8-Biarylchromen-4-one inhibitors of the DNA-dependent protein kinase (DNA-PK)

Article abstract of DOI:10.1016/j.bmcl.2008.07.066

The synthesis and biological evaluation of libraries of 8-biarylchromen-4-ones enabled the elucidation of structure-activity relationships for inhibition of the DNA-dependent protein kinase (DNA-PK), with 8-(3-(thiophen-2-yl)phenyl)chromen-4-one and 8-(3-(thiophen-3-yl)phenyl)chromen-4-one being especially potent inhibitors.

Full text of DOI:10.1016/j.bmcl.2008.07.066

Bioorganic & Medicinal Chemistry Letters 18 (2008) 4885–4890
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
8-Biarylchromen-4-one inhibitors of the DNA-dependent protein kinase
(DNA-PK)
Marine Desage-El Murr a, Celine Cano a, Bernard T. Golding a, Ian R. Hardcastle a, Marc Hummersome b,
Mark Frigerio b, Nicola J. Curtin a, Keith Menear b, Caroline Richardson b, Graeme C. M. Smith b,
Roger J. Griffin a,
*
a Northern Institute for Cancer Research, School of Natural Sciences-Chemistry, Bedson Building, Newcastle University, Newcastle Upon Tyne NE1 7RU, UK
b KuDOS Pharmaceuticals Ltd, 410 Cambridge Science Park, Milton Road, Cambridge CB4 0PE, UK
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis and biological evaluation of libraries of 8-biarylchromen-4-ones enabled the elucidation of
structure–activity relationships for inhibition of the DNA-dependent protein kinase (DNA-PK), with 8-(3-
(thiophen-2-yl)phenyl)chromen-4-one and 8-(3-(thiophen-3-yl)phenyl)chromen-4-one being especially
potent inhibitors.
Received 5 June 2008
Revised 16 July 2008
Accepted 16 July 2008
Available online 20 July 2008
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
DNA-PK
Kinase inhibitor
DNA-repair
Chromenones
Cancer
The cellular response to DNA double-strand break (DSB) forma-
tion is an essential component of normal cell survival, following
exposure to DNA-damaging chemicals and ionising radiation.1
The DNA-dependent protein kinase (DNA-PK), a member of the
phosphatidylinositol (PI) 3-kinase-related kinase (PIKK) family,
plays an important role in DNA DSB repair via the non-homologous
end-joining (NHEJ) pathway.2–4 The ability of DNA-PK to detect
and signal the repair of DNA damage may also protect cancer cells
from the cytotoxic effects of DNA-damaging cancer therapies.
Accordingly, inhibition of DNA-PK has been demonstrated to
potentiate the cytotoxicity of ionising radiation and a number of
DSB-inducing antitumour agents in vitro.5,6 A major objective of
our research is the development of potent and selective DNA-PK
inhibitors, suitable for clinical evaluation as chemo- and radio-sen-
sitisers in the treatment of cancer.
R
O
O
O
N
O
N
O
O
1; R = H
2
4
; R = Ph
S
O
In the absence of suitable structural biology information for
DNA-PK, inhibitor design has been guided by a combination of
homology modelling, utilising the known crystal structure of PI
3-kinase,7 and pharmacophore mapping based on the non-selec-
tive DNA-PK inhibitor LY294002 (1).8 These initial studies enabled
the elucidation of structure–activity relationships (SARs) for DNA-
PK inhibition, and the discovery of potent and selective chrome-
none inhibitors, exemplified by NU7026 (2).9 Encouraged by the
O
O
N
3
potency and kinase-selectivity of 2 and related compounds, a sys-
tematic variation of the substitution pattern on the chromenone
* Corresponding author. Tel./fax: +44 191 222 8591.
E-mail address: r.j.griffin@ncl.ac.uk (R.J. Griffin).
0960-894X/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2008.07.066

4886
M. Desage-El Murr et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4885–4890
of which were found to exhibit potency comparable with the
benchmark DNA-PK inhibitor NU7441 (3).
distal
ring
Our previous studies have utilised the chromenone triflate
derivative 6 as a key reagent for the preparation of 8-substituted
chromenone libraries, employing Suzuki–Miyaura palladium-
catalysed cross-coupling reactions.9–11,13 This strategy was also
amenable for the synthesis of the target substituted 8-biaryl-3-
ylchromenones (5) through analogous reactions on the triflate
building block 11, which was readily accessible from 6 (Scheme 1).
For ease of manipulation, and with a view to preparing 11 on a con-
venient scale, initial studies utilised 3-benzyloxyphenylboronic
acid (7), which was available from 3-bromophenol by standard
methods. Coupling of 7 with 6 gave the chromenone 8 in high
yield, and subsequent removal of the benzyl protecting group by
hydrogenation afforded the required phenol 9.
proximal
ring
R
morpholine
ring
O
N
O
O
chromen-4-one
scaffold
Figure 1. Structural components of 8-(biaryl-3-yl)chromen-4-one inhibitors of
Consistent with the known tolerance of the Suzuki–Miyaura
reaction to a wide range of aryl substituents,14 we subsequently
found that the commercially available 3-hydroxyphenylboronic
acid pinacol ester (10) could be coupled to 6, to give 9 directly
and in comparable yield. Smooth conversion of 9 into 11 was read-
ily achieved with the mild triflating agent N-phenyltriflimide. Prior
to undertaking library synthesis, a coupling reaction was con-
ducted between triflate (11) and phenylboronic acid under previ-
ously optimised reaction conditions (Pd(PPh3)4, K2CO3, dioxane,
reflux). The resulting 8-biphenylchromenone (4) proved identical
to that prepared previously by an alternative method.11 Analogous
cross-coupling reactions were conducted with 11 and 10 commer-
cially available arylboronic acids in a GreenHouseTM reactor (Rad-
leys) to furnish the target arylchromenones (5a–5j) (Scheme 1).
Given the commercial availability of 1,3-phenylene-bis-boronic
acid (12), the possibility of undertaking a ‘one-pot’ double Suzuki–
Miyaura coupling reaction with chromenone triflate (6) and the
appropriate bromoheterocycle (ArBr) was also investigated.15
Again, a model reaction conducted with 12, the triflate 6 and bro-
mobenzene under microwave conditions (5 min at 150 °C) con-
firmed the viability of the reaction, with 4 being obtained in
good yield (60%). This approach was utilised for the preparation
of compounds (5k–5x) as shown in Scheme 1. Replacement of
the 8-phenyl substituent of 4 by a substituted thienyl, thiazolyl
or pyridyl group was achieved by coupling either the chromenone
triflate (6) or the chromenone-8-boronate (13) with the appropri-
ate heterocyclic boronic acid or dihaloheterocycle, respectively, as
DNA-PK.
pharmacophore was undertaken, employing
a solution-phase
multiple-parallel synthesis approach for the preparation of focused
compound libraries.10,11 NU7441 (3) was prominent amongst sev-
eral hits emanating from the library screen, with an independent
resynthesis confirming the high potency (IC50 = 12 nM) and DNA-
PK selectivity of this chromenone. Preclinical antitumour studies
with 3 have demonstrated that this DNA-PK inhibitor potentiates
the in vivo cytotoxicity of topoisomerase inhibitors in a human tu-
mour model.12
Although less potent than NU7441 as a DNA-PK inhibitor, the 8-
(biphenyl-3-yl)chromen-4-one derivative (4; IC50 = 180 nM) was
also identified from the library as a potentially interesting struc-
tural lead. In particular, the non-planar biphenyl motif of 4 offered
the opportunity to probe alternative regions of the ATP-binding
domain of the kinase. In this letter, we report the results of studies
designed to elucidate preliminary SARs for the 8-biarylchrome-
none pharmacophore varied with respect to three parameters as
shown in Figure 1, namely the substituent R on the 30-phenyl
group, and the nature of the ‘proximal’ and ‘distal’ aryl groups.
For this purpose, the core chromen-4-one scaffold and the 2-mor-
pholin-4-yl substituent were retained. A solution-phase multiple-
parallel synthesis approach was employed for the preparation of
libraries of the required 8-biarylchromenones, several members
R
R
R
n
Het
O
X
O
O
R1
O
O
N
O
O
N
O
O
N
i; 7 or 10
vi
v; 12
7; R = OBn, R1 = B(OH)2
5k 5x
-
6; X = OTf
14; R = Br or Cl
vii
13; X = B(OH)2
8; R = OBn
9; R = OH
15a-15v; R = Ar
n = 1 or 2
O
ii
10; R = OH, R1 =
12; R = R1 = B(OH)2
B
O
iii
iv
11; R = OTf
4, 5a-5j; R = Ar
Scheme 1. Reagents and conditions: (i) (6 and 7) cat. Pd(PPh3)4, K2CO3, dioxane, reflux, 88%, (6 and 10) cat. PdCl2(dppf), Cs2CO3, THF, 90%; (ii) H2, Pd/C, MeOH, 93%; (iii)
PhNTf2, Et3N, THF, 77%; (iv) cat. Pd(PPh3)4, K2CO3, ArB(OH)2, dioxane, reflux, 10–30%; (v) (6 and 12) ArBr, cat. PdCl(PPh3)2, Na2CO3, DME/H2O/EtOH (7:3:2), microwave, 150 °C,
5 min, 50–70%; (vi) cat. Pd(PPh3)4, K2CO3, ArB(OH)2 or aryl halide, DMF, 150 °C, 10 min, 40–90%; (vii) cat. Pd(PtBu3)2, K2CO3, ArB(OH)2, DMF, 150 °C, 5 min, 50–60%. [See Tables
1 and 2 for the nature of R in 5a–5x and 15a–15v, respectively.]

M. Desage-El Murr et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4885–4890
4887
Table 1
Chemical structures and DNA-PK inhibitory activity of 2-morpholin-4-yl-8-phenyl-chromen-4-ones bearing a 30-aryl substituent
R
O
O
O
N
a
a
Compound
R
IC50
1.6
(
lM)
Compound
R
IC50 (lM)
Me
1 (LY294002)
H
5l
0.36
1.13
S
Me
S
4
0.18
5m
Me
HO
5a
5b
5c
5d
5e
5f
0.14
0.83
1.48
1.10
1.0
5n
5o
5p
5q
5r
0.64
0.38
0.27
1.82
0.49
3.01
S
MeO
NC
O
O
OHC
HN
S
N
HO
N
H2N
1.9
5s
N
Me
CO2H
N
5g
5h
0.51
0.26
5t
1.23
0.88
N
Me
H2N
H2N
CO2Me
5u
N
N
N
5i
5j
0.018
0.02
5v
5w
5x
0.51
3.05
1.57
S
S
N
N
N
Me
5k
1.07
S
a
IC50 values were determined in accordance with Ref. 9.

4888
M. Desage-El Murr et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4885–4890
Table 2
Chemical structures and DNA-PK inhibitory activity of 2-morpholin-4-yl-chromen-4-ones bearing 8-heteroaryl substituents
Ar
O
O
O
N
a
a
Compound
Ar
IC50
(
lM)
Compound
Ar
IC50 (lM)
N
Ph
15a
0.40
0.09
15l
0.17
1.55
S
S
N
Ph
Ph
Ph
15b
15c
15m
15n
S
S
N
S
N
0.09
2.46
0.34
0.67
1.26
0.30
S
15d
15e
15o
15p
N
S
HN
S
N
S
O
N
15f
15g
15h
0.71
0.37
0.48
15q
15r
15s
1.24
2.37
1.41
N
N
S
Ph
S
N
N
N
S
N
N
S
N
N
S
N
15i
0.24
15t
1.75
S
N

M. Desage-El Murr et al. / Bioorg. Med. Chem. Lett. 18 (2008) 4885–4890
4889
Table 2 (continued)
a
a
Compound
Ar
IC50
(l
M)
Compound
Ar
IC50 (lM)
O
N
N
15j
0.24
15u
0.61
4.45
H
S
S
N
N
N
N
15k
0.51
15v
N
a
IC50 values were determined in accordance with Ref. 9.
shown in Scheme 1. Subsequent Suzuki–Miyaura arylation of the
intermediate 14 with a range of arylboronic acids afforded the tar-
get heterobiaryl-3-ylchromenones (15a–15v). In all cases, reaction
progress was monitored by LC–MS analysis and the products were
purified by semi-preparative HPLC. The DNA-PK inhibitory activity
of both compound libraries is summarised in Tables 1 and 2.
Results and discussion. Our ongoing programme to develop
inhibitors of DNA damage-activated kinases as radio- and chemo-
potentiators in cancer therapy has resulted in the identification
of a number of potent and kinase-selective inhibitors of DNA-PK,
most notably NU7441 (3). That significant inhibitory activity also
resides in the simple 8-biphenylchromenone (4) was surprising,
in light of our previous structure–activity studies indicating that
an extended planar aromatic system at the chromenone 8-position
is a prerequisite for potent DNA-PK inhibition. Previous studies had
also demonstrated that chromenones bearing a biphenyl-3-yl
group (e.g., 4) were more active than the corresponding biphe-
nyl-2-yl and biphenyl-4-yl isomers.11 This is consistent with the
likely disposition of the 3-phenyl group of 4 relative to the termi-
nal aryl ring of the dibenzothiophen-4-yl substituent in 3 within
the ATP-binding pocket. The fact that 4 is approximately 10-fold
more potent than the parent 8-phenylchromenone LY294002 (1,
posal, the weak activity of the analogous pyrrole derivative (5q)
was unexpected. Perhaps not surprisingly, replacement of the pen-
dant phenyl ring of 4 by a pyridyl or pyrimidinyl heterocycle (5u–
5x) resulted in a loss of potency.
The activity of chromenones bearing heteroaryl groups at the 8-
position is summarised in Table 2. Replacement of the 8-phenyl
substituent of 4 by a thiophen-2-yl group (15a and 15b) did not
improve DNA-PK inhibitory activity, although the 4-phenylthio-
phen-2-yl derivative (15b), together with the bithiophene ana-
logue (15c), proved the most potent member of this series
(IC50 = 90 nM). With the exception of the indolyl derivative (15e),
larger heterocyclic groups on the thiophene ring were detrimental
to potency, and where direct comparisons were possible (15a with
15g, and 15c with 15i), replacement of thiophen-2-yl by thiazol-2-
yl was not beneficial. Substitution of the 8-phenyl ring of 4 by a
2- or 4-pyridyl group (15m and 15n) resulted in a reduction in po-
tency, and derivatives bearing other heterocycles on the pyridyl
ring (15o–15v) were all also less active.
In summary, we have identified a novel series of 8-biarylchro-
men-4-ones as inhibitors of DNA-PK that exhibit a range of poten-
cies against the isolated enzyme. Notably, 8-(3-(thiophen-2-
yl)phenyl)chromenone (5i), the most potent of these inhibitors,
was also found to potentiate the cell killing of 2 Gy of ionising radi-
ation by a factor of 1.6 when used at concentration of 500 nM in a
Hela cervical carcinoma cell-based assay.12 This demonstrates that
5i is cell permeable, and that cellular inhibition of DNA-PK is
achievable with this compound at pharmacologically relevant con-
centrations. Overall, the studies described in this letter have fur-
ther elucidated an understanding of SARs for DNA-PK inhibition,
and will provide a platform for ongoing efforts to optimise potency
and in vitro/in vivo activity for this chemotype.
IC50 = 1.6
lM) strongly suggests that the 3-phenyl substituent of
4 is making additional binding interactions within the ATP-binding
domain of DNA-PK. The overall objective of this study was thus to
probe this putative binding interaction further, with a view to
delineating SARs and improving potency.
For the small series of 4-substituted biphenyl-3-ylchromenones
evaluated (5a–5f), it is evident that substitution did not improve
activity, and with the exception of the 4-hydroxy derivative (5a),
which proved to be equipotent with the biphenyl-3-ylchromenone
(4), a 5- to 10-fold reduction in potency was observed (Table 1). A
modest improvement in activity over compounds 5b–5f was ob-
served for the 3,5-disubstituted derivatives (5g and 5h), but both
compounds were less potent than 4. By contrast, replacement of
the 3-phenyl group of 4 by an isosteric thiophen-2-yl (5i) or a thio-
phen-3-yl (5j) substituent improved DNA-PK inhibitory activity
approximately 10-fold, with 5i and 5j exhibiting IC50 values of
18 nM and 20 nM, respectively. Interestingly, the introduction of
a methyl group onto the thiophene ring (5k–5n) proved detrimen-
tal to inhibitory activity, with a 50-fold reduction in potency being
observed for derivatives bearing a methyl group ortho to the het-
eroatom (5k and 5m). This is consistent with the evidence adduced
previously indicating limited steric tolerance at this position. The
high potency exhibited by the thiophene derivatives 5i and 5j
implies that a small electron-rich aryl ring is a prerequisite for
DNA-PK inhibitory activity, and this is supported by the activity
of phenol 5a. However, although the reduced potency of the corre-
sponding 3-furanylchromenones (5o and 5p), the thiazole (5r) and
the imidazole derivatives (5s and 5t) is consistent with this pro-
Acknowledgment
The authors thank Cancer Research UK for financial support.
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