2
K. N. More et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
NH2
N
study. This preliminary result encouraged us to pursue modifica-
H2N
H2N
N
N
tion of meridianin C to find more potent inhibitors against the
pim kinases. Previous reports and modeling study suggested that
pyrazine-2-yl group at 5-position of indole in meridianin C is suit-
able to make interaction with Lys67 in pim kinase pocket.12 Substi-
tution of 6-ethoxypyrazine at C-5 of indole improved inhibitory
activity against Pim-1 kinase more than an order of magnitude
(Table 1). To further improve the potency and hydrophilicity of
7a, 6-ethoxy group on pyrazine was replaced with aminoalkyl
groups using either amine or ether linkage. Addition of 2-amino-
ethyl group by amine linkage provided the compounds showing
significantly improved potency against Pim-1 kinase. The activity
gain may be attributed by two factors. First, the difference in
basicity of pyrazine depending on the type of atom at 2-position
of pyrazine may affect the interaction with Lys67. Alkoxy group
at 2-position of pyrazine is less basic than amino group by 2 pKb
units.25 Secondly, the interaction of aminoalkyl substituent with
carbonyl and/or carboxyl groups of enzyme is influenced by the
position of amino group. Two methylene units between amino
group of aminoalkyl substituent and nitrogen linker were found
better than three methylene units. The potency of compound
against Pim-3 kinase was similar to Pim-1 and showed very close
substituent dependency. However, the inhibitory activity of
compound against Pim-2 kinase was lower by more than an order
of magnitude compared to other isozymes. Replacement of 2-ami-
nopyrimidine ring at C-3 position of indole with substituted ketone
group considerably decreased the Pim-1 potency by more than 50
fold (Tables 1 and 2. 7b vs 7k, 7f vs 7l). The effect of 2-aminopy-
rimidine on the potency could be due to both hydrophobic interac-
tion26 and hydrogen bond interaction with enzyme,16 as
demonstrated by docking study (Fig. 2).
R
N
N
N
R2
F
Br
N
R
N
N
N
H
R1
H
(A)
(B)
(C)
Figure 1. Structures of meridianin
C and synthesized indole analogues. (A)
synthesized meridianin analogues, (B) meridianin C, (C) 3,5-disubstituted indole
as an inhibitor of pan-pim kinases.
presented as very potent and selective inhibitor of pan-pim kinases
(Fig. 1).12
Since meridianin C (3,5-disubstituted indole) exhibited a single
digit micromolar potency for Pim-1, we sought to optimize its scaf-
fold for in vitro potency against Pim-1 as well as other family
members such as Pim-2 and 3. Further modification and structure
activity relationship towards meridianin C core gave us potent
compounds with single digit nanomolar potency.
The general synthetic route for synthesis of 3-(2-aminopyrimi-
dine)-5-substituted indole series is depicted in Scheme 1. The pro-
tection of 5-bromoindole with tosyl chloride under basic
conditions followed by acetylation at C-3 position by general Fri-
edel–Crafts acylation provided compound 3. Reaction of compound
3 with N,N-dimethylformamide diethyl acetal in N,N-dimethyl-
formamide provided enaminone 4, which was used in the last step
for the synthesis of 2-aminopyrimdine ring at C-3 position of the
indole.23 Pinacol ester was introduced at C-5 position by Miyaura
borylation.24 Compound 6 was obtained by palladium catalyzed
Suzuki type coupling of compound 5 with 6-substituted-2-chloro-
pyrazine (Het-Cl). Cyclization to 2-aminopyrimidine with simulta-
neous cleavage of tosyl group using guanidine and K2CO3
facilitated the preparation of final compound. Synthesis of
compound 7k and 7l was done by removal of tosyl group from
compound 6f and 6b, respectively.
A molecular modeling based on the reported X-ray structure
(PDB code: 3BGQ, Human Pim-1 kinase in complex with an triazo-
lopyridazine inhibitor VX2) showed that compound 7f was bound
in the ATP binding pocket of Pim-1 (Fig. 2).27 As expected, one
nitrogen atom of pyrazine forms a hydrogen bonding interaction
(3.13 Å) with the e-amino group in the side chain of Lys67. 2-Ami-
nopyrimidine rotates about 4 degree relative to the indole ring,
which makes its amino group to form a hydrogen bond (3.28 Å)
with the backbone carbonyl group of Pro123. In addition, there
Meridianin C bearing 2-aminopyrimidine at C-3 and bromine at
C-5 position was chosen as starting point in our study. Meridianin
C was found to exhibit IC50 of 1.04
l
M in Pim-1 kinase inhibition
O
NH2
NH
N
N
(a)
N
(b)
Ts
N
Ts
Br
Br
NH
Br
O
1
2
3
N
N
X
R
O
N
N
O
(f)
N
7a-j
N
Ts
(c)
(d)
(e)
N
Ts
N
Ts
O
O
N
N
X
B
O
N
R
(g)
Br
NH
4
N
N
X
5
6a-j
R
N
Cl
N
Cl
X
Cl
(h)
R
X : O, NH or NCH3
7k, 7l
N
N
Het-Cl
Scheme 1. Synthesis of compounds 7a–l. Reagents and experimental conditions: (a) NaH, TsCl, DMF, 0 °C to RT; (b) AcCl, AlCl3, CS2, RT under N2; (c) DMF-DEA, DMF,
microwave; (d) bis(pinacolato)diboron, PdCl2(dppf), KOAc, 1,4-dioxane, microwave; (e) Het-Cl, PdCl2(PPh3)2, 2 M K2CO3, 1,4-dioxane/EtOH (2:3), microwave; (f) guanidine
carbonate, K2CO3, 2-methoxyethanol, microwave; (g) K2CO3, 2-methoxyethanol, microwave; (h) NaH, ROH, THF, microwave or RNH2 (or RNHCH3), K2CO3, DMF, RT.