Discovery and SAR of substituted 3-oxoisoindoline-4-carboxamides as potent inhibitors of poly(ADP-ribose) polymerase (PARP) for the treatment of cancer

Article abstract of DOI:10.1016/j.bmcl.2009.12.042

Through conformational restriction of a benzamide by formation of a seven-membered hydrogen-bond with an oxindole carbonyl group, a series of PARP inhibitors was designed for appropriate orientation for binding to the PARP surface. This series of compound

Full text of DOI:10.1016/j.bmcl.2009.12.042

Bioorganic & Medicinal Chemistry Letters 20 (2010) 1023–1026
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery and SAR of substituted 3-oxoisoindoline-4-carboxamides as
potent inhibitors of poly(ADP-ribose) polymerase (PARP) for the treatment
of cancer
a
a
a
a
a
b
Viraj B. Gandhi a, , Yan Luo , Xuesong Liu , Yan Shi , Vered Klinghofer , Eric F. Johnson , Chang Park ,
*
Vincent L. Giranda a, Thomas D. Penning a, Gui-Dong Zhu a
a Cancer Research, GPRD, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064, USA
b Structural Biology, GPRD, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Through conformational restriction of a benzamide by formation of a seven-membered hydrogen-bond
with an oxindole carbonyl group, a series of PARP inhibitors was designed for appropriate orientation
for binding to the PARP surface. This series of compounds with a 3-oxoisoindoline-4-carboxamide core
structure, displayed modest to good activity against PARP-1 in both intrinsic and cellular assays. SAR
studies at the lactam nitrogen of the pharmacophore have suggested that a secondary or tertiary amine
is important for cellular potency. An X-ray structure of compound 1e bound to the protein confirmed the
formation of a seven-membered intramolecular hydrogen bond. Though revealed previously in peptides,
this type of seven-membered intramolecular hydrogen bond is rarely observed in small molecules. Lar-
gely due to the formation of the intramolecular hydrogen bond, the 3-oxoisoindoline-4-carboxamide core
structure appears to be planar in the X-ray structure. An additional hydrogen bond interaction of the
piperidine nitrogen to Gly-888 also contributes to the binding affinity of 1e to PARP-1.
Received 20 October 2009
Revised 5 December 2009
Accepted 10 December 2009
Available online 14 December 2009
Keywords:
PARP inhibitor
Inhibitor of poly(ADP-ribose) polymerase
3-Oxoisoindoline-4-carboxamide
Ó 2009 Elsevier Ltd. All rights reserved.
Poly(ADP-ribose) polymerases (PARPs) constitute a family of
cell signaling enzymes abundant in all eukaryotes.1 Poly(ADP-ri-
bose) polymerase-1 and 2 (PARP-1 and 2), key members of the
PARP family, play a significant role in preservation of genomic
integrity through DNA repair and control of RNA transcription.2
In the event of a stimuli caused during pathophysiological pro-
cesses such as inflammation, stroke, myocardial infarction, neuro-
degenerative disorder or radiation therapy, PARP is activated to
catalyze the synthesis of poly(ADP-ribose) chains from nicotin-
amide adenine dinucleotide (NAD+) through automodification or
ribosylation to aid the process of cell repair.3 Thus PARPs are crit-
ical regulatory components and act as gatekeepers in maintaining
cell fidelity.4 The formation of ADP-ribose polymers is crucial in the
repair of damaged DNA caused by radiation therapy or chemother-
apeutic agents. Consequently, PARP-1 and PARP-2 interfere with
the aforementioned therapies and diminish its effectiveness in
treating cancer. Moreover, it has been observed that PARP expres-
sion is enhanced in a number of hematological and solid tumors as
compared to normal cells. The dual role played by the enzyme
therefore suggests that pharmacological modulation of its activity
could increase the efficacy of DNA-damaging antitumor drugs.5–12
Additionally, tumors with deficiencies in DNA-repair genes such as
BRCA-1 or BRCA-2 have displayed acute sensitivity towards inhibi-
tion of PARP-1, suggesting a potential utilization of PARP inhibitors
as single agents in treating certain mutations of breast cancer.13
PARP-1 is therefore regarded as a valuable target in the exploration
of new cancer treatment regimens.
Figure 1 is a schematic presentation of the X-ray structure of
NAD+ binding to PARP-1, and formation of multiple hydrogen bond
Gly-863
N
O
H
H
Ser-904
O
N
NH2
N
H
H
O
N
N
O
P
O
P
N+
O
O
N
O
O
O- O-
HO
HO
OH
OH
OH
Tyr-907
Figure 1. A schematic presentation of the X-ray structure of NAD+ binding to PARP-
1. The cycled part of structure (red) showed critical hydrogen bond interactions of
NAD+ with PARP-1.
* Corresponding author.
E-mail address: viraj.b.gandhi@abbott.com (V.B. Gandhi).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.12.042

1024
V. B. Gandhi et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1023–1026
Br
O
Br
O
KHCO3/MeI
Zn(CN)2
OH
O
O
N
DMF
18 h
Pd(PPh3)4
DMF
80 ºC/20 h
N
2
3
H
O
N
N
O
O
NH2R/THF
O
O
Br
NBS/AIBN
Figure 2. The molecular structure of o-HBDI. (Picture adopted from Journal of the
American Chemical Society, 2007, 129, 4534, and copyright granted from American
Chemical society (License number 2312091023118).)
-20 ºC - r.t/ 18 h
CHCl3
65 ºC/18 h
4
5
interactions between the carboxamide moiety and two key amino
acid residues in the PARP active site, Gly-863 and Ser-904.5–7 Most
known PARP inhibitors mimic the NAD+ binding mode, by confor-
mational restriction of a primary amide or by incorporation of an
amide into a lactam.5–12 Formation of a more planar structure as
shown in the circle of Figure 1 is more likely optimal for the
binding.
N
O
NH2
O
O
PPA/ 110 ºC/ 18 h
N
R
N R
or
30% HBr in CH3COOH
CH3COOH/ 2h
6
1
Restricted rotation of a functional group has been frequently
achieved through formation of five- or six-membered ring intra-
molecular hydrogen bond.5–7 An intramolecular hydrogen bond
incorporating in a larger ring has been rarely observed, primarily
in peptide type of structures. Recently, a seven-membered-ring
hydrogen-bonding system 4-(2-hydroxybenzylidene)-1,2-dime-
thyl-1H-imidazol-5(4H)-one (o-HBDI) has been reported,14 and
formation of the intramolecular hydrogen bond is believed to be
critical to an excited-state intramolecular proton transfer. The
structure of the seven-membered ring hydrogen bond has been
confirmed by an X-ray structure and the ring structure incorporat-
ing the hydrogen bond appears to be planar (Fig. 2).
To our surprise, no significant difference on the binding energy
was observed in peptides among 7–10-membered-ring intramo-
lecular hydrogen bond, ranging from 5 to 10 kcal/mol.15,16 This
binding energy is also in the same range as compared to five- or
six-membered intramolecular hydrogen bonds. Since a seven-
membered ring intramolecular hydrogen bond contributes to the
same level of orientation restriction as compared to that of smaller
rings, we have designed 3-oxoisoindoline-4-carboxamide 1 in
which the carboxamide is restricted into the shown orientation
as in Figure 3, through formation of seven-membered ring hydro-
gen bond.
Scheme 1 outlines the general procedure used in the synthesis
of our PARP inhibitors. Commercially available 2-bromo-6-methyl-
benzoic acid 2 was treated with methyl iodide and potassium
bicarbonate in DMF to provide the methyl ester 3 in nearly quan-
titative yields. Benzonitrile 4 was obtained in excellent yield
through a palladium-catalyzed cyanation of aryl bromide 3 and
zinc cyanide as the cyanide source. Free radical bromination of 4
with AIBN and NBS yielded 5 in good yield. The penultimate trans-
formation to provide the cyclized oxoisoindoline core 6 was
achieved by treating 5 with selected alkyl and aryl primary amines
in THF. Carboxamides 1a–1q were obtained through hydrolysis of
nitrile 6 in PPA. Alternatively, the nitrile to amide transformation
can be accomplished by heating in 30% HBr in acetic acid.
Scheme 1.
Table 1 outlines a variety of PARP analogs synthesized employ-
ing the procedure as described above in Scheme 1. Alkyl analogs 1a
and 1b showed modest intrinsic potencies but poor cellular activ-
ity. Introduction of a cyclic amine, in particular, 1d and 1e resulted
in significant boost in PARP activity, with enzymatic activity of
59 nM and 33 nM and cellular activity of 160 nM and 51 nM,
respectively. Based on the initial observation it was evident that
a basic amino functionality was beneficial for improved cellular
activity. To this end, compounds 1f–1i, containing 3-piperidine,
2-pyrrolidine, 3- and 4-methyl piperidines were synthesized. How-
ever, this modification resulted in ca. fivefold to 10-fold drop in po-
tency as compared to piperidine 1e demonstrating that the
presence and position of the basic amino functionality had signifi-
cant impact on the PARP activity, both in an enzyme and cellular
assays. In an attempt to understand the binding mode of these
compounds in the active site and explore other potential interac-
tions, substituted phenyl and aryl analogs 1j–1q were synthesized.
Generally, these analogs showed a similar enzyme and cellular po-
tency profile compared to aforementioned cycloalkyl analogs.
However, 4-substituted phenyl analogs were significantly more
potent compared to the 3-substituted analogs. For instance, com-
pound 1l with a para substitution has enzymatic potency of
53 nM, while 1m is significantly less potent at 391 nM. Similarly,
analog 1o, para-substituted piperidine, is 10-fold more potent in
comparison with the meta-substituted piperidine analog 1n. A
comparison of analogs 1n and 1p indicated that addition of second
nitrogen in the ring results in a slight drop in enzymatic potency
while 1p gained more than fivefold boost in cellular potency. Final-
ly, incorporation of a pyridine-3-methyl functionality 1q led to loss
of PARP activity. Selected compounds (e.g., 1e) were tested against
the closely-related PARP-2 enzyme, and displayed equal potency as
PARP-1.
A versatile characteristic of our novel 3-oxoisoindoline-4-car-
boxamide class of PARP inhibitors is the favorable orientation of
the carboxamide moiety relative to the 3-oxo substitution. As
shown in Figure 3, the nitrogen of the carboxamide moiety and
the 3-oxo substitution are involved in a seven-membered intramo-
lecular hydrogen bonding interaction thereby restricting the free
rotation of the amide functionality. The locked conformation of
the molecule is presumably responsible for its optimal interaction
with the PARP enzyme and its inhibition. The favorable orientation
of the amide enables the 3-oxoisoindoline-4-carboxamide moiety
H
N
H
O
O
R
N
1
Figure 3. Structure of designed scaffold 1.

V. B. Gandhi et al. / Bioorg. Med. Chem. Lett. 20 (2010) 1023–1026
1025
Table 1
SAR of 2-substituted 3-oxoisoindoline-4-carboxamide
O
NH2
O
N R
1
Compound
R
PARP-1a
Cellulara
(Ki, nM)
(EC50, nM)
a
b
CH3
i-Bu
123
112
ND
>1000
c
d
e
172
59
>1000
160
NH
NH
NH
33
51
Figure 4. X-ray co-crystal structure of PARP-1 and 1e.18
f
g
140
377
384
ND
374
ND
In conclusion, through rational design of a seven-membered
ring intramolecular hydrogen bond utilizing a carbonyl group
and a carboxamide functionality, we have identified 3-oxoisoindo-
line-4-carboxamide as a lead inhibitor of poly(ADP-ribose) poly-
merase-1 (PARP-1). Preliminary SAR study of this series of PARP
inhibitors indicates that the presence and position of an amine
functionality has significant impact both on enzymatic and cellular
potency. An X-ray co-crystal structure of PARP-1 with 1e con-
firmed the critical multiple hydrogen bonding interactions of the
4-carboxamide with two PARP residues, as well as intramolecu-
larly with the 3-carbonyl to lock the amide in an optimal planar
conformation. Additional hydrogen bond interaction to Gly-888
was also achieved through an appropriately-oriented piperidine
nitrogen.
N
H
h
NH
i
j
164
261
ND
ND
N
H
k
l
70
53
372
53
N
N
Acknowledgment
N
m
n
391
32
ND
The authors are grateful to the Abbott analytical department for
acquisition of 1H NMR and MS.
120
NH
NH
References and notes
o
314
ND
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Gly-888 and lastly the p-stacking interaction between the oxoiso-
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