Quinolinone and pyridopyrimidinone inhibitors of DNA-dependent protein kinase

Article abstract of DOI:10.1039/b705095j

8-Substituted 2-morpholin-4-yl-quinolin-4-ones and 9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones with selected aryl and heteroaryl groups as the substituent have been synthesised as potential inhibitors of DNA-dependent protein kinase. A multiple-parallel approach, employing Suzuki cross-coupling methodology, was utilised in the preparation of 8-substituted 2-morpholin-4-yl-quinolin-4-ones. For this purpose 8-bromo-2-morpholin-4-yl- quinolin-4-one was required as an intermediate. This compound was obtained by adapting a literature route in which thermal cyclocondensation of (2-bromoanilino)-morpholin-4-yl-5-methylene-2,2-dimethyl[1,3]dioxane-4,6-dione afforded 8-bromo-2-morpholin-4-yl-quinolin-4-one. A multiple-parallel approach, employing Suzuki cross-coupling methodology, was also utilised to prepare 9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones using 9-hydroxy-2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-one O- trifluoromethanesulfonate as an intermediate. 8-Substituted 2-morpholin-4-yl- quinolin-4-ones and 9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones were both inhibitors of DNA-dependent protein kinase. When the substituent was dibenzothiophen-4-yl, dibenzofuran-4-yl or biphen-3-yl, IC₅₀ values in the low nanomolar range were observed. Interestingly, the pyridopyrimidinones and quinolinones were essentially equipotent with the corresponding 8-substituted 2-morpholin-4-yl-chromen-4-ones previously reported (I. R. Hardcastle, X. Cockcroft, N. J. Curtin, M. Desage El-Murr, J. J. J. Leahy, M. Stockley, B. T. Golding, L. Rigoreau, C. Richardson, G. C. M. Smith and R. J. Griffin, J. Med. Chem., 2005, 48, 7829-7846). The Royal Society of Chemistry.

Full text of DOI:10.1039/b705095j

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PAPER
www.rsc.org/obc | Organic & Biomolecular Chemistry
Quinolinone and pyridopyrimidinone inhibitors of DNA-dependent
protein kinase†
Olivier R. Barbeau,a Celine Cano-Soumillac,a Roger J. Griffin,a Ian R. Hardcastle,a Graeme C. M. Smith,c
Caroline Richardson,c William Clegg,b Ross W. Harringtonb and Bernard T. Golding*a,b
Received 3rd April 2007, Accepted 18th June 2007
First published as an Advance Article on the web 5th July 2007
DOI: 10.1039/b705095j
8-Substituted 2-morpholin-4-yl-quinolin-4-ones and 9-substituted 2-morpholin-4-yl-
pyrido[1,2-a]pyrimidin-4-ones with selected aryl and heteroaryl groups as the substituent have been
synthesised as potential inhibitors of DNA-dependent protein kinase. A multiple-parallel approach,
employing Suzuki cross-coupling methodology, was utilised in the preparation of 8-substituted
2-morpholin-4-yl-quinolin-4-ones. For this purpose 8-bromo-2-morpholin-4-yl-quinolin-4-one was
required as an intermediate. This compound was obtained by adapting a literature route in which
thermal cyclocondensation of (2-bromoanilino)-morpholin-4-yl-5-methylene-2,2-dimethyl[1,3]dioxane-
4,6-dione afforded 8-bromo-2-morpholin-4-yl-quinolin-4-one. A multiple-parallel approach, employing
Suzuki cross-coupling methodology, was also utilised to prepare 9-substituted 2-morpholin-4-yl-
pyrido[1,2-a]pyrimidin-4-ones using 9-hydroxy-2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-one
O-trifluoromethanesulfonate as an intermediate. 8-Substituted 2-morpholin-4-yl-quinolin-4-ones and
9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones were both inhibitors of DNA-dependent
protein kinase. When the substituent was dibenzothiophen-4-yl, dibenzofuran-4-yl or biphen-3-yl, IC50
values in the low nanomolar range were observed. Interestingly, the pyridopyrimidinones and
quinolinones were essentially equipotent with the corresponding 8-substituted 2-morpholin-4-
yl-chromen-4-ones previously reported (I. R. Hardcastle, X. Cockcroft, N. J. Curtin, M. Desage
El-Murr, J. J. J. Leahy, M. Stockley, B. T. Golding, L. Rigoreau, C. Richardson, G. C. M. Smith and
R. J. Griffin, J. Med. Chem., 2005, 48, 7829–7846).
2g and 9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-
ones 3a–3f. The 8-substituent in 2a–2g and the 9-substituent in
Introduction
DNA-dependent protein kinase (DNA-PK), a member of the
3a–3f are selected aryl and heteroaryl groups, some of which are
phosphatidyl inositol (PI3)-kinase related kinase (PIKK) family,
known to confer high potency in 8-substituted 2-morpholin-4-
is a multi-component serine/threonine protein kinase that plays a
yl-chromenones. We show that the 8-substituted 2-morpholin-
key role in the repair of mammalian DNA double-strand breaks
(DSBs).1 Selective DNA-PK inhibitors could be useful for defining
4-yl-quinolin-4-ones 2a–2g and 9-substituted 2-morpholin-
4-yl-pyrido[1,2-a]pyrimidin-4-ones 3a–3f are equipotent with
the role of DNA-PK. In addition, by impeding DNA DSB repair,
the corresponding 8-aryl 2-morpholin-4-yl-chromen-4-ones
(1a–1g).2
DNA-PK inhibitors have potential application as radio- and
chemo-potentiators in the treatment of cancer. 8-Aryl-substituted
2-morpholin-4-yl-chromenones (e.g. 1a–1g) have been shown to
be potent and selective inhibitors of DNA-PK.2 Structure–activity
studies and homology modelling have shown the importance of
the morpholinyl oxygen atom and chromenone carbonyl oxygen
Results and discussion
as hydrogen bond donors to specific groups in the protein target.2,3
To probe whether alterations to the core heterocycle at positions
other than the carbonyl group would be tolerated we have
synthesised 8-substituted 2-morpholin-4-yl-quinolin-4-ones 2a–
Synthesis of 8-substituted 2-morpholin-4-yl-quinolin-4-ones
Previous investigators4,5 have demonstrated the synthesis of
2,6-disubstituted quinolin-4-ones from 5-(bis-methylsulfanyl-
methylene)-2,2-dimethyl[1,3]dioxane-4,6-dione 4, which is easily
prepared from Meldrum’s acid (2,2-dimethyl[1,3]dioxane-4,6-
dione).6 Sequential displacement of the methylsulfanyl groups
of 4 with amines, one of which was p-bromoaniline, to give e.g.
5, followed by heating of this intermediate in diphenyl ether or
polyphosphoric acid gave 2,6-disubstituted quinolin-4-ones, e.g. 6.
The mechanism of the cyclisation step in this remarkable sequence
was proposed to involve an imidoylketene.7 To access 8-substituted
2-morpholin-4-yl-quinolin-4-ones 2a–2g we envisaged using 8-
bromo-2-morpholin-4-yl-quinolin-4-one 7 as an intermediate for
aNorthern Institute for Cancer Research, Bedson Building, Newcastle
University, Newcastle upon Tyne, UK NE1 7RU
bSchool of Natural Sciences-Chemistry, Bedson Building, Newcastle Univer-
sity, Newcastle upon Tyne, UK NE1 7RU
cKuDOS Pharmaceuticals Ltd, 327 Cambridge Science Park, Milton Rd,
Cambridge, UK CB4 4WG. E-mail: b.t.golding@ncl.ac.uk; Fax:+44 (0)191
2226929; Tel: +44 (0)191 2226647
† Electronic supplementary information (ESI) available: Experimental
procedures for compounds 2b–2g and 3b–3f and crystal structures of
compounds 10a and 16. See DOI: 10.1039/b705095j
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Suzuki couplings, with 7 being derived from 4 in an analogous
manner to that described3,4 for 2,6-disubstituted quinolin-4-ones.
We have found an improved methodology leading to substi-
tuted quinolin-4-ones from 4 and have uncovered some further
mechanistic features. Reaction of 4 with 2-bromoaniline proceeded
smoothly in 2,2,2-trifluoroethanol (TFE) solvent at reflux giv-
ing 5-[2-bromoanilino-(methylthio)-methylene]-2,2-dimethyl-4,6-
dione 8 in 88% yield. This step was accomplished even more
efficiently by prior oxidation of 4 with 3-chloroperbenzoic acid
(mCPBA), which enabled reaction with 2-bromoaniline in TFE to
occur at room temperature. Reaction of 8 with 2 mole equiv. of
morpholine in TFE at reflux gave (2-bromoanilino)-morpholin-4-
yl-5-methylene-2,2-dimethyl[1,3]dioxane-4,6-dione 9 (64%). In an
analogous manner to that described for e.g. 5,3,4,8 heating 9 at reflux
in diphenyl ether gave 8-bromo-2-morpholin-4-yl-quinolin-4-one
7 (84%). A one-pot conversion of 8 into 7 was achieved by initial
heating of 8 with 2 mole equivalents of morpholine in diphenyl
10a. When the temperature was raised from 100 ◦C to 200 ◦C
10a was obtained within 30 minutes. Finally heating at 250 ◦C
for 3 hours gave 7. The experiments performed (summarised in
Scheme 1) show that efficient formation of 7 from 8 in a one-pot
system occurs better with 2 mole equiv. rather than 1 mole equiv. of
morpholine and that although 9 is an intermediate, the immediate
precursor of 7 is 10a/10b.
Suzuki reactions were performed on compound 7 leading
to 8-substituted 2-morpholin-4-yl-quinolin-4-ones 2a–2g. The
couplings were optimised using a 24-reaction GreenHouseTM
,
which led to the selection of potassium carbonate as base with
tetrakis(triphenylphosphine)palladium(0) as catalyst in dioxane
solvent.
Synthesis of 9-substituted
2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones
◦
ether a◦t 90 C for 18 hours, followed by raising the temperature
A
key intermediate for the synthesis of 9-substituted 2-
to 260 C for 3 hours.8 In an attempt to conduct this sequence at
lower temperature we found that heating 8 with 2 mole equiv. of
morpholine at 90 ◦C for 18 hours and then at 200 ◦C for 1 hour led
to a different product, which was identified as 3-(2-bromoanilino)-
3-morpholin-4-yl-propionamide morpholinamide 10a by crystal
structure analysis‡ (Fig. 1, Supplementary Information†). Heating
10a in diphenyl ether at 250 ◦C gave 7 (47%).
morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones 3a–3f was 2,9-
dihydroxypyrido[1,2-a]pyrimidin-4-one 12, which was prepared
by heating 2-amino-3-hydroxypyridine with either diethyl mal-
onate or di-(2,4,6-trichlorophenyl) malonate (Scheme 2).9,10 This
method followed closely the published syntheses of 12, the
parent 2-hydroxypyrido[1,2-a]pyrimidin-4-one 13 and related
compounds.9,11,12 The 2-hydroxyl group of 12 was selectively
chlorinated by reaction with an excess of phosphorus oxy-
chloride to afford 2-chloro-9-hydroxypyrido[1,2-a]pyrimidin-4-
one 14. Displacement of the chloro function of 14 with mor-
pholine gave 9-hydroxy-2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-
4-one 15, quantitatively. Compound 15 has been reported in the
patent literature as a precursor to inhibitors of phosphoinositol
3-kinase, but without details of its preparation.13 Finally, 15 was
converted into the corresponding 9-O-triflate 16 by reaction with
trifluoromethanesulfonic (triflic) anhydride. The structure of 16
was unambiguously determined as 9-hydroxy-2-morpholin-4-yl-
pyrido[1,2-a]pyrimidin-4-one O-trifluoromethanesulfonate by an
X-ray crystal structure analysis‡ (Figs. 2 and 3, Supplemen-
tary Information†). 9-Substituted 2-morpholin-4-yl-pyrido[1,2-
a]pyrimidin-4-ones 3a–3f were obtained from 16 by Suzuki reac-
tions employing the same optimised combination of base, catalyst
and solvent [K2CO3, tetrakis(triphenylphosphine)palladium(0)
and dioxane], as described above for the preparation of 2a–2g.
These experiments show that whilst 7 can undoubtedly be
derived directly from 9 without addition of further morpholine,3,4,8
there is an alternative pathway to 7 via 10a. The latter may proceed
via an electrocyclic process, involving the tautomer 10b, which
leads to an intermediate that expels morpholine. The fact that 7
can also arise by thermolysis of 10a/10b raises the question as
to whether the precursor of 7 in the one-pot reaction described
using 2 mole equivalents of morpholine is 9 or 10a/10b. Actually,
heating 8 with 1 mole equivalent of morpholine in diphenyl ether at
100 ◦C gave 9, from which 7 was derived by raising the temperature
to 250 ◦C. However, under these conditions a by-product was
11. Monitoring a one-pot reaction by HPLC showed that 8 with
2 mole equivalents of morpholine at 100 ◦C gave exclusively 9
after 1 hour. Heating the mixture at 150 ◦C for 16 hours afforded
‡ CCDC reference numbers 647931 and 647932. For crystallographic data
in CIF or other electronic format see DOI: 10.1039/b705095j
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Scheme 1 Reagents and conditions: (i) mCPBA, DCM, rt; (ii) 2-bromoaniline, TFE, rt; (iii) morpholine (2 equiv.), TFE or Ph2O, 90 ◦C; (iv) Ph2O, 260 ◦C;
(v) morpholine (2 equiv.), Ph2O, 200 ◦C; (vi) Ph2O, 250 ◦C; (vii) polyphosphoric acid, 130 ◦C; (viii) cat. Pd(PPh3)4, K2CO3, ArB(OH)2, dioxane, 90 ◦C.
Scheme 2 Reagents and conditions: (i) 3-hydroxy-2-aminopyridine, PhBr, reflux; (ii) POCl3, reflux; (iii) morpholine, ethanol, reflux; (iv) Tf2O, NEt3,
DCM, −78 ◦C; (v) ArB(OH)2, K2CO3, cat. Pd(PPh3)4, dioxane, 90 ◦C (see Scheme 1 for definition of Ar groups in 3a–3f).
An alternative route to 3a–3f was explored via 9-bromo-2-
hydroxypyrido[1,2-a]pyrimidin-4-one 17. 2-Amino-3-bromopyri-
dine was prepared from 2-aminopyridine as described.14 However,
reaction of 2-amino-3-bromopyridine with diethyl malonate gave
only 10% yield of 17. In another approach 2-phenylacetonitrile
was converted into 2-amino-3-phenylpyridine in the manner
described.15 Heating this compound with diethyl malonate gave
only 21% 2-hydroxy-9-phenylpyrido[1,2-a]pyrimidin-4-one 18 and
so this route was also discontinued.
Inhibition of DNA-dependent protein kinase by 8-substituted
2-morpholin-4-yl-quinolin-4-ones and 9-substituted
2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones
Compounds 2a–2g and 3a–3f were evaluated as inhibitors
of DNA-PK using the published assay,16 and the results are
summarised in Table 1. The DNA-PK inhibitory activities of
the analogous 8-substituted chromen-4-ones are included for
comparison. The results show that replacing the 1-oxygen of
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Table 1 Inhibition of DNA-PK by 8-substituted 2-morpholin-4-yl-quinolin-4-ones (2a–2g) and 9-substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-
4-ones (3a–3f)
DNA-PK inhibitory activity (IC50)/nMa
Ar
2a
4900
3a
733
1ab
1600
2b
2c
1200
1260
3b
3c
390
1b
1c
300
800
1420
2d
49
3d
89
1d
176
2e
2f
28
28
3e
3f
59
13
—
1e
1f
40
12
2g
2430
—
1g
600
a DNA-PK inhibitory activity was determined as described in reference 16. The values for the 8-substituted chromenones are taken from reference 2.
b LY294002.
8-aryl-2-morpholin-4-yl-chromenones with NH (2a–2g) does not
significantly affect the biological activity, indicating that neither
the 1-oxygen nor 1-NH make significant productive hydrogen
bonding interactions with DNA-PK. This analysis assumes that
the tautomer indicated for the 8-substituted 2-morpholin-4-yl-
quinolin-4-ones 2a–2g is preferred over the alternative 19. This
statement is based on analogy with quinolin-4-one17 and similar
heterocycles18 for which the pyridone-like tautomer is preferred
in polar solvents, although the population of the enol tautomer
may be increased by appropriate hydrogen bonding interactions.19
Replacing the 5-C of 8-aryl-2-morpholin-4-yl-chromenones with
N (3a–3f) also had no significant effect on the biological activity.
As with the 8-aryl-2-morpholin-4-yl-chromenones, the optimum
8- or 9-substituents were dibenzothiophen-4-yl, dibenzofuran-4-yl
and biphen-3-yl, all of which gave IC50 values in the low nanomolar
range.
Conclusions
This paper presents efficient methods for preparing a variety of
8-substituted 2-morpholin-4-yl-quinolin-4-ones and 9-substituted
2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones using the Suzuki
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reaction on the readily available precursors 8-bromo-2-morpholin-
4-yl-quinolin-4-one and 9-hydroxy-2-morpholin-4-yl-pyrido[1,2-
a]pyrimidin-4-one O-trifluoromethanesulfonate, respectively.
Quinolin-4-ones 2a–2g and pyrido[1,2-a]pyrimidin-4-ones 3a–3f
with 8-/9-substituents dibenzothiophen-4-yl, dibenzofuran-4-yl
and biphen-3-yl were especially potent against DNA-dependent
protein kinase. These and previous results2,3,16,18 indicate that three
structural motifs are principally associated with low nanomolar
(IC50) potency against DNA-PK: i) a bicyclic heterocyclic core
containing a carbonyl group; ii) a morpholin-4-yl substituent
on the core and iii) a bulky aromatic/heteroaromatic system
(e.g. dibenzothiophen-4-yl) attached to the core. There is an
optimum substitution pattern: 2-morpholinyl and 4-one in all
cases; 8-aryl (or heteroaryl) for chromenones 1 and quinolinones
2 {or the corresponding 9-aryl (or heteroaryl) for pyrido[1,2-
a]pyrimidinones 3}. Remarkably, there is little difference in
potency between the three structurally distinct heterocyclic cores
described in this paper. We are continuing our studies with the
aim of obtaining a clinical candidate from the structural types
studied or closely related analogues or derivatives thereof.
8-Bromo-2-morpholin-4-yl-1H-quinolin-4-one (7)
Method A. A solution of 5-[(2-bromoanilino)morpholin-
4-yl-methylene]-2,2-dimethyl[1,3]dioxane-4,6-dione (103 mg,
0.25 mmol) in diphenyl ether (0.7 mL) was stirred and heated at
reflux for 3 h. After cooling, petrol was added and the precipitated
product was collected by suction. The crude product was purified
by medium pressure chromatography using DCM–methanol (95 :
5 v/v) as eluent. The title compound was obtained as a brown
solid (65 mg, 84%).
Method B. A solution of 5-[2-bromoanilino(methylthio)-
methylene]-2,2-dimethyl-4,6-dione (163 mg, 0.44 mmol) and mor-
pholine (40 lL, 0.44 mmol) in diphenyl ether (2.5 mL) was stirred
and heated at 90 ◦C for 18 h. The temperature was raised to
260 ◦C over 3 h. After cooling, petrol was added and the product
was collected by suction. The crude product was purified by
chromatography (see Method A) to give the title compound as
a brown solid (87 mg, 65%).
Method C. A solution of 3-(2-bromophenylimino)-1,3-di-
morpholin-4-yl-propan-1-one (200 mg, 0.50 mmol) in diphenyl
ether (1 mL) was stirred and heated to 250 ◦C for 1 h. After work-
up as described in Method A the title compound was obtained
as a brown solid (84 mg, 54%): Rf = 0.25 (MeOH–DCM 1 : 19);
Experimental
◦
mp: 96 C; IR (cm−1) 3395, 2959, 2849, 1617, 1577, 1487, 1421,
Materials and methods
1384, 1327, 1263, 1229, 1188, 1152, 1111, 1066, 999, 902, 785; 1H
NMR, (300 MHz, CDCl3) d 3.72 (4H, t, J = 5.0 Hz, 2 × CH2N-
morpholine), 3.75 (4H, t, J = 5.0 Hz, 2 × CH2O–morpholine),
5.95 (1H, s, H-3), 7.04 (1H, dd, J = 8.0 and 8.0 Hz, H-6), 7.69
(1H, dd, J = 1.3 and 8.0 Hz, H-7), 8.09 (1H, dd, J = 1.3 and
8.0 Hz, H-5); 13C NMR, (75 MHz, CDCl3) d 46.4, 66.6, 92.5,
114.5, 123.0, 123.5, 124.7, 134.5, 138.0, 156.1, 172.6; MS (ES+)
m/z 308.98, 310.98. HRMS calcd for C13H1379BrN2O2 [M + H]+
309.0233, found 309.0233.
Chemicals and solvents were obtained from reputable suppliers
(for further details see ref. 20). Solvents were either dried by stan-
dard techniques or purchased as anhydrous. Triethylamine was
dried by distillation from calcium hydride and stored over potas-
sium hydroxide, under nitrogen. Chromatography, spectroscopy
and combustion analyses were performed as described.2,3,16,20
Where the inclusion of dichloromethane (DCM) was necessary
to obtain a good agreement between calculated and found values
for C, H and N, the requisite amount of DCM was observed
in the 1H NMR spectrum of the compound concerned. A
GreenHouseTM 24-position reactor (Radley’s, Cambridge, UK)
was used for Suzuki reactions, which were all run under an argon
atmosphere.
5-[2-Bromoanilino(methylthio)methylene]-2,2-dimethyl-4,6-dione
(8)
Method A. A solution of 5-(bis-methylsulfanylmethylene)-
2,2-dimethyl[1,3]dioxane-4,6-dione (900 mg, 3.6 mmol) and 2-
bromoaniline (624 mg, 3.6 mmol) in 2,2,2-trifluoroethanol
(3.6 mL) was stirred and heated at reflux for 18 h. After cooling,
the solvent was removed to afford crude product that was
recrystallised from methanol to yield the title compound (1.19 g,
88%) as white crystals.
5-(Bis-methylsulfanylmethylene)-2,2-dimethyl[1,3]dioxane-4,6-
dione (5)
To 2,2-dimethyl-1,3-dioxane-4,6-dione (Meldrum’s acid) (4.09 g,
28.4 mmol) in DMSO (14 mL) were added triethylamine (7.9 mL,
56.8 mmol) and carbon disulfide (1.7 mL, 28.4 mmol) in quick
succession. The mixture was stirred vigorously at room temper-
ature for 1 h. After cooling (ice-bath), iodomethane (3.5 mL,
56.8 mmol) was slowly added to the reaction mixture. When the
addition was complete the mixture was allowed to warm up to
room temperature and was stirred for a further 4 h before being
diluted with ice-cold water (25 mL). Trituration of the mixture
precipitated the product, which was filtered off and washed with
petrol. The crude product (2.76 g, 45%) was obtained as a yellow
solid that was pure e◦nough for use in subsequent reactions: mp:
118 ◦C (lit.6 116–118 C); IR (cm−1) 3728, 1668, 1373, 1302, 1264,
1199; 1H NMR, (300 MHz, CDCl3) d 1.54 (6H, s, 2 × CH3); 2.58
(6H, s, 2 × CH3S); 13C NMR, (75 MHz, CDCl3) d 21.9; 27.2; 103.3;
160.3; 194.0.
Method B. A solution of 5-(bis-methylsulfanylmethylene)-2,2-
dimethyl[1,3]dioxane-4,6-dione (100 mg, 0.40 mmol) and mCPBA
(75% pure, 102 mg, 0.44 mmol) in DCM (2 mL) was stirred at
room temperature for 1 h. The solvent was removed and the
residual solid was taken up in TFE (2 mL). 2-Bromoaniline
(45 lL, 0.4 mmol) was added and the mixture was stirred at room
temperature for 1.5 h. The solvent was removed and the residue
was taken up in DCM. The resulting solution was washed with an
aqueous saturated sodium bicarbonate solution, dried (MgSO4),
and the DCM was removed. The crude product was purified by
medium pressure chromatography using DCM as eluent. The title
compound was obtained as a white solid (145 mg, 98%): Rf = 0.31
(DCM); mp: 159 ◦C; IR (cm−1) 2990, 1706, 1653, 1535, 1370, 1199;
1H NMR, (300 MHz, CDCl3) d 1.69 (6H, s, 2 × CH3), 2.15 (3H, s,
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CH3S), 7.18 (1H, ddd, J = 2.0, 8.0 and 8.0 Hz, H-4ꢀ), 7.35 (2H,
m, H-5ꢀ and H-6ꢀ), 7.61 (1H, dd, J = 2.0 and 8.0 Hz, H-3ꢀ), 12.51
(1H, s, NH); 13C NMR, (75 MHz, CDCl3) d 18.8, 26.9, 87.5, 103.3,
120.5, 127.8, 128.5, 129.9, 133.6, 136.9, 163.9, 178.7; MS (ES−)
m/z 369.97, 371.98; HRMS calcd for C14H1479BrNO4S [M + H]+
371.9900, found 371.9903.
7.12 (1H, dd, J = 8.0 and 7.9 Hz, H-6), 7.73 (1H, d, J = 7.9 Hz,
H-7), 8.17 (1H, d, J = 8.0 Hz, H-5), 8.46 (1H, s, NH); 13C NMR,
(75 MHz, CDCl3) d 15.0, 106.7, 112.0, 124.7, 124.8, 125.9, 135.2,
138.0, 168.0, 173.0; MS (ES+) m/z 269.91, 271.91; HRMS calcd
for C10H879BrNOS [M + H]+ 269.9583, found 269.9582.
2,9-Dihydroxypyrido[1,2-a]pyrimidin-4-one (12)
5-[(2-Bromoanilino)morpholin-4-yl-methylene]-2,2-
dimethyl[1,3]dioxane-4,6-dione (9)
Method A. A mixture of di-(2,4,6-trichlorophenyl) malonate21
(17.3 g, 37.5 mmol) and 3-hydroxy-2-aminopyridine (4.12 g,
37.5 mmol) in bromobenzene (37 mL) was heated at reflux for 3 h.
After cooling, the mixture was filtered and the resulting solid was
washed with ethanol. The solid was solubilised in a 1 M sodium
hydroxide solution and acetic acid was added to precipitate the
product (6.53 g, 98%) as a pale yellow solid.
A
solution of 5-[2-bromoanilino(methylthio)methylene]-2,2-
dimethyl-4,6-dione (234 mg, 0.6 mmol) and morpholine (110 lL,
1.3 mmol) in 2,2,2-trifluoroethanol (1 mL) was stirred and heated
at reflux for 18 h. After cooling, the solvent and excess of
morpholine were removed. The residual solid was recrystallised
from methanol to yield the title compound as white crystals
Method B. To 3-hydroxy-2-aminopyridine (5.21 g, 47.3 mmol)
in o-xylene (16 mL) was added diethyl malonate (10.8 mL,
71 mmol) and the mixture was heated at 170 ◦C for 18 h. The
reaction was worked up as described in Method A to give the
product (6.0 g, 82%) as a pale yellow solid: Rf = 0.11 (MeOH–
DCM, 3 : 17); mp: 320 ◦C (dec.) (lit.9: 310 ◦C); kmax (EtOH)/nm
252; IR (cm−1) 2862, 1688, 1564, 1374, 1295, 1102, 783; 1H NMR,
(300 MHz, DMSO) d 5.22 (1H, s, H-3), 7.12 (1H, t, J = 7 Hz,
H-7), 7.27 (1H, d, J = 8 Hz, H-8), 8.43 (1H, d, J = 7 Hz, H-6);
13C NMR, (75 MHz, DMSO) d 103.3, 116.5, 117.1, 119.0, 144.0,
148.8, 157.3, 157.5; MS (ES+) m/z 179.02.
◦
(158 mg, 64%): Rf = 0.05 (DCM); mp: 212–213 C; IR (cm−1)
1627, 1342, 1305, 1100, 1022, 934; 1H NMR, (300 MHz, CDCl3)
d 1.77 (6H, s, 2 × CH3), 3.24 (4H, dd, J = 4.5 and 4.9 Hz, 2 ×
CH2N-morpholine), 3.66 (4H, dd, J = 4.5 and 4.9 Hz, 2 × CH2O-
morpholine), 7.18 (2H, m, H-4ꢀ and H-6ꢀ), 7.40 (1H, dd, J = 8.0
and 8.0 Hz, H-5ꢀ), 7.69 (1H, dd, J = 1.4 and 8.0 Hz, H-3ꢀ), 9.62
(1H, s, NH); 13C NMR, (75 MHz, CDCl3) d 26.8, 51.1, 65.6, 87.5,
102.8, 120.5, 127.2, 128.9, 129.0, 134.9, 138.5, 164.9, 178.7; MS
(ES−) m/z 409.01, 411.01.
3-(2-Bromophenylimino)-1,3-dimorpholin-4-yl-propan-1-one (10a)
2-Chloro-9-hydroxypyrido[1,2-a]pyrimidin-4-one (14)
A
solution of 5-[2-bromoanilino(methylthio)methylene]-2,2-
dimethyl-4,6-dione (8.86 g, 23.8 mmol) and morpholine (4.15 mL,
47.6 mmol) in diphenyl ether (40 mL) was stirred and heated at
90 ◦C for 18 h. The temperature was raised to 200 ◦C over 1 hour.
After cooling, petrol was added and the precipitated solid was
collected by suction. The crude product was purified by medium
pressure chromatography using DCM–methanol (95 : 5, v/v) as
eluent to give the title compound as white crystals (5.09 g, 54%):
Rf = 0.27 (MeOH–DCM 1 : 19); mp: 159–160 ◦C; IR (cm−1) 2950,
2901, 2854, 1631, 1600, 1448, 1413, 1358, 1298, 1253, 1222, 1166,
Method A. 2,9-Dihydroxypyrido[1,2-a]pyrimidin-4-one (1.071 g,
6.02 mmol) was cautiously dissolved in phosphorus(III) oxychlo-
ride (7.5 mL, 80.4 mmol) to give a solution that was heated at
reflux for 48 h. After cooling, the reaction mixture was poured
carefully into ice-cold water (100 mL) and the pH was adjusted
to 7 by addition of a saturated solution of sodium carbonate.
The aqueous layer was extracted with DCM. The combined
organic layers were dried (MgSO4) and the solvent was removed
to yield a brown solid. The crude product was purified by medium
pressure chromatography using DCM as eluent to furnish the title
compound as a white solid (712 mg, 60%).
1
1105, 1064, 1024, 964, 950, 912; H NMR, (300 MHz, CDCl3) d
2.98 (2H, m, CH2-morpholine), 3.18 (2H, m, CH2-morpholine),
3.31 (2H, m, CH2-morpholine), 3.45 (2H, m, CH2-morpholine),
3.53 (6H, m, 2 × CH2-morpholine and CH2), 3.71 (4H, m, 2 ×
CH2 morpholine), 6.70 (1H, dd, J = 7.8 Hz and 1.5 Hz, H-6), 6.77
(1H, ddd, J = 7.5, 7.5 and 1.5 Hz, H-4), 7.11 (1H, ddd, J = 7.8, 7.5
Method B. 3-Hydroxy-2-aminopyridine (444 mg, 4.03 mmol)
and di-(2,4,6-trichlorophenyl) malonate (1.865 g, 4.03 mmol)
were cautiously dissolved in phosphorus(III) oxychloride (3.2 mL,
34.3 mmol). The reaction was continued as described in Method
A to give crude product that was purified by medium pressure
chromatography using DCM–methanol (98 : 2 v/v) as eluent. The
title compound was obtained as a white solid (251 mg, 32%): Rf =
and 1.4 Hz, H-5), 7.45 (1H, dd, J = 7.5 Hz and 1.4 Hz, H-3); 13
C
NMR, (75 MHz, CDCl3) d 33.7, 42.7, 46.4, 46.4, 66.6, 67.0, 67.1,
117.5, 123.8, 123.9, 128.4, 133.1, 149.4, 154.9, 166.4; MS (ES+)
m/z 396.01, 398.01; HRMS calcd for C17H2279BrN3O3 [M + H]+
396.0917, found 396.0914.
◦
0.34 (MeOH–DCM, 1 : 19); mp: 162 C; IR (cm−1) 3103, 1684,
1630, 1511, 1458, 1297, 1105; 1H NMR, (300 MHz, CDCl3) d 6.4
(1H, s, H-3), 7.11 (1H, dd, J = 7.3 and 7.4 Hz, H-7), 7.25 (1H,
d, J = 7.4 Hz, H-8), 8.51 (1H, d, J = 7.3 Hz, H-6); 13C NMR,
(75 MHz, CDCl3) d 103.3, 116.5, 117.1, 119.0, 144.0, 148.8, 157.3,
157.5; MS (ES+) m/z 196.93 M+; HRMS calcd for C8H5ClN2O2
[M + H]+ 197.0112, found 197.0114.
8-Bromo-2-methylsulfanyl-1H-quinolin-4-one (11)
To polyphosphoric acid (160 mg, 1.13 mmol) stirred at
◦
130 C was added 5-[2-bromoanilino(methylthio)methylene]-2,2-
dimethyl-4,6-dione (191 mg, 0.54 mmol). Work-up as in method B
for the preparation of 8-bromo-2-morpholin-4-yl-1H-quinolin-4-
one gave the title compound as a yellow solid (118 mg, 0.44 mmol,
82%): Rf = 0.34 (MeOH–DCM 1 : 19); mp: 108–110 ◦C; IR (cm−1)
9-Hydroxy-2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-one (15)
1
3356, 2167, 2021, 1616, 1551, 1492, 1426, 1334, 1097, 933; H
2-Chloro-9-hydroxypyrido[1,2-a]pyrimidin-4-one (142 mg,
0.72 mmol) and morpholine (315 lL, 3.6 mmol) were taken
NMR, (300 MHz, CDCl3) d 2.53 (3H, s, CH3), 6.21 (1H, s, H-3),
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up in ethanol (5 mL) and the resulting solution was heated at
reflux for 18 h with vigorous stirring. The solvent was removed
to give a yellow solid that was recrystallised from ethanol to
afford the title compound as white crystals (174 mg, 97%): Rf =
0.27 (MeOH–DCM 1 : 19); mp: 245 ◦C; kmax (EtOH)/nm 267;
2-Morpholin-4-yl-8-phenyl-4-yl-1H-quinolin-4-one (2a). Phe-
nylboronic acid (59 mg, 0.49 mmol) and 8-bromo-2-morpholin-
4-yl-1H-quinolin-4-one (100 mg; 0.32 mmol) gave crude product
that was purified by medium pressure chromatography using ethyl
acetate–methanol (98 : 2 v/v) as eluent. Further purification was
achieved by HPLC (aqueous methanol as eluent) to afford the title
compound as a cream solid (65 mg, 65%): Rf = 0.11 (EtOAc); mp:
109–110 ◦C (dec.); kmax (EtOH)/nm 251; IR (cm−1) 3410, 2954,
2846, 1613, 1576, 1488, 1422, 1364, 1227, 1111, 996, 903, 800,
1
IR (cm−1) 3302, 1690, 1644, 1551, 1427, 1224, 1110; H NMR,
(500 MHz, CDCl3) d 3.56 (4H, m, 2 × NCH2-morpholine), 3.75
(4H, m, 2 × OCH2-morpholine), 5.55 (1H, s, H-3), 6.80 (1H, dd,
J = 7.0 and 7.0 Hz, H-7), 7.02 (1H, dd, J = 7.0 Hz and 1.3 Hz,
H-8), 7.33 (1H, s, OH), 8.37 (1H, dd, J = 7.0 Hz and 1.3 Hz,
H-6); 13C NMR, (125 MHz, CDCl3) d 45.3, 67.0, 82.2, 113.5,
114.2, 119.1, 143.0, 147.5, 159.0, 161.0; MS (ES+) m/z 248.08;
Anal. Calcd for 0.8 mol C12H12N3O3 + 0.2 mol H2O: C, 57.25, H,
5.40, N, 16.69. Found: C, 57.58, H, 5.12, N, 16.58.
1
700; H NMR, (300 MHz, CDCl3) d 3.05 (4H, m, 2 × NCH2-
morpholine), 3.68 (4H, m, 2 × OCH2-morpholine), 5.68 (1H, s,
H-3), 7.26 (1H, dd, J = 7.0 and 7.0 Hz, H-6), 7.40 (3H, m), 7.51
(2H, m), 7.82 (1H, m, H-7), 8.23 (1H, d, J = 7.0 Hz, H-5); 13C
NMR, (75 MHz, CDCl3) d 46.7, 66.4, 92.9, 123.4, 125.8, 129.6,
132.5, 135.6, 137.0, 154.3, 178.9; MS (ES+) m/z 307.13; Anal.
Calcd for 0.73 mol C19H18N2O2 + 0.27 mol DCM: C, 69.21, H,
5.73, N, 8.18. Found: C, 69.21, H, 5.60, N, 8.58%.
9-Hydroxy-2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-one
9-O-triflate (16)
A 3-necked flask equipped with a thermometer, addition funnel
and magnetic stirrer, was charged with 9-hydroxy-2-morpholin-
4-yl-pyrido[1,2-a]pyrimidin-4-one (2.11 g, 8.5 mmol) in DCM
(70 mL). The mixture was cooled to −30 ◦C and triethylamine
(3.6 mL, 25.6 mmol) was added. After 5 min, trifluoromethanesul-
fonic anhydride (2.1 mL, 12.8 mmol) in DCM (10 mL) was added
dropwise to the reaction mixture, via the funnel. The addition took
place over 30 min, and the temperature of the reaction mixture
9-Substituted 2-morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-ones
(3a–3f)
Typical procedure. A similar procedure to that detailed above
for 8-substituted 2-morpholin-4-yl-quinolin-4-ones was also used
for reactions of arylboronic acids with 9-hydroxy-2-morpholin-4-
yl-pyrido[1,2-a]pyrimidin-4-one 9-O-triflate. The crude product
was purified as described below for compound 3a and in the
Supplementary Information† for compounds 3b–3f.
◦
was kept below −20 C during this time. After 3 h, the reaction
mixture was washed with a saturated solution of sodium carbonate
(50 mL) and extracted with DCM (3 × 30 mL). The combined
organic layers were dried (MgSO4) and the solvent was removed
to yield a brown solid. The crude product was purified by medium
pressure chromatography using DCM as eluent to furnish the title
compound as an orange solid (2.91 g, 90%): Rf = 0.42 (MeOH–
2-Morpholin-4-yl-9-phenyl-4-yl-pyrido[1,2-a]pyrimidin-4-one
(3a). Phenylboronic acid (36 mg, 0.26 mmol) and 9-hydroxy-2-
morpholin-4-yl-pyrido[1,2-a]pyrimidin-4-one 9-O-triflate (100 mg,
0.26 mmol) gave crude product that was recrystallised from ethyl
acetate. The title compound was obtained as a brown solid (71 mg,
88%): Rf = 0.22 (MeOH–DCM 1 : 19); mp: 187–188 ◦C; kmax
(EtOH)/nm 275; IR (cm−1) 2862, 1681, 1539, 1494, 1427, 1219,
◦
DCM, 1 : 19); mp: 146–147 C; kmax (EtOH)/nm 271; IR (cm−1)
1
1705, 1644, 1551, 1189, 1112, 939, 769; H NMR, (300 MHz,
1
1106, 756; H NMR, (300 MHz, CDCl3) d 3.47 (4H, m, 2 ×
CDCl3) d 3.56 (4H, m, 2 × NCH2-morpholine), 3.71 (4H, m, 2 ×
OCH2-morpholine), 5.53 (1H, s, H-3), 6.80 (1H, dd, J = 8.0 and
7.0 Hz, H-7), 7.46 (1H, dd, J = 8.0 Hz and 1.3 Hz, H-8), 8.79 (1H,
dd, J = 7.0 Hz and 1.3 Hz, H-6); 13C NMR, (75 MHz, CDCl3)
d 45.2, 66.9, 81.8, 110.2, 112.6, 116.9, 121.1, 125.3, 127.9, 128.1,
141.5, 145.8, 158.1, 160.4; MS (ES+) m/z 380.16; Anal. Calcd for
C13H12F3N3O5S: C, 41.16, H, 3.19, N, 11.08. Found: C, 41.29, H,
2.88, N, 10.93%.
NCH2-morpholine), 3.65 (4H, m, 2 × OCH2-morpholine), 5.57
(1H, s, H-3), 6.91 (1H, dd, J = 7.0 and 7.1 Hz, H-7), 7.33–
7.41 (3H, m), 7.55–7.59 (3H, m), 8.90 (1H, dd, J = 7.0 Hz and
1.6 Hz, H-6); 13C NMR, (75 MHz, CDCl3) d 44.9, 66.9, 81.3, 112.9,
127.5, 128.3, 128.6, 130.2, 136.0, 136.7, 137.2, 149.5, 159.5, 160.9;
MS (ES+) m/z 308.14; Anal. Calcd for 2.0 mol C19H18N2O2S +
0.1 mol H2O: C, 68.34, H, 5.73, N, 13.28. Found: C, 68.72, H, 5.17,
N, 13.08%.
8-Substituted 2-morpholin-4-yl-quinolin-4-ones (2a–2g)
Acknowledgements
Typical procedure. The arylboronic acid (1 mol equiv.) and
potassium carbonate (1.9 mol equiv.) were dispersed in diox-
ane (4 mL per mmol boronic acid). Concurrently, 8-bromo-2-
morpholin-4-yl-1H-quinolin-4-one (0.67 mol equiv.) and tetrakis-
(triphenylphosphine)palladium (3 mol%) were taken up in dioxane
(6 mL per mmol bromo compound). Argon was bubbled into both
mixtures, which were sonicated for 15 min. The mixtures were
combined, stirred and heated at 95 ◦C for 48 h. After cooling,
DCM (10 mL per mmol boronic acid) was added. The resulting
solution was washed with water and brine (10 mL each per mmol
boronic acid), dried (MgSO4), and the solvent was removed. The
crude product was purified as described below for compound 2a
and in the Supplementary Information† for compounds 2b–2g.
The authors thank Cancer Research UK for support of this
research, EPSRC for funding of crystallography, including the
UK National Crystallography Service, and CCLRC for access to
synchrotron facilities. We are grateful to the EPSRC National
Mass Spectrometry Service Centre at Swansea University for
selected mass spectra.
References
1 G. C. M. Smith and S. P. Jackson, Genes Dev., 1999, 13, 916.
2 J. J. J. Leahy, B. T. Golding, R. J. Griffin, I. R. Hardcastle, C.
Richardson, L. Rigoreau and G. C. M. Smith, Bioorg. Med. Chem.
Lett., 2004, 14, 6083–6087; I. R. Hardcastle, X. Cockcroft, N. J. Curtin,
2676 | Org. Biomol. Chem., 2007, 5, 2670–2677
This journal is
The Royal Society of Chemistry 2007
©

View Article Online
M. Desage El-Murr, J. J. J. Leahy, M. Stockley, B. T. Golding, L.
Rigoreau, C. Richardson, G. C. M. Smith and R. J. Griffin, J. Med.
Chem., 2005, 48, 7829–7846.
3 J. J. Hollick, L. J. M. Rigoreau, C. Cano-Soumillac, X. Cockcroft,
N. J. Curtin, M. Frigerio, B. T. Golding, S. Guiard, I. R. Hardcastle,
I. Hickson, M. Hummersone, K. A. Menear, N. M. B. Martin, I.
Matthews, D. R. Newell, R. Ord, C. Richardson, G. C. M. Smith and
R. J. Griffin, J. Med. Chem., 2007, 50, 1958–1972.
12 H. Reimlinger, F. Billiau, M. A. Peiren and R. Merenyi, Chem. Ber.,
1972, 105, 108–114.
13 P. Thompson, A. D. Robertson, S. Jackson, V. Kenche, C. Yaip and H.
Parbaharan, PCT Int. Appl., 2001, WO 2001053266.
14 T. Iwaki, A. Yasuhara and T. Sakamoto, J. Chem. Soc., Perkin Trans.
1, 1999, 1505–1510.
15 S. Ohba, T. Sakamoto and H. Yamanaka, Heterocycles, 1990, 31, 1301–
1308.
4 F. Ye, B. Chen and X. Huang, Synthesis, 1989, 317–320.
5 B. Erb, B. Rigo, B. Pirotte and D. Couturier, J. Heterocycl. Chem.,
2002, 39, 15–28.
6 X. Huang and B. Chen, Synthesis, 1986, 967–968; G. A. Hunter and
H. McNab, J. Chem. Soc., Perkin Trans. 1, 1995, 1209–1214.
7 B. E. Fulloon and C. Wentrup, J. Org. Chem., 1996, 61, 1363–1368.
8 B. Chen, X. Huang and J. Wang, Synthesis, 1987, 482–483.
9 T. Kappe, M. A. A. Chirazi and E. Ziegler, Monatsh. Chem., 1972, 103,
426–434.
16 R. J. Griffin, G. Fontana, B. T. Golding, S. Guiard, I. R. Hardcastle,
J. J. J. Leahy, N. Martin, C. Richardson, L. Rigoreau, M. Stockley and
G. C. M. Smith, J. Med. Chem., 2005, 48, 569–585.
17 A. De La Cruz, J. Elguero, P. Goya, A. Martinez and W. Pleiderer,
Tetrahedron, 1992, 48, 6135–6150.
18 M. J. Mphahlele and A. M. El-Nahas, J. Mol. Struct., 2004, 688, 129–
136.
19 E. Murguly, T. B. Norsten and N. Branda, J. Chem. Soc., Perkin Trans.
2, 1999, 2789–2794 and references cited therein.
10 R. A. Glennon, M. E. Rogers and M. K. El-Said, J. Heterocycl. Chem.,
20 S. Rodriguez, M. Desage El-Murr, B. T. Golding, R. J. Griffin and
I. R. Hardcastle, Org. Lett., 2006, 8, 5927–5929.
1980, 17, 337–340.
11 A. D. Dunn and R. Z. Norrie, Z. Chem., 1990, 30, 98–99.
21 T. Kappe, Monatsh. Chem., 1967, 98, 874–886.
This journal is
The Royal Society of Chemistry 2007
Org. Biomol. Chem., 2007, 5, 2670–2677 | 2677
©