Discovery of thienopyrimidine-based FLT3 inhibitors from the structural modification of known IKKβ inhibitors

Article abstract of DOI:10.1016/j.bmcl.2014.04.058

Inactivation of the NF-κB signaling pathway by inhibition of IKKβ is a well-known approach to treat inflammatory diseases such as rheumatoid arthritis and cancer. Thienopyrimidine-based analogues were designed through modification of the known IKKβ inhibitor, SPC-839, and then biologically evaluated. The resulting analogues had good inhibitory activity against both nitric oxide and TNF-α, which are well-known inflammatory responses generated by activated NF-κB. However, no inhibitory activity against IKKβ was observed with these compounds. The thienopyrimidine-based analogues were subsequently screened for a target kinase, and FLT3, which is a potential target for acute myeloid leukemia (AML), was identified. Thienopyrimidine-based FLT3 inhibitors showed good inhibition profiles against FLT3 under 1 μM. Overall, these compounds represent a promising family of inhibitors for future development of a treatment for AML.

Full text of DOI:10.1016/j.bmcl.2014.04.058

Bioorganic & Medicinal Chemistry Letters xxx (2014) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of thienopyrimidine-based FLT3 inhibitors from the
structural modification of known IKKb inhibitors
Chun-Ho Park a,c, , Chulho Lee b, , Jee Sun Yang b, Bo-Young Joe c, Kwangwoo Chun c, Hyuntae Kim b,d
,
Hye Yun Kim e, Jong Soon Kang f, Jangik I. Lee g, Myung-Hwa Kim c, , Gyoonhee Han b,d,
⇑
⇑
a Graduate Program in Biomaterials Science & Engineering, Yonsei University, Seodaemun-gu, Seoul 120-749, Republic of Korea
b Translational Research Center for Protein Function Control, Department of Biotechnology, Yonsei University, Seodaemun-gu, Seoul 120-749, Republic of Korea
c Division of New Drug Development, R&D Center, Jeil Pharmaceutical Co., Ltd 117-1, Keungok-Ri, Baekam-Myun, Cheoin-Gu, Yongin-City, Kyunggi-Do 449-861, Republic of Korea
d Department of Biomedical Sciences (WCU Program), Yonsei University, Seodaemun-gu, Seoul 120-749, Republic of Korea
e Center for Neuro-Medicine of Brain Science Institute, Korea Institute of Science and Technology, Hwarangno 14-gil 5, Seongbuk-gu, Seoul 136-791, Republic of Korea
f Bioevaluation Center, Korea Research Institute of Bioscience and Biotechnology, Ochang, Cheongwon, Chungbuk 363-883, Republic of Korea
g College of Pharmacy, Yonsei University, 85 Songdogwahak-ro, Yeonsu-gu, Incheon 406-840, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Inactivation of the NF-jB signaling pathway by inhibition of IKKb is a well-known approach to treat
inflammatory diseases such as rheumatoid arthritis and cancer. Thienopyrimidine-based analogues were
designed through modification of the known IKKb inhibitor, SPC-839, and then biologically evaluated.
Received 19 March 2014
Revised 15 April 2014
Accepted 16 April 2014
Available online xxxx
The resulting analogues had good inhibitory activity against both nitric oxide and TNF-
a, which are
well-known inflammatory responses generated by activated NF- B. However, no inhibitory activity
j
against IKKb was observed with these compounds. The thienopyrimidine-based analogues were subse-
quently screened for a target kinase, and FLT3, which is a potential target for acute myeloid leukemia
(AML), was identified. Thienopyrimidine-based FLT3 inhibitors showed good inhibition profiles against
Keywords:
FLT3
IKKb
Anti-inflammation
Thienopyrimidine
Acute myeloid leukemia (AML)
FLT3 under 1
lM. Overall, these compounds represent a promising family of inhibitors for future devel-
opment of a treatment for AML.
Ó 2014 Elsevier Ltd. All rights reserved.
FLT3 (Fms-like tyrosine kinase 3), a receptor tyrosine kinase, is a
member of receptor III tyrosine kinase family including PDGFR, KIT
and FMS.1 FLT3 is normally expressed on the surface of hematopoi-
etic progenitor cells by immature hematopoietic cell and it has
important roles in normal stem cell development and the immune
system.2,3 Abnormal overexpression and mutation of FLT3 is often
observed in patients with leukemia. Specifically, diverse mutations
of FLT3, such as D835V, D835Y, and ITD (internal tandem duplica-
tion), were reported in AML.4 AML cells induce molecular defects,
which promote leukemic proliferation, defect in differentiation
and resistance to apoptosis.5 Increased transcriptional level of
FLT3 is observed in AML, and then this increase of expression level
may influence to phosphorylation of FLT3.6 The phosphorylation
and activation of FLT3 receptor may cause the activation of a
down-stream kinase pathway, such as Ras/Mitogen-activated pro-
tein kinase (MAPK), which can induce an abnormal cell growth and
gene regulation.3,7,8 For these reasons, FLT3 is recently regarded as
a one of important target for treatment of AML patients.
Our primary research purpose was to discover the IKKb inhibi-
tors for the treatment of inflammatory diseases. I
is considered as a promising target for diverse diseases related
with NF- B pathway including inflammatory diseases and autoim-
mune diseases such as rheumatoid arthritis (RA).9–14 Recently, new
structural and functional data for IKK and IKKb have revealed that
IKKb is a useful target of the NF- B activation pathway for the
jB kinase b (IKKb)
j
a
j
development of anti-inflammatory and anti-cancer therapeutic
agents.15–17 We have designed and synthesized thienopyrimidine
analogues as potent IKKb inhibitors, however those analogues did
not have IKKb inhibitory activities. From an assay, FLT3 has been
discovered as a new target for thienopyrimidine analogues. Herein,
we describe this novel series of thienopyrimidine-based FLT3
inhibitors made through the modification of IKKb inhibitors.
Toward the discovery of small molecule inhibitors of IKKb, we
have performed fragment-based virtual screening from known
⇑
Corresponding authors at present address: Korea Drug Development Fund, 14th
Fl, KT&G Seodaemun Tower, 21-1, Migeun-dong, Seodaemun-gu, Seoul, Republic of
Korea. Tel.: +82 2 261 3630; fax: +82 2 361 3609 (M.-H.K.); tel.: +82 2 2123 2882;
fax: +82 2 362 7265 (G.H.).
E-mail addresses: mhkim@kddf.org (M.-H. Kim), gyoonhee@yonsei.ac.kr (G.
Han).
Designates co-first authorship.
http://dx.doi.org/10.1016/j.bmcl.2014.04.058
0960-894X/Ó 2014 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058

2
C.-H. Park et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
IKKb active compounds libraries22,23 and the thienopyrimidine
core has been discovered. We have designed and synthesized a
novel series of thienopyrimidine derivatives through structural
modifications of the thienopyrimidine core and the quinazoline
analogue SPC-839, and then biologically evaluated these deriva-
tives (Fig. 1). We introduced thienopyrimidine as a core instead
of quinazoline and diversified its functional groups, using alkyl,
substituted aromatic, and heterocyclic groups at positions 2, 4
and 5 of the thienopyrimidine core.
Early attempts to synthesize thienopyrimidine-based analogues
1 began with modifications of the 3-methyl-1H-pyrrole-2,5-dione
moiety at C4 position, followed by substitutions of moderate sizes
at the C2 and C5 positions. The synthetic routes for the newly
synthesized thienopyrimidine derivatives are outlined in
Schemes 1–3.
Compounds 8a–e in Table 1 were synthesized from 2-acetyl-
thiophene according to Scheme 1. Knoevenagel condensation of
2-acetylthiophene with malononitrile in the presence of NH4OAc
and AcOH yielded 2-[1-(thiophen-2-yl)ethylidene]malononitrile
2,18 which was treated with elemental sulfur and piperidine to pro-
duce thiophene 3a,19 Compound 3a was heated with formamide at
180 °C to produce thienopyrimidine 4a, which was converted to
chloride 5a by Sandmeyer reaction under CuCl2 and t-BuONO.20,21
5-(Thiophen-2-yl)thieno[2,3-d]pyrimidin-4-amine derivatives 8a–
e was then prepared in a stepwise fashion by first treating 5a with
hydrazine hydrate in tetrahydrofuran followed by reacting with
the appropriate furans 7a–e.
The analogues 13a–m were prepared from two synthetic meth-
ods. Compounds 3b–e were heated with commercially available
substituted cyanide under acidic condition at 110 °C to produce
thienopyrimidine 4b–e, which were converted to chloride 5b–e
by the Sandmeyer reaction under CuCl2 and t-BuONO according
to method 1 of Scheme 2. Substituted 2-aminothiophen-3-carbox-
ylate compounds 10a–i were obtained by the Gewald reaction as
depicted in method 2 of Scheme 2. Compounds 11a–i were conve-
niently synthesized by heating carboxylates 10a–i with an appro-
priate carbonitrile under acidic conditions, which were
chlorinated using POCl3 at 100 °C to produce 4-chloro-thieno[2,3-
d]pyrimidine 12a–i. Thieno[2,3-d]pyrimidin-4-amine derivatives
13a–m were then prepared in a stepwise fashion by first treating
5a–d or 12a–i with hydrazine hydrate in tetrahydrofuran, followed
by reacting with 3-methylfuran-2,5-dione.
Synthesis of analogues 17a–d, containing a phenol group
substituted with terminal amine units at the C5-position of thie-
no[2,3-d]pyrimidine, is described in Scheme 3. Compounds 16a–
d were synthesized from chlorides 14a–d, which were obtained
from phenols 12h and 12i through the Mitsunobu reaction accord-
ing to the same method described above for the preparation of
3-methyl-1H-pyrrole-2,5-dione derivatives 13a–m. Finally, the N-
substituted amine salt compounds 17a–d were prepared using
4 M HCl in 1,4-dioxane to enhance water solubility.
Our initial studies investigated structure–activity relationships
that optimized inhibitory activity against production of nitric oxide
O
O
(NO) and TNF-
by activated NF-
a summary of results of NO and TNF-
a
, two well-known inflammatory responses caused
B in LPS-treated RAW264.7 cells.24 Table 1 shows
inhibition assay by the thi-
j
N
N
N
HN
4
a
R2
5
O
HN
O
enopyrimidine derivatives with various pyrrole substituents at the
C4 position. Among those analogues, we found that thienopyrimidine
8b, with 3-methyl-1H-pyrrole-2,5-dione moiety, displayed significant
O
N
2
S
S
N
R1
N
activity in the NO inhibition assay (IC50 = 2.49
lM), but did not show
subsequent activity in the TNF- inhibition assay. Except for com-
pound 8b, most of C4-substituted analogues, such as 8a with 1H-
a
SPC-839
Thienopyrimidine-based analogues (1)
pyrrole-2,5-dione and 8c with 3,4-dimethyl-1H-pyrrole-2,5-dione,
Figure 1. Known IKKb inhibitor, SPC-839 and thienopyrimidine-based analogues 1.
N
N
O
a
b
N
N
N
+
+
S
S
S
NH2
S
S
S
2
3
NH2
S
S
NH2
N
HN
N
Cl
N
e
c
d
N
N
S
S
S
N
4
5
6
O
A
A
N
N
f
S
HN
+
O
O
O
O
S
N
7a-e
8a-e
Scheme 1. (a) NH4OAc, AcOH, toluene, reflux; (b) piperidine, EtOH, 80 °C; (c) HCONH2, 180 °C; (d) CuCl2, t-BuONO, THF/MeCN, 70 °C; (e) H2N-NH2ÁH2O, THF, 80 °C; (f) CHCl3,
80 °C.
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058

C.-H. Park et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
3
Method 1
N
NH2
R1
Cl
N
R1
S
a
N
b
N
N
R1
+
NH2
R2
S
R2
N
R2
S
3b-e
4b-e
5b-e
Method 2
N
O
O
O
O
c
d
N
+
S
R1
+
O
R1
9a-i
OH
O
Cl
N
R1
S
R1
O
N
e
f
N
R1
N
+
R2
NH2
S
N
R2
R2
S
10a-i
11a-i
12a-i
O
NH2
N
N
Cl
N
HN
N
HN
R1
S
R1
S
R1
S
g
h
O
N
N
+
O
R2
R2
O
N
R2
O
5b-e or 12a-i
6b-n
13a-m
Scheme 2. (a) 4 M HCl, 1,4-dioxane, 110 °C; (b) CuCl2, t-BuONO, THF/MeCN, 70 °C; (c) NH4OAc, AcOH, toluene, reflux; (d) piperidine, EtOH, 80 °C; (e) 4 M HCl, 1,4-dioxane,
110 °C; (f) POCl3, 100 °C; (g) H2N-NH2ÁH2O, THF, 80 °C; (h) CHCl3, 80 °C.
OH
OR
OR
NH2
N
Cl
N
Cl
N
HN
N
a
b
N
N
+
O
O
S
S
S
S
S
S
O
12h-i
14a-d
15a-d
OR
S
O
OR
O
N
N
N
N
HN
HN
c
d
O
O
HCl
S
S
S
N
N
16a-d
17a-d
Scheme 3. (a) R-OH, PPh3, DIAD, THF; (b) H2N-NH2ÁH2O, THF, 80 °C; (c) CHCl3, 80 °C; (d) 4 M HCl, 1,4-dioxane.
did not exhibit NO inhibitory activity. To maintain NO inhibitory
activity in downstream reactions, we fixed 3-methyl-1H-pyrrole-
2,5-dione moiety in the C4 position, and then proceeded with the
additional steps.
aromatic, phenyl and heterocyclic, such as furan, thiophene and
pyridine) at C2 and C5 position of thienopyrimidine core structure.
Table 2 summarizes the inhibitory activity against IKKb of the
tested compounds. Most of thienopyrimidine analogues showed
Next, we modified the C2 and C5 positions of thienopyrimidine
analogues based on their inhibitory activity of cellular NO and
good inhibition profile against NO and TNF-
3.36 M, respectively). SPC-839, a well-known IKKb inhibitor, gave
an inferior inhibition profile (4.41 and 3.32 M, respectively)
a (average = 3.13 and
l
TNF-a. We introduced various substituents, (methyl, monocyclic
l
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058

4
C.-H. Park et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
Table 1
Structure–activity-relationship of compounds varied at the C4-position
O
X
N
S
HN
N
A
X =
N
O
R1
S
Compound
R1
X
Cell-based assaya
IKKb kinase assay
Lance ULight system ( M) Radioisotope system (lM)
NO IC50
>10
(
l
M)
TNF-a
IC50
(lM)
l
O
N
8a
H
NA
NT
NT
O
O
N
8b
8c
8d
H
H
H
2.49
>10
>10
>10
NA
NA
NT
NT
NT
NT
NT
NT
O
O
N
O
O
N
O
O
N
N
8e
H
—
>10
NA
NT
NT
NT
O
SPC-839
—
4.41
3.32
0.062b
NT: Not tested, NA: Not active.
a
Values are means of three experiments.
The value was reported from Celgene.
b
compared to most of the thienopyrimidine analogues. In particular,
13k, 2-thiophene at C2 position and phenyl at C5 position showed
are implicated in the inflammatory response, including IKKb, and
performed a kinase panel assay (KinaseProfiler™, Merck Millipore,
UK) with compounds 13k and 17a. As shown in Table 3, represen-
tative thienopyrimidine analogues 13k and 17a showed no inhibi-
tion profile against IKKb (116% and 84% remaining activities with
excellent NO and TNF-
respectively). However, most of the analogues did not demonstrate
significant inhibition of IKKb (IC50 = 59.45 ꢀ 365.9 M), while
these analogues showed good inhibitory activities against NO
and TNF- . For example, compound 17c, with relatively poor NO
and TNF- inhibitory activity (7.54 and 9.88 M, respectively),
showed very low IKKb inhibitory activity about 365.9 M. In
contrast, compound 13k, with potent inhibition against NO and
TNF- (0.75 and 1.29 M, respectively), also showed poor IKKb
inhibitory activity over 50 M. Overall, we found that these
derived compounds demonstrated poor IKKb inhibition regardless
of their ability to inhibit NO and TNF- . To confirm the IKKb
a inhibitory activity (0.75 and 1.29 lM,
l
10
inhibitory activity against FLT3 (8% and 4% remaining activities
in 10 M, respectively).
lM treatment). However, these analogues have significant
a
a
l
l
l
Once we identified FLT3 as a possible target kinase for the thi-
enopyrimidine-based analogues, we selected 10 analogues with
good NO and TNF-a inhibition to test FLT3 kinase inhibition with
a FLT3-specific assay. As shown Table 4, most of thienopyrimidine
a
l
l
analogues showed good inhibition profiles against FLT3
a
(IC50 = 0.065 ꢀ 0.750
of inhibitory profiles with well-known FLT3 inhibitors, AC220 and
MLN518 (IC50 = 0.120 and 0.102 M, respectively). Among those
tested compounds, 13i25 with methyl group at C5 position showed
the best FLT3 inhibitory activity (IC50 = 0.065 M). 13h with hydro-
gen at C5 position showed less inhibitory activity (IC50 = 0.175 M)
lM). These analogues showed a similar range
inhibition profile of thienopyrimidine analogues, we performed a
second IKKb kinase assay using a radioisotope system (Merck
Millipore). However, most analogues showed no IKKb inhibitory
activity by this second assay, and a few analogues showed very
l
l
low inhibitory activity, with IC50 >50
Even though thienopyrimidine-based analogues showed no
IKKb inhibitory activity, they could inhibit NO and TNF- . These
lM.
l
than 13i. Compounds with the hydroxyl group at the meta- or para-
position relative to the phenyl group (13l and 13m) at C5 position
a
results imply that thienopyrimidine analogues may target another
kinase in the signaling cascade. To identify the target kinase of the
thienopyrimidine-based analogues, we selected 22 kinases, which
(0.488 and 0.750
than the unsubstituted phenyl group (13k, 0.208
appropriate bulky groups such as a piperidinylethoxy group on
l
M, respectively) showed less inhibitory activity
l
M). However,
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058

C.-H. Park et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
5
Table 2
Structure–activity-relationships of compounds varied at the C2- and C5-positions
O
N
N
R1
S
HN
O
N
R2
Compound
R1
R2
Cell-based assaya
M) TNF- IC50
IKKb kinase assay
M) Lance ULight system ( M) Radioisotope system (lM)
NO IC50
(l
a
(l
l
13a
13b
13c
13d
13e
13f
13g
13h
13i
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
H
Methyl
CH2COOEt
Phenyl
3-Hydroxy-phenyl
4-Hydroxy-phenyl
Thiophen-2-yl
Phenyl
4-Pyridinyl
Cyclopropanyl
Furan-2-yl
2.19
3.13
2.11
3.12
5.25
0.71
1.13
>10
NT
NT
NT
NT
NT
NT
NT
>50
>50
>50
>50
NT
NT
NT
NT
NT
NT
NT
NT
NT
NA
NA
NA
NA
NA
NA
4.35
3.96
1.24
1.39
8.95
4.22
5.23
1.29
2.67
1.32
3-Methoxy phenyl 3.47
4-Methoxy phenyl 2.79
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
6.98
1.92
5.07
0.75
1.23
0.82
13j
13k
13l
13m
O
17a
17b
17c
Thiophen-2-yl
Thiophen-2-yl
Thiophen-2-yl
2.15
3.26
7.54
2.92
2.38
9.88
NT
NT
N
HCl
O
NT
>50
>50
N
HCl
O
O
>50
N
HCl
O
N
17d
Thiophen-2-yl
1.68
2.14
NT
NA
HCl
SPC-839
Average (13a–17d)
—
—
—
—
4.41
3.14
3.32
3.36
0.062b
—
NT
—
NT: Not tested, NA: Not active.
a
Values are means of three experiments.
The value was reported from Celgene.
b
Table 3
Remaining kinase activity profiles of kinases by 13k and 17a (%)
Kinase
Compound (10
l
M)a
Kinase
Compound (10 lM)
13k
17a
13k
17a
BTK(h)
FLT3(h)
76
8
119
116
103
87
94
121
86
92
102
100
4
98
84
111
93
117
112
74
81
94
MKK4(m)
MKK6(h)
MKK7b(h)
MSK1(h)
PKA(h)
ROCK-II(h)
SAPK2a(h)
SAPK2b(h)
SAPK3(h)
SAPK4(h)
TYK2(h)
113
99
122
96
125
99
92
98
109
90
112
96
108
102
126
96
96
103
99
IKKa
(h)
IKKb(h)
JAK1(h)
JNK1
JNK2
a
a
1(h)
2(h)
JNK3(h)
Lck(h)
Lyn(h)
MAPKAP-K2(h)
100
87
86
a
Remaining was determined by using the KinaseProfiler service at Millipore.
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058

6
C.-H. Park et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
Table 4
FLT3 kinase assay of thienopyrimidine-based analogues
a
Compound
FLT3 kinase assay IC50
(
lM)
Compound
FLT3 kinase assay IC50 (lM)
13h
13i
13j
13k
13l
AC220
0.175 0.229
0.065 0.125
0.673 0.113
0.208 0.161
0.488 0.214
0.120 0.030
13m
17a
17b
17c
17d
0.750 0.109
0.133 0.084
0.540 0.098
0.175 0.056
0.085 0.190
0.102 0.303
MLN518
a
IC50 was determined by using the KinaseProfiler service at Millipore. Values are means of duplicates.
Table 5
Remaining kinase activity and tumor growth inhibition profiles of 13i
Kinasea (%)
GI50
(lM)
b
Compound
FLT3(h)
FLT3(D835Y)(h)
K562
MV4-11
HL60
THP1
13i
5
1
2
0
0.257 0.1189
0.003 0.0298
0.264 0.1459
0.111 0.2463
a
Remaining activity% in 10
Growth inhibition was measured by XTT assay. Values are means of four experiments.
l
M. Values are means of duplicates.
b
the meta- or para-positions (17a and 17d) relative to the phenyl
058. These data include MOL files and InChiKeys of the most
important compounds described in this article.
group at the C5 position (0.133 and 0.085
better FLT3 inhibitory activities.
lM, respectively) showed
The most active compound 13i has been selected to evaluate
the inhibitory profiles against wild type and mutant FLT3. As
shown in Table 5, 13i showed excellent inhibitory activity against
References and notes
1. Plowman, G. D.; Sudarsanam, S.; Bingham, J.; Whyte, D.; Hunter, T. Proc. Natl.
Acad. Sci. U.S.A. 1999, 96, 13603.
2. Rosnet, O.; Buhring, H. J.; deLapeyriere, O.; Beslu, N.; Lavagna, C.; Marchetto, S.;
Rappold, I.; Drexler, H. G.; Birg, F.; Rottapel, R.; Hannum, C.; Dubreuil, P.;
Birnbaum, D. Acta Haematol-Basel 1996, 95, 218.
wild type FLT3 with about 5% remaining activity at 10
lM, as well
as 2% remaining activity at 10 M against D835Y mutant FLT3. In
l
addition, 13i was evaluated for cell growth inhibition activities
using four leukemia cell lines: MV4-11, THP1, HL60 and K562.
Among these 4 different leukemia cell lines, 13i showed better
inhibitory activities against MV4-11 and THP1 with wild type
3. Gilliland, D. G.; Griffin, J. D. Blood 2002, 100, 1532.
4. Yokota, S.; Kiyoi, H.; Nakao, M.; Iwai, T.; Misawa, S.; Okuda, T.; Sonoda, Y.; Abe,
T.; Kahsima, K.; Matsuo, Y.; Naoe, T. Leukemia 1997, 11, 1605.
5. Frelin, C.; Imbert, V.; Griessinger, E.; Peyron, A. C.; Rochet, N.; Philip, P.;
Dageville, C.; Sirvent, A.; Hummelsberger, M.; Berard, E.; Dreano, M.; Sirvent,
N.; Peyron, J. F. Blood 2005, 105, 804.
6. Ozeki, K.; Kiyoi, H.; Hirose, Y.; Iwai, M.; Ninomiya, M.; Kodera, Y.; Miyawaki, S.;
Kuriyama, K.; Shimazaki, C.; Akiyama, H.; Nishimura, M.; Motoji, T.; Shinagawa,
K.; Takeshita, A.; Ueda, R.; Ohno, R.; Emi, N.; Naoe, T. Blood 2004, 103, 1901.
7. Takahashi, S.; McConnell, M. J.; Harigae, H.; Kaku, M.; Sasaki, T.; Melnick, A. M.;
Licht, J. D. Blood 2004, 103, 4650.
8. Takahashi, S.; Harigae, H.; Kaku, M.; Sasaki, T.; Licht, J. D. Biochem. Biophys. Res.
Commun. 2004, 316, 85.
9. Yang, L.; Cohn, L.; Zhang, D. H.; Homer, R.; Ray, A.; Ray, P. J. Exp. Med. 1998, 188,
1739.
10. Barnes, P. J.; Adcock, I. M. Trends Pharmacol. Sci. 1997, 18, 46.
11. Akama, K. T.; Albanese, C.; Pestell, R. G.; Van Eldik, L. J. Proc. Natl. Acad. Sci.
U.S.A. 1998, 95, 5795.
12. Mercurio, F.; Manning, A. M. Oncogene 1999, 18, 6163.
13. Rayet, B.; Gelinas, C. Oncogene 1999, 18, 6938.
and mutant FLT3 (IC50 = under 0.1 and 0.111
than the inhibitory activities against FLT-ITD negative K562 and
HL60 cell lines (0.257 and 0.264 M, respectively).
lM, respectively)
l
Thienopyrimidine-based FLT3 inhibitors were designed from the
well-known IKKb inhibitor, SPC-839. However, the resulting ana-
logues appear to target a different kinase, FLT3. We evaluated thie-
nopyrimidine analogues with good NO and TNF-
profiles, well-known inflammatory responses that indicate acti-
vated NF- B. However, thienopyrimidine analogues exhibited no
a inhibition
j
IKKb inhibitory activity. Instead, we have identified several thieno-
pyrimidine analogues that act to inhibit FLT3. Furthermore, these
analogues demonstrated inhibitory activities against mutant FLT3
as well. The most active compound, 13i had good inhibitory profiles
on wild and mutant FLT3, and it also had potent inhibitory activities
against AML cell lines with/without FLT3-ITD mutation. Thus, these
analogues could represent a promising class of drugs for AML. Future
studies will focus on optimizing these thienopyrimidine analogues.
14. Schneider, A.; Martin-Villalba, A.; Weih, F.; Vogel, J.; Wirth, T.; Schwaninger, M.
Nat. Med. 1999, 5, 554.
15. Karin, M.; Yamamoto, Y.; Wang, Q. M. Nat. Rev. Drug Disc. 2004, 3, 17.
16. Blake, S. M.; Swift, B. A. Curr. Opin. Pharmacol. 2004, 4, 276.
17. Strnad, J.; Burke, J. R. Trends Pharmacol. Sci. 2007, 28, 142.
18. Mowry, D. T. J. Am. Chem. Soc. 1945, 67, 1050.
19. Hassan, A. Y.; Ghorab, M. M.; Nassar, O. M. Phosphorus, Sulfur Silicon Relat. Elem.
1998, 134, 77.
20. Nguyen, P.; Corpuz, E.; Heidelbaugh, T. M.; Chow, K.; Garst, M. E. J. Org. Chem.
2003, 68, 10195.
Acknowledgments
21. Wollein, G.; Troschütz, R. J. Heterocycl. Chem. 2002, 39, 1195.
22. Sun, H. P.; Zhu, J.; Chen, F. H.; Zhang, S. L.; Zhang, Y.; You, Q. D. Eur. J. Med.
Chem. 2011, 46, 3942.
23. Xie, H. Z.; Liu, L. Y.; Ren, J. X.; Zhou, J. P.; Zheng, R. L.; Li, L. L.; Yang, S. Y. J.
Biomol. Struct. Dyn. 2011, 29, 165.
24. Lee, J.; Rhee, M. H.; Kim, E.; Cho, J. Y. Mediators Inflamm. 2012, 11.
25. 3-Methyl-1-[5-methyl-2-(thiophen-2-yl)thieno[2,3-d]pyrimidin-4-ylamino]-
1H-pyrrole-2,5-dione (13i): 1H NMR (400 MHz, CDCl3) d 7.78 (d, 1H, J = 3.6 Hz),
7.40 (s, 1H), 7.37 (d, 1H, J = 5.2 Hz), 7.07 (t, 1H, J = 4.4 Hz), 6.87 (s, 1H), 6.62 (s,
1H), 2.58 (s, 3H), 2.26 (s, 3H); ESI (m/z) 357 (MH+); HRMS (ESI) calcd for
C16H12N4O2S2 [MH+]: 357.0480, found: 357.0464.
This research was supported by Grants from the Translational
Research Center for Protein Function Control (2009-0083522), the
Ministry of Health & Welfare (A120478), the National Research
Foundation funded by
Ministry of
Education (NRF-
2013R1A1A2008165) and the Ministry of Science, ICT & Future Plan-
ning (NRF-2006-511-C00060 and NRF-2013M3A6A4072536),
Republic of Korea, and the Yonsei University Research Fund of 2012.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2014.04.
Please cite this article in press as: Park, C.-H.; et al. Bioorg. Med. Chem. Lett. (2014), http://dx.doi.org/10.1016/j.bmcl.2014.04.058