Synthesis and structure-activity relationships of novel poly(ADP-ribose) polymerase-1 inhibitors

Article abstract of DOI:10.1016/j.bmcl.2005.10.099

A series of novel pyrrolocarbazoles was synthesized as potential PARP-1 inhibitors. Pyrrolocarbazole 1 was identified as a potent PARP-1 inhibitor (IC₅₀ = 36 nM) from our internal database. Synthesis of analogs around this template with the aid

Full text of DOI:10.1016/j.bmcl.2005.10.099

Bioorganic & Medicinal Chemistry Letters 16 (2006) 938–942
Synthesis and structure–activity relationships of novel
poly(ADP-ribose) polymerase-1 inhibitors
Ming Tao,* Chung Ho Park, Ron Bihovsky, Gregory J. Wells, Jean Husten,
Mark A. Ator and Robert L. Hudkins
Cephalon, Inc., 145 Brandywine Parkway, West Chester, PA 19380-4245, USA
Received 19 September 2005; revised 26 October 2005; accepted 28 October 2005
Available online 15 November 2005
Abstract—A series of novel pyrrolocarbazoles was synthesized as potential PARP-1 inhibitors. Pyrrolocarbazole 1 was identified as
a potent PARP-1 inhibitor (IC50 = 36 nM) from our internal database. Synthesis of analogs around this template with the aid of
modeling studies led to the identification of the truncated imide 14. Compound 14 (IC50 = 40 nM), with deleted B-ring, was found
to be an equipotent PARP-1 inhibitor.
Ó 2005 Elsevier Ltd. All rights reserved.
Poly(ADP-ribose) polymerase-1 (PARP-1) is a nuclear
enzyme that catalyzes the synthesis of poly(ADP-ribose)
chains from NAD+ in response to single-strand DNA
breaks as part of the DNA repair process.1,2 The
PARP-1 enzyme is comprised of three functional re-
gions: an N-terminal DNA binding domain containing
two zinc fingers, a linker region, and a C-terminal cata-
lytic domain. Upon activation in response to DNA dam-
age, PARP-1 synthesis and degradation consumes
massive amounts of NAD+, which leads to depletion
of ATP energy stores, and ultimately necrotic cell death.
PARP-1 has been implicated in many important patho-
physiological processes such as stroke, myocardial ische-
mia, diabetes, shock, and traumatic CNS injury.3
PARP-1 inhibition in tumor cells may potentiate radio-
therapy and cancer chemotherapeutic agents targeting
DNA due to its involvement in processes related to
DNA repair.3 Therefore, potent, selective, soluble
PARP-1 inhibitors might be therapeutically useful in
the treatment of neurodegenerative disorders and
cancers.
inhibitors have been disclosed in the literature, many
suffer from development problems such as toxicity, poor
solubility, or poor pharmacokinetic profiles. In search of
novel PARP-1 inhibitors, we identified pyrrolocarbazole
1 (IC50 = 36 nM) as a potent inhibitor from our internal
database.6 Described here are the synthesis and evalua-
tion of the structure–activity relationships around this
novel pyrrolocarbazole PARP-1 template.
6
H
5
7
N
A
O
O
4
3
D
B
E
C
2
N
H
1
1
The synthesis of the pyrrolocarbazole imides is illustrat-
ed in Schemes 1 and 2 . In Scheme 1, protection of the
indole nitrogen with carbon dioxide, followed by 2-lith-
iation and addition to the cycloketone, provided the 2-
substituted indole–alcohol intermediate. Acid catalyzed
elimination of the alcohol at room temperature to the
diene, Diels–Alder reaction with neat maleimide, fol-
lowed by DDQ oxidation at 60 °C in toluene gave the
pyrrolocarbazole targets 1–3. Alternatively, as shown
in Scheme 2, the indole diene was prepared using an
intramolecular Wittig reaction.7 The indole diene react-
ed with maleimide, followed by DDQ oxidation to form
the pyrrolocarbazole imides 4–9.
In the literature, a variety of scaffolds, which mimic and
bind to the nicotinamide site of NAD+, have been
reported as inhibitors of PARP-1.4 The X-ray crystal
structures of 3-aminobenzamides and several other clas-
ses have been reported.5 Although a variety of PARP-1
Keywords: Pyrrolocarbazoles; PARP-1 inhibitors.
*
Corresponding author. E-mail: mtao@cephalon.com
Present address: Key Synthesis LLC, 804 Primrose Lane, Wynne-
wood, PA 19096, USA.
0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2005.10.099

M. Tao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 938–942
939
R2
H
R2
N
O
R2
O
O
R1
a
b
R1
R1
N
N
N
H
O
O
c
H
H
H
H
H
N
H
N
N
O
O
O
O
O
e
d
R2
R2
R1, R2 =
-CH2CH2CH2CH2
-
R1
R1
N
N
N
H
H
H
3
1, 2
Scheme 1. Reagents and conditions: (a) BuLi/THF, À78 °C, CO2(g), t-BuLi/THF, then, the ketone, 60–80%; (b) HCl, rt, 80–90%; (c) 190 °C, neat,
1 h, 50%; (d) DDQ, toluene, 60 °C, 75–85%; (e) Pd/C/hexene, reflux.
O
COCl
R1
R2
NH2
Br-
P
C6H56
R2
R2
NH
b
C6H5
C H5
Br-
P
C6H56
R1
+
C6H5
C H5
R1
a
N
H
+
H
N
H
H
N
N
O
O
O
O
O
O
d
R2
c
R2
R1
N
H
R1
N
H
4-9
Scheme 2. Reagents and conditions: (a) pyridine, CH2Cl2, reflux, 30 min, 76%; (b) potassium t-butoxide/toluene, reflux, 30 min, 48%; (c) neat,
190 °C, 61%; (d) DDQ, toluene, 40 °C, 40%.
The pyrrolocarbazole lactam regioisomers were pre-
pared using ethyl cis-b-cyanoacrylate as a dienophile
in a thermal Diels–Alder reaction with 2-cyclopentenyl
indole to form mainly cis-tetrahydrocarbazole regioi-
somers. DDQ oxidation and reductive cyclization of
5- and 7-cyano-esters using RaneyÒ nickel in DMF
produced lactams 10 and 11 after fractional recrystalli-
zations from DMF and acetone (Scheme 3). The benzo-
furan (12) and benzothiophene (13) analogs were
prepared in a similar manner for compounds 1–3, as
outlined in Scheme 4.
Truncated analogs were prepared as shown in Scheme 5.
The triisopropylsilyl-protected 2-cyclopentenyl pyrrole
diene was reacted with neat dimethyl acetylenedicarb-
oxylate at 150 °C to form the indole diester.8 Hydrolysis
of the diester to the diacid, anhydride formation, and
subsequent transamination with (TMS)2NH gave 14
NC
CO2Et
EtO2C
CN
NC
CO2Et
b
+
a
N
H
N
H
N
H
H
N
H
N
O
O
NC
CO2Et
EtO2C
CN
c
+
+
N
H
N
N
N
H
H
H
11
10
Scheme 3. Reagents and conditions: (a) neat, 200–205 °C, 1.5 h, >95%; (b) DDQ, toluene, 35–40 °C, 20 h, 25%; (c) RaneyÒ Nickel/DMF, 45 psi,
7 days; 28%.

940
M. Tao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 938–942
H
H
N
O
a
N
O
O
O
O
b
X
X
X
X = O, S
12, 13
Scheme 4. Reagents and conditions: (a) conditions for benzothiophene: (i)—BuLi/ether, 0 °C; (ii)—TsOH, reflux for 5 min, 47%; conditions for
benzofuran: (iii)—BuLi/ether, 0 °C; (iv)—TsOH, 40 °C for 30 min, 36%; (b) tetrachloroquinone, neat, 190 °C, 23–29%.
H
N
R2
O
O
MeO2C
N
CO2Me
R2
MeO2C
CO2Me
R1
R2
N
b-d
Si(OiPr)3
a
N
H
R1
R1
R1, R2 = H, cyclopentyl
Si(OiPr)3
14, 15
O
CO2Me
CO2Me
MeO2C
CO2Me
g-i
NH
e-f
O
16
Scheme 5. Reagents and conditions: (a) neat, 150 °C, 64 h, 21–35%; (b) 10 N NaOH in EtOH, reflux, 3 h, 96%; (c) acetic anhydride, 73 h, 66%; (d)
(TMS)2NH/MeOH, DMF, 73 °C, 4 h, 88%; (e) neat, rt, 18 h, 40%; (f) DDQ/toluene, rt, 1 h, 82%; (g) 5 N NaOH/MeOH, rt, 1 h, 66%; (h) Ac2O, 4 h,
85%; (i) NH2CONH2, neat, 150 °C, 30 min, 60%.
and 15. Compound 16 lacking the indole ring was pre-
pared by the same procedure as 14 and 15 but 1-vinyl-
cyclopentene was utilized as the starting material and
transamination was accomplished with urea (Scheme 5).
13 are inactive for PARP-1, indicating that the indole
nitrogen in compound 1 is important for activity.
Carbazole 1 was truncated to further explore the contri-
bution of the different rings. The indole imide 14, which
lacks phenyl ring B, displayed an equally potent PARP-
1 activity (IC50 = 40 nM), compared to the lead com-
pound 1 (IC50 = 36 nM). However, further truncated
compound 15, in which pyrrole ring C is also absent,
is a weaker PARP-1 inhibitor. Indole imide 16, where
both phenyl ring B and the cyclopentane ring have been
removed, is almost 20-fold less potent (IC50 = 750 nM).
The activity of 16 compared to, 15 reveals the impor-
tance of the indole N–H group and represents the min-
imum pharmacophore for retaining PARP-1 activity in
the series.
The pyrrolocarbazole analogs were evaluated as inhib-
itors of recombinant human poly(ADP-ribose) poly-
merase-1 as shown in Tables 1–4. As mentioned
earlier, pyrrolocarbazole 1 inhibits PARP-1 with the
IC50 value of 36 nM. Modification of the ring size
led to a loss of potency. The cyclohexyl analog 2
and fused phenyl analog 3 displayed IC50 values
>10 lM, indicating that the cyclopentyl ring is critical
for activity. The fused furano analog 4 is also a weak
PARP-1 inhibitor as shown in Table 1. Deleting the
cyclopentyl ring (5) or replacement with alkyl and
dialkyl groups led to a significant loss in potency.
The dimethyl analog 6 displayed modest PARP-1
activity (IC50 = 700 nM), while the other alkyl and
dialkyl analogs showed weaker activity.
A molecular docking study of carbazole 1 to the catalyt-
ic domain of chicken PARP-1 was conducted and is
illustrated in Figure 1. A binding model for PARP-1
was derived using the coordinates for 4-amino-1,8-naph-
thalimide bound at the NAD+ site of the catalytic frag-
ment of chicken PARP-1 (PDB 2PAX).5a Compound 1
was docked and minimized in the model. The key inter-
actions identified and supported by the SAR were
hydrogen-bond donor–acceptor interactions with the
imide/lactam 7-oxo C@O and NH with backbone
carbonyl and amino of Gly863. Ser904 is involved in a
H-bond with the 7-oxo C@O. The indole N–H shares
a hydrogen-bond with the carboxyl side chain of
Glu988. The aromatic p-stacking interactions occur
between carbazole rings B and D, and the aryl groups
As shown in Table 2, both 5-oxo (11) and 7-oxo (10) lac-
tam pyrrolocarbazoles were synthesized to explore the
role of the imide functionality. The 7-oxo lactam 10 is
2.5-fold less potent (IC50 = 90 nM) than the imide 1,
while the 5-oxo analog 11 is essentially inactive
(IC50 = 10 lM). The 7-oxo lactam isomer 10 is >100-
fold more potent than the 5-oxo isomer 11, indicating
that the 7-oxo carbonyl is required to bind to PARP-
1. The role of the indole N–H was evaluated by synthe-
sizing the benzofuran and benzothiophene analogs 12
and 13. As shown by the data in Table 3 both 12 and

M. Tao et al. / Bioorg. Med. Chem. Lett. 16 (2006) 938–942
941
Table 1. PARP-1 inhibition by pyrrolocarbazoles9
Table 3. Benzofuran and benzothiophene imides
H
H
N
N
O
O
O
O
R2
X
N
R1
H
a
PARP-1 IC50 (nM)
Compound
X
12
13
O
S
>10,000
>10,000
a,b
PARP-1 IC50 (nM)
Compound
R1
R2
a Values of duplicate determinations were within 2-fold of each other.
1
36
Table 4. PARP-1 inhibition by truncated analogs
2
3
ꢀ10,000
>10,000
a
PARP-1 IC50 (nM)
Compound
Structure
H
N
O
O
O
14
40
O
4
5
ꢀ10,000
ꢀ10,000
N
H
H
N
O
H
H
15
16
750
6
7
8
9
CH3
CH3
H
CH3
H
700
N
H
H
5000
N
O
O
2220
CH3
ꢀ2000
>10,000
a Values of duplicate determinations were within 2-fold of each other.
Et
nPr
a 4-Amino-1,8-naphthalimide (IC50 = 26 nM).
b Values of duplicate determinations were within 2-fold of each other.
Table 2. PARP-1 inhibition by pyrrolocarbazole lactams
a
PARP-1 IC50 (nM)
Compound
Structure
H
N
5
O
10
90
N
H
H
N
O
7
11
ꢀ10,000
N
H
a Values of duplicate determinations were within 2-fold of each other.
of Tyr896 and Tyr907. The cyclopentyl fits closely into a
fold formed by the Lys861 side chain, Ala898, Trp861,
and Asn987 which locks the inhibitor in the pocket.
Figure 1. Important interactions of compound 1 with PARP-1.
ing of our internal library. Structural modification to
the core identified the key pharmacophore elements
necessary for PARP-1 inhibition. The cyclopentyl ring
In summary, we identified a novel pyrrolocarbazole
PARP-1 inhibitor (1) through high throughput screen-

942
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required. The des-aryl compound 14, with a lower
molecular weight, would be anticipated to have im-
proved physical chemical properties and represents a
novel small molecule PARP-1 scaffold.
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Acknowledgments
The authors acknowledge the support of and discussions
with Drs. John Mallamo, Ed Bacon, and Jeffry Vaught.
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