ¼ 0:020 lM for 31). There is no obvious ratio-
nale for these observations. Meanwhile, we also exam-
ined if there is any correlation between the basicity of
the heteroaryls (pKa of the protonated heteroaryls) and
the GSK-3b inhibitory potency. Since some of the most
basic heteroaryls such as pyrrole 6 (Ki ¼ 0:006 lM),
imidazole 17 (29.9% inhibition @ 1 lM) imidazole 27
(Ki ¼ 5:54 lM) displayed a wide range of potency, some
of the less basic heteroaryls such as pyrimidine 31
(Ki ¼ 0:020 lM), pyrimidine 32 (21.5% inhibition @
3.1. Cellular activity
Among the multiple cellular processes in which GSK-3
has been implicated, the ability to phosphorylate and
inhibit GS was the first regulatory process discovered
and is perhaps the most thoroughly studied.1 Therefore,
the cell-based assay examining GS activation represents
a direct functional assay to measure the cellular activity
of GSK-3 inhibitors. As described above, LiCl is a
known inhibitor of GSK-3b. LiCl increases GS activity
in isolated rat adipocytes and hepatocytes,33 and pro-
duces intracellular effects similar to that achieved by
GSK-3 inhibition.34;35
All potent compounds

3172
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
Table 1. Inhibition constants at GSK-3ba
H
N
O
O
Ar
N
N
HO
Compd
Ar
GSK-3b
% Inhibition @ 1 lM ( SEM)
Ki ꢀ SEMb (lM)
6
7
0.006 0.001
N
H
0.019 0.001
N
CH3
8
9
0.050 0.014
0.089 0.012
O
O
NH
17
21
22
29.9 4.7
N
N
0.048 0.009
0.285 0.032
N
CH3
CH3
N
N
N
23
27
0.098 0.006
5.54 1.09
N
CH3
N
N
(CH2)3OH
N
28
1.9 0.27
N
(CH2)2OH
CH3
29
30
28.5 2.5
8.5 3.2
N
H3C
N
O
N
S
31
32
0.020 0.001
N
N
21.5 1.1
N
N
N
33
0.025 0.001
OCH3
N
34
35
0.006 0.001
0.084 0.007
N
O
OCH3
O

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3173
Table 1 (continued)
Compd
Ar
GSK-3b
Ki ꢀ SEMb (lM)
% Inhibition @ 1 lM ( SEM)
N
36
37
0.057 0.003
O
0.029 0.005
0.081 0.013
0.098 0.013
S
38
39
N
N
O
40
55.5 5.3
a Assay details were described in the Experimental section.
b SEM: standard error mean.
Table 2. Inhibition constants at GSK-3ba
T1=2 ¼ 41 min; 34, T1=2 ¼ 28 min; 37, T1=2 ¼ 35 min, and
64, T1=2 ¼ 44 min, Table 4) to high metabolic stability
(21, T1=2 ¼ 83 min; 31, 33, 65, T1=2 > 100 min, and 66,
T1=2 ¼ 97 min) in HLM. It is interesting to observe that
while N-methyl pyrazole 21 showed high metabolic
stability (T1=2 ¼ 83 min), the corresponding N-methyl
pyrrole 7 showed much lower metabolic stability
(T1=2 ¼ 17 min). On the other hand, in the series of di-
H
N
O
O
R
N
N
HO
methoxypyrimidine, the hydroxypropyl 34 (T1=2
¼
28 min) and cyanopropyl 64 (T1=2 ¼ 44 min) are meta-
Compd
R
GSK-3b
bolically more stable than phenylpropyl 48 (T1=2
<
Ki ꢀ SEMb (lM) % Inhibition @
1 lM ( SEM)
2 min) or phenoxypropyl 60 (T1=2 ¼ 2 min). Therefore,
although it is difficult to predict the metabolic stability
of compounds in HLM in general, our designed
molecular template, the combination of hydroxypropyl
side chain and 7-azaindolylmaleimide, seems to be
metabolically stable in HLM.
41
42
44
OH
CN
4.0 2.8
1.0 0.7
n-C4H9
0.129 0.028
a Assay details were described in the Experimental section.
b SEM: standard error mean.
3.3. Kinase selectivity
(Ki < 0:050 lM) and LiCl were tested for the ability to
increase GS activity in human embryonic kidney
(HEK293) cells. In general, all of the compounds
exhibited much greater potency (EC50 ¼ 0.032–1.7 lM,
Table 4) than LiCl (EC50 > 3000 lM) in the HEK293
cells. The 6–12-fold difference in potencies between the
EC50 of the GS cell assay and the Ki of the GSK-3b
kinase assay for most of the compounds may attribute
to the differences in the species tested, that is, human
cells versus recombinant rabbit enzyme. The good GS
activity demonstrated that these maleimides should have
good cell permeabilities.
Since many reported specific inhibitors of protein ki-
nases were found to be nonspecific when reexamined
against a large panel of protein kinases by Cohen and
co-workers,36 we decided to evaluate the selectivity of
our compounds against a broad panel of 80 protein
kinase assays at UBI (Upstate Biotech Inc.). Three po-
tent compounds (21, 33, and 34) that covered a broad
spectrum in GSK-3b inhibitory potency, GS cell-based
activity and metabolic stability in HLM were selected
for these kinase selectivity studies. In the presence of
100 lM ATP, compounds 21, 33, and 34 inhibited GSK-
3b kinase activity by 92%, 88%, and 96% at a concen-
tration of 10 lM, respectively (Table 5). Interestingly,
compound 21 only showed moderate inhibitions in the
CDK, PKB, and RsK assays and exhibited very weak
inhibitory activities at the other 74 kinases. Compound
34 also only showed moderate inhibitions at CDK
3.2. Metabolic stability in human liver microsomes
We were pleased to note that most of the compounds
tested exhibited acceptable metabolic stability (6,

3174
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
Table 3. Inhibition constants at GSK-3ba
H
N
O
O
Ar
N
N
(CH2)nR
Compd
n
R
Ar
GSK-3b
Ki ꢀ SEMb (lM)
% Inhibition @ 1 lM
OCH3
N
48
3
Ph
0.006 0.001
N
OCH3
N
49
55
3
2
Ph
Ph
0.178 0.004
0.063 0.009
N
OCH3
N
N
OCH3
OCH3
N
56
60
64
1
3
3
Ph
61.4 3.3
N
OCH3
OCH3
N
OPh
CN
0.026 0.003
0.018 0.001
N
OCH3
OCH3
N
N
N
OCH3
65
66
3
3
CN
CN
0.045 0.004
0.051 0.009
N
N
N
CH3
a Assay details were described in the Experimental section.
b SEM: standard error mean.
at PKC/GSK-3b.32a;38 Therefore, it was not unusual to
see that both 21 and 34 displayed moderate inhibitions
at CDK isoenzymes. However, it was surprised to
observe very weak inhibitions at the PKC isoenzymes
for both 21 and 34 considering the structure similarity
between them and bisindolylmaleimides. Finally and
remarkably, compound 33 exhibited very low activity
against all of the other 79 kinases and thus representing
a ‘specific inhibitor’ of GSK-3b.
Table 4. Glycogen synthase activity in HEK293 cells and metabolic
stability in human liver microsomesa
Compd
EC50 SEM (lM)
T1=2 (min)
6
0.043 0.005
0.165 0.025
0.290 0.021
1.700 0.284
0.168 0.028
0.032 0.013
0.075 0.011
0.035 0.011
0.053 0.008
0.181 0.031
0.150 0.042
0.391 0.031
0.350 0.041
>3000
41
17
7
8
10
83
21
31
33
34
37
48
60
64
65
66
LiCl
>100
>100
28
35
<2
4. Conclusion
2
44
Two approaches were developed to synthesize the novel
7-azaindolyl-heteroarylmaleimides. The first approach
was based upon the palladium-catalyzed Suzuki cross-
coupling or Stille cross-coupling of 2-chloro-maleimide
5 with various arylboronic acids or arylstannanes. The
second approach was based upon the condensation of
ethyl 7-azaindolyl-3-glyoxylate 12 with various acet-
>100
97
––
a Assay details were described in the Experimental section.
assays and exhibited very weak inhibitions at the other
75 kinases. It was reported recently that some potent
CDK inhibitors could also inhibit GSK-3b,37 and bis-
indolylmaleimides (Scheme 1) displayed similar potency
amides.
The
hydroxypropyl-substituted
7-aza-
indolylmaleimide template was first used to screen
different heteroaryls attached to the maleimide.

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3175
Table 5. Activities at 80 protein kinasesa
Table 5 (continued)
Protein kinase
Activity (% of control)
33 34
12
Protein kinase
Activity (% of control)
21
21
80
33
34
GSK3b (h)
Abl (m)
8
90
66
99
97
76
97
87
89
64
39
56
46
60
73
81
67
82
84
89
97
86
99
89
76
82
98
88
117
99
103
122
105
65
80
88
107
98
112
93
92
93
83
86
68
104
87
102
73
74
93
95
88
45
89
92
105
57
105
91
120
52
93
51
75
4
PRK2 (h)
ROCK-II (h)
ROCK-II (r)
Rsk1 (r)
100
111
101
86
91
104
106
84
78
69
98
91
95
99
77
83
85
74
89
95
67
85
82
98
97
64
49
28
92
93
99
95
68
106
91
89
96
AMPK (r)
Arg (m)
100
83
101
70
Aurora-A (h)
Axl (h)
Blk (m)
Rsk2 (h)
Rsk3 (h)
72
58
86
89
79
84
SAPK2a (h)
SAPK2b (h)
SAPK3 (h)
SAPK4 (h)
SGK (h)
69
93
Bmx (h)
65
96
55
CAMKII (r)
CAMKIV (h)
CDK1/cyclinB (h)
CDK2/cyclinA (h)
CDK3/cyclinE (h)
CDK5/p35 (h)
CDK6/cyclinD3
CDK7/cyclinH
CHK1 (h)
CHK2 (h)
CK1 (y)
103
87
102
98
68
61
27
11
91
79
70
64
Syk (h)
TrkB (h)
17
93
96
68
78
37
Yes (h)
ZAP-70 (h)
55
82
80
82
88
a Protein kinase were assayed with 10 lM of 21, 33, and 34 in the
presence of 100 lM ATP. Activities were given as the mean per-
centage of that in control incubations (averages of duplicate deter-
minations). Assay details were described in the Experimental section.
75
79
82
92
63
57
CK2 (h)
71
92
c-RAF (h)
CSK
cSRC (h)
87
89
75
Replacement of hydroxypropyl with different chain
lengths and different functional groups were studied
next. Many compounds synthesized were demonstrated
to have high potency at GSK-3b and good GS activity
in HEK293 cells. More importantly, most of these
potent GSK-3b inhibitors possess good to excellent
metabolic stability in human liver microsomes. Three
representative compounds (21, 33, and 34) were
demonstrated to have good selectivity against a panel of
80 kinase assays. Among them, compound 33 exhibited
very weak inhibitions at the other 79 kinase assays, and
behaved as a GSK-3b ‘specific inhibitor’. The high
inhibitory potency, selectivity, cellular activities, and
metabolic stability in HLM suggested that these 7-aza-
indolyl-heteroarylmaleimides may render them as valu-
able pharmacological tools in elucidating the complex
roles of GSK-3b in cell signaling pathways and the
potential usage for the treatment of elevated level of
GSK-3b involved diseases.
104
87
90
82
Fes (h)
FGFR3 (h)
Fyn (h)
IGF-1R
80
58
100
93
106
80
IKKa
IKKb
IR
93
82
101
115
79
91
94
JNK1a1 (h)
JNK2a2 (h)
JNK3 (r)
88
121
79
121
63
Lck (h)
Lyn (h)
Lyn (m)
68
89
55
79
90
94
62
88
MAPK1 (h)
MAPK2 (h)
MAPK2 (m)
MAPKAP-K2 (h)
MEK1 (h)
MKK4 (m)
MKK6 (h)
MKK7b (h)
MSK1 (h)
p70S6K (h)
PAK2 (h)
PDGFRa
PDGFRb
PDK1 (h)
PKA (b)
96
106
97
100
100
103
96
95
100
90
89
80
106
87
70
67
5. Experimental
5.1. Chemistry
113
94
108
86
106
89
107
84
5.1.1. General information. 1H NMR spectra were
measured on a Bruker AC-300 (300 MHz) spectrometer
using tetramethylsilane as an internal standard. Ele-
mental analyses were obtained by Quantitative Tech-
nologies Inc. (Whitehouse, New Jersey), and the results
were within 0.4% of the calculated values unless other-
wise mentioned. Melting points were determined in
open capillary tubes with a Thomas–Hoover appara-
tus and were uncorrected. Electrospray mass spectra
(MS-ES) were recorded on a Hewlett Packard 59987A
spectrometer. High resolution mass spectra (HRMS)
were obtained on a Micromass Autospec. E. spec-
trometer. The term ‘DMAP’ refers to dimethylamino-
pyridine, ‘TFA’ refers to trifluoroacetic acid, ‘NMP’
refers to 1-methyl-2-pyrrolidinone, ‘DPPF’ refers to
94
96
83
96
PKA (h)
95
95
101
94
PKBa (h)
PKBb (h)
PKBc (h)
PKCa (h)
PKCb II (h)
PKCc (h)
PKCd (h)
PKCe (h)
PKCg (h)
PKCl (h)
PKCh (h)
PKD2 (h)
PRAK (h)
63
95
136
89
95
92
97
101
83
64
129
102
119
69
102
118
108
77
90
61
97
74
77
90

3176
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
1,10-bis(diphenylphosphino)ferrocene, ‘Pd2(dba)3’ refers
to tris(dibenzylideneacetone)dipalladium(0)-chloroform
adduct, ‘DPPA’ refers to diphenylphosphoryl azide,
‘TBAF’ refers to tetrabutylammonium fluoride.
0.06 mmol) in 1:1 CH2Cl2/DMF (2 mL) at 23 ꢁC under
nitrogen. After 1 h the reaction mixture was concen-
trated and a crude product was purified by column
chromatography (SiO2) to give 0.023 g (73%) of com-
1
pound 5 as a yellow solid: H NMR (400 MHz, CDCl3)
d 10.14 (s, 1H), 8.69 (dd, J ¼ 8:1, 1.5 Hz, 1H), 8.4 (dd,
J ¼ 4:6, 1.5 Hz, 1H), 8.36 (s, 1H), 7.67 (m, 4H), 7.41 (m,
6H), 7.25 (dd, J ¼ 8:1, 4.6 Hz, 1H), 4.67 (t, J ¼ 6:8 Hz,
2H), 3.74 (t, J ¼ 6:0 Hz, 2H), 2.24 (m, 2H), 1.04 (s, 9H);
MS (ES) m=z 544 (M+Hþ).
5.1.2. 2-(1H-Pyrrolo[2,3-b]pyridin-3-yl)-acetamide (2).
Potassium carbonate (3.4 g, 24.7 mmol) was added to a
solution of compound 1 (7.75 g, 49.4 mmol) in DMSO
(15 mL) at 0 ꢁC, followed by dropwise addition of
hydrogen peroxide (8.4 mL, 74 mmol; 30% solution in
H2O). The resulting solution was stirred for 10 min,
CH2Cl2 was added and the reaction mixture was then
filtered and concentrated. CH2Cl2 (100 mL) was added
followed by Et2O (20 mL) and the mixture cooled. The
resulting precipitate was filtered off to give 7.212 g (84%)
5.1.6. General procedure for the synthesis of 6, 8, 29, 31,
36–38, and 40 (method A). 4-[1-(3-Hydroxypropyl)-1H-
pyrrolo[2,3-b]pyridin-3-yl]-[3,30-bi-1H-pyrrole]-2,5-dione
(6). 1-(Triisopropyl)silyl-pyrrole-3-boronic acid (0.053
mL, 0.2 mmol) was added to a solution of compound
5 (0.054 g, 0.1 mmol), Pd2(dba)3 (5 mg, 0.005 mmol),
Pd(tBu3P)2 (5 mg, 0.01 mmol), and potassium fluoride
(20 mg, 0.34 mmol) in THF (1 mL) at 23 ꢁC under
nitrogen. The mixture was stirred at 23 ꢁC for 18 h,
then diluted with EtOAc (10 mL), filtered through Cel-
ite, and concentrated. The product was purified by col-
umn chromatography (SiO2) to give 0.044 g (60%) of
silyl-protected product as a yellow solid: 1H NMR
(300 MHz, CDCl3) d 8.28 (d, J ¼ 4:9 Hz, 1H), 7.74 (s,
1H), 7.66 (m, 4H), 7.36 (m, 7H), 7.17 (m, 1H), 6. 89 (dd,
J ¼ 8:3, 4.9 Hz, 1H), 6.62 (m, 1H), 6.37 (m, 1H), 4.53 (t,
J ¼ 6:8 Hz, 2H), 3.7 (t, J ¼ 6:0 Hz, 2H), 2.2 (m, 2H),
1.37 (sep, J ¼ 7:4 Hz, 3H),1.09 (s, 9H), 1.05 (s, 9H), 1.02
(s, 9H); MS (ES) m=z 731 (M+Hþ).
1
of compound 2 as a yellow solid: H NMR (300 MHz,
DMSO) d 11.44 (br s, 1H), 8.19 (d, J ¼ 4:7, 1.5 Hz, 1H),
7.95 (dd, J ¼ 7:9, 1.5 Hz, 1H), 7.39 (br s, 1H), 7.29 (s,
1H), 7.06 (dd, J ¼ 7:9, 4.9 Hz, 1H), 6.86 (br s, 1H), 3.50
(s, 2H); MS (ES) m=z 176 (M+Hþ).
5.1.3.
2-{1-[3-(tert-Butyl-diphenyl-silanyloxy)-propyl]-
1H-pyrrolo[2,3-b]pyridin-3-yl}-acetamide (3). Cesium
carbonate (0.45 g, 1.37 mmol) and (3-bromo-propoxy)-
tert-butyl-diphenyl-silane (0.19 g, 0.5 mmol) were added
to a solution of compound 2 (0.08 g, 0.46 mmol) in
DMF (2 mL) and the resulting mixture was stirred at
70 ꢁC. After 6 h, the reaction mixture was filtered
through Celite, diluted with EtOAc (10 mL), and wa-
shed with water (5 · 10 mL). The organic layer was then
dried (MgSO4) and concentrated. The crude product
was purified by column chromatography (SiO2) to give
TBAF (0.12 mL, 1 M solution in THF, 0.12 mmol) as
added dropwise to a solution of this silyl-ether in THF
(1 mL) under nitrogen. After 18 h, the mixture was
concentrated and purified by column chromatography
(SiO2) to give 0.017 g (84%) of compound 6 as an orange
1
0.132 g (60%) of compound 3 as a clear oil: H NMR
(400 MHz, CDCl3) d 8.31 (dd, J ¼ 4:6, 1.3 Hz, 1H), 7.85
(dd, J ¼ 4:6, 1.3 Hz, 1H), 7.64 (m, 4H), 7.38 (m, 6H),
7.05 (m, 2H), 4.42 (t, J ¼ 6:6 Hz, 2H), 3.64 (t,
J ¼ 5:9 Hz, 2H), 3.62 (s, 2H), 2.09 (m, 2H), 1.08 (s, 9H);
MS (ES) m=z 472 (M+Hþ).
1
solid: H NMR (300 MHz, acetone-d6) d 10.47 (s, 1H),
9.57 (s, 1H), 8.3 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.9 (s, 1H),
7.56 (m, 1H), 7.49 (dd, J ¼ 7:9, 1.5 Hz, 1H), 7.01 (dd,
J ¼ 7:9, 4.7 Hz, 1H), 6.75 (m, 1H), 6.22 (m, 1H), 4.55 (t,
J ¼ 6:6 Hz, 2H), 4.02 (t, J ¼ 5:7 Hz, 1H), 3.56 (q,
J ¼ 5:8 Hz, 2H), 2.11 (m, 2H); MS (ES) m=z 337
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 337.1301, found
337.1292.
5.1.4.
3-{1-[3-(tert-Butyl-diphenyl-silanyloxy)-propyl]-
1H-pyrrolo[2,3-b]pyridin-3-yl}-4-hydroxy-pyrrole-2,5-di-
one (4). Potassium tert-butoxide (0.69 mL, 0.69 mmol;
1 M solution in THF) was added dropwise to a solution
of compound 3 (0.13 g, 0.35 mmol) and diethyl oxalate
(0.101 g, 0.69 mmol) in TMF (2 mL) at 0 ꢁC under
nitrogen. After 20 min the reaction mixture was con-
centrated and a crude product was purified by column
chromatography (SiO2) to give 0.117 g (80%) of com-
5.1.7. General procedure for the synthesis of 7, 9, 21, 23,
30, 32–35, and 39 (method B). 4-[1-(3-Hydroxypropyl)-
1H-pyrrolo[2,3-b]pyridin-3-yl]-10-methyl-10H-[3,30]bipyr-
rolyl-2,5-dione (7). To a solution of compound 5 (54 mg,
0.1 mmol) and Pd(tBu3P)2 (5 mg, 0.01 mmol) in THF
(2 mL) was added 1-methyl-3-tributylstannanyl-1H-
pyrrole (51 mg, 0.13 mmol), at 23 ꢁC under nitrogen. The
reaction mixture was refluxed for 18 h. Upon cooling,
the mixture was diluted with EtOAc (10 mL) and wa-
shed with H2O, KF, brine, and dried. After concentra-
tion, the crude product was purified by column
chromatography (SiO2) to give 35 mg (60%) of silyl-
1
pound 4 as a yellow solid: H NMR (300 MHz, CDCl3)
d 9.44 (s, 1H), 8.69 (dd, J ¼ 7:9, 1.3 Hz, 1H), 8.33 (dd,
J ¼ 4:7, 1.5 Hz, 1H), 8.20 (s, 1H), 7.69 (m, 4H), 7.41 (m,
6H), 7.14 (dd, J ¼ 7:9, 4.5 Hz, 1H), 4.6 (t, J ¼ 6:8 Hz,
2H), 3.73 (t, J ¼ 6:0 Hz, 2H), 2.2 (m, 2H), 1.07 (s, 9H);
MS (ES) m=z 526 (M+Hþ).
5.1.5.
3-{1-[3-(tert-Butyl-diphenyl-silanyloxy)-propyl]-
protected product as a
yellow solid: 1H NMR
1H-pyrrolo[2,3-b]pyridin-3-yl}-4-chloro-pyrrole-2,5-dione
(5). Oxalyl chloride (0.015 mL, 0.18 mmol) was added in
one portion to a solution of compound 4 (0.03 g,
(300 MHz, CDCl3) d 8.34 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.74
(s, 1H), 7.64 (m, 4H), 7.35 (m, 9H), 6.97 (dd, J ¼ 7:9,
4.7 Hz, 1H), 6.43 (m, 1H), 6.12 (m, 1H), 4.53 (m, 2H),

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3177
4.01 (m, 1H), 3.72 (m, 2H), 3.66 (s, 3H), 2.18 (m, 2H),
1.09 (s, 9H); MS (ES) m=z 589 (M+Hþ).
5.1.11. {1-[3-(tert-Butyl-dimethyl-silanyloxy)-propyl]-1H-
pyrrolo[2,3-b]pyridin-3-yl}-oxo-acetic acid ethyl ester
(12). To
1.46 mmol) and Cs2CO3 (2.38 g, 7.30 mmol) in DMF
(5 mL) was added DMF (2 mL) solution of
(3-bromopropoxy)-tert-butyldimethylsilane (1.85 g,
a mixture of compound 11 (318 mg,
To a solution of this silyl-ether (35 mg, 0.06 mmol) in
THF (2 mL) was added TBAF (1 M solution in THF,
1.5 equiv) dropwise under nitrogen. After 18 h, the
mixture was concentrated and purified by column
chromatography (SiO2) to give 15 mg (71%) of com-
a
7.30 mmol) at 80 ꢁC. After it was stirred at 80 ꢁC for
10 min, the mixture was cooled, diluted with EtOAc, and
filtered through Celite. The filtrate was washed with
water (4 · 25 mL), dried (Na2SO4), and concentrated.
The residue was chromatographed on silica gel, eluting
gradually with hexane/EtOAc to give 457 mg (80%) of
compound 12 as a white crystalline solid: 1H NMR
(300 MHz, CDCl3) d 8.61 (dd, J ¼ 6:3, 1.5 Hz, 1H), 8.46
(s, 1H), 8.35 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.22 (m, 1H), 4.43
(t, J ¼ 6:8 Hz, 2H), 4.38 (q, J ¼ 7:1 Hz, 2H), 3.57 (t,
J ¼ 5:7 Hz, 2H), 2.05 (m, 2H), 1.38 (t, J ¼ 7:1 Hz, 3H),
0.87 (s, 9H), 0.00 (s, 6H); MS (ES) m=z 391 (M+Hþ).
1
pound 7 as an orange solid: H NMR (300 MHz, ace-
tone-d6) d 9.54 (s, 1H), 8.30 (dd, J ¼ 4:7, 1.5 Hz, 1H),
7.89 (s, 1H), 7.50 (dd, J ¼ 8:1, 1.7 Hz, 1H), 7.44 (m, 1H),
7.02 (dd, J ¼ 7:9, 4.7 Hz, 1H), 6.60 (m, 1H), 6.11 (dd,
J ¼ 2:8, 1.7 Hz, 1H), 4.54 (m, 2H), 4.01 (m, 1H), 3.70 (s,
3H), 3.55 (m, 2H), 2.11 (m, 2H); MS (ES) m=z 351
(M+Hþ). Anal. Calcd for C19H18N4O3Æ0.1H2O: C,
64.80; H, 5.21; N, 15.91. Found: C, 64.77; H, 5.08; N,
15.82.
5.1.8. 3-Furan-3-yl-4-[1-(3-hydroxypropyl)-1H-pyrrolo-
[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione (8). Method A:
using 3-furanboronic acid (2 equiv) and following the
same procedure as in the preparation of 6 gave 8 (59%)
5.1.12. [1-(2-Trimethylsilanylethoxymethyl)-1H-imida-
zol-2-yl]acetonitrile (14). To a water solution (20 mL)
of KCN (5.46 g, 83.9 mmol) at 0 ꢁC was added com-
pound 13 (2.3 g, 9.32 mmol) in EtOH (40 mL) dropwise.
Once addition was completed, the mixture was stirred at
23 ꢁC for 4 h. The solution was filtered and the precipi-
tate was washed with 95% EtOH (100 mL). The filtrate
was then concentrated to small volume and water added
(20 mL). After extraction of the aqueous layer with
CHCl3 (4 · 50 mL), the combined organic layers were
concentrated to give a dark oil, which was purified by
column chromatography (SiO2) to give 0.813 g (40%) of
compound 14 as a pale oil: 1H NMR (300 MHz, CDCl3)
d 6.95 (s, 2H), 5.26 (s, 2H), 3.89 (s, 2H), 3.45 (br t,
J ¼ 8:4 Hz, 2H), 0.87 (br t, J ¼ 8:4 Hz, 2H), )0.06 (s,
9H); MS (ES) m=z 260 (M+Naþ).
1
as an orange solid: H NMR (300 MHz, acetone-d6) d
8.34 (d, J ¼ 4:5, 1.5 Hz, 1H), 8.16 (m, 1H), 8.01 (s, 1H),
7.53 (m, 2H), 7.11 (dd, J ¼ 8:1, 4.7 Hz, 1H), 6.47 (dd,
J ¼ 2:1, 0.8 Hz, 1H), 4.56 (m, 2H), 3.59 (m, 2H), 2.13
(m, 2H); MS (ES) m=z 338 (M+Hþ).
5.1.9. 3-Furan-2-yl-4-[1-(3-hydroxypropyl)-1H-pyrrolo-
[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione (9). Method B:
using 2-(tributylstannyl)furan (1.5 equiv) and following
the same procedure as in the preparation of 7 gave 9
(64%): 1H NMR (300 MHz, CDCl3) d 8.31 (dd, J ¼ 4:7,
1.3 Hz, 1H), 7.96 (s, 1H), 7.57 (s, 1H), 7.51 (dd, J ¼ 8:1,
1.5 Hz, 1H), 7.39 (d, J ¼ 1:1 Hz, 1H), 7.31 (d,
J ¼ 3:6 Hz, 1H), 7.03 (dd, J ¼ 8:1, 4.7 Hz, 1H), 6.58 (dd,
J ¼ 3:6, 1.7 Hz, 1H), 4.54 (m, 2H), 3.50 (m, 2H), 2.07
(m, 2H); MS (ES) m=z 338 (M+Hþ); FAB-HRMS
(M+Hþ). Calcd 338.1141, found 338.1153.
5.1.13. 2-[1-(2-Trimethylsilanylethoxymethyl)-1H-imida-
zol-2-yl]acetamide (15). To a DMSO solution (3 mL) of
compound 14 (0.813 g, 3.4 mmol) at 0 ꢁC was added
K2CO3 (0.2 g, 1.7 mmol) in one portion followed by
H2O2 (0.5 mL, 5.1 mmol) dropwise. After 5 min, MeOH
(5 mL) was added and the mixture was filtered, con-
centrated, and the DMSO was removed by a nitrogen
stream. The product compound 15 (0.648 g, 74%) was
then obtained by recrystallization from Et2O as pale
5.1.10. Oxo-(1H-pyrrolo[2,3-b]pyridin-3-yl)-acetic acid
ethyl ester (11). To a THF solution (20 mL) of 7-azain-
dole 10 (2.30 g, 19.5 mmol) was added EtMgBr
(21.5 mmol, 1 M solution in THF), and the mixture was
heated to gentle reflux for 1 h, and cooled to 20 ꢁC.
Diethyl oxolate (8.0 mL, 58.5 mmol) was dissolved in
THF (50 mL) and cooled to )40 ꢁC, and the freshly
prepared Grignard reagent was introduced slowly via a
cannula. After the addition was complete, the mixture
was heated to 70 ꢁC for 1.5 h, and cooled to 20 ꢁC. It was
quenched with 5 mL of saturated NaHCO3, and diluted
with water. The aqueous layer was extracted with
EtOAc. The organic layers were combined, dried
(MgSO4), and concentrated. The residue was purified by
flash chromatography on silica gel, eluting gradually
with hexane/EtOAc to give 1 g (23%) of compound 11 as
1
crystals: H NMR (300 MHz, CD3OD) d 7.18 (s, 1H),
6.90 (s, 1H), 5.37 (s, 2H), 3.77 (s, 2H), 3.53 (br t,
J ¼ 7:9 Hz, 2H), 0.89 (br t, J ¼ 8:2 Hz, 2H), )0.02 (s,
9H); MS (ES) m=z 256 (M+Hþ).
5.1.14. 3-[1-[3-(tert-Butyl-dimethylsilanyloxy)propyl]-1H-
pyrrolo[2,3-b]pyridin-3-yl]-4-[1-(2-trimethylsilanylethoxy-
methyl)-1H-imidazol-2-yl]pyrrole-2,5-dione (16). To a
THF solution (0.4 mL) of compound 15 (0.126 g,
0.493 mmol) and compound 12 (0.214 g, 0.548 mmol) at
0 ꢁC was added potassium tert-butoxide (1.1 mL, 1 M
solution in THF, 1.1 mmol) dropwise under nitrogen.
After 15 min the mixture was allowed to warm to
23 ꢁC and stirred for 30 min. The crude product was
then partially concentrated and purified by column
1
a pale yellow solid: H NMR (300 MHz, CDCl3) d 11.8
(s, 1H), 8.75 (dd, J ¼ 7:9, 1.5 Hz, 1H), 8.70 (s, 1H), 8.45
(dd, J ¼ 4:8, 1.5 Hz, 1H), 7.35 (dd, J ¼ 7:9, 4.8 Hz, 1H),
4.44 (q, J ¼ 7:1 Hz, 2H), 1.46 (t, J ¼ 7:1 Hz, 3H); MS
(ES) m=z 219 (M+Hþ).

3178
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
chromatography (SiO2) to give 0.06 g (21%) of com-
pound 16 as a yellow oil: 1H NMR (400 MHz, CDCl3) d
8.51 (s, 1H), 8.31 (m, 2H), 7.32 (m, 1H), 6.91 (dd, J ¼ 8,
4.8 Hz, 1H), 6.57 (d, J ¼ 7:7 Hz, 1H), 5.21 (s, 2H), 4.50
(t, J ¼ 6:8 Hz, 2H), 3.7 (t, J ¼ 5:7 Hz, 2H), 3.44 (br t,
J ¼ 8:2 Hz, 2H), 2.15 (m, J ¼ 6:2 Hz, 2H), 0.97 (s, 9H),
0.79 (br t, J ¼ 8:2 Hz, 2H), 0.12 (s, 6H), )0.08 (s, 9H);
MS (ES) m=z 583 (M+Hþ).
J ¼ 8:1, 4.6 Hz, 1H), 6.47 (dd, J ¼ 8:1, 1.5 Hz, 1H), 6.45
(d, J ¼ 1:8 Hz, 1H), 4.43 (m, 2H), 3.61 (m, 2H), 3.47 (s,
3H), 2.07 (m, 2H), 0.87 (s, 9H), 0.00 (s, 6H); MS (ES)
m=z 466 (M+Hþ).
5.1.17. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-(1-methyl-1H-pyrazol-3-yl)-1H-pyrrole-2,5-dione
(21). TBAF (1.3 mL, 1 M solution in THF, 1.3 mmol)
was added to a solution of compound 19 (0.35 g,
0.75 mmol) in THF (15 mL) at 23 ꢁC dropwise under
nitrogen. After 18 h, the mixture was concentrated and
the product was purified by column chromatography
(SiO2) followed by recrystallization to give 0.22 g (82%)
5.1.15. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-(1H-imidazol-2-yl)-1H-pyrrole-2,5-dione (17) . To
a CH2Cl2 solution (2 mL) of compound 16 (0.048 g,
0.082 mmol) at 20 ꢁC was added TFA (1 mL). After 20 h
toluene (5 mL) was added and the mixture was concen-
trated. The crude product was purified by column
chromatography (SiO2) to give 0.008 g (29%) of com-
pound 17 as a yellow solid: 1H NMR (400 MHz,
CD3OD) d 8.40 (s, 1H), 8.24 (dd, J ¼ 4:8, 1.5 Hz, 1H),
7.28 (m, 2H), 7.12 (dd, J ¼ 8, 1.5 Hz, 1H), 6.98 (dd,
J ¼ 7:9, 4.6 Hz, 1H), 4.48 (t, J ¼ 6:8 Hz, 2H), 3.61 (t,
J ¼ 6:2 Hz, 2H), 2.10 (m, J ¼ 6:4 Hz, 2H); MS (ES) m=z
338 (M+Hþ); FAB-HRMS (M+Hþ). Calcd 338.1253,
found 338.1261.
of compound 21 as
a
yellow solid: 1H NMR
(300 MHzMHz, DMSO-d6) d 11.05 (s, 1H). 8.45 (s, 1H),
8.27 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.78 (d, J ¼ 2:3 Hz, 1H),
7.27 (dd, J ¼ 8:0, 1.5 Hz, 1H), 7.05 (dd, J ¼ 8:1, 4.7 Hz,
1H), 6.67 (d, J ¼ 2:3 Hz, 1H), 4.4 (t, J ¼ 7 Hz, 2H), 3.79
(s, 3H), 3.46 (t, J ¼ 6 Hz, 2H), 1.99 (m, 2H); MS (ES)
m=z 352 (M+Hþ). Anal. Calcd for C18H17N5O3Æ0.1H2O:
C, 61.22; H, 4.91; N, 19.83. Found: C, 61.03; H, 4.75; N,
19.78.
Compound 21 was also prepared by an alternative
route. Method B: using 2-1-methyl-3-tributylstannanyl-
1H-pyrazole (1.5 equiv) and following the same proce-
dure as in the preparation of 7 gave 21 (75%) as a yellow
solid.
5.1.16. 3-{1-[3-(tert-Butyl-dimethyl-silanyloxy)-propyl]-
1H-pyrrolo[2,3-b]pyridin-3-yl}-4-(1-methyl-1H-pyrazol-3-
yl)-pyrrole-2,5-dione 19 and 3-{1-[3-(tert-Butyl-dimethyl-
silanyloxy)-propyl]-1H-pyrrolo[2,3-b]pyridin-3-yl}-4-(2-
methyl-2H-pyrazol-3-yl)-pyrrole-2,5-dione (20). Cesium
carbonate (3.5 g, 10.8 mmol) and iodomethane (0.51 g,
3.6 mmol) were added to a solution of 18 (0.45 g,
3.6 mmol) in DMF (5 mL) at 23 ꢁC under nitrogen. The
mixture was warmed to 70 ꢁC and stirred for 3 h. After
cooling, the mixture was diluted with EtOAc (20 mL),
filtered through Celite, and washed with water
(4 · 10 mL). The organic layer was dried (MgSO4), fil-
tered, and concentrated to give a crude product (0.46 g)
as a white solid. After chromatography, the isolated
product was shown to be a 2:1 mixture of 2-(1-methyl-
1H-pyrazol-3-yl)-acetamide and 2-(2-methyl-2H-pyra-
zol-3-yl)-acetamide.
5.1.18. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-(2-methyl-2H-pyrazol-3-yl)-pyrrole-2,5-dione (22).
TBAF (1.3 mL, 1 M solution in THF, 1.3 mmol) was
added to a solution of 20 (92 mg, 0.20 mmol) in THF
(15 mL) at 23 ꢁC dropwise under nitrogen. After 18 h,
the mixture was concentrated and the crude product was
then recrystallized (CH2Cl2/hexane) to give 64 mg (91%)
1
of compound 22 as a yellow solid: H NMR (300 MHz,
CDCl3) d 8.28 (m, 2H), 7.62 (d, J ¼ 1:9 Hz, 1H), 6.94
(dd, J ¼ 8:5, 5.1 Hz, 1H), 6.59 (dd, J ¼ 8:1, 1.3 Hz, 1H),
6.51 (d, J ¼ 1:7 Hz, 1H), 4.53 (m, 2H), 4.13 (m, 1H),
3.54 (s, 3H), 3.43 (m, 2H), 2.07 (m, 2H); MS (ES) m=z
352 (M+Hþ); FAB-HRMS (M+Hþ). Calcd 352.1410,
found 352.1406.
Potassium tert-butoxide (6.6 mL, 6.6 mmol; 1 M solu-
tion in THF) was added dropwise to a solution of the
above mixture of products and compound 12 (1.36 g,
3.47 mmol) in THF (20 mL) at 0 ꢁC under nitrogen.
After warming to 23 ꢁC, the reaction mixture was stirred
for 2 h and then concentrated. The crude reaction mix-
ture was purified by column chromatography (SiO2) to
give a yellow solid, which was then recrystallized
(EtOAc/hexanes) to give 0.36 g (21%) of compound 19
5.1.19. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-(1-methyl-1H-pyrazol-4-yl)-pyrrole-2,5-dione (23).
Method B: using 1-methyl-4-tributylstannanyl-1H-pyr-
azole (1.5 equiv) and following the same procedure as in
the preparation of 7 gave 23 (31%): 1H NMR (300 MHz,
acetone-d6) d 8.38 (s, 1H), 8.29 (m, 1H), 8.07 (s, 1H),
7.51 (s, 1H), 6.9 (dd, J ¼ 8:1, 4.7 Hz, 1H), 6.67 (d,
J ¼ 2:3 Hz, 1H), 6.42 (s, 1H), 4.56 (m, 2H), 3.51
(m, 5H), 2.05 (m, 2H); MS (ES) m=z 352 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 352.1410, found 352.1424.
1
and 0.1 g (6%) of compound 20. For compound 19: H
NMR (400 MHz, CDCl3) d 8.32 (s, 1H), 8.30 (dd,
J ¼ 4:8, 1.7 Hz, 1H), 7.42 (d, J ¼ 2:2 Hz, 1H), 7.37
(s, 1H), 7. 09 (dd, J ¼ 7:9, 1.5 Hz, 1H), 6.93 (dd, J ¼ 7:9,
4.6 Hz, 1H), 6.73 (d, J ¼ 2:2 Hz, 1H), 4.47
(t, J ¼ 7:0 Hz, 2H), 3.84 (s, 3H), 3.71 (t, J ¼ 5:9 Hz,
2H), 2.15 (m, 2H), 0.92 (s, 9H), 0.07 (s, 6H); MS (ES)
m=z 466 (M+Hþ). For compound 20: 1H NMR
(300 MHz, CDCl3) d 8.72 (s, 1H), 8.30 (s, 1H), 8.25 (dd,
J ¼ 4:7, 1.4 Hz, 1H), 7.56 (d, J ¼ 2:0 Hz, 1H), 6.83 (dd,
5.1.20. 2-{1-[3-(tert-Butyl-dimethyl-silanyloxy)-propyl]-
1H-imidazol-4-yl}-acetamide (25). To a mixture of
compound 24 (115 mg, 0.92 mmol) and Cs2CO3 (450 mg,
1.38 mmol) in DMF (2.0 mL), (3-bromopropoxy)-tert-

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3179
butyldimethylsilane (350 mg, 1.38 mmol) was added.
The resulting mixture was heated to 80 ꢁC for 5.5 h, and
it was then cooled to 20 ꢁC. The mixture was diluted
with EtOAc and filtered through Celite. The filtrate was
washed with water, dried (Na2SO4), and concentrated
under reduced pressure to give 100 mg (36%) of com-
5.1.23. 3-[1-(2-Hydroxyethyl)-1H-imidazol-4-yl]-4-[1-(3-
hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]-1H-pyr-
role-2,5-dione (28). KOt-Bu (1.2 mL, 1.20 mmol, 1 M in
THF) at 0 ꢁC was added to a THF (1.1 mL) solution of
oxolate 12 (234 mg, 0.60 mmol) and imidazole 26
(153 mg, 0.54 mmol). The mixture was stirred at 0 ꢁC for
10 min, and then warmed to 20 ꢁC for 1.5 h. It was
concentrated and the resulting residue was chromato-
graphed on silica gel, eluting with EtOAc/hexane to give
161 mg of silylated product as a red chip: 1H NMR
(400 MHz, DMSO-d6) d 10.93 (s, 1H), 8.29 (s, 1H), 8.24
(d, J ¼ 4:5 Hz, 1H), 7.91 (s, 1H), 7.61 (s, 1H), 7.59
(d, J ¼ 8:1 Hz, 1H), 7.00 (m, 1H), 4.39 (t, J ¼ 7:1 Hz,
2H), 4.13 (t, J ¼ 4:3 Hz, 2H), 3.83 (t, J ¼ 4:5 Hz, 2H),
3.64 (t, J ¼ 5:8 Hz, 2H), 2.03 (t, J ¼ 6:4 Hz, 2H), 0.87
(s, 9H), 0.81 (s, 9H), 0.01 (s, 6H), )0.01 (s, 6H); MS (ES)
m=z 610 (M+Hþ).
pound 25 as
a
sticky yellow oil: 1H NMR
(300 MHz, CDCl3) d 7.38 (s, 1H), 6.70 (s, 1H), 3.98 (t,
J ¼ 6:9 Hz, 2H), 3.52 (t, J ¼ 5:6 Hz, 2H), 3.46 (s, 2H),
1.87 (m, 2H), 0.86 (s, 9H), 0.00 (s, 6H); MS (ES) m=z 298
(M+Hþ).
5.1.21. 2-{1-[2-(tert-Butyl-dimethyl-silanyloxy)-ethyl]-1H-
imidazol-4-yl}-acetamide (26). DMF (1.0 mL) solution of
(2-bromoethoxy)-tert-butyldimethylsilane (301 mg, 1.26
mmol) was added to a mixture of compound 24 (105 mg,
0.84 mmol) and Cs2CO3 (411 mg, 1.26 mmol) in DMF
(2.0 mL). The mixture was heated to reflux for 5 h, and it
was then cooled to 20 ꢁC. The mixture was diluted with
EtOAc and filtered through Celite. The filtrate was
washed with water, dried (Na2SO4), and concentrated
under reduced pressure to give 149 mg (63%) of
TBAF (0.56 mL, 0.56 mmol, 1 M in THF) at 20 ꢁC was
added to a THF (2.0 mL) solution of the above silylated
product. The mixture was stirred at 20 ꢁC for 2 h, and
then concentrated under reduced pressure. The residue
was purified by column chromatography eluting with
MeOH/CH2Cl2 to give 90 mg (44%) of compound 28 as
an orange red solid after recrystallized from MeOH/
EtOAc: 1H NMR (400 MHz, DMSO-d6) d 10.96 (s, 1H),
8.35 (s, 1H), 8.26 (d, J ¼ 4:3 Hz, 1H), 7.90 (s, 1H), 7.68
(s, 1H), 7.61 (d, J ¼ 8:0 Hz, 1H), 7.04 (dd, J ¼ 7:9,
4.6 Hz, 1H), 5.00 (t, J ¼ 5:1 Hz, 1H), 4.66 (d,
J ¼ 4:6 Hz, 1H), 4.40 (t, J ¼ 6:9 Hz, 2H), 4.08 (t,
J ¼ 5:1 Hz, 2H), 3.68 (q, J ¼ 4:6 Hz, 2H), 3.48 (d,
J ¼ 4:9 Hz, 2H), 1.99 (m, 2H); MS (ES) m=z 382
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 382.1516, found
382.1517.
1
compound 26 as an oil: H NMR (300 MHz, CDCl3)
d 7.51 (s, 1H), 6.86 (s, 1H), 4.03 (t, J ¼ 4:7 Hz, 2H), 3.86
(t, J ¼ 4:8 Hz, 2H), 3.54 (s, 2H), 0.89 (s, 9H), 0.00 (s,
6H); MS (ES) m=z 284 (M+Hþ).
5.1.22. 3-[1-(3-Hydroxypropyl)-1H-imidazol-4-yl]-4-[1-(3-
hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]-1H-pyr-
role-2,5-dione (27). KOt-Bu (0.24 mL, 0.24 mmol, 1 M
solution in THF) at 0 ꢁC was added to a THF (0.25 mL)
solution of oxolate compound 12 (48 mg, 0.12 mmol)
and imidazole 25 (33 mg, 0.11 mmol). After the mixture
was stirred at 0 ꢁC for 15 min, it was warmed to 20 ꢁC for
1 h. After the solvent was removed under reduced
pressure, the residue was chromatographed on silica gel,
eluting with Hex/EtOAc to give 32 mg of silylated
product as an orange red oil: 1H NMR (300 MHz,
CDCl3) d 8.30 (dd, J ¼ 4:7, 1.3 Hz, 1H), 8.25 (s, 1H),
7.74 (s, 1H), 7.54 (dd, J ¼ 8:0, 1.4 Hz, 1H), 7.44 (s, 1H),
6.96 (dd, J ¼ 8:0, 4.7 Hz, 1H), 4.47 (t, J ¼ 7:0 Hz, 2H),
4.12 (t, J ¼ 6:8 Hz, 2H), 3.72 (t, J ¼ 5:8 Hz, 2H), 3.60
(t, J ¼ 5:6 Hz, 2H), 2.14 (m, 2H), 1.98 (m, 2H), 0.92 (s,
9H), 0.91 (s, 9H), 0.07 (s, 6H), 0.00 (s, 6H); MS (ES) m=z
624 (M+Hþ).
5.1.24. 3-(3,5-Dimethyl-isoxazol-4-yl)-4-[1-(3-hydroxy-
propyl)-1H-pyrrolo-[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione
(29). Method A: using 3,5-dimethylisoxazole-4-boronic
acid (2 equiv) and following the same procedure as in the
1
preparation of 6 gave 29 (10%): H NMR (300 MHz,
acetone-d6) d 8.33 (dd, J ¼ 4:6, 1.5 Hz, 1H), 8.24 (s, 1H),
7.24 (dd, J ¼ 8:1, 1.7 Hz, 1H), 7.03 (dd, J ¼ 8:1, 4.8 Hz,
1H), 4.56 (m, 2H), 3.53 (m, 2H), 2.11 (m, 2H), 2.08 (s,
3H), 2.02 (s, 3H); MS (ES) m=z 367 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 367.1406, found 367.1398.
5.1.25. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-thiazol-2-yl-pyrrole-2,5-dione (30). Method B:
using 2-tributylstannylthiazole (1.5 equiv) and following
the same procedure as in the preparation of 7 gave 30
TBAF (0.4 mL, 0.40 mmol, 1 M solution in THF) at
20 ꢁC was added to a THF (1.0 mL) solution of the
above silylated product. After the mixture was stirred
for 30 min, the solvent was removed under reduced
pressure. The residue was crystallized from MeOH/
EtOAc to give 21 mg (48%) of compound 27 as an or-
1
(9%): H NMR (300 MHz, acetone-d6) d 8.89 (s, 1H),
8.33 (d, J ¼ 2:1 Hz, 1H), 7.94 (m, 2H), 7.71
(d, J ¼ 2:3 Hz, 1H), 7.1 (dd, J ¼ 8:1, 4.7 Hz, 1H), 4.58
(m, 2H), 3.99 (m, 1H), 3.61 (m, 2H), 2.14 (m, 2H); MS
(ES) m=z 355 (M+Hþ); FAB-HRMS (M+Hþ). Calcd
355.0865, found 355.0865.
1
ange solid: H NMR (300 MHz, DMSO-d6) d 10.9 (s,
1H), 8.33 (s, 1H), 8.25 (d, J ¼ 4:4 Hz, 1H), 7.84 (s, 1H),
7.69 (s, 1H), 7.55 (d, J ¼ 7:5 Hz, 1H), 7.02 (m, 1H), 4.65
(s, 1H), 4.64 (s, 1H), 4.39 (t, J ¼ 6:7 Hz, 2H), 4.10
(t, J ¼ 6:6 Hz, 2H), 3.47 (d, J ¼ 5:8 Hz, 2H), 3.38
(d, J ¼ 5:5 Hz, 2H), 1.98 (t, J ¼ 6:2 Hz, 2H), 1.87
(t, J ¼ 6:1 Hz, 2H); MS (ES) m=z 396 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 396.1672, found 396.1668.
5.1.26. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-pyrimidin-5-yl-pyrrole-2,5-dione (31). Method A:
using pyrimidine-5-boronic acid (2 equiv) and following

3180
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
the same procedure as in the preparation of 6 gave 31
1
(29%) as an orange solid: H NMR (300 MHz, acetone-
d6) d 9.11 (s, 1H), 8.86 (s, 2H), 8.29 (m, 2H), 6.95 (m,
2H), 4.58 (m, 2H), 3.55 (m, 2H), 2.11 (m, 2H); MS (ES)
m=z 348 (M)Hþ); FAB-HRMS (M+Hþ). Calcd
350.1253, found 350.1265.
1.5 Hz, 1H), 7.65 (dd, J ¼ 8:1, 7.4 Hz, 1H), 7.45 (dd,
J ¼ 8:5, 4.1 Hz, 1H), 6.52 (m, 2H), 4.44 (m, 2H), 3.46
(m, 2H), 1.99 (m, 2H); MS (ES) m=z 399 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 399.1457, found 399.1456.
5.1.32. 3-(2-Benzofuranyl)-4-[1-(3-hydroxypropyl)-1H-
pyrrolo[2,3-b]pyridin-3-yl]-1H-pyrrole-2,5-dione
(37).
5.1.27. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-pyrimidin-2-yl-pyrrole-2,5-dione (32). Method B:
using 2-tributylstannylpyrimidine (1.5 equiv) and fol-
lowing the same procedure as in the preparation of 7
Method A: using benzofuran-2-boronic acid (2 equiv)
and following the same procedure as in the preparation
of 6 gave 37 (57%) as an orange solid: 1H NMR
(300 MHz, DMSO-d6) d 11.31 (s, 1H), 8.32 (dd, J ¼ 4:7,
1.5 Hz, 1H), 8.27 (s, 1H), 7.77 (m, 1H), 7.65 (s, 1H), 7.57
(dd, J ¼ 7:9, 1.3 Hz, 1H), 7.29 (m, 3H), 7.02 (dd,
J ¼ 7:9, 4.5 Hz, 1H), 4.47 (t, J ¼ 6:9 Hz, 2H), 3.5 (t,
J ¼ 6:0 Hz, 2H), 2.05 (m, 2H); MS (ES) m=z 388
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 388.1297, found
388.1311.
1
gave 32 (70%): H NMR (300 MHz, CD3OD) d 8.85 (d,
J ¼ 4:9 Hz, 2H), 8.32 (s, 1H), 8.21 (d, J ¼ 4:0 Hz, 1H),
7.49 (m, 1H), 6.87 (dd, J ¼ 8:1, 4.8 Hz, 1H), 6.68 (d,
J ¼ 7:9 Hz, 1H), 4.47 (m, 2H), 3.56 (m, 2H), 2.08 (m,
2H); MS (ES) m=z 350 (M+Hþ); FAB-HRMS (M+Hþ).
Calcd 350.1253, found 350.1262.
5.1.28. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-pyrazin-2-yl-pyrrole-2,5-dione (33). Method B:
using 2-tributylstannylpyrazine (1.5 equiv) and follow-
ing the same procedure as in the preparation of 7 gave
5.1.33. 3-Benzo[b]thiophen-2-yl-4-[1-(3-hydroxypropyl)-
1H-pyrrolo-[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione
(38).
Method A: using benzothiophene-2-boronic acid
(2 equiv) and following the same procedure as in the
1
1
33 (46%): H NMR (300 MHz, acetone-d6) d 9.01 (s,
preparation of 6 gave 38 (40%): H NMR (300 MHz,
1H), 8.58 (m, 1H), 8.42 (s, 1H), 8.27 (dd, J ¼ 4:4, 1.8 Hz,
1H), 6.94 (m, 2H), 4.54 (m, 2H), 3.57 (m, 2H), 2.11 (m,
2H); MS (ES) m=z 350 (M+Hþ); FAB-HRMS (M+Hþ).
Calcd 350.1253, found 350.1258.
acetone-d6) d 8.52 (dd, J ¼ 8:1, 1.5 Hz, 1H), 8.39 (m,
2H), 8.32 (dd, J ¼ 4:7, 1.5 Hz, 1H), 8.18 (s, 1H), 7.83 (m,
1H), 7.39 (m, 2H), 7.27 (dd, J ¼ 8:1, 4.7 Hz, 1H), 6.97
(dd, J ¼ 8:1, 4.7 Hz, 1H), 4.59 (m, 2H), 3.58 (m, 2H),
2.14 (m, 2H); MS (ES) m=z 404 (M+Hþ); FAB-HRMS
(M+Hþ). Calcd 404.1069, found 404.1056.
5.1.29. 3-(2,4-Dimethoxy-pyrimidin-5-yl)-4-[1-(3-hydr-
oxypropyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]-pyrrole-2,5-di-
one (34). Method B: using 2,4-dimethoxy-5-tributyl-
stannanyl-pyrimidine 43 (1.5 equiv) and following the
same procedure as in the preparation of 7 gave 34 (36%):
1H NMR (300 MHz, CD3OD) d 8.48 (s, 1H), 8.26 (dd,
J ¼ 4:7, 1.5 Hz, 1H), 8.13 (s, 1H), 7.20 (dd, J ¼ 7:9,
1.3 Hz, 1H), 6.96 (dd, J ¼ 8:1, 4.7 Hz, 1H), 4.49 (m, 2H),
4.01 (s, 3H), 3.56 (m, 2H), 3.42 (s, 3H), 2.10 (m, 2H);
MS (ES) m=z 410 (M+Hþ).
5.1.34. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-(4,5,6,7-tetrahydro-pyrazolo[1,5-a]pyridin-2-yl)-
pyrrole-2,5-dione (39). Method B: using 2-tributylst-
annanyl-4,5,6,7-tetrahydro-pyrazolo[1,5-a]pyridine (1.5
equiv) and following the same procedure as in the
1
preparation of 7 gave 39 (71%): H NMR (300 MHz,
CDCl3) d 8.33 (s, 1H), 8.27 (dd, J ¼ 4:7, 1.3 Hz, 1H),
7.53 (s, 1H), 7.19 (dd, J ¼ 8:1, 1.5 Hz, 1H), 6.98 (dd,
J ¼ 8:1, 4.7 Hz, 1H), 6.51 (s, 1H), 4.50 (m, 2H), 4.00
(m, 2H), 3.48 (m, 2H), 2.85 (m, 2H), 2.06 (m, 4H), 1.89
(m, 2H); MS (ES) m=z 392 (M+Hþ); FAB-HRMS
(M+Hþ). Calcd 392.1723, found 392.1710.
5.1.30. 3-(5,6-Dihydro-[1,4]dioxin-2-yl)-4-[1-(3-hydroxy-
propyl)-1H-pyrrolo-[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione
(35). Method B: using tributyl-(5,6-dihydro-[1,4]dioxin-
2-yl)-stannane (1.5 equiv) and following the same pro-
1
cedure as in the preparation of 7 gave 35 (49%): H
5.1.35. 3-(4-Dibenzofuranyl)-4-[1-(3-hydroxypropyl)-1H-
pyrrolo[2,3-b]pyridin-3-yl]-1H-pyrrole-2,5-dione
NMR (300 MHz, CDCl3) d 8.32 (dd, J ¼ 4:7, 1.3 Hz,
1H), 7.93 (dd, J ¼ 7:9, 1.3 Hz, 1H), 7.76 (s, 1H), 7.39 (s,
1H), 7.16 (dd, J ¼ 7:9, 4.9 Hz, 1H), 4.49 (m, 2H), 4.14
(m, 2H), 3.89 (m, 2H), 3.46 (m, 2H), 2.05 (m, 2H); MS
(ES) m=z 356 (M+Hþ); FAB-HRMS (M+Hþ). Calcd
356.1247, found 356.1246.
(40).
Method A: using 4-dibenzofuranboronic acid (2 equiv)
and following the same procedure as in the preparation
of 6 gave 40 (34%) as a yellow solid: 1H NMR
(300 MHz, acetone-d6) d 8.28 (s, 1H), 8.18 (dd, J ¼ 7:7,
1.3 Hz, 1H), 8.02 (ddd, J ¼ 8:3, 4.5, 1.5 Hz, 2H), 7.74
(dd, J ¼ 7:5, 1.1 Hz, 1H), 7.51 (m, 1H), 7.32 (m, 2H),
7.17 (m, 1H), 6.64 (dd, J ¼ 7:9, 1.5 Hz, 1H), 6.53 (dd,
J ¼ 8:1, 4.7 Hz, 1H), 4.50 (t, J ¼ 6:6 Hz, 2H), 3.49 (m,
2H), 2.04 (m, 2H); MS (ES) m=z 438 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 438.1454, found 438.1455.
5.1.31. 3-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-quinolin-8-yl-pyrrole-2,5-dione (36). Method A:
using 8-quinolineboronic acid (2 equiv) and following
the same procedure as in the preparation of 6 gave 36
1
(50%) as an orange solid: H NMR (300 MHz, acetone-
d6) d 8.69 (dd, J ¼ 4:0, 1.7 Hz, 1H), 8.37 (dd, J ¼ 8:5,
5.1.36. 3-Hydroxy-4-[1-(3-hydroxypropyl)-1H-pyrrolo-
[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione (41). TBAF (0.29
1.7 Hz, 1H), 8.13 (s, 1H), 8.08 (m, 2H), 7.78 (dd, J ¼ 7:0,

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3181
mL, 0.29 mmol, 1 M in THF) at 20 ꢁC was added to a
THF (10 mL) solution of 4 (0.1 g, 0.19 mmol). The
mixture was stirred at 20 ꢁC for 18 h, and then concen-
trated under reduced pressure. The residue was purified
by column chromatography eluting with MeOH/CH2Cl2
to give 0.05 g (90%) of compound 41 as a yellow solid:
1H NMR (400 MHz, CD3OD) d 9.27 (dd, J ¼ 8:1,
1.1 Hz, 1H), 8.46 (dd, J ¼ 5:7, 0.8 Hz, 1H), 8.26 (s, 1H),
7.57 (dd, J ¼ 8:1, 5.9 Hz, 1H), 4.57 (m, 2H), 3.56 (m,
2H), 2.12 (m, 2H); MS (ES) m=z 288 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 288.0984, found 288.0984.
of 45 (0.412 g, 1.3 mmol) in DMF (10 mL) at 0 ꢁC was
added (CO2Et)2 (0.38 g, 2.6 mmol), and then KOt-Bu
(2.6 mL, 2.6 mmol, 1 M in THF) dropwise. The resulting
red solution was stirred for 15 min and concentrated.
The product was purified by column chromatography to
1
give 0.393 g (81%) of 46 as a yellow solid: H NMR
(300 MHz, acetone-d6) d 9.11 (s, 1H), 8.75 (m, 1H), 8.10
(m, 1H), 7.97 (s, 1H), 7.12 (m, 5H), 6.88 (m, 1H), 4.15
(m, 2H), 2.45 (m, 2H), 2.04 (m, 2H); MS (ES) m=z 374
(M+Hþ).
5.1.41. 3-Chloro-4-[1-(3-phenyl-propyl)-1H-pyrrolo[2,3-
b]pyridin-3-yl]-pyrrole-2,5-dione (47). To a solution of
46 (0.393 g, 1.05 mmol) in DMF and CH2Cl2 (1:1) was
added (COCl)2 (0.4 g, 3.15 mmol) in one portion at 0 ꢁC.
The reaction was followed by TLC until starting mate-
rial disappeared (ꢁ1 h), then NaHCO3 solution was
added. The mixture was diluted with EtOAc and washed
with water, dried, concentrated, and chromatographed
on silica gel to give 0.372 g (89%) of 47 as a yellow solid:
1H NMR (300 MHz, CDCl3) d 8.46 (dd, J ¼ 8:1, 1.5 Hz,
1H), 8.43 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.75 (s, 1H), 7.24 (m,
7H), 4.41 (m, 2H), 2.70 (m, 2H), 2.30 (m, 2H); MS (ES)
m=z 366 (M+Hþ).
5.1.37. 4-[1-(3-Hydroxypropyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-2,5-dioxo-2,5-dihydro-1H-pyrrole-3-carbonitrile (42).
To a solution of 5 (54 mg, 0.1 mmol) in DMF (3 mL)
under N2 was added Pd(Ph3P)4 (12 mg, 0.01 mmol), LiCl
(13 mg, 0.3 mmol), and Zn(CN)2 (23 mg, 0.2 mmol). The
reaction mixture was heated to 120 ꢁC and stirred for 3 h.
After cooling, the reaction mixture was diluted with
EtOAc, filtered through Celite, washed with water (3·),
dried with MgSO4, filtered, and concentrated to give
6 mg (11%) of silylated product as a yellow solid. This
solid was dissolved in THF (2 mL), TBAF (17 lL,
0.01 mmol) was added and the mixture was stirred for 8 h
then concentrated and purified by column chromatog-
1
raphy to give 3 mg (92%) of 42 as a yellow solid: H
NMR (300 MHz, acetone-d6) d 8.72 (s, 1H), 8.65 (dd,
J ¼ 8:1, 1.5 Hz, 1H), 8.37 (dd, J ¼ 4:8, 1.5 Hz, 1H), 7.29
(dd, J ¼ 8:1, 4.6 Hz, 1H), 4.52 (m, 2H), 3.47 (m, 2H),
2.03 (m, 2H); MS (ES) m=z 297 (M+Hþ); FAB-HRMS
(M+Hþ). Calcd 297.0988, found 297.0978.
5.1.42. 3-(2,4-Dimethoxy-pyrimidin-5-yl)-4-[1-(3-phenyl-
propyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione
(48). To a solution of 47 (30 mg, 0.082 mmol) and
Pd(tBu3P)2 (4 mg, 0.008 mmol) in THF (1 mL) was ad-
ded the organo-tin 43 (53 mg, 0.123 mmol) at 23 ꢁC un-
der nitrogen. The reaction mixture was refluxed for 18 h.
Upon cooling, the mixture was diluted with EtOAc
(10 mL) and washed with H2O, KF, brine, and dried.
After concentration, the crude product was purified by
column chromatography (SiO2) to give 6 mg (16%) of
5.1.38. 3-Butyl-4-[1-(3-hydroxypropyl)-1H-pyrrolo[2,3-
b]pyridin-3-yl]-pyrrole-2,5-dione (44). Method B: using
2,4-dimethoxy-5-tributylstannanyl-pyrimidine 43 (1.5
equiv) and following the same procedure as in the
preparation of 7 gave 34 (36%, see above) and 44 (21%):
1H NMR (300 MHz, CDCl3) d 8.35 (d, J ¼ 4:6 Hz, 1H),
8.06 (d, J ¼ 8:1, 1H), 7.65 (s, 1H), 7.21 (dd, J ¼ 8:1,
4.8 Hz, 1H), 4.51 (m, 2H), 3.47 (m, 2H), 2.64 (m, 2H),
2.04 (m, 2H), 1.62 (m, 2H), 1.37 (m, 2H), 0.88 (t,
J ¼ 7:3 Hz, 3H); MS (ES) m=z 328 (M+Hþ); FAB-
HRMS (M+Hþ). Calcd 328.1661, found 328.1675.
1
compound 48: H NMR (300 MHz, CDCl3) d 8.50 (s,
1H), 8.31 (dd, J ¼ 4:7, 1.5 Hz, 1H), 8.09 (s, 1H), 7.59 (s,
1H), 7.31 (m, 2H), 7.20 (m, 4H), 7.05 (dd, J ¼ 7:9,
1.3 Hz, 1H), 6.90 (dd, J ¼ 7:9, 4.7 Hz, 1H), 4.41 (m, 2H),
4.04 (s, 3H), 3.43 (s, 3H), 2.69 (m, 2H), 2.29 (m, 2H);
MS (ES) m=z 470 (M+Hþ); FAB-HRMS (M+Hþ).
Calcd 470.1829, found 470.1838.
5.1.43. 3-[1-(3-Phenyl-propyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-4-pyrimidin-5-yl-pyrrole-2,5-dione (49). Method A:
using pyrimidine-5-boronic acid (2 equiv) and following
the same procedure as in the preparation of 48 gave 49
5.1.39. 2-[1-(3-Phenyl-propyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-acetamide (45). To a solution of the amide 2
(0.5 g, 2.7 mmol) in DMF (10 mL) was added Cs2CO3
(2.65 g, 8.1 mmol) and 3-phenyl-propyl bromide (0.8 g,
4.05 mmol). The reaction was heated at 70 ꢁC for 2 h.
After cooling down, the solution was diluted with
EtOAc and washed with H2O. The organic layer was
dried (MgSO4), concentrated, and chromatographed on
silica to give 0.412 g (49%) of compound 45 as a white
1
(36%) as a yellow solid: H NMR (300 MHz, CDCl3) d
9.20 (s, 1H), 8.89 (s, 2H), 8.33 (dd, J ¼ 4:7, 1.5 Hz,
1H), 8.16 (s, 1H), 7.23 (m, 6H), 6.90 (dd, J ¼ 7:9, 4.7 Hz,
1H), 6.74 (dd, J ¼ 8:1, 1.3 Hz, 1H), 4.42 (m, 2H),
2.73 (m, 2H), 2.35 (m, 2H); MS (ES) m=z 410
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 410.1617, found
410.1620.
1
solid: H NMR (300 MHz, CDCl3) d 8.36 (dd, J ¼ 4:7,
1.5 Hz, 1H), 7.88 (dd, J ¼ 7:9, 1.5 Hz, 1H), 7.22 (m, 6H),
7.10 (m, 1H), 4.32 (m, 2H), 3.70 (s, 2H), 2.66 (m, 2H);
MS (ES) m=z 316 (M+Hþ).
5.1.44. 3-Chloro-1-methyl-4-(1H-pyrrolo[2,3-b]pyridin-3-
yl)-pyrrole-2,5-dione (50).
A solution of EtMgBr
(26.6 mL, 79.8 mmol, 3 M in Et2O) was added dropwise
under argon to a well-stirred solution of 7-azaindole
5.1.40. 3-Hydroxy-4-[1-(3-phenyl-propyl)-1H-pyrrolo-
[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione (46). To a solution

3182
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
(9 g, 76 mmol) in dry toluene (270 mL) at room tem-
perature. After 1 h, a solution of the 2,3-dichloro-N-
methylmaleimide (4.5 g, 38 mmol) in toluene (240 mL)
was slowly added. After 15 min, anhydrous CH2Cl2
(300 mL) was added and the reaction mixture was
heated at 50 ꢁC for 24 h. Hydrolysis was performed by a
saturated solution of NH4Cl till pH 7. After extraction
with EtOAc (2 · 400 mL), the combined organic layers
were dried over MgSO4, filtered, and the solvent was
removed under reduced pressure. Compound 50 was
precipitated from methanol, filtered, washed with
methanol, and dried under vacuum. Compound 50 was
obtained as an orange solid (2.12 g, 21%): 1H NMR
(300 MHz, DMSO-d6) d 12.71 (s, 1H), 8.34 (m, 2H), 8.21
(s, 2H), 7.21 (m, 1H), 2.98 (s, 3H); MS (ES) m=z 262
(M+Hþ).
5.1.48. 3-(1-Benzyl-1H-pyrrolo[2,3-b]pyridin-3-yl)-4-(2,4-
dimethoxy-pyrimidin-5-yl)-1-methyl-pyrrole-2,5-dione (54).
Compound 52 (51 mg, 0.145 mmol), Pd2(dba)3 (14.5 mg,
0.014 mmol), organo-tin 43 (93.6 mg, 0.218 mmol), and
P(tBu)3 (14.3 mg, 0.028 mmol) was mixed in anhydrous
THF and DMF (10:1 by volume). The mixture was sealed
in a tube and microwaved for 350 s at 200 ꢁC. The reaction
mixture was concentrated and the product was purified by
1
column chromatography to give 47mg (71%) of 54: H
NMR (300MHz, CDCl3) d 8.49 (s, 1H), 8.32 (dd, J ¼ 4:7,
1.5 Hz, 1H), 8.02 (s, 1H), 7.32–7.29 (m, 5H), 7.09 (dd,
J ¼ 8:0, 1.5 Hz, 1H), 6.93–6.89 (m, 1H), 5.55 (s, 2H), 4.04
(s, 3H), 3.34 (s, 3H), 3.14 (s, 3H); MS (ES) m=z 456
(M+Hþ).
5.1.49. 3-(2,4-Dimethoxy-pyrimidin-5-yl)-4-(1-phenethyl-
1H-pyrrolo[2,3-b]pyridin-3-yl)-pyrrole-2,5-dione (55). To
a solution of 53 (27 mg, 0.058 mmol) in EtOH (2 mL)
was added KOH aqueous solution (1 mL, 10 N) and the
reaction was stirred for 2 h. Water (5 mL) was added and
the mixture was acidified with 10% citric acid. After
extraction with CH2Cl2 (three times), the organic layers
were dried and concentrated to give the crude anhydride
(23 mg). To this crude anhydride in anhydrous DMF
(1.5 mL) was added HMDS (93.6 mg, 0.58 mmol) in
MeOH (0.8 mL). The reaction mixture was heated at
80 ꢁC for 2 h and then cooled down slowly. The solvent
was removed and the residue was purified by column
chromatography to give 9.2 mg (35%) of compound 55:
1H NMR (300 MHz, CDCl3) d 8.52 (s, 1H), 8.35 (dd,
J ¼ 4:7, 1.5 Hz, 1H), 7.98 (s, 1H), 7.28–7.23 (m, 3H),
7.14–7.12 (m, 2H), 6.99–6.95 (m, 2H), 4.64
(t, J ¼ 7:2 Hz, 2H), 4.05 (s, 3H), 3.41 (s, 3H), 3.24
(t, J ¼ 7:2 Hz, 2H); MS (ES) m=z 456 (M+Hþ).
5.1.45. 3-Chloro-1-methyl-4-(1-phenethyl-1H-pyrrolo[2,3-
b]pyridin-3-yl)-pyrrole-2,5-dione (51). To a solution of 50
(250 mg, 0.96 mmol) in anhydrous DMF (15 mL) was
added NaH (5 equiv, 60% dispersion in oil) under N2.
After stirring 10 min, (2-bromo-ethyl)-benzene (355 mg,
1.92 mmol) was added and the reaction mixture was
heated at 70 ꢁC for 90 min. Water (15 mL) was added at
20 ꢁC followed by EtOAc (50 mL). The aqueous layer
was acidified with 1 N HCl, extracted with EtOAc (three
times), and organic layers were combined, dried, and
concentrated. The product was purified by column
1
chromatography to give 37 mg (11%) of 51: H NMR
(300 MHz, CDCl3) d 8.51 (dd, J ¼ 8:1, 1.5 Hz, 1H), 8.43
(dd, J ¼ 4:7, 1.5 Hz, 1H), 7.90 (s, 1H), 7.26–7.20 (m,
3H), 7.14–7.06 (m, 3H), 4.62 (t, J ¼ 7:2 Hz, 2H), 3.22 (t,
J ¼ 7:2 Hz, 2H), 3.14 (s, 3H); MS (ES) m=z 366
(M+Hþ).
5.1.50. 3-(1-Benzyl-1H-pyrrolo[2,3-b]pyridin-3-yl)-4-(2,4-
dimethoxy-pyrimidin-5-yl)-pyrrole-2,5-dione (56). Using
compound 54 and following the same procedure as in
the preparation of 55 gave 56 (26%): 1H NMR
(300 MHz, CDCl3) d 8.50 (s, 1H), 8.33 (dd, J ¼ 4:7,
1.5 Hz, 1H), 8.02 (s, 1H), 7.46 (s, 1H), 7.33–7.29 (m,
5H), 7.09 (dd, J ¼ 8:0, 1.5 Hz, 1H), 6.94–6.90 (m, 1H),
5.56 (s, 2H), 4.04 (s, 3H), 3.34 (s, 3H); MS (ES) m=z 442
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 442.1516, found
442.1532.
5.1.46.
3-(1-Benzyl-1H-pyrrolo[2,3-b]pyridin-3-yl)-4-
chloro-1-methyl-pyrrole-2,5-dione (52). Using benzyl
bromide (3 equiv) and following the same procedure as
in the preparation of 51 gave 52 (76%) as a yellow solid:
1H NMR (300 MHz, CDCl3) d 8.50 (dd, J ¼ 8:1, 1.5 Hz,
1H), 8.44 (dd, J ¼ 4:7, 1.5 Hz, 1H), 8.14 (s, 1H), 7.32–
7.28 (m, 5H), 7.26–7.21 (m, 1H), 5.58 (s, 2H), 3.14 (s,
3H); MS (ES) m=z 352 (M+Hþ).
5.1.47. 3-(2,4-Dimethoxy-pyrimidin-5-yl)-1-methyl-4-(1-
phenethyl-1H-pyrrolo[2,3-b]pyridin-3-yl)-pyrrole-2,5-dione
(53). Compound 51 (38 mg, 0.1 mmol), Pd2(dba)3
(10.4 mg, 0.01 mmol), organo-tin 43 (64 mg, 0.15 mmol),
and P(tBu)3 (10.2 mg, 0.02 mmol) were mixed in anhy-
drous THF and DMF (10:1 by volume). The mixture
was sealed in a tube and microwaved for 350 s at 200 ꢁC.
The reaction mixture was concentrated and the product
was purified by column chromatography to give 27 mg
(55%) of 53: 1H NMR (300 MHz, CDCl3) d 8.46 (s, 1H),
8.31 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.91 (s, 1H), 7.31–7.23
(m, 3H), 7.18–7.15 (m, 2H), 7.01 (dd, J ¼ 8:0, 1.5 Hz,
1H), 6.91–6.87 (m, 1H), 4.62 (t, J ¼ 7:3 Hz, 2H), 4.07 (s,
3H), 3.44 (s, 3H), 3.24 (t, J ¼ 7:1 Hz, 2H); MS (ES) m=z
470 (M+Hþ).
5.1.51. 2-[1-(3-Phenoxy-propyl)-1H-pyrrolo[2,3-b]pyri-
din-3-yl]-acetamide (57). Using 3-phenoxyl-propyl bro-
mide (1.5 equiv) and following the same procedure as in
the preparation of 45 gave 57 (27%): 1H NMR
(300 MHz, CDCl3) d 8.33 (dd, J ¼ 4:6, 1.3 Hz, 1H), 7.89
(dd, J ¼ 7:9, 1.3 Hz, 1H), 7.26 (m, 2H), 7.16 (s, 1H), 7.08
(dd, J ¼ 7:9, 4.6 Hz, 1H), 6.94 (m, 1H), 6.86 (m, 2H),
4.49 (m, 2H), 3.92 (m, 2H), 3.64 (s, 2H), 2.36 (m, 2H);
MS (ES) m=z 310 (M+Hþ).
5.1.52. 3-Hydroxy-4-[1-(3-phenoxy-propyl)-1H-pyrrolo-
[2,3-b]pyridin-3-yl]-pyrrole-2,5-dione (58). Using com-
pound 57 and following the same procedure as in the

D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
3183
1
preparation of 46 gave 58 (53%) as a yellow solid: H
NMR (300 MHz, CDCl3) d 8.50 (m, 1H), 8.41 (m, 1H),
8.17 (s, 1H), 7.25 (m, 4H), 7.21 (m, 1H), 6.86 (m, 2H),
4.62 (m, 2H), 3.98 (m, 2H), 2.44 (m, 2H); MS (ES) m=z
364 (M+Hþ).
following the same procedure as in the preparation of 48
1
gave 64 (43%): H NMR (300 MHz, CDCl3) d 8.46 (s,
1H), 8.29 (dd, J ¼ 4:7, 1.5 Hz, 1H), 8.07 (s, 1H), 7.07
(dd, J ¼ 7:9, 1.5 Hz, 1H), 6.92 (dd, J ¼ 7:9, 4.7 Hz, 1H),
4.50 (s, 2H), 4.04 (s, 3H), 3.48 (s, 3H), 2.36 (m, 4H); MS
(ES) m=z 419 (M+Hþ).
5.1.53. 3-Chloro-4-[1-(3-phenoxy-propyl)-1H-pyrrolo[2,3-
b]pyridin-3-yl]-pyrrole-2,5-dione (59). Using compound
58 and following the same procedure as in the prepa-
5.1.59. 4-[3-(2,5-Dioxo-4-pyrazin-2-yl-2,5-dihydro-1H-
pyrrol-3-yl)-pyrrolo[2,3-b]pyridin-1-yl]-butyronitrile (65).
Method B: using 2-tributylstannylpyrazine (1.5 equiv)
and following the same procedure as in the preparation
of 64 gave 65 (68%): 1H NMR (300 MHz, CDCl3) d 9.04
(s, 1H), 8.57 (s, 2H), 8.30 (m, 2H), 7.82 (s, 1H), 6.89 (dd,
J ¼ 7:9, 4.4 Hz, 1H), 6.75 (d, J ¼ 7:9 Hz, 1H), 4.52 (m,
2H), 2.44 (m, 2H), 2.37 (m, 2H); MS (ES) m=z 359
(M+Hþ); FAB-HRMS (M+Hþ). Calcd 359.1256, found
359.1257.
1
ration of 47 gave 59 (78%) as a yellow solid: H NMR
(300 MHz, CDCl3) d 8.39 (dd, J ¼ 4:9, 1.5 Hz, 1H), 7.97
(dd, J ¼ 7:9, 1.5 Hz, 1H), 7.25 (m, 4H), 7.14 (dd,
J ¼ 7:9, 4.9 Hz, 1H), 6.95 (m, 1H), 6.86 (m, 2H), 4.53
(m, 2H), 3.95 (m, 2H), 2.35 (m, 2H); MS (ES) m=z 382
(M+Hþ).
5.1.54. 3-(2,4-Dimethoxy-pyrimidin-5-yl)-4-[1-(3-phen-
oxy-propyl)-1H-pyrrolo[2,3-b]pyridin-3-yl]-pyrrole-2,5-di-
one (60). Method B: using compound 59 and following
the same procedure as in the preparation of 48 gave 60
5.1.60.
4-{3-[4-(1-Methyl-1H-pyrazol-3-yl)-2,5-dioxo-
2,5-dihydro-1H-pyrrol-3-yl]-pyrrolo[2,3-b]pyridin-1-yl}-
butyronitrile (66). Method B: using 2-1-methyl-3-tribut-
ylstannanyl-1H-pyrazole (1.5 equiv) and following the
same procedure as in the preparation of 64 gave 66
(33%): 1H NMR (300 MHz, CDCl3) d 8.31 (m, 2H), 7.45
(m, 2H), 7.22 (dd, J ¼ 8:1, 1.5 Hz, 1H), 6.99 (dd,
J ¼ 8:1, 4.8 Hz, 1H), 6.80 (d, J ¼ 2:4 Hz, 1H), 4.51 (s,
2H), 3.85 (s, 3H), 2.44 (m, 2H), 2.36 (s, 2H); MS (ES)
m=z 361 (M+Hþ); FAB-HRMS (M+Hþ). Calcd
361.1413, found 361.1419.
1
(23%): H NMR (400 MHz, CDCl3) d 8.49 (s, 1H), 8.32
(d J ¼ 4:6 Hz, 1H), 8.12 (s, 1H), 8.02 (s, 1H), 7.28 (m,
2H), 7.08 (d, J ¼ 7:9 Hz, 1H), 6.92 (m, 4H), 4.61 (m,
2H), 4.06 (s, 3H), 4.01 (m, 2H), 3.43 (s, 3H), 2.44 (m,
2H); MS (ES) m=z 486 (M+Hþ); FAB-HRMS (M+Hþ).
Calcd 486.1778, found 486.1775.
5.1.55. 2-[1-(3-Cyano-propyl)-1H-pyrrolo[2,3-b]pyridin-
3-yl]-acetamide (61). Using 3-cyano-propyl bromide
(1.5 equiv) and following the same procedure as in the
1
preparation of 45 gave 61 (42%): H NMR (300 MHz,
5.2. Biology
CDCl3) d 8.33 (dd, J ¼ 4:7, 1.5 Hz, 1H), 7.91 (dd,
J ¼ 7:9, 1.5 Hz, 1H), 7.20 (s, 1H), 7.13 (dd, J ¼ 7:9,
4.7 Hz, 1H), 4.47 (m, 2H), 3.71 (s, 2H), 2.38 (m, 2H),
2.29 (m, 2H); MS (ES) m=z 243 (M+Hþ).
5.2.1. GSK-3 kinase assay. Compounds were tested for
the ability to inhibit recombinant rabbit GSK-3b (New
England Biolabs) using the following protocol. Protein
phosphatase inhibitor-2 (PPI-2, Calbiochem) phos-
phorylation was measured using a standard filtration
assay (MultiScreen-DV/Millipore). Briefly, the test
compounds were added to a reaction mixture containing
PPI-2 (45 ng), GSK-3b (0.75 units), and 33P-ATP (1 lCi)
in 50 mM Tris–HCl (pH 8.0), 10 mM MgCl2, 0.1% BSA,
1 mM DTT, and 100 lM activated Sodium Orthovana-
date. The total ATP concentration was 25 lM. After
90 min incubation at 30 ꢁC, the phosphorylated PPI-2
was precipitated using one volume of 20% trichloro-
acetic acid (TCA). Filter plates were subsequently wa-
shed with 10% TCA and radioactivity was quantified
using a TopCount Scintillation Counter (Packard). IC50
values were determined from at least three separate
experiments and the Ki values SEM were calculated
using the Cheng-Prussof equation.
5.1.56. 4-[3-(4-Hydroxy-2,5-dioxo-2,5-dihydro-1H-pyr-
rol-3-yl)-pyrrolo[2,3-b]pyridin-1-yl]-butyronitrile
(62).
Using compound 61 and following the same procedure
as in the preparation of 46 gave 62 (73%) as a yellow
solid: 1H NMR (300 MHz, CD3OD) d 8.65 (dd, J ¼ 8:1,
1.7 Hz, 1H), 8.17 (dd, J ¼ 4:8, 1.5 Hz, 1H), 7.69 (s, 1H),
7.06 (dd, J ¼ 7:9, 4.8 Hz, 1H), 4.38 (m, 2H), 2.43 (m,
2H), 2.21 (m, 2H); MS (ES) m=z 297 (M+Hþ).
5.1.57. 4-[3-(4-Chloro-2,5-dioxo-2,5-dihydro-1H-pyrrol-
3-yl)-pyrrolo[2,3-b]pyridin-1-yl]-butyronitrile (63). Using
compound 62 and following the same procedure as in
the preparation of 47 gave 63 (77%) as an orange solid:
1H NMR (300 MHz, CD3OD) d 8.52 (dd, J ¼ 8:1,
1.5 Hz, 1H), 8.39 (dd, J ¼ 4:7, 1.5 Hz, 1H), 8.32 (s, 1H),
7.26 (dd, J ¼ 8:1, 4.7 Hz, 1H), 4.53 (m, 2H), 2.51 (m,
2H), 2.28 (m, 2H); MS (ES) m=z 315 (M+Hþ).
5.2.2. Protein kinase selectivity panel (Upstate Biotech
Inc.). Protein kinase selectivity assays were performed as
previously described.36;39 Briefly, protein kinases were
assayed for their ability to phosphorylate the appropri-
ate peptide/protein substrates in the presence of 10 lM
compound. Assays were done using 100 lM ATP and
were linear with respect to time.
5.1.58. 4-{3-[4-(2,4-Dimethoxy-pyrimidin-5-yl)-2,5-dioxo-
2,5-dihydro-1H-pyrrol-3-yl]-pyrrolo[2,3-b]pyridin-1-yl}-
butyronitrile (64). Method B: using compound 63 and

3184
D. J. O’Neill et al. / Bioorg. Med. Chem. 12 (2004) 3167–3185
5.2.3. Glycogen synthase assay. Compounds were tested
for the ability to increase glycogen synthase (GS)
activity in living cells (HEK293 cells). To do this, cell
extracts were prepared from cells treated with com-
pounds or vehicle and GS activity measured using a
modified protocol.15b Briefly, cells were serum (and
glucose) starved for 3 h and treated with the appropriate
compounds for 90 min at 37 ꢁC. Cells were then washed,
scraped, and collected by centrifugation prior to lysis
using three freeze/thaw cycles. Lysates were then clari-
fied by centrifugation and the supernatants assayed for
GS activity. To do this, 14C-UDP glucose incorporation
into glycogen was measured in the absence or presence
of glucose 6-phosphate. The EC50 for GS activation was
then determined and compared with lithium.
1; MAPK2, mitogen-activated protein kinase 2; MAP-
KAP-K2, MAPK-activated protein kinase 2; MEK1,
MAPK/ERK kinase 1; MKK4, MAPK kinase 4;
MKK6, MAPK kinase 6; MKK7b, MAPK kinase 7b;
MSK1, mitogen- and stress-activated protein kinase 1;
PAK2, p21-activated protein kinase-2; PDGFR, plate-
let-derived growth factor receptor; PDK1, 3-phospho-
inositide-dependent protein kinase 1; PKA, camp-
dependent protein kinase; PKBa, protein kinase B a
(also called Akt); PKBb, protein kinase B b; PKC,
protein kinase C; PRAK, p38-regulated/activated
kinase; PRK2, protein kinase c-related protein kinase-2;
c-Raf, cellular product of raf proto-oncogene; RsK,
ribosomal S6 kinase; SAPK2b, stress-activated protein
kinase 2b (also known as p38b 2); P70S6K, p70 ribo-
somal protein S6 kinase; c-SRC, cellular product of src
oncogene, Syk, splenic tyrosine kinase; TrkB, tyrosine
receptor kinase-B; Yes, cellular product of the yes proto-
oncogene; ZAP-70, zeta-associated protein kinase-70.
m ¼ mouse, h ¼ human.
5.2.4. Metabolism, human liver microsomes. Human liver
microsomes were obtained from Xenotech (Kansas City,
KS). The microsomal reaction composition was pre-
pared as: microsomes, 1 mg/mL; NADPH, 1 mM;
potassium phosphate (pH 7.4), 100 mM; magnesium
chloride, 10 mM; test compound, 5 lM, and equilibrated
at 37 ꢁC for 3 min. The reaction was initiated by addition
of NADPH, and then incubated in a shaking water bath
at 37 ꢁC. Aliquots (100 lL) were withdrawn in duplicate
at 0, 15, 30, and 60 min and combined with 400 lL of
ice-cold 50/50 acetonitrile/dH2O to terminate the reac-
tion. Several controls (testosterone, propranolol, and
atenolol) were run simultaneously with the test com-
pounds. The percent compound remaining in the
incubation mixture were plotted as a function of time; a
first-order exponential equation was fit to the observed
data. The elimination half-lives associated with the dis-
appearance of test and control compounds were deter-
mined to compare their relative metabolic stability. The
T1=2 of testosterone ¼ 5.4 min, propranolol ¼ 29 min, and
atenolol > 100 min.
Acknowledgements
We thank Earl Danser for sending the compounds to
UBI for selectivity testing.
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