Synthesis and molecular modelling studies of 8-arylpyrido[30,20:4,5] thieno[3,2-d]pyrimidin-4-amines as multitarget Ser/Thr kinases inhibitors

Article abstract of DOI:10.1016/j.ejmech.2014.12.038

This paper reports the design and synthesis of a novel series of 8-arylpyrido[3g2,2g2:4,5]thieno[3,2-d]pyrimidin-4-amines via microwave-assisted multi-step synthesis. A common precursor of the whole series, 3-amino-5-bromothieno[2,3-b]pyridine-2-carbonitrile, was rapidly synthesized in one step from commercially-available 5-bromo-2-chloronicotinonitrile. Formylation with DMF-DMA led to (E)-Ng2-(5-bromo-2-cyanothieno[2,3-b]pyridin-3-yl)-N,N-dimethylformimidamide (4) which was conveniently functionalized at position 8 by palladium-catalyzed Suzuki-Miyaura cross-coupling to introduce a heteroaromatic ring. High-temperature formamide-mediated cyclization of the cyanoamidine intermediate gave seventeen 8-arylpyrido[3g2,2g2:4,5]thieno[3,2-d]pyrimidin-4-amines. The inhibitory potency of the final products was evaluated against five protein kinases (CDK5/p25, CK1δ/, GSK3α/β, DYRK1A and CLK1) and revealed that 8-(2,4-dichlorophenyl)pyrido[3g2,2g2:4,5]thieno[3,2-d]pyrimidin-4-amine 1g specifically inhibits CK1 δ and CLK1 (220 and 88 nM, respectively) while its 7-(2,4-dichlorophenyl)pyrido[3g2,2g2:4,5]thieno[3,2-d]pyrimidin-4-amine isomer 10 showed no activity on the panel of tested kinases. Molecular modelling of 10 and 1g in the ATP binding sites of CK1 δ/and CLK1 showed that functionalization at position 7 of pyrido[3g2,2g2:4,5]thieno[3,2-d]pyrimidin-4-amines is likely to induce a steric clash on the CK1 δ/epies; P-loop and thus a complete loss of inhibitory activity.

Full text of DOI:10.1016/j.ejmech.2014.12.038

European Journal of Medicinal Chemistry 92 (2015) 124e134
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and molecular modelling studies of 8-arylpyrido[30,20:4,5]
thieno[3,2-d]pyrimidin-4-amines as multitarget Ser/Thr kinases
inhibitors
,
,
1
b
b
Yvonnick Loidreau a 1, Emmanuel Deau a , Pascal Marchand , Marie-Renee Nourrisson ,
ꢀ
Cedric Loge , Gael Coadou , Nadege Loaec d, Laurent Meijer d, Thierry Besson a
Normandie Univ, COBRA, UMR 6014 & FR 3038; Univ Rouen; INSA Rouen; CNRS, Batiment IRCOF, 1 rue Tesniere, 76821 Mont St Aignan Cedex, France
a
c
,
, *
ꢀ
ꢀ b
€
ꢁ
€
a
^
ꢁ
b
ꢀ
ꢀ
ꢀ
ꢀ
Universite de Nantes, Nantes Atlantique Universites, Laboratoire de Chimie Therapeutique, Cibles et Medicaments des Infections et du Cancer, IICiMed
UPRES EA 1155, UFR des Sciences Pharmaceutiques et Biologiques, 1 rue Gaston Veil, 44035 Nantes, France
c Protein Phosphorylation & Human Disease Group, Station Biologique, 29680 Roscoff, France
d Manros Therapeutics, Centre de Perharidy, 29680 Roscoff, France
a r t i c l e i n f o
a b s t r a c t
This paper reports the design and synthesis of a novel series of 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyr-
imidin-4-amines via microwave-assisted multi-step synthesis. A common precursor of the whole series,
3-amino-5-bromothieno[2,3-b]pyridine-2-carbonitrile, was rapidly synthesized in one step from
commercially-available 5-bromo-2-chloronicotinonitrile. Formylation with DMF-DMA led to (E)-N0-(5-
bromo-2-cyanothieno[2,3-b]pyridin-3-yl)-N,N-dimethylformimidamide (4) which was conveniently
functionalized at position 8 by palladium-catalyzed Suzuki-Miyaura cross-coupling to introduce a het-
eroaromatic ring. High-temperature formamide-mediated cyclization of the cyanoamidine intermediate
gave seventeen 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines. The inhibitory potency of the
Article history:
Received 30 October 2014
Received in revised form
10 December 2014
Accepted 21 December 2014
Available online 23 December 2014
Keywords:
Microwave-assisted chemistry
Alzheimer disease
Pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-
amines
final products was evaluated against five protein kinases (CDK5/p25, CK1
CLK1) and revealed that 8-(2,4-dichlorophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine 1g
specifically inhibits CK1 /ε and CLK1 (220 and 88 nM, respectively) while its 7-(2,4-dichlorophenyl)
pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine isomer 10 showed no activity on the panel of tested
kinases. Molecular modelling of 10 and 1g in the ATP binding sites of CK1 /ε and CLK1 showed that
functionalization at position 7 of pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines is likely to induce a
steric clash on the CK1 /ε P-loop and thus a complete loss of inhibitory activity.
© 2014 Elsevier Masson SAS. All rights reserved.
d/ε, GSK3a/b, DYRK1A and
d
Kinase inhibitors
d
d
1. Introduction
amino acids. Our research groups are mainly invested in the syn-
thesis of C,N,S- or C,N,O-containing heterocyclic precursors of
bioactive molecules able to modulate the activity of kinases in
signal transduction [4e8] and especially Ser/Thr kinases (CDK5,
GSK3, CLK1 and CK1) and dual-specificity kinases (DYRK1A) [4e9].
These eukaryotic kinases belong to a larger family known as the
CMGC group of serine/threonine kinases. Since these five kinases
are all involved to some extent in Alzheimer's disease (AD), a multi-
kinase inhibitor able to target more than one of them may provide a
highly pertinent therapeutic strategy for patients suffering of this
neurodegenerative disease. Although recent works seem to
demonstrate the interest of multi-target-directed ligands (MTDLs)
[10], it still remains unknown whether single or multi-selective
inhibitors are most suited for Alzheimer's disease drug develop-
ment. This uncertainty mainly lies in the fact that it still remains
Protein kinases catalyze protein phosphorylation, a key cellular
regulatory mechanism which is frequently deregulated in human
diseases. All major diseases, including cancer, neurodegenerative
disorders and cardiovascular diseases have been closely associated
with kinases' deregulations [1e3]. Consequently, protein kinases
and their potential inhibitors represent valuable targets for the
pharmaceutical industry in its search for new therapeutic agents.
Most kinases act on both serine and threonine, others act on
tyrosine, and a number (dual-specificity kinases) act on all three
* Corresponding author.
E-mail address: thierry.besson@univ-rouen.fr (T. Besson).
These authors contributed equally to this work.
1
http://dx.doi.org/10.1016/j.ejmech.2014.12.038
0223-5234/© 2014 Elsevier Masson SAS. All rights reserved.

Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
125
unresolved to what extent each of these five kinases plays a more
prominent role in Alzheimer's disease than the others and, thus,
which one should preferably be targeted.
Suzuki-Miyaura cross-coupling reaction with various hetero-
arylboronic acids in order to insert the aryl ring in position 8. Final
cyclization into pyrimidin-4-amine would be achieved by nucleo-
philic attack of ammonia generated via thermal degradation of
formamide. The order of this synthetic sequence was defined after
the preliminary exploration of a similar synthesis [12a]. It was
discovered that the most effective route to access such derivatives
implied the use of formamide in the last step of the sequence.
Experimentally, 5-bromo-2-chloronicotinonitrile (2) was
treated with freshly prepared 3-mercaptopropionitrile [12] and
aqueous potassium hydroxide, in dimethylformamide (DMF) at
0 ꢀC. After 1 h of stirring, bromoacetonitrile was added to the cold
reaction mixture to give the expected 3-amino-5-bromothieno[2,3-
b]pyridine-2-carbonitrile (3) in high yield (91%). Then, (E)-N0-(5-
In the course of our work, N-arylbenzothieno[3,2-d]pyrimidin-
4-amines and their pyrido and pyrazino analogues were described
[6e8]. These original heteroaromatics provide new means to target
and inhibit some of the above-mentioned kinases in the nanomolar
range. It was found that N-arylpyrido[30,20:4,5]thieno[3,2-d]pyr-
imidin-4-amine series (see I in Scheme 1) turned out to be
particularly promising for the development of new pharmacolog-
ical inhibitors of CK1, CLK1 and DYRK1A kinases. Among these
compounds, II (Scheme 1) was identified as the most active product
with low IC50 values for CK1 (31 nM) and CLK1 (680 nM). This
compound also displayed a micromolar value for DYRK1A (2.8 mM)
[6b]. Pursuing our research on the design of original kinase in-
hibitors, we wished to synthesize and investigate the biological
potential of a new series (series 1) of C8-substituted pyrido
[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines (Scheme 1).
The choice to introduce heteroaryl substituents in position 8 of
the tricyclic structure was guided by our previous endeavour to
synthesize C7-substituted pyrido[20,30:4,5]furo[3,2-d]pyrimidin-4-
amine derivatives [7] and some of their pyrido[20,30:4,5]furo[3,2-
d]pyrimidin-4-amine analogues (III in Scheme 1), most of which
showed moderate affinities for the five targeted Ser/Thr kinases
(CDK5, CK1, CLK1, DYRK1A and GSK3).
This article describes the development of a simple and reliable
synthetic protocol that allowed preparation of a library of new C8-
substituted pyrido[30,20:4,5]thieno[3,2-d]pyrimidines (series 1) for
which interesting kinase inhibitory activities were observed. The
main part of the chemistry described in this paper was carried out
bromo-2-cyanothieno[2,3-b]pyridin-3-yl)-N,N-dimethyl
for-
mimidamide intermediate (4) was prepared in quantitative yield by
reaction of cyanoenamine 3 with N,N-dimethylformamide dime-
thylacetal (DMF-DMA) after 30 min of microwave heating (90 ꢀC)
(Scheme 3).
Introduction of phenyl substituents prior to the pyrimidin-4-
amine ring formation was accomplished by Suzuki cross-coupling
of various heteroarylboronic acids with intermediate 4 in usual
conditions of temperature and time to give (E)-N0-(5-aryl-2-cyano)
thieno[2,3-b]pyridin-3-yl)-N,N-dimethylformimidamide
series
(5aeq) in very good yields (Table 1). Strong microwave-assisted
heating (185 ꢀC) of these highly reactive intermediates with
formamide allowed synthesis of novel 8-(het)arylpyrido[30,20:4,5]
thieno[3,2-d]pyrimidin-4-amines (1aeq) in good overall yields
(55e100%) [13].
In order to confirm the real impact of the functionalization in
position 8 of series 1, a short study was undertaken. We wished to
synthesize an isomeric derivative of the best inhibitor of the pre-
vious series. We focused our attention on the preparation of a
regioisomer of pyrido[30,20:4,5]thieno[3,2-d]pyrimidine 1g. The
target molecule (see 10 in Scheme 4) would also bear a 2,4-
dichlorophenyl substituent at position 7 of the ring instead of po-
sition 8. While the pyrimidinic moiety would remain unaffected, a
change in the spatial constraints of the pyridinic ring may induce
important alterations of the biological activity, hence proving the
importance of the substitution in position 7 or 8.
under microwave irradiation in
a combinatorial chemistry
approach. Since we thought it would be more appropriate to test all
brain isoforms at once as they are expressed in the brain, the target
tissue, the evaluation of kinase inhibition was performed on a panel
of five Ser/Thr kinases and their isoforms. Docking studies were
achieved in hope to understand and confirm the interactions of the
best products in the ATP binding sites.
2. Chemistry
After preliminary investigations, the chosen synthetic route was
adapted from previous work on regioselective cross-coupling re-
actions of boronic acids with dihaloheterocycles [14,15]. Commer-
cial 2,6-dichloropyridine-3-carbonitrile (6) was thus chosen as the
starting material to access isomer 10. Microwave-assisted Suzuki-
Miyaura cross-coupling of 6 with 2,4-dichlorophenylboronic acid,
sodium carbonate, and Pd(PPh3)4, in a dioxane/water (3:1, v/v) at
100 ꢀC (400 W) yielded 2-chloro-6-(2,4-dichlorophenyl)nic-
otinonitrile (7) in excellent 87% yield after 2 h of reaction (Table 1).
The rest of the synthesis was achieved according to the route
described in Scheme 3. Accordingly, 6-substituted 2-
The synthetic route to accomplish the synthesis of 8-arylpyrido
[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine derivatives (1) is depic-
ted in Scheme 2. This multi-step synthetic pathway was inspired by
our previous works on the efficacy of formamide degradation.
Microwave-assisted degradation of formamide is well known for
generating ammonia in situ and cyclizing cyanoamidines into pyr-
imidin-4-amine rings via a nitrogen atom introduction, from
anthranilonitrile, or its benzofuro or benzothieno analogue [11].
The cyanoamidine (4) could be synthesized in two steps from 5-
bromo-2-chloronicotinonitrile (2). This intermediate could be
converted into the expected tricyclic pyrimidin-4-amine after a
Scheme 1. Structures of 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine targets (series 1), their pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine and N-phenyl analogues (I
and II) [6], and furo- or thieno[3,2-d]pyrimidin-4-amine isomers (III) [8].

126
Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
Scheme 2. Synthetic route envisioned for access to the target 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines (series 1).
Scheme 3. Synthesis of 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines (1aeq); for detailed yields see Table 1.
chloronicotinonitrile 7 was successively treated with mercapto-
propionitrile and bromoacetonitrile in aqueous potassium hy-
droxide and DMF at 0 ꢀC to give the expected 3-amino-6-(2,4-
dichlorophenyl)thieno[2,3-b]pyridine-2-carbonitrile (8) in modest
28% yield.
cases, a 600e800 W irradiation was enough to efficiently reach the
programmed temperature. This parameter was mainly monitored
via a contactless-infrared pyrometer which was calibrated in con-
trol experiments with a fibre-optic contact thermometer.
Then, (E)-N0-[2-cyano-6-(2,4-dichlorophenyl)thieno[2,3-b]pyr-
idin-3-yl]-N,N-dimethyl formimidamide intermediate (9) was
prepared in 73% yield by reaction of cyanoenamine 8 with DMF-
DMA after 30 min of microwave heating at 90 ꢀC. Strong micro-
wave heating (185 ꢀC) of the highly reactive intermediate 9 in the
presence of formamide allowed convenient synthesis (87% yield) of
the expected 7-(2,4-dichlorophenyl)pyrido[30,20:4,5]thieno[3,2-d]
pyrimidin-4-amine (10), the regioisomeric product of 1g.
At this stage of the study, the synthesis of seventeen 8-
substituted pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines was
performed with success. Suzuki cross-coupling reactions were used
to functionalize pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines
with n heteroaryl ring in position 8. An isomer 10 of one of the
most active compound 1g was also prepared via a preliminary and
regioselective cross-coupling reaction.
Note that microwave heating was mainly performed at atmo-
spheric pressure in a controlled multimode cavity with a micro-
wave power delivery system ranging from 0 to 1200 W. Concerning
the technical aspect, the choice of a reactor able to work at atmo-
spheric pressure was guided by our previous experience in
microwave-assisted heterocyclic syntheses, especially in the
chemistry of quinazolines [16]. Open vessel microwave experi-
ments have some advantages, such as the possibility of easier scale-
up and the possibility to use common laboratory glassware. Our
choice was also guided by a recent work describing the tendency of
pressure to accumulate when a product as DMF-DMA was heated
into pressurized vials, especially under microwaves [11]. In most
3. Biological activities
The final products (1aeq) and 10 were tested on five different
in vitro kinase assays (CDK5/p25 (cyclin-dependent kinase), CK1
d/ε
(casein kinase 1), GSK3 (glycogen synthase kinase 3), DYRK1A
a/b
(dual-specificity tyrosine phosphorylation regulated kinase) and
CLK1 (cdc2-like kinase 1) to evaluate their inhibition potency
[17e20]. All compounds were first tested at a final concentration of
10
considered as inactive (IC50 > 10
than 50% inhibition at 10 M were next tested over a wide range of
concentrations (usually 0.01e10 M), and IC50 values were deter-
mined from the doseeresponse curves (Sigma-Plot). Harmine
(entry 20 in Table 2) is a -carboline alkaloid known to be a potent
mM. Compounds showing less than 50% inhibition were
m
M). Compounds displaying more
m
m
b
inhibitor of DYRK1A [21]. It was also tested as positive control and
its IC50 value was compared to those obtained for the compounds
under study. Results of the most active compound (see II in Scheme
1) in the preceding study [6b] were obtained in the same assays and
were added to Table 2 (entry 19).
Results given in Table 2 demonstrate that none of the tricyclic
derivatives prepared in this work showed affinity against CDK5/p25
and GSK3. On a general aspect, the biological activity of 8-
arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines (1aeq) and
one of their isomer, 10, was oriented towards three kinases of the
initial panel: CK1, CLK1 and DYRK1A.
Undoubtedly, the most active molecules prepared in this study
were series 1iel and compounds 1g and 1p. These compounds

Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
127
Table 1
showed
submicromolar
activities
against
M < IC50 < 0.19 mM)
CK1
Synthesis of N,N-dimethylformimidamides 5aeq and final thieno[3,2-d]pyrimidines
1aeq.
(0.19 M < IC50 < 0.50
m
mM) and CLK1 (0.071
m
with a slight preference for the second one. The DYRK1A IC50 values
obtained for all these compounds are situated mainly in the
(Het)Aryl
micromolar range (1.1
(entry 7) and 1k (entry 11) for which submicromolar IC50 values
(0.53 and 0.33 M, respectively) were observed.
mM < IC50 < 11 mM) except for compounds 1g
m
Compound Nꢀ
Yield (%)a,b,c
99
Product Nꢀ
Yield (%)b,c,d
87
Comparison of the results obtained for compounds 1aef (en-
tries 1e6), 1meo (entries 13e15), and 1q (entry 17) with those
obtained for 1g (entry 7) and 1iel (entries 9e12), demonstrates
that the presence of two substituting groups on the phenyl ring in
position 8 seems to have a beneficial effect on the inhibitory ac-
tivity. Without being able to establish a general rule, the presence
of substituents in positions 2 and 4 of the aromatic ring in position
8 of the main core has a rather beneficial effect compared to para-
substituted derivatives.
5a
1a
5b
5c
99
79
1b
1c
93
85
Biological results obtained for compound 1p, (entry 16: 0.19
mM
5d
94
1d
78
and 0.18 M for CK1 and CLK1, respectively) are remarkable. 1p is
m
the only derivative with a thiophenic ring in place of the phenyl
group that can inhibit the activity of CK1 and CLK1, indicating that
other thiophene-substituted derivatives could be bioisosteres of
their phenyl-substituted analogues of this study. Values given in
Table 2 (entry 16) are similar to those obtained for the 2,4-
disubstituted phenyl leaders (1g and series 1ie1l). Compared to
the unsubstituted phenyl precursor 1a (entry 1), the thiophenic
derivative 1p may be considered as slightly more active although
5e
5f
98
55
1e
1f
92
99
5g
90
1g
73
their IC50 values against CLK1 (0.18 and 0.19 mM) were similar.
Biological data for the isomeric derivative 10 (entry 18) and
those obtained for 1g (entry 7) prove that changing the 2,4-
dichlorophenyl substituent from position 8 (series 1) to position 7
(compound 10) dramatically decreased the affinity of the most
active inhibitor of this series against kinase CK1. Product 10
completely lost its affinity against CK1 and showed a quite selective
5h
5i
64
70
1h
1i
64
66
IC50 value against CLK1 (0.1
mM), whilst the affinity measured for
DYRK1A (IC50 ¼ 6.1
mM) was found modest and not really
5j
67
88
99
1j
66
79
61
significant.
All these results demonstrate that it is difficult to define any role
for the various substituents of the aryl group located at position 8.
Noteworthy, the presence of two halogen atoms on the aromatic
rings (1g and 1iel, Table 2) resulted in submicromolar affinity for
CK1 and also for CLK1 in a large number of the active products,
although the observed affinities were lower for DYRK1A. On the
contrary, the presence of one substituent on the N-aryl moiety
decreased the inhibitory activity on all the other tested kinases.
CK1 is a Ser/Thr protein kinase family consisting of multiple
5k
5l
1k
1l
isoforms (
a
,
b
,
g
1ꢁ3,
d and ε) which are highly homologous within
5m
5n
5o
14
22
64
1m
1n
1o
65
55
93
their kinase domains. Recently, several studies have highlighted the
overactivity of some CK1 isoforms in neurodegenerative diseases,
especially in tauopathies such as Alzheimer disease (AD) [22]. All
these results demonstrated that the two CK1
seem to be the most interesting to target and to modulate as a
therapeutic strategy [23]. It was also highlighted that CK1 /ε iso-
d and CK1ε isoforms
d
forms are also key regulators of the Wnt signalling pathway and
consequently, may have a regulation role in cell survival and cancer
progression. As the involvement of CK1 in cell regulation and
neurodegenerative diseases pathogenesis has recently been iden-
tified, there has been relatively little work regarding inhibitors of
this family of protein kinases [24].
5p
5q
79
54
1p
1q
87
67
CLK1 is one of the four isoforms (CLK1-4) of the cdc2-like kinase
family. In humans, the highest levels of CLK1 expression were found
in the brain. It was described that inhibitors of CLK1 could prove to
be useful agents in disease phenotypes characterized by abnormal
mRNA splicing. In this sense CLK inhibitors may alter the splicing of
microtubule-associated protein tau implicated in Alzheimer's dis-
ease and Parkinson0s disease [25].
a
Reactions were performed for 90 min at 150 ꢀC and at atmospheric pressure
under microwave irradiation (800 W) on a 1.0 mmol scale from 4 with 1.5 equiv of
appropriate boronic acid.
b
Yield of isolated product.
Microwave reactor: RotoSYNTH™ from Milestone S.r.l, Italy.
Reactions were performedfor 30 minat 185 ꢀC and at atmospheric pressure under
c
d
microwaves (200 W) on a 0.2 mmol scale from 5aeq with 40 equiv of formamide.

128
Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
2,4-dichlorophenyl-
1) HS-(CH2)2-CN (1.2 eq.)
KOH (2 eq.)
DMF, 0°C, 15 min
boronic acid (1.1 eq.)
Pd(PPh3)4 (30 mol %)
Na2CO3 (3 eq.)
CN
Cl
NH2
CN
CN
Cl
N
N
Cl
Dioxane-H2O
100 °C (µw)
2h
S
Cl
N
2) BrCH2CN (1.5 eq.)
0 °C, r.t., 1h
Cl
Cl
Cl
6
7 (87%)
8 (28%)
N
N
N
DMF-DMA (10 eq.)
NH2CHO (40 eq.)
N
N
Cl
S
90 °C (µw)
30 min
185 °C (µw)
1h
N
Cl
CN
NH2
S
Cl
Cl
9 (73%)
10 (83%)
Scheme 4. Multistep synthesis of 7-(2,4-dichlorophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amine (10).
The third kinase targeted by the compounds described in this
such a situation would seem to be partially avoided with CLK1
(Fig. 2c).
study was DYRK1A. Evidence for the role of overexpressed DYRK1A
in various neurodegenerative diseases and Down syndrome is now
well established and it has become an attractive drug target for
numerous research groups [26].
5. Conclusion
A chemical library of seventeen novel 8-arylpyrido[30,20:4,5]
thieno[3,2-d]pyrimidin-4-amines 1aeq was rapidly achieved using
microwave-assisted synthesis. Good control of the reaction pa-
rameters allowed efficient heating of the reaction mixture, result-
ing in short reaction times and high yields. The inhibitory potency
of the final products against five kinases involved in AD was eval-
uated. Also, a 7-substituted isomer (10) of one of the most active
compound (1g) was successfully prepared and evaluated biologi-
cally. Our study demonstrates that several molecules described in
this paper are particularly promising for the development of new
multi-target inhibitors of CK1 and CLK1 kinases. Although the af-
finity of these compounds for DYRK1A is not negligible, it still needs
optimization to provide a synergetic effect to these molecules. The
most active compounds 1g and 1p showed submicromolar inhibi-
tion and a relative selectivity for CK1 and CLK1 over the other tested
enzymes. Molecular modelling proved to be a very useful tool to
understand the role of the heteroaryl substituents in the affinity
against the panel of tested kinases. Functionalization of position 8
of pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines with 2,4-
disubstituted phenyl groups induced an increased binding affinity
with the ATP binding sites of CK1 and CLK1. Contrariwise, substi-
tution at position 7 led to a steric clash on the P-loop of CK1
resulting in a complete loss of inhibitory activity against CK1 while
partially sparing the activity on CLK1.
Recently, low benefits of drugs which focused their activity
against a single target encouraged new studies in the direction of
multi-targeting strategies. Therefore, developing a drug which has
micromolar or submicromolar affinities on a panel of two or three
kinases seems to be of great interest. Such inhibitors should be able
to regulate the targeted kinases without affecting the basic activity
in cells. In this sense the new series described in this paper may
lead to the discovery of valuable multi-kinases inhibitors.
4. Molecular modelling
Molecular modelling studies were performed to understand the
nature of interactions governing the binding mode of these mole-
cules in the active sites of CK1 and CLK1. To this end, compounds 1g
and 10 were docked into crystal structures of CLK1 in complex with
debromohymenialdisine at 1.70 Å resolution (pdb code: 1Z57) [27]
and/or CK1d in complex with PF4800567 at 2.07 Å resolution (pdb
code: 4HNF) [28]. Interestingly, CK1 inhibitor PF4800567 (Fig. 1)
also displays a pyrimidin-4-amine group that makes classical H-
bond interactions with the hinge region of the kinase. These in-
teractions are similar to those established by the adenine moiety of
ATP in the active site, with the exception of the meta-chlorophenyl
moiety of PF4800567 which binds in the hydrophobic pocket
adjacent to the CK1d gatekeeper residue Met82 that could likely
account for the high selectivity over other kinases [29,30]. Thereby,
even if different binding modes in the ATP site could be involved
with our molecules, we applied distance constraint between the
common aminopyrimidine core and the hinge domains formed
6. Experimental section
6.1. Chemistry
either by the CLK1 residues Glu242 and Leu244 or by the CK1
d
residues Glu83 and Leu85. In this way, a similar binding pose was
found with the 2,4-dichlorophenyl group (1g) oriented towards the
phosphate binding loop (P-loop) (Fig. 2), a highly flexible region of
Melting points of powder compounds were measured on an
STUART-Advanced apparatus. IR spectra were recorded on a Per-
kinElmer Spectrum 100 Series FT-IR spectrometer. 1H, 13C NMR
spectra were recorded on a Bruker DXP 300 spectrometer at 300,
75 MHz, respectively and a Bruker AVANCE 400 MHz high resolu-
tion NMR spectrometer at 400,100 MHz, respectively. Multiplicities
were abbreviated as follows: singlet (s), doublet (d), triplet (t),
multiplet (m), and broad (br). Reactions were monitored by TLC
analysis using Merck silica gel 60F-254 thin layer plates. Column
chromatography was carried out on silica gel Merck 60 (70e230
mesh ASTM). Elemental analyses were found within 0.4% of the
kinases [31]. On CLK1, this group can establish additional
p-p in-
teractions with Phe172, a conserved aromatic residue that may
greatly contribute to the biological activity compared to non-
substituted compound II (Fig. 2a), while on CK1, the correspond-
ing amino acid (Phe20) is more distant from the active site and may
not have any impact on the binding affinity (Fig. 2b). Also, the loss
of activity observed with analogue 10 on CK1 may result from a
steric clash between the P-loop and the displaced 7-aryl group,

Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
129
Table 2
programmed temperature after a ramp period of 2 min.
Compounds 1aeg and their precursors (2, 3, 4 and 5aeg) were
described in a previous paper [12a].
Kinase activity (IC50
)
of 8-arylpyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-amines
(1aeq).a,b
Entry
Compound
CDK5/p25
CK1
d
/ε
CLK1
DYRK1A
GSK3a/b
6.1.1. General procedure for the synthesis of (E)-N0-(5-aryl-2-cyano)
thieno[2,3-b]pyridin-3-yl)-N,N-dimethylformimidamide derivatives
(5heq)
1
2
3
4
5
6
7
8
1a
1b
1c
1d
1e
1f
1g
1h
1i
1j
1k
1l
1m
1n
1o
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
0.63
>10
1.1
0.19
2.5
0.28
1.1
0.69
2.4
0.088
1.7
0.13
0.11
0.19
0.071
0.91
0.5
0.52
0.18
0.95
0.1
11
>10
>10
1.1
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
0.51
0.91
0.88
0.22
1.2
0.22
0.22
0.38
0.5
A mixture of (E)-N0-(5-bromo-2-cyanothieno[2,3-b]pyridin-3-
yl)-N,N-dimethyl formimi-damide 4 (0.160 g, 0.52 mmol), appro-
priate boronic acid (1.5 equiv), sodium carbonate (0.110 g,
1.04 mmol) and Pd(PPh3)4 (0.016 g) in DMF (8 mL) was irradiated at
150 ꢀC (800 W) for 90 min. On completion (followed by thin-layer
chromatography), the reaction was cooled to room temperature
and H2O was added. The powder was filtered off, washed with
EtOAc and the organic layer was evaporated in vacuo. The crude
powder was purified by silica gel column chromatography using a
gradient of PE/EtOAc (100:0 to 0:100, v/v) as the eluent to give the
desired compounds.
1.9
>10
0.53
6.8
1.1
1.4
0.33
2
2.1
1.4
3.8
9
10
11
12
13
14
15
16
17
18
19
20
1.1
0.64
0.53
0.19
0.57
>10
0.031
1p
1q
1.3
1.3
6.1
11
10
IIc
0.68
0.026
6.1.1.1. (E)-N0-(2-cyano-5-(4-(trifluoromethyl)phenyl)thieno[2,3-b]
pyridin-3-yl)-N,N-dimethylformimidamide (5h). Yellow powder
(0.124 g, 64%); mp 146e148 ꢀC; IR nmax (cmꢁ1): 2201 (CN), 1628,
1616, 1588, 1537, 1486, 1416, 1386, 1362, 1329, 1251, 1167, 1107, 1072,
Harmined
1.5
0.029
a
IC50 values are reported in
m
M. The most significant results (IC50 ꢂ 0.5
mM) are
presented in bold.
b
Kinases activities were assayed in triplicate. Typically, the standard deviation of
single data points was below 10%.
1050, 1014, 983, 912, 836, 761; 1H NMR (300 MHz, DMSO-d6):
d 9.14
c
(d, 1H, J ¼ 2 Hz, H-4), 8.46 (d, 1H, J ¼ 2 Hz, H-6), 8.31 (s, 1H, NCHN),
Data given in Ref. [6b].
Data given in Ref. [21].
d
8.07 (d, 2H, J ¼ 9 Hz, H-ar), 7.88 (d, 2H, J ¼ 9 Hz, H-ar), 3.16 (s, 3H,
Fig. 1. Structures of CK1 inhibitor PF4800567 (a) and the design concept of 8-(het)arylpyrido[30,2’:4,5]thieno[3,2-d]pyrimidin-4-amines (b).
theoretical values. Mass spectra were performed by the Mass
Spectrometry Laboratory of the University of Rouen. Mass spectra
(EI) were recorded with a Waters ZQ 2000 and a Waters LCP 1er XR
spectrometer.
NCH3), 3.11 (s, 3H, NCH3); 13C NMR (75 MHz, DMSO-d6):
d 158.9,
156.3, 155.3, 149.7, 140.6, 131.4, 129.8, 128.8, 128.3 (2C), 128.2, 127.9,
125.9 (2C), 115.7, 86.0, 34.3 (2C); HRMS calcd for C18H14F3N4S
[MþH]þ 375.0891 found 375.0886.
Microwave experiments were conducted in a commercial mi-
crowave reactor especially designed for synthetic chemistry.
RotoSYNTH™ (Milestone S.r.l. Italy) is a multi-mode cavity with a
microwave power delivery system ranging from 0 to 1200 W. The
temperatures of the reactions were mainly monitored via contact-
less infrared pyrometer which was calibrated in control experi-
ments with a fibre-optic contact thermometer protected in a Teflon
coated ceramic well inserted directly in the reaction mixture. Open
vessel experiments were carried out in a 50e250 mL round bottom
flask fitted with a reflux condenser. The vessel contents were stir-
red by means of an adjustable rotating magnetic plate located
below the floor of the microwave cavity and a Teflon-coated mag-
netic stir bar inside the vessel. Temperature and power profiles
were monitored in both cases through the EASY-Control software
provided by the manufacturer. Time indicated in the various pro-
tocols is the time measured when the mixtures reached the
6.1.1.2. (E)-N0-(5-(2,4-Difluorophenyl)-2-cyanothieno[2,3-b]pyridin-
3-yl)-N,N-dimethyl-formimidamide (5i). White powder (0.125 g,
70%); mp 215e217 ꢀC; IR nmax (cmꢁ1): 2197 (CN), 1632, 1615, 1590,
1540, 1512, 1421, 1359, 1266, 1145, 1111, 1097, 1044, 974, 966, 962,
847, 730; 1H NMR (300 MHz, DMSO-d6):
d
8.94 (d,1H, J ¼ 2 Hz, H-4),
8.32 (s, 1H, NCHN), 8.32 (d, 1H, J ¼ 2 Hz, H-6), 7.82e7.77 (m, 1H, H-
ar), 7.54e7.46 (m, 1H, H-ar), 7.34e7.28 (m, 1H, H-ar), 3.19 (s, 3H,
NCH3), 3.13 (s, 3H, NCH3); 13C NMR (75 MHz, DMSO-d6):
d 158.5,
156.4, 155.2, 150.7, 132.5, 131.5, 127.7, 127.2, 115.7, 112.6, 112.3, 105.0,
104.7, 104.3, 85.7, 34.2 (2C); HRMS calcd for C17H13F2N4S [MþH]þ
343.0829 found 343.0820.
6.1.1.3. (E)-N0-(5-(4-Chloro-2-fluorophenyl)-2-cyanothieno[2,3-b]
pyridin-3-yl)-N,N-dimethyl formimidamide (5j). Pale yellow powder
(0.125 g, 67%); mp 222e224 ꢀC; IR nmax (cmꢁ1): 2189 (CN), 1626,

130
Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
Fig. 2. (a) Docking model of compound 1g into the ATP site of CLK1; (b) Docking model of compound 1g into the ATP site of CK1; (c) Docking model of compound 10 into the ATP
site of CLK1 superimposed with CK1. Active site pocket is highlighted in cyan (MOLCAD surface; program SYBYL-X 1.3). Hydrogens bonds are indicated as yellow dotted lines. P-
loops are presented as green (CLK1) or red (CK1) ribbons. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)
1613, 1584, 1537, 1473, 1458, 1414, 1405, 1386, 1370, 1264, 1248,
6.1.1.6. (E)-N0-(5-(4-Bromophenyl)-2-cyanothieno[2,3-b]pyridin-3-
yl)-N,N-dimethylformimi-damide (5m). White powder (0.028 g,
14%); mp 84e186 ꢀC; IR nmax (cmꢁ1): 2188 (CN), 1625, 1536, 1476,
1458, 1408, 1362, 1249, 1108, 1008, 978, 914, 870, 818, 759; 1H NMR
1210, 1108, 1037, 979, 894, 925, 759; 1H NMR (300 MHz, DMSO-d6):
d
8.93 (t, 1H, J ¼ 2 Hz, H-4), 8.32 (t, 1H, J ¼ 2 Hz, H-6), 8.29 (s, 1H,
NCHN), 7.76 (dd, 1H, J1 ¼ 8 Hz, J2 ¼ 9 Hz, H-ar), 7.65 (dd, 1H,
J1 ¼ 2 Hz, J2 ¼ 9 Hz, H-ar), 7.47 (dd,1H, J1 ¼ 2 Hz, J2 ¼ 8 Hz, H-ar), 3.15
(s, 3H, NCH3), 3.09 (s, 3H, NCH3); HRMS calcd for C17H13ClFN4S
[MþH]þ 359.0533 found 359.0548.
(300 MHz, DMSO-d6):
d
9.09 (d, 1H, J ¼ 2 Hz, H-4), 8.39 (d, 1H,
J ¼ 2 Hz, H-6), 8.30 (s, 1H, NCHN), 7.81 (d, 2H, J ¼ 9 Hz, H-ar), 7.73 (d,
2H, J ¼ 9 Hz, H-ar), 3.16 (s, 3H, NCH3), 3.11 (s, 3H, NCH3); 13C NMR
(75 MHz, DMSO-d6):
d 158.5, 156.4, 155.5, 149.6, 135.7, 132.0 (2C),
6.1.1.4. (E)-N0-(5-(2-Chloro-4-fluorophenyl)-2-cyanothieno[2,3-b]
pyridin-3-yl)-N,N-dimethyl-formimidamide (5k). Pale yellow pow-
der (0.163 g, 88%); mp 208e210 ꢀC; IR nmax (cmꢁ1): 2201 (CN),1628,
1600, 1588, 1544, 1539, 1503, 1476, 1462, 1382, 1361, 1254, 1205,
1108, 1043, 969, 893, 866, 825; 1H NMR (300 MHz, DMSO-d6):
131.7, 129.4, 129.3 (2C), 127.9, 121.9, 115.8, 86.0, 34.3 (2C); HRMS
calcd for C17H14BrN4S [MþH]þ 385.0123 found 385.0124.
6.1.1.7. (E)-N0-(5-(4-Nitrophenyl)-2-cyanothieno[2,3-b]pyridin-3-
yl)-N,N-dimethylformimi-damide (5n). Yellow powder (0.041 g,
22%); mp 208e210 ꢀC; IR nmax (cmꢁ1): 2196 (CN), 1627, 1591, 1545,
1515, 1480, 1464, 1437, 1359, 1343, 1257, 1093, 1015, 975, 919, 868,
d
8.79 (d, 1H, J ¼ 2 Hz, H-4), 8.27 (s, 1H, NCHN), 8.23 (d, 1H, J ¼ 2 Hz,
H-6), 7.68e7.62 (m, 2H, H-ar), 7.39 (td, 1H, J1 ¼ 3 Hz, J2¼9 Hz, H-ar),
3.13 (s, 3H, NCH3), 3.06 (s, 3H, NCH3); 13C NMR (75 MHz, DMSO-d6):
851, 792, 759, 753, 737; 1H NMR (300 MHz, DMSO-d6):
d 9.25 (d,1H,
d
158.5, 156.4, 155.2, 151.3, 133.5, 132.8, 132.2, 130.7, 127.4, 117.3,
J ¼ 2 Hz, H-4), 8.57 (d, 1H, J ¼ 2 Hz, H-6), 8.43 (d, 2H, J ¼ 9 Hz, H-ar),
8.38 (s, 1H, NCHN), 8.21 (d, 2H, J ¼ 9 Hz, H-ar), 3.22 (s, 3H, NCH3),
3.18 (s, 3H, NCH3); HRMS calcd for C17H14N5O2S [MþH]þ 352.0868
found 352.0868.
116.9, 115.8, 115.2, 114.8, 85.8, 34.4 (2C); HRMS calcd for
C
17H13ClFN4S [MþH]þ 359.0533 found 359.0537.
6.1.1.5. (E)-N0-(5-(2-Fluoro-4-methoxyphenyl)-2-cyanothieno[2,3-b]
pyridin-3-yl)-N,N-dimethylformimidamide
(5l). White powder
6.1.1.8. (E)-N0-(5-(Benzo[d][1,3]dioxol-5-yl)-2-cyanothieno[2,3-b]
pyridin-3-yl)-N,N-dimethyl formimidamide (5o). Yellow powder
(0.116 g, 64%); mp 194e196 ꢀC; IR nmax (cmꢁ1): 2182 (CN), 1621,
1471, 1446, 1369, 1324, 1252, 1229, 1109, 1097, 1052, 1034, 1017, 977,
(0.184 g, 99%); mp 180e182 ꢀC; IR nmax (cmꢁ1): 2201 (CN), 1617,
1589, 1536, 1478, 1461, 1425, 1382, 1365, 1292, 1263, 1164, 1130,
1114, 1030, 992, 913, 860, 810; 1H NMR (300 MHz, DMSO-d6):
d 8.90
(t, 1H, J ¼ 2 Hz, H-4), 8.29 (s, 1H, NCHN), 8.26 (t, 1H, J ¼ 2 Hz, H-6),
7.64 (dd, 1H, J1 ¼8 Hz, J2 ¼ 9 Hz, H-ar), 7.07e6.95 (m, 2H, H-ar), 3.85
(s, 3H, OCH3), 3.15 (s, 3H, NCH3), 3.09 (s, 3H, NCH3); 13C NMR
931, 870, 859, 799, 756; 1H NMR (300 MHz, DMSO-d6):
d 9.03 (d,1H,
J ¼ 2 Hz, H-4), 8.30 (d, 1H, J ¼ 2 Hz, H-6), 8.29 (s, 1H, NCHN), 7.45 (d,
1H, J ¼ 2 Hz, H-ar), 7.30 (dd, 1H, J1 ¼ 2 Hz, J2 ¼ 8 Hz, H-ar), 7.07 (d,
1H, J ¼ 8 Hz, H-ar), 6.11 (s, 2H, CH2), 3.15 (s, 3H, NCH3), 3.11 (s, 3H,
(75 MHz, DMSO-d6):
d 161.1, 157.8, 156.4, 155.0, 150.7, 131.8, 131.0,
128.0, 127.7, 116.5, 115.7, 111.3, 102.3, 102.0, 85.7, 55.9, 34.5 (2C);
NCH3); 13C NMR (75 MHz, DMSO-d6):
d 157.6, 156.2, 155.3, 149.6,
HRMS calcd for C18H16FN4OS [MþH]þ 355.1029 found 355.1027.
148.2, 147.5, 132.8, 130.5, 128.8, 127.8, 121.2, 115.8, 108.9, 107.7, 101.4,

Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
131
85.7, 34.2 (2C); HRMS calcd for C18H15N4O2S [MþH]þ 351.0916
8.74 (d, 1H, J ¼ 2 Hz, H-7), 8.59 (s, 1H, H-2), 7.82 (ddd, 1H, J 1, 2, 8 Hz,
H-ar), 7.75 (br s, 2H, NH2), 7.66 (dd,1H, J 3, 8 Hz, H-ar), 7.49 (dd,1H, J
found 351.0909.
2, 3 Hz, H-ar); 13C NMR (75 MHz, DMSO-d6):
d 160.8, 158.5, 155.7,
6.1.1.9. (E)-N0-(2-Cyano-5-(thiophen-2-yl)thieno[2,3-b]pyridin-3-
yl)-N,N-dimethylformimi- damide (5p). Light brown powder
(0.128 g, 79%); mp 175e176 ꢀC; IR nmax (cmꢁ1): 2191 (CN), 1627,
1539,1527,1482,1467,1409,1387,1370,1353,1304,1259,1237,1114,
153.2, 151.2, 134.1, 132.3, 130.9, 127.8, 127.0, 125.6, 123.8, 117.1, 116.6,
113.1; HRMS calcd for
331.0238.
C
15H9ClFN4S [MþH]þ 331.0220 found
1035, 980, 901, 849, 755; 1H NMR (300 MHz, DMSO-d6):
d
9.12 (d,
6.1.2.4. 8-(2-Chloro-4-fluorophenyl)pyrido[30,20:4,5]thieno[3,2-d]
pyrimidin-4-amine (1k). Brown powder (0.052 g, 79%);
mp > 300 ꢀC; IR nmax (cmꢁ1): 3118, 1658, 1574, 1503, 1392, 1250,
1H, J ¼ 2 Hz, H-4), 8.29 (d, 1H, J ¼ 2 Hz, H-6), 8.28 (s, 1H, NCHN), 7.79
(dd,1H, J1 ¼1 Hz, J2 ¼ 3 Hz, H-ar), 7.70 (dd,1H, J1 ¼1 Hz, J2 ¼ 5 Hz, H-
ar), 7.23 (dd,1H, J1 ¼3 Hz, J2 ¼ 5 Hz, H-ar), 3.16 (s, 3H, NCH3), 3.12 (s,
1205, 902, 852, 785; 1H NMR (300 MHz, DMSO-d6):
d 8.87 (d, 1H,
3H, NCH3); 13C NMR (75 MHz, DMSO-d6):
d
157.6, 156.3,155.1, 148.3,
J ¼ 2 Hz, H-9), 8.63 (d, 1H, J ¼ 2 Hz, H-7), 8.58 (s, 1H, H-2), 7.76e7.67
138.9, 128.8, 128.0, 127.2 (2C), 127.1, 125.7, 115.7, 85.9, 34.3 (2C);
(m, 4H, H-ar and NH2), 7.43 (dd, 1H, J 2, 3 Hz, H-ar); 13C NMR
HRMS calcd for C15H13N4S2 [MþH]þ 313.0568 found 313.0581.
(75 MHz, DMSO-d6): d 160.5, 158.5, 155.6, 153.1, 151.7, 133.4, 132.5,
131.6, 130.7, 127.2, 117.5, 117.1, 115.2, 114.9, 113.0; HRMS calcd for
6.1.1.10. (E)-N0-(2-Cyano-5-(pyrimidin-5-yl)thieno[2,3-b]pyridin-3-
yl)-N,N-dimethylformimi damide (5q). Grey powder (0.087 g, 54%);
mp 229e230 ꢀC; IR nmax (cmꢁ1): 2200 (CN), 1625, 1536, 1462, 1415,
1384, 1258, 1117, 1044, 989, 900, 873, 759, 724; 1H NMR (300 MHz,
C
15H9ClFN4S [MþH]þ 331.0220 found 331.0227.
6.1.2.5. 8-(2-Fluoro-4-methoxyphenyl)pyrido[30,20:4,5]thieno [3,2-d]
pyrimidin-4-amine (1l). Light brown powder (0.044 g, 61%); mp
296e298 ꢀC; IR nmax (cmꢁ1): 3121, 1650, 1627, 1594, 1570, 1542,
1519, 1468, 1372, 1317, 1297, 1252, 1226, 1162, 1133, 1073, 1047, 860,
DMSO-d6):
d
9.32 (d, 2H, J ¼ 1 Hz, H-ar), 9.28 (d, 1H, J ¼ 1 Hz, H-ar),
9.20 (d, 1H, J ¼ 2 Hz, H-4), 8.62 (d, 1H, J ¼ 2 Hz, H-6), 8.32 (s, 1H,
NCHN), 3.16 (s, 3H, NCH3), 3.12 (s, 3H, NCH3); 13C NMR (75 MHz,
786; 1H NMR (300 MHz, DMSO-d6):
8.66 (d, 1H, J ¼ 2 Hz, H-7), 8.58 (s, 1H, H-2), 7.74 (br s, 2H, NH2), 7.70
(ddd, 1H, J 1, 2, 9 Hz, H-ar), 7.05 (dd, 1H, J 4, 9 Hz, H-ar), 6.98 (dd, 1H,
J 2, 4 Hz, H-ar), 3.86 (s, 3H, OCH3); 13C NMR (75 MHz, DMSO-d6):
d
8.97 (d, 1H, J ¼ 2 Hz, H-9),
DMSO-d6):
d
159.3,157.8,156.3,155.4 (2C),155.3,149.5,130.3,130.2,
127.8, 127.0, 115.7, 86.1, 34.2 (2C); HRMS calcd for C15H13N6S
[MþH]þ 309.0922 found 309.0931.
d
161.0, 159.8, 158.6, 155.7, 153.2, 151.1, 131.5, 130.5, 128.1, 127.5,
6.1.2. General procedure for the synthesis of 8-arylpyrido [30,20:4,5]
thieno[3,2-d]pyrimidin-4-amines (1heq)
116.4, 113.2, 111.3, 102.5, 102.1, 55.9; HRMS calcd for C16H12FN4OS
[MþH]þ 327.0716 found 327.0703.
Formamide (2.0 mL) was added to the corresponding for-
mimidamide (0.20 mmol). The mixture was irradiated at various
temperature (200 W) for 30 min. On completion (followed by
GCeMS chromatography), the reaction was cooled to room tem-
perature and H2O was added. The powder was filtered off, washed
with H2O and dried. The crude powder was purified by silica gel
column chromatography using DCM/EtOAc (100:0 to 0:100, v/v) as
the eluent to give the desired compounds.
6.1.2.6. 8-(4-Bromophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-
4-amine (1m). Light brown powder (0.046 g, 65%); mp > 300 ꢀC; IR
nmax (cmꢁ1): 3142, 1674, 1576, 1540, 1520, 1371, 1249, 1077, 1008,
862, 813, 785, 762; 1H NMR (300 MHz, DMSO-d6):
d 9.16 (d, 1H,
J ¼ 2 Hz, H-9), 8.83 (d, 1H, J ¼ 2 Hz, H-7), 8.60 (s, 1H, H-2), 7.88 (d,
2H, J ¼ 8 Hz, H-ar), 7.76 (br s, 2H, NH2), 7.75 (d, 2H, J ¼ 8 Hz, H-ar);
13C NMR (75 MHz, DMSO-d6):
d 160.6, 158.5, 155.5, 153.5, 150.0,
135.9, 132.2 (2C), 131.9, 129.3 (2C), 128.6, 128.0, 121.9, 102.3; HRMS
6.1.2.1. 8-(4-(Trifluoromethyl)phenyl)pyrido[30,20:4,5]thieno [3,2-d]
pyrimidin-4-amine (1h). Kaki powder (0.044 g, 64%), mp > 300 ꢀC;
IR nmax (cmꢁ1): 3137, 1627, 1558, 1543, 1517, 1478, 1317, 1288, 1268,
1251, 1165, 1111, 1066, 1042, 1014, 962, 862, 843, 787, 778; 1H NMR
calcd for C15H10BrN4S [MþH]þ 356.9810 found 356.9805.
6.1.2.7. 8-(4-Nitrophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-
amine (1n). Light brown powder (0.036 g, 55%); mp > 300 ꢀC; IR
nmax (cmꢁ1): 3112, 1660, 1568, 1518, 1373, 1342, 1270, 1248, 1103,
(300 MHz, DMSO-d6):
d
9.21 (d, 1H, J ¼ 2 Hz, H-9), 8.90 (d, 1H,
J ¼ 2 Hz, H-7), 8.60 (s, 1H, H-2), 8.14 (d, 2H, J ¼ 8 Hz, H-ar), 7.90 (d,
1071, 1027, 913, 847, 784, 753; 1H NMR (300 MHz, DMSO-d6):
d 9.25
2H, J ¼ 8 Hz, H-ar), 7.74 (br s, 2H, NH2); 13C NMR (75 MHz, DMSO-
(d, 1H, J ¼ 2 Hz, H-9), 8.95 (d, 1H, J ¼ 2 Hz, H-7), 8.60 (s, 1H, H-2),
d6): d 161.0, 158.5, 155.6, 153.3, 150.2, 140.7, 131.5, 129.2, 128.8, 128.3
8.36 (d, 2H, J ¼ 8 Hz, H-ar), 8.21 (d, 2H, J ¼ 8 Hz, H-ar), 7.78 (br s, 2H,
(2C), 128.1, 127.9, 126.0 (2C), 113.1; HRMS calcd for C16H10F3N4S
NH2); 13C NMR (75 MHz, DMSO-d6):
d 158.5, 155.7, 153.3, 147.2,
[MþH]þ 347.0578 found 347.0581.
143.1,132.6,130.9,129.4,128.6,127.9, 124.2,120.8,119.7,118.1,113.2;
HRMS calcd for C15H10N5O2S [MþH]þ 324.0555 found 324.0540.
6.1.2.2. 8-(2,4-Difluorophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyr-
imidin-4-amine (1i). Yellow powder (0.042 g, 66%), mp > 300 ꢀC; IR
nmax (cmꢁ1): 3111, 1663, 1596, 1572, 1544, 1529, 1514, 1422, 1372,
1313, 1287, 1271, 1257, 1228, 1153, 955, 921, 885, 810, 786, 731; 1H
6.1.2.8. 8-(Benzo[d][1,3]dioxol-5-yl)pyrido[30,20:4,5]thieno[3,2-d]pyr-
imidin-4-amine (1o). Light brown powder (0.060 g, 93%);
mp > 300 ꢀC; IR nmax (cmꢁ1): 3137, 1627, 1572, 1539, 1509, 1470,
1444, 1377, 1243, 1111, 1073, 1037, 978, 931, 806, 785; 1H NMR
NMR (300 MHz, DMSO-d6):
d
8.98 (d, 1H, J ¼ 2 Hz, H-9), 8.71 (d, 1H,
J ¼ 2 Hz, H-7), 8.57 (s, 1H, H-2), 7.85 (ddd, 1H, J 1, 2, 8 Hz, H-ar), 7.74
(br s, 2H, NH2), 7.48 (ddd, 1H, J 1, 3, 8 Hz, H-ar), 7.27 (ddd, 1H, J 1, 2,
(300 MHz, DMSO-d6):
d
9.09 (d, 1H, J ¼ 2 Hz, H-9), 8.74 (d, 1H,
J ¼ 2 Hz, H-7), 8.59 (s, 1H, H-2), 7.72 (br s, 2H, NH2), 7.51 (d, 1H,
J ¼ 1 Hz, H-ar), 7.37 (dd, 1H, J1¼1 Hz, J2 ¼ 8 Hz, H-ar), 7.08 (d, 1H,
J ¼ 8 Hz, H-ar), 6.12 (br s, 2H, CH2); 13C NMR (75 MHz, DMSO-d6):
3 Hz, H-ar); 13C NMR (75 MHz, DMSO-d6):
d 160.5, 159.0, 155.5,
153.3, 151.2, 132.5, 130.7, 127.7, 127.2, 121.3, 113.0, 112.6, 112.3, 105.1,
104.8; HRMS calcd for
315.0524.
C
15H9F2N4S [MþH]þ 315.0516 found
d 158.5, 155.4, 153.3, 150.0, 148.2, 147.6, 132.8, 130.6, 128.2, 127.8,
126.1, 121.1, 113.1, 108.9, 107.6, 101.4; HRMS calcd for C16H11N4O2S
[MþH]þ 323.0603 found 323.0599.
6.1.2.3. 8-(4-Chloro-2-fluorophenyl)pyrido[30,20:4,5]thieno[3,2-d]
pyrimidin-4-amine (1j). Light brown powder (0.043 g, 66%);
mp > 300 ꢀC; IR nmax (cmꢁ1): 3107, 1672, 1568, 1542, 1525, 1497,
1466, 1396, 1371, 1310, 1282, 1260, 1243, 1208, 1086, 899, 855, 812,
6.1.2.9. 8-(4-Nitrophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-4-
amine (1p). Brown powder (0.049 g, 87%); mp 254e255 ꢀC; IR nmax
(cmꢁ1): 1636, 1572, 1545, 1520, 1475, 1436, 1427, 1379, 1291, 1258,
785; 1H NMR (300 MHz, DMSO-d6):
d
9.00 (d, 1H, J ¼ 2 Hz, H-9),
1207, 898, 786, 745, 709; 1H NMR (300 MHz, DMSO-d6):
d 9.18 (d,

132
Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
1H, J ¼ 2 Hz, H-9), 8.74 (d, 1H, J ¼ 2 Hz, H-7), 8.59 (s, 1H, H-2), 7.68
(dd, 1H, J1¼2 Hz, J2 ¼ 5 Hz, H-ar), 7.76e7.73 (m, 3H, H-ar and NH2),
filtered and evaporated in vacuo. Purification by silica gel column
chromatography using PE/EtOAc (5:5, v/v) as the eluent gave the
desired compound 9 as a yellow powder (0.056 g, 73%); mp
216e218 ꢀC; IR nmax (cmꢁ1): 2195 (CN), 1631, 1574, 1558, 1477, 1434,
1415, 1378, 1340, 1100, 1083, 1039, 1028, 975, 852, 843, 820, 769; 1H
7.26 (d, 1H, J ¼ 2 Hz, H-ar); 13C NMR (75 MHz, DMSO-d6):
d 159.6,
158.5, 155.5, 153.0, 148.5, 139.0, 128.9, 127.8, 127.1 (2C), 126.6, 125.6,
113.2; HRMS calcd for
285.0277.
C
13H9N4S2 [MþH]þ 285.0269 found
NMR (300 MHz, DMSO-d6):
d
8.35 (d, 1H, J ¼ 8 Hz, H-4), 8.29 (s, 1H,
NCHN), 7.81 (m, 2H, H-5 and H-ar), 7.71 (d, 1H, J ¼ 8 Hz, H-ar), 7.61
(dd, 1H, J1¼2 Hz, J2 ¼ 8 Hz, H-ar), 3.16 (s, 3H, CH3), 3.10 (s, 3H, CH3);
6.1.2.10. 8-(Pyrimidin-5-yl)pyrido[30,20:4,5]thieno[3,2-d]pyrimidin-
4-amine (1q). Light brown powder (0.038 g, 67%); mp > 300 ꢀC; IR
nmax (cmꢁ1): 1652, 1578, 1417, 1376, 1254, 1129, 1077, 908, 864, 787,
13C NMR (75 MHz, DMSO-d6):
d 160.9, 159.0, 156.4, 155.1, 136.8,
134.6, 133.1, 132.4, 132.3, 129.6, 127.8, 126.9, 121.7, 115.9, 85.9, 34.2
723; 1H NMR (300 MHz, DMSO-d6):
d
9.39 (d, 2H, J ¼ 1 Hz, H-ar),
(2C); HRMS calcd for
375.0235.
C
17H13Cl2N4S [MþH]þ 375.0238 found
9.29 (s, 1H, J ¼ 1 Hz, H-ar), 9.27 (d, 1H, J ¼ 2 Hz, H-9), 9.04 (d, 1H,
J ¼ 2 Hz, H-7), 8.61 (s, 1H, H-2), 7.79 (br s, 2H, NH2); HRMS calcd for
C
13H9N6S [MþH]þ 281.0609 found 281.0598.
6.1.3.4. 7-(2,4-Dichlorophenyl)pyrido[30,20:4,5]thieno[3,2-d]pyr-
imidin-4-amine (10). Formamide (2 mL) was added to (E)-N0-(2-
cyano-6-(2,4-dichlorophenyl)thieno[2,3-b]pyridin-3-yl)-N,N-
dimethylformimidamide (9) (0.050 g, 0.07 mmol). The mixture was
irradiated at 185 ꢀC (200 W) for 1 h. On completion (followed by
GCeMS chromatography), the reaction was cooled to room tem-
perature and H2O was added. The powder was filtered off, washed
with H2O and dried. The crude powder was purified by silica gel
column chromatography using DCM/EtOAc (100:0 to 0:100, v/v) as
the eluent to give the desired compound 10 as a pale yellow powder
(0.038 g, 83%); mp > 300 ꢀC; IR nmax (cmꢁ1): 3136, 1664, 1575, 1523,
1477, 1442, 1402, 1382, 1340, 1291, 1105, 1089, 1029, 868, 844, 787,
6.1.3. Synthesis of 7-(2,4-dichlorophenyl)pyrido[30,20:4,5]thieno
[3,2-d]pyrimidin-4-amine (10)
6.1.3.1. 2-Chloro-6-(2,4-dichlorophenyl)nicotinonitrile
(7). A
mixture of 2,6-dichloropyridine-3-carbonitrile (6) (0.1 g,
0.58 mmol), 2,4-dichlorophenylboronic acid (0.12 g, 0.63 mmol),
sodium carbonate (0.184 g, 1.73 mmol) and Pd(PPh3)4 (0.034 g) in
dioxane (3 mL)/H2O (1 mL) was irradiated at 100 ꢀC (400 W) for 2 h.
On completion (followed by thin-layer chromatography), the re-
action was cooled to room temperature and H2O was added. The
powder was filtered off, washed with EtOAc and the organic layer
was evaporated in vacuo. The crude powder was purified by silica
gel column chromatography using a gradient of PE/EtOAc (100:0 to
0:100, v/v) as the eluent to give the desired compound 7 as a yellow
powder (0.143 g, 87%); mp 188e189 ꢀC; IR nmax (cmꢁ1): 2238 (CN),
1587, 1578, 1556, 1432, 1349, 1300, 1172, 1142, 1090, 1043, 1030, 881,
757; 1H NMR (300 MHz, DMSO-d6):
d
8.73 (d,1H, J ¼ 8 Hz, H-9), 8.59
(s, 1H, H-2), 7.92 (d, 1H, J ¼ 8 Hz, H-8), 7.85 (d, 1H, J ¼ 2 Hz, H-ar),
7.78 (m, 3H, H-ar and NH2), 7.63 (dd, 1H, J1 ¼ 2 Hz, J2 ¼ 8 Hz, H-ar);
13C NMR (75 MHz, DMSO-d6):
d 160.9, 158.4, 157.0, 155.6, 136.9,
134.5, 133.2, 132.3, 131.6, 129.5, 127.8, 127.7, 126.9, 121.9, 112.9;
869, 850, 817, 792; 1H NMR (300 MHz, DMSO-d6):
d
8.62 (d, 1H,
HRMS calcd for C15H9Cl2N4S [MþH]þ 346.9925 found 346.9931.
J ¼ 8 Hz, H-4), 7.98 (d, 1H, J ¼ 8 Hz, H-5), 7.86 (d, 1H, J ¼ 2 Hz, H-ar),
7.69 (d, 1H, J ¼ 9 Hz, H-ar), 7.63 (dd, 1H, J1¼2 Hz, J2 ¼ 9 Hz, H-ar); 13
C
6.2. In vitro kinase preparation and assays [17]
NMR (75 MHz, DMSO-d6):
d 158.5, 150.7, 144.4, 135.5, 134.8, 133.0,
132.2, 129.7, 128.0, 124.0, 115.0, 108.8; HRMS calcd for C12H6Cl3N2
6.2.1. Buffers
[MþH]þ 282.9597 found 282.9598.
Buffer A: MgCl2 (10 mM),
aminoethylether)-N,N,N0,N0-tetraacetic acid (EGTA), 1 mM dithio-
threitol (DTT), 25 mM TriseHCl pH 7.5, 50 g heparin/mL.
Buffer B: -Glycerophosphate (60 mM), 30 mM p-nitro-
phenylphosphate, 25 mM 3-(N-morpholino)propanesulfonic acid
(Mops) (pH 7.2), 5 mM EGTA, 15 mM MgCl2, 1 mM DTT, 0.1 mM
sodium vanadate.
1 mM ethylene glycol-bis(2-
6.1.3.2. 3-Amino-6-(2,4-dichlorophenyl)thieno[2,3-b]pyridine-2-
carbonitrile (8). A solution of potassium hydroxide (82 mg,
1.46 mmol) in H2O (0.4 mL) was added dropwise to a stirred and
cold solution (ice bath) containing 2-chloro-6-(2,4-dichlorophenyl)
m
b
nicotinonitrile
mercaptopropionitrile (0.051 g, 0.58 mmol) in DMF (4 mL). The
cold mixture was stirred for 15 min and bromoacetonitrile (52 L,
(7)
(0.138
g,
0.49
mmol)
and
3-
m
6.2.2. Kinase preparations and assays
0.73 mmol) was added dropwise. After 1 h at 0 ꢀC, the mixture was
poured onto iced water. The crude product was collected by
filtration and purified with silica gel column chromatography using
a gradient of PE/EtOAc (100:0 to 0:100, v/v) as the eluent to give the
desired compound 8 as a yellow powder (0.043 g, 28%); mp (neat)
285e286 ꢀC; IR (neat) nmax (cmꢁ1): 2192 (CN), 1641, 1577, 1532,
1477, 1449, 1407, 1383, 1347, 1209, 1142, 1107, 1091, 1031, 861, 839,
Kinase activities were assayed in triplicates in buffer A or B, for
30 min at 30 ꢀC, at a final adenosine triphosphate (ATP) concen-
tration of 15
mM. Blank values were substracted and activities
expressed in % of the maximal activity, i.e., in the absence of in-
hibitors. Controls were performed with appropriate dilutions of
dimethylsulfoxide (DMSO). IC50 values were calculated from dos-
eeresponse curves established by Sigma-Plots. The GSK-3, CK1,
DYRK1A and CLK1 peptide substrates were obtained from Proteo-
genix (Oberhausbergen, France).
813, 772; 1H NMR (300 MHz, DMSO-d6):
d
8.61 (d,1H, J ¼ 8 Hz, H-4),
7.80 (m, 2H, H-5 and H-ar), 7.69 (d, 1H, J ¼ 9 Hz, H-ar), 7.59 (dd, 1H,
J1¼2 Hz, J2 ¼ 9 Hz, H-ar), 7.43 (br s, 2H, NH2); 13C NMR (75 MHz,
DMSO-d6):
d
159.2, 156.3, 150.2, 136.8, 134.5, 133.0, 132.3, 131.5,
6.2.2.1. CDK5/p25. (Human, recombinant) was prepared as previ-
129.5, 127.7, 123.3, 121.0, 115.6, 71.8; HRMS calcd for C14H8Cl2N3S
ously described [18]. Its kinase activity was assayed in buffer A,
[MþH]þ 319.9816 found 319.9812.
with 1 mg of histone H1/mL, in the presence of 15
mM [
g-
33P] ATP
(3000 Ci/mmol; 10 mCi/mL) in a final volume of 30
mL. After 30 min
6.1.3.3. (E)-N0-(2-Cyano-6-(2,4-dichlorophenyl)thieno[2,3-b]pyridin-
incubation at 30 ꢀC, 25
mL aliquots of supernatant were spotted onto
3-yl)-N,N-dimethyl formimidamide (9). A mixture of 3-amino-6-
sheets of Whatman P81 phosphocellulose paper, and 20 s later, the
filters were washed eight times (for at least 5 min each time) in a
solution of 10 mL phosphoric acid/L of water. The wet filters were
counted in the presence of 1 mL ACS (Amersham) scintillation fluid.
(2,4-dichlorophenyl)thieno[2,3-b]pyridine-2-carbonitrile
8
(0.066 g, 0.20 mmol) and DMF-DMA (2 mL) was irradiated at 90 ꢀC
(800 W) for 30 min. On completion, the solution was cooled to
room temperature and crude products were extracted with EtOAc.
The organic layers were washed with cold H2O, dried over Na2SO4,
6.2.2.2. GSK-3a/b. (Porcine brain, native) was assayed, as described

Y. Loidreau et al. / European Journal of Medicinal Chemistry 92 (2015) 124e134
133
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(GS-1: YRRAAVPPSPSLSRHSSPHQpSEDEEE) (pS stands for phos-
phorylated serine) [19].
b) P. Cohen, Protein kinases e the major drug targets of the twenty-first
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as dual cyclin-dependent kinase 1 (CDK1)/glycogen synthase kinase-3 (GSK-3)
inhibitors: synthesis, biological evaluation and molecular modelling studies,
Eur. J. Med. Chem. 43 (2008) 1469e1477;
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CDK5/p25 but using the CK1-specific peptide substrate
RRKHAAIGpSAYSITA [20].
ꢀ
ꢀ
b) A. Testard, C. Loge, B. Leger, J.-M. Robert, O. Lozach, M. Blairvacq, L. Meijer,
6.2.2.4. DYRK1A. (Rat, recombinant, expressed in E. coli as a
glutathione transferase (GST) fusion protein) was purified by af-
finity chromatography on glutathione-agarose and assayed, as
described for CDK5/p25 using Woodtide (KKISGRLSPIMTEQ)
ꢀ
V. Thiery, T. Besson, Thiazolo[5,4-f]quinazolin-9-ones, inhibitors of glycogen
synthase kinase-3, Bioorg. Med. Chem. Lett. 16 (2006), 4319e3423;
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20 (2005) 557e568.
(1.5 mg/assay) as a substrate.
[5] a) M. Antoine, M. Gerlach, E. Günther, T. Schuster, M. Czech, I. Seipelt,
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fusion protein) was assayed in buffer A (þ0.15 mg BSA/ml) with RS
ꢀ
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b) A. Breteche, P. Marchand, M.-R. Nourrisson, P. Hautefaye, G. De Nanteuil,
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c) M. Antoine, M. Czech, M. Gerlach, E. Günther, T. Schuster, P. Marchand,
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[2,3-b]pyrazines, Synthesis (2011) 794e806.
6.3. Molecular modelling
[6] a) Y. Loidreau, P. Marchand, C. Dubouilh-Benard, M.-R. Nourrisson, M. Duflos,
Molecular modelling studies were performed using SYBYL-X 1.3
software [31] running on a Dell precision T3400 workstation. The
three-dimensional structure of compounds 1g and 10 was built
from a standard fragments library and optimized using the Tripos
force field [32] including the electrostatic term calculated from
Gasteiger and Hückel atomic charges. Powell's method available in
Maximin2 procedure was used for energy minimization until the
gradient value was smaller than 0.001 kcal/mol.Å. The crystal
structures of CLK1 in complex with debromohymenialdisine at
€
N. Loaec, L. Meijer, T. Besson, Synthesis and biological evaluation of N-aryl-7-
methoxybenzo[b]furo[3,2-d]pyrimidin-4-amines and their N-arylbenzo[b]
thieno[3,2-d]pyrimidin-4-amine analogues as dual inhibitors of CLK1 and
DYRK1A kinases, Eur. J. Med. Chem. 59 (2013) 283e295;
b) Y. Loidreau, P. Marchand, C. Dubouilh-Benard, M.-R. Nourrisson, M. Duflos,
€
O. Lozach, N. Loaec, L. Meijer, T. Besson, Synthesis and biological evaluation of
N-arylbenzo[b]thieno[3,2-d]pyrimidin-4-amines and their pyrido and pyr-
azino analogues as Ser/Thr kinase inhibitors, Eur. J. Med. Chem. 58 (2012)
171e183.
[7] Y. Loidreau, P. Marchand, C. Dubouilh-Benard, M.-R. Nourrisson, M. Duflos,
T. Besson, First synthesis of 4-aminopyrido[2',3':4,5]furo[3,2-d]pyrimidines,
Tetrahedron Lett. 53 (2012) 944e947.
1.70 Å resolution (pdb code: 1Z57) [27] and CK1d in complex with
€
[8] E. Deau, Y. Loidreau, P. Marchand, M.-R. Nourrisson, N. Loaec, L. Meijer,
PF4800567 at 2.07 Å resolution (pdb code: 4HNF) [28] were used as
templates for docking. The original ligands as wsell as the water
molecules were removed from the coordinates set. Flexible docking
of compounds 1g and 10 into ATP-binding site was performed using
GOLD software [33]. A distance range of 1.5e3.0 Å between the N3
atom of inhibitors and the main chain NH of Leu244/Leu85 (CLK1/
CK1) from the hinge region was set as a constraint. The most stable
docking model was selected according to the best scored confor-
mation predicted by the GoldScore scoring function.
V. Levacher, T. Besson, Synthesis of novel 7-substituted pyrido[2',3':4,5]furo
[3,2-d]pyrimidin-4-amines and their N-aryl analogues and evaluation of their
inhibitory activity against Ser/Thr Kinases, Bioorg. Med. Chem. Lett. 23 (2013)
6784e6788.
ꢀ
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[9] D. Hedou, R. Guillon, C. Lecointe, C. Loge, E. Chosson, T. Besson, Novel syn-
thesis of angular thiazolo[5,4-f] and [4,5-h]quinazolines, preparation of their
linear thiazolo[4,5-g] and [5,4-g]quinazoline analogs, Tetrahedron 69 (2013)
3182e3191.
[10] For recent papers and revues on the interest of MTDLs in Alzheimer’s disease
treatment see: a) C. Schmitt, P. Miralinaghi, M. Mariano, R.W. Hartmann,
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and DYRK1A Inhibitors, Eur. J. Med. Chem. 80 (2014) 352e363;
Acknowledgements
ꢀ
This work was supported by a PhD grant from the Region Haute-
c) M. Bajda, N. Guzior, M. Ignasik, B. Malawska, Multi-target-directed ligands
in Alzheimer's disease treatment, Curr. Med. Chem. 18 (2011) 4949e4975;
d) A. Cavalli, M.L. Bolognesi, A. Minarini, M. Rosini, V. Tumiatti, M. Recanatini,
C. Melchiorre, Multi-target-directed ligands to combat neurodegenerative
diseases, J. Med. Chem. 51 (2008) 347e372.
Normandie (YL) and the ISCE-CHEM program. We thank the LABEX
SynOrg (ANR-11-LABX-0029) and AI-Chem Channel program for
financial support. TB acknowledges Milestone S.r.l. (Italy) for
providing the microwave reactors, financial and technical support.
This research was supported by grants from the “Fonds Unique
[11] a) Y. Loidreau, T. Besson, Microwave-assisted thermal decomposition of
formamide: a tool for coupling a pyrimidine ring with an aromatic partner,
Tetrahedron 67 (2011) 4852e4857;
ꢀ
Inter-ministeriel” (FUI) PHARMASEA and TRIAD projects (LM), the
ꢁ
“Association France-Alzheimer (Finistere)” (LM), and the European
b) I. Nouira, I.K. Kostakis, C. Dubouilh, E. Chosson, M. Iannelli, T. Besson,
Decomposition of formamide assisted by microwaves, a tool for synthesis of
nitrogen-containing heterocycles, Tetrahedron Lett. 49 (2008) 7033e7036;
c) I.K. Kostakis, H. Elomri, E. Seguin, M. Iannelli, T. Besson, Rapid synthesis of
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Union 7th Framework Program Knowledge-Based Bio-economy
(FP7-KBBE)- BlueGenics 2012 grant (LM).
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2014.12.038.
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