Design, synthesis, crystallographic studies, and preliminary biological appraisal of new substituted triazolo[4,3-b]pyridazin-8-amine derivatives as tankyrase inhibitors

Article abstract of DOI:10.1021/jm401356t

Searching for selective tankyrases (TNKSs) inhibitors, a new small series of 6,8-disubstituted triazolo[4,3-b]piridazines has been synthesized and characterized biologically. Structure-based optimization of the starting hit compound NNL (3) prompted us to the discovery of 4-(2-(6-methyl-[1,2,4] triazolo[4,3-b]pyridazin-8-ylamino)ethyl)phenol (12), a low nanomolar selective TNKSs inhibitor working as NAD isostere as ascertained by crystallographic analysis. Preliminary biological data candidate this new class of derivatives as a powerful pharmacological tools in the unraveling of TNKS implications in physiopathological conditions.

Full text of DOI:10.1021/jm401356t

Brief Article
pubs.acs.org/jmc
Design, Synthesis, Crystallographic Studies, and Preliminary
Biological Appraisal of New Substituted Triazolo[4,3‑b]pyridazin-8-
amine Derivatives as Tankyrase Inhibitors
Paride Liscio,†,∥ Andrea Carotti,† Stefania Asciutti,‡ Tobias Karlberg,§ Daniele Bellocchi,† Laura Llacuna,‡
Antonio Macchiarulo,† Stuart A Aaronson,‡ Herwig Schuler,§ Roberto Pellicciari,†,∥
̈
and Emidio Camaioni*,†
†Dipartimento di Chimica e Tecnologia del Farmaco, Universita
̀
degli Studi di Perugia, Via del Liceo 1, 06123 Perugia, Italy
‡Mount Sinai School of Medicine, Dept. Oncological Sciences, 1425 Madison Avenue, New York, New York 10029 United States
§Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S-17177 Stockholm, Sweden
∥TES Pharma, via P. Togliatti 22bis, 06073 Localita
̀
Terrioli, Corciano, Italy
S
* Supporting Information
ABSTRACT: Searching for selective tankyrases (TNKSs)
inhibitors, a new small series of 6,8-disubstituted triazolo[4,3-
b]piridazines has been synthesized and characterized bio-
logically. Structure-based optimization of the starting hit
compound NNL (3) prompted us to the discovery of 4-(2-
(6-methyl-[1,2,4]triazolo[4,3-b]pyridazin-8-ylamino)ethyl)-
phenol (12), a low nanomolar selective TNKSs inhibitor
working as NAD isostere as ascertained by crystallographic analysis. Preliminary biological data candidate this new class of
derivatives as a powerful pharmacological tools in the unraveling of TNKS implications in physiopathological conditions.
TNKS-dependent PARsylation and thus promoting β-catenin
phosphorylation and degradation. Recently, they have been also
cocrystallized with TNKS-2.4,5 While 1 (XAV-939) binds in the
classical nicotinamide binding site,4 2 (IWR-1) occupies an
accessory pocket making interaction with the so-called D-loop.5
A thorough review of TNKS inhibitors as well as their
pharmacological implications are however reported else-
where.6−8 As a continuation of our research project devoted
to the design and synthesis of new inhibitors of the PARP’s
family,9,10 we have recently focused our attention to the
discovery of new selective TNKS-1 and TNKS-2 inhibitors.
The Structural Genomics Consortium (SGC) released
several crystal structures of the catalytic domain of TNKS-2
in complex with new ligands.4,10 Among all new deposited
structures, our attention was attracted by the cocrystal of
TNKS-2 and N-(4-chlorophenethyl)-6-methyl-[1,2,4]triazolo
[4,3-b] pyridazin-8-amine (NNL, 3, PDB code 3P0Q).10
Interestingly, although 3 (NNL) is missing the amide feature,
all the interactions formed by the classical PARP inhibitors that
bind in the canonical site were conserved (Figure 1S of
Supporting Information, (SI)). Herein, with the aim to define
structure−activity relationships around this unexplored scaffold,
we have synthesized a small library of new triazolopyridazine
derivatives bearing different amine in position C-8 with or
without a methyl or ethyl group in position C-6. To further
INTRODUCTION
■
Tankyrases (TNKS-1 and TNKS-2) belong to the PARP
superfamily. Because of their ability in the transfer of
polyADPribose chain (PAR) to targeted proteins, they are
also referred to as ADP-ribosyltransferases ARTD-5 and
ARTD-6, respectively.1 TNKS-1 and TNKS-2 share high
sequence and structural homology and overlapping functions.
In 2009, two independent works reported the first selective
TNKSs inhibitors endowed with Wnt pathway disruption
properties through axin stabilization. By using a standard TCF/
β-catenin-dependent reporter assay, Huang et al.2 identified
XAV-939 (1, Chart 1) as the first selective TNKSs inhibitor
Chart 1. Chemical Structure of Parent TNKSs Inhibitors
(IC50: TNKS-1, 0.011 μM; TNKS-2, 0.004 μM) while by using
a similar reporter-based screening approach, Chen et al.3
discovered that structurally distinct small molecules, including
IWR-1 (2, Chart 1), were equally able to disrupt Wnt signaling
via TNKSs inhibition (IC50: TNKS-1, 0.131 μM; TNKS-2,
0.056 μM). These two TNKSs inhibitors block Wnt target gene
expression stabilizing Axin-1 and -2 proteins by preventing their
Received: September 4, 2013
Published: February 16, 2014
© 2014 American Chemical Society
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Brief Article
investigate the influence of the nitrogen atoms of this
heterocycle on the interaction with the enzyme binding site,
the scaffold of the most active compound was simplified by the
preparation of the corresponding 8-amino-sustituted-imidazo-
[1,2-a]pyridine, -[1,2,4]triazolo[1,5-a]pyridine, and -quinoline
derivatives, thus reducing the endocyclic nitrogen atoms from 4
to 1. Finally, all the new compounds were tested for their
capability to inhibit in vitro TNKS-1 and TNKS-2, and the
most promising compound was further characterized bio-
logically.
Scheme 2. Synthesis of [1,2,4]Triazolo[4,3-b]pyridazine
Derivatives
a
RESULTS AND DISCUSSION
■
The synthesis of the s-triazolo[b]pyridazine nucleus was first
reported in 1959 by Steck and co-workers.11 Indeed, 8-
chlorine-6-alkyl-[1,2,4]triazolo[4,3-b]pyridazine derivatives 4
and 5 (Scheme 1) were obtained in high yields following a
a
Reagents and conditions: (a) DPPA, t-BuOH, dioxane, 110 °C; (b)
TFA/DCM 1/1, rt; (c) NH2NH2·H2O, 105 °C; (d) CHO2H, reflux;
(e) H2, Pd/C 10%, DIPEA, EtOH, 40 psi; (f) isoamylnitrite, CH2I2,
CH3CN, reflux; (g) 4-methoxyphenethylamine, DMF, 105 °C; (h)
BBr3, DCM, rt.
Scheme 1. General Synthesis of 6-Alkyl-[1,2,4]triazolo[4,3-
b]pyridazine Derivatives
a
demethylation with BBr3, the compound 33, in overall good
yield. The preparation of other nitrogen-containing hetero-
cycles 40−41, 45−46, and 49−50 has been carried out
following the synthetic routes reported in Scheme 3.
8-Nitro-1,2,4-triazolo[4,3-a]pyridine 36 was prepared in high
yield starting from commercially available 2-chloro-3-nitro
pyridine 34 as already described.18,19 Dimroth rearrangement of
intermediate 36 followed by hydrogenation of the nitro group
furnished the corresponding isomer 8-amino-1,2,4-triazolo[2,3-
a
Reagents and conditions: (a) R2NH2, DMF, 105 °C; (b) BBr3, DCM,
rt; (c) BzCl, Py, rt.
similar approach of that already reported11 (Scheme 1S, SI).
They were then submitted to nucleophilic substitution
reactions with suitable amines, thus furnishing the correspond-
ing final compounds 3, 6−11, 14−20, and 22−23 (Scheme 1).
Derivatives 11 and 23 bearing a methoxy group in para-
position of the distal phenyl ring were demethylated by
treatment with boron tribromide to obtain the desired hydroxyl
derivatives 12 and 24, respectively, in high yields, while this
reaction on p-methoxy benzylamino compound 18 afforded the
8-amino-6-methyl-[1,2,4]triazolo[4,3-b]pyridazine derivative
21 (Scheme 1).
Scheme 3. Synthesis of Other Nitrogen Containing
a
Heterocycles 40−41, 45−46, and 49−50
C-6 unsubstituted derivatives 32 and 33 were prepared
following the synthetic procedure depicted in Scheme 2. 3,6-
Dichloro-4-pyridazine carboxylic acid 25 was easily synthesized
in three steps as previously described12,13 (see Scheme 2S, SI).
Amino replacement of the carboxyl group of this latter
intermediate was accomplished in two steps via Curtius
rearrangement of the acid 25 and by subsequent deprotection
of the so formed tert-butoxy carbonyl amide 26. Selective
exchange of one halogen atom was accomplished by treatment
of the dichloro derivative 2714 with hydrazine hydrate. 6-
Chloro-3-hydrazino-pyridazin-4-ylamine 2815 was refluxed in
formic acid, affording the key intermediate 6-chloro-[1,2,4]-
triazolo[4,3-b]pyridazin-8-ylamine 29 in acceptable yields.16
Removal of the chlorine atom in C-6 position of derivative 29
was affected quantitatively by hydrogenation over a palladium
catalyst at 40 psi, furnishing the corresponding [1,2,4]triazolo-
[4,3-b]pyridazin-8-ylamine 30.17 Because of the low reactivity
of the previous amine toward acylation reaction, the classical
Sandmayer procedure was applied, converting in high yield
compound 30 to its 8-iodo derivative 31. Nucleophilic
substitution of this latter intermediate with 4-methoxypheneth-
yl amine afforded the compound 32 and, by subsequent
a
Reagents and conditions: (a) NH2NH2·H2O, MeOH, rt; (b)
CHO2H, 150 °C; (c) NaHCO3(ss), rt; (d) H2, Pd/C 10%, EtOH,
20 psi, 50 °C; (e) 4-methoxyphenylacethyl chloride, Et3N, DCM; (f)
LiAlH4, Et2O or DCM, rt; (g) BBr3, DCM, rt.
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Journal of Medicinal Chemistry
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a]pyridine 38 in overall 60% yield for two steps.19 Coupling of
derivative 38 with 2-(4-methoxyphenyl)acetyl chloride fur-
nished the corresponding amide 39 in 79% yield. The latter
amide was easily reduced with lithium aluminum hydride
providing the phenethyl amine derivative 40, which was then
submitted to the usual demethylation reaction with boron
tribromide, furnishing the desired compound 4-(2-([1,2,4]-
triazolo[1,5-a]pyridin-8-ylamino)ethyl)phenol 41 in high yield.
The synthesis of the N-(4-methoxyphenethyl)H-imidazo[1,2-
a]pyridin-8-amine (45) and 4-(2-(H-imidazo[1,2-a]pyridin-8-
ylamino)ethyl)-phenol (46) was carried out in a similar
procedure of that described before. Indeed, H-imidazo[1,2-
a]pyridin-8-amine (43) was synthesized in good yield following
a literature protocol20,21 and starting from commercially
available 2,3-diaminopyridine (42). 43 was then submitted, as
previously described, to the acylation reaction with 2-(4-
methoxyphenyl)acetyl chloride, providing the corresponding
amide 44, and to subsequent amide reduction, furnishing the
phenethylamine derivative 45 in high yield. Finally, demethy-
lation reaction of 45 provided the desired hydroxyl-derivative
46 in 43% yield. Derivatives 49 and 50 were prepared in three
steps as already described, starting from commercial 8-
aminoquinoline (47).
All newly synthesized compounds 3, 6−24, 32−33, 40−41,
45−46, and 49−50 were tested for their ability to inhibit in
vitro TNKS activity at 1 μM concentration, and the results are
reported in Table 1. One was used as reference compound,
showing full inhibition (100%) of both tankyrases at a
concentration of 1 μM. The hit compound 3, previously
cocrystallized with TNKS-2, inhibited at 1 μM TNKS-1 and -2
enzymatic activities by 75 and 52%, respectively. The
corresponding unsubstituted phenethylamino derivative 6 was
found to be less active, while the 2-naphthyl analogue 7 was
more active, displaying TNKSs inhibition potency of 81 and
74%, respectively. Replacement of the distal phenyl ring with
other heterocycles such as 2-thienyl or 2-pyridinyl moieties (8
and 9, respectively) gave contrasting results. While the presence
of a thiophene decreased the inhibitory activity versus both
enzymes (compound 8, Table 1), the pyridine ring maintained
the activity profile only on TNKS-1 (compound 9). Further
substituents were introduced in the para-position of the distal
phenyl ring in order to modulate the inhibition activity and to
increase the water solubility. Electron-donor groups like
methoxy, as in derivative 11, increased the inhibition activity
values to 87 and 50% at TNKS-1 and -2, respectively. On the
contrary, small electron-withdrawing groups such as fluorine
(10) did not elicit interesting changes in activity. Replacing the
methoxy group with a hydroxy moiety provided derivative 12
that resulted at 1 μM in full inhibition of TNKS-1 and 82% of
inhibition at TNKS-2. Benzoylation of the hydroxy group of
derivative 12 as in compound 13 was tolerated, whereas
replacement of this hydroxyl with an amine group was found to
be ineffective, as derivative 14 was less potent. Replacement of
the phenyl ring with N-methyl piperazine (15) and morpholine
(16) led to complete loss of the inhibitory activity. The nature
of the linker between the distal aromatic moiety and the
triazolo[4,3-b]pyridazine ring is crucial for the activity vs both
TNKSs. Indeed, reducing the linker length to a methylene
bridge (17 and 18) as well as increasing the spacer length with
a propylene group (19) afforded to very weak inhibitors. The
same trend was observed when the flexibility of the spacer was
blocked by introduction of an indane ring as in derivative 20 or
when the phenylethyl side chain was removed (21). The
Table 1. Inhibition Data of Novel Derivatives at 1 μM
Concentration against Recombinant Human TNKS-1 and -2
% TNKS
a
inhibition
compd
R1
R2
X
Y
1
2
1
100
75
23
81
36
72
65
87
100
63
45
na
na
8
99
52
12
74
24
25
25
50
82
43
31
na
na
5
3
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Me
Et
(CH2)2-Ph-p-Cl
(CH2)2-Ph
6
7
(CH2)2-2-Naph
(CH2)2-2-Th
8
9
(CH2)2-2-Py
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
32
33
40
41
45
46
49
50
(CH2)2-Ph-p-F
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
(CH2)2-Ph-p-OCOPh
(CH2)2-Ph-p-NH2
(CH2)2-4-N-mepiperazine
(CH2)2-N-morpholine
CH2-Ph
CH2-Ph-p-OCH3
(CH2)3-Ph
na
na
16
na
71
na
81
32
48
na
38
na
8
na
7
2-indanyl
6
H
9
(CH2)2-2-Th
33
na
40
18
33
8
Et
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
(CH2)2-Ph-p-OCH3
(CH2)2-Ph-p-OH
Et
H
H
N
N
N
N
N
N
20
na
13
na
43
CH
CH
na
22
a
Data are from experiments conduct in duplicate at concentration of 1
μM; na, not active.
methyl group in position C-6 (derivatives 11, 12) appears to be
the optimal substituent as its ethyl replacement or removal,
such as in compounds 23−24 and 32−33, respectively, led to
less active species. This was also highlighted by the super-
position analysis of our crystal and the 3W51 pdb complex
from Larsson et al.,22 where the two methyl groups occupy the
same pocket (Figure 2S, SI). The triazolo[4,3-b]pyridazine
nucleus of compound 12 is crucial for activity, given that
simplified analogues 41, 46, and 50 were less active
(computational studies and Figure 3S, SI). Among all new
derivatives, 4-(2-(6-methyl-[1,2,4]triazolo[4,3-b]pyridazin-8-
ylamino)ethyl)phenol (compound 12), bearing a p-hydroxy
phenyl ring in the side chain, was the most potent TNKSs
inhibitor of the series and was selected for further biological
characterization.
The selectivity of 12 was assessed at a concentration of 10
μM on several members of the PARP superfamily (PARP 1−3,
6−8, 10−12) and compared with AZD22816 (Olaparib), which
displayed >70% inhibition in all tested PARPs, as reported in
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Journal of Medicinal Chemistry
Brief Article
Figure 1 (Table 1S, SI) and in literature.22 At this
concentration, 12 fully inhibited both TNKSs whereas it was
Figure 1. Selectivity panel of compound 12 toward several members of
the PARP superfamily. Data are from experiments conduct in duplicate
at 10 μM, AZD2281 was used as positive control.
Figure 2. (A) TOP/RL TCF-luciferase analysis. (B) Cell growth
inhibition of DLD-1 colon tumor cells. Compound 12 was compared
with standard inhibitors (1 and 2) in Wnt activated DLD-1 cells
(DMSO is a negative control). Data are mean SEM from at least
three independent experiments.
a weak inhibitor on other PARPs (for further details see Table
1S, SI). Furthermore, as shown in Table 2, 12 exhibited a clean
selectivity trend toward PARP-1 and -2 with respect to the
reference compound 1.
Table 2. IC50 (μM) and PARP Selectivity of 1 and 12
compd
PARP1
PARP2
TNKS1
TNKS2
a
a
a
a
1
0.12
>10
0.046
>10
0.011
0.008
12
0.012
0.2
a
Data are from ref 22.
Finally, full dose−response curves of 12 against TNKS-1 and
-2 were performed with resulting IC50s of 0.012 0.002 and
0.2 0.1 μM, respectively (Table 2; Figure 4S, SI).
Taking into account the excellent potency and the
improvement on the selectivity profile, we deemed it
appropriate to further investigate compound 12 in cellular
assays involving Wnt pathway signaling.
Figure 3. Crystal structure of 12 with the catalytic domain of TNKS-2.
Electron density around 12 displayed as mesh contoured at 1.5 σ (0.39
eÅ−3). Hydrogen bonding interactions from 12 to active site residues
are highlighted in green. The figure was prepared using PyMOL
(http://www.pymol.org).
Previous studies have shown that Axin stabilization by TNKS
inhibitors can inhibit TCF signaling and, subsequently, cell
proliferation of Wnt activated DLD-1 cancer cells.2 To assess
the effect of our inhibitor 12 on TCF-dependent transcriptional
activity, we subjected mass cultures of DLD-1 colorectal cancer
cells to a 24 h treatment with increasing concentrations of
compound 12 (Figure 2A). The new ligand inhibited dose-
dependently TCF reporter. While at low doses (1 μM)
reference compounds 1 and 2 displayed high inhibitory
activities, at 10 and 20 μM all compounds showed a similar
pattern of inhibition. Furthermore, good long-term growth
inhibition on DLD-1 was also observed (Figure 2B).
A crystal structure of the TNKS-2 PARP domain in complex
with the compound 12 at 1.95 Å resolution was also obtained
(Figure 3). Resulting electron density for this ligand was of
excellent quality, which allowed for an accurate placement of
12. The interactions observed for compound 3 mimicking the
nicotinamide in NAD+ were also present for compound 12.
The hydroxyphenylethyl chain is directed toward the solvent
and the aromatic ring structure is neatly flanked by hydro-
phobic side chains (Pro1034, Phe1035, Tyr1050, and Ile1075),
whereas the hydroxyl makes interactions, via two waters, with
residues on both sides of the pocket.
CONCLUSION
■
Herein we have synthesized a small library of new [1,2,4]-
triazolo[4,3-b]pyridazin-8-amine derivatives substituted either
in position 6 or 8 as potential selective TNKSs inhibitors. On
the basis of the biological results, starting from the selected “hit
compound” 3, a first structure−activity relationship profile was
drawn. Thus, the absence of an amide moiety in an anti
conformation as the classical pharmacophoric group of PARP
inhibitors confers to this new class of derivatives high TNKSs
specificity. Compound 12 resulted the most active and selective
derivative, endowed with low nanomolar values of IC50 on
TNKSs proteins and devoid of any PARPs cross-reactivity.
Furthermore, it was also assessed as Wnt pathway diruptor. In
parallel to this work, Shultz et al.23 discovered that [1,2,4]-
triazol-3-ylamine derivatives are also able to inhibit TNKSs by
binding in a similar manner as our molecule. These new classes
of compounds are promising isosteres of nicotinamide
derivatives and can be considered new powerful pharmaco-
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Journal of Medicinal Chemistry
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(4) Karlberg, T.; Markova, N.; Johansson, I.; Hammarstrom, M.;
̈
logical tools in the unravelling of TNKS implications in
physiopathological conditions.
Schutz, P.; Weigelt, J.; Schuler, H. Structural basis for the interaction
̈
̈
between tankyrase-2 and a potent Wnt-signaling inhibitor. J. Med.
Chem. 2010, 53, 5352−5325.
EXPERIMENTAL SECTION
Chemistry. All procedures are fully described in the SI. All tested
compounds were found to have >95% purity determined by HRMS
(HPLC/Q-TOF) analyses.
■
(5) Narwal, M.; Venkannagari, H.; Lehtio, L. Structural basis of
̈
selective inhibition of human tankyrases. J. Med. Chem. 2012, 55,
1360−1367.
(6) Liscio, P.; Camaioni, E.; Carotti, A.; Pellicciari, R.; Macchiarulo,
A. From Polypharmacology to Target Specificity: The Case of PARP
Inhibitors. Curr. Top. Med. Chem. 2013, 13, 2939−2954.
(7) Riffell, J. L.; Lord, C. J.; Ashworth, A. Tankyrase-targeted
therapeutics: expanding opportunities in the PARP family. Nature Rev.
Drug Discovery 2012, 11, 923−936.
Biology. All PARP assays were done by following the BPS PARP
assay kit protocols (BPS Bioscience Inc., San Diego, USA) using
human recombinant proteins. To determine TCF-luciferase reporter
activity, cells were transduced with TOP TCF reporter lentivirus-
expressing firefly luciferase together with renilla luciferase lentivirus
(1:20) used to normalize for infection efficiency. Twenty-four h after
infection, cells were lysed and analyzed utilizing the dual luciferase
reporter assay system. Luciferase reporter activity was calculated by
dividing TOP/RL ratio. For colony growth assay, 5 × 103 DLD-1 cells
were treated daily with increasing concentrations of compound 1, 2, or
12 dissolved in dimethyl sulfoxide. At 10 days, cells were fixed in 10%
methanol/acetic acid solution and stained with 1% crystal violet.
(8) Lehtio, L.; Chi, N. W.; Krauss, S. Tankyrases as drug targets.
̈
FEBS J. 2013, 15, 3576−3593.
(9) Pellicciari, R.; Camaioni, E.; Gilbert, A. M.; Macchiarulo, A.;
Bikker, A. J.; Shah, F.; Bard, J.; Costantino, G.; Gioiello, A.; Robertson,
G. M.; Sabbatini, P.; Venturoni, F.; Liscio, P.; Carotti, A.; Bellocchi, D.;
Cozzi, A.; Wood, A.; Gonzales, C.; Zaleska, M. M.; Ellingboe, J. W.;
Moroni, F. Discovery and characterization of novel PARP-1 inhibitors
endowed with neuroprotective properties: from TIQ-A to HYDAM-
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ASSOCIATED CONTENT
* Supporting Information
■
S
(10) Wahlberg, E.; Karlberg, T.; Kouznetsova, E.; Markova, N.;
Macchiarulo, A.; Thorsell, A. G.; Pol, E.; Frostell, A.; Ekblad, T.; Oncu,
̈
Whole experimental data of the synthesis; computational and
crystallographic studies; biological tests. This material is
available free of charge via the Internet at http://pubs.acs.org.
D.; Kull, B.; Robertson, G. M.; Pellicciari, R.; Schuler, H.; Weigelt, J.
̈
Family-wide chemical profiling and structural analysis of PARP and
tankyrase inhibitors. Nature Biotechnol. 2012, 30, 283−288.
(11) Steck, E. A.; Brundage, P. R. Some s-triazolo[b]pyridazine. J.
Am. Chem. Soc. 1959, 81, 6289−6290.
Accession Codes
Coordinates and structure factors of compound 12 in complex
with the catalytic domain of TNKS-2 have been deposited to
the protein data bank with accession: 4m7b.
(12) Linholter, S.; Kristensen, A. A. B.; Rosenorn, R.; Nielsen, S. E.;
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AUTHOR INFORMATION
Corresponding Author
*Phone: +390755855129. Fax: +390755855161. E-mail:
emidio.camaioni@unipg.it.
■
Author Contributions
The manuscript was written through contributions of all
authors. All authors have given approval to the final version of
the manuscript.
Notes
The authors declare no competing financial interest.
(17) Yanai, M.; Kinoshita, T.; Takeda, S.; Nishimura, M.; Kuraishi, T.
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ABBREVIATIONS USED
■
ARTD, ADP-ribosyltransferase; PARP, poly(ADP-ribose)
polimerase; TNKS, tankyrase; SGC, structural genomic
consortium; PDB, Protein Data Bank
(18) Reich, M.; Fabio, P. F.; Lee, V. J.; Kuck, N. A.; Testa, R. T.
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