S. Lin et al. / Bioorg. Med. Chem. Lett. xxx (2015) xxx–xxx
3
Table 1
Br
24
SAR on the position of Ar2 on indazole
O
N
Br
OMe
NIS/DMF
F3C
O
60 oC, 4 h
60%
N
H
Br
NaH/DMF, rt, 2 h
68%
OMe
25
23
OMe
5
6
Ar2
N
F3C
N
OH
7
H
N
F3C
OMe
B(OH)2
Pd(dppf)Cl
I
O
OMe
O
2
N
Br
Na CO , DME,
O
2
3
N
Br
Compound
Position
Ar2
hGCGR
binding (nM)
hGCGR
cAMP (nM)
60 oC, 12 h
40%
O
OMe
26
OR'
13a
13b
13c
13d
13e
13f
C-5
C-6
C-7
C-5
C-6
C-7
4-MeO-Ph
4-MeO-Ph
4-MeO-Ph
4-Me-Ph
4-Me-Ph
4-Me-Ph
10
4.0
1700
7.0
2.0
26
11
2700
17
5.0
27
28
R' = Me
R' = H
aq. NaOH
F3C
quantitative
780
3800
OMe
β-Ala-OEt
N
Br
OEt
EDC, HOBt
DIEA, CH Cl
H
N
2
2
Table 2
hGCGR activity of indole-, 7-azaindole-, and indazole-based GRAs
O
78%
O
29
F3C
Ar -B(OH)
Pd(dppf)Cl
1)
2
2
OMe
2
Na CO , DME, 60 oC, 16 h
2
3
Ar2
N
OH
O
H
2) aq. NaOH, MeOH
72-83%
N
O
30
Compound
X/Y
Ar2
hGCGR
hGCGR
binding (nM)
cAMP (nM)
Scheme 3. Synthesis of indole GRA 30.
13b
30aa
13e
30ba
16a
22
N/CH
CH/CH
N/CH
CH/CH
N/CH
CH/N
4-MeO-Ph
4-MeO-Ph
4-Me-Ph
4-Me-Ph
4-CF3O-Ph
4-CF3O-Ph
4.0
8.0
2.0
4.0
1.0
12
11
130
5.0
48
5.6
360
affinity of compounds for the human glucagon receptor (hGCGR)
was determined by their ability to displace [125I] Glucagon from
hGCGR using a membrane preparation from a CHO cell line
expressing the human GCGR (CHO hGCGR). The functional activity
was assessed by the ability of these compounds to inhibit gluca-
gon-induced cAMP production in a cell line expressing hGCGR.
Both binding and functional activities are expressed as values of
IC50. All data reported are from the more active enantiomers tested
unless otherwise noted.
Based on the structure homology of MK-0893 and indazole
GRAs designed, it is obvious that the C-3 aryl (Ar1) of 5 is critical
for the activity against the hGCGR. However, it is not as clear as
to where the second aryl substituent (Ar2) should be placed on
the indazole ring. We therefore set out to compare all C-5, C-6,
and C-7 positions of indazole. As shown in Table 1, we were
pleased to find that all indazole-based GRAs were active in vitro
against the hGCGR. However, it is clear from the SAR that C-7 is
the least preferred position for the second aryl substitution (13c
and 13f). While both C-5 and C-6 afforded GRAs with potent activ-
ity, C-6 position seems to be more preferred with an edge over C-5
position (13b vs 13a, and 13e vs 13d).
With the identification of the C-6 position being the preferred
position for the 2nd aryl substitution, we set out to find out how
the indazole core compared to other 6/5 ring systems such as
indole or azaindole (Table 2). Interestingly, while head to head
comparison of C-3/C-6 substituted indazoles with corresponding
indoles showed very similar activity in binding to the hGCGR,
indoles exhibited ca. 5-fold weaker potency in the cAMP functional
assay (13b vs 30a, and 13e vs 30b). Indazole 16a was found to be a
very potent GRA in both binding assay (IC50 = 1 nM) and cAMP
functional assay (IC50 = 5.6 nM). However, moving the nitrogen at
a
Compounds were tested as a racemic mixture.
indazole 2 position to 7 position, in another word, replacement
of indazole core of 16a with a 7-azaindole core, resulted in signif-
icant loss of hGCGR activity (binding IC50 = 12 nM, cAMP
IC50 = 360 nM for 7-azaindole indole 22). Obviously, the indazole
core is the most preferred scaffold for hGCGR activity among the
three heterocycles studied.
Aryl substitution on the C-3 position of indazole was then
examined for their hGCGR activity (Table 3). In general, most of
the aryl groups, including substituted phenyl, pyridinyl, or
pyrimidinyl groups, are tolerated. Evidently, phenyl groups substi-
tuted with simple halides, a methoxyl group, or a trifluoromethyl
group is more likely to provide GRAs with good potency (14a–f,
and 14l–m). A polar substitution on the phenyl group is usually
less preferred (14g, 14p, and 14q), and often resulted in weaker
functional activity (14g and 14q). Similarly, placement of substi-
tuted pyridines (14h–k) and pyrimidine (14r) generally afforded
compounds with weaker hGCGR activity. Not surprisingly, 14a,
an analog of 16a with a 1-MeO-4-CF3O-phen-2-yl group at C-3
position, exhibited best functional activity (cAMP IC50 = 11 nM)
among all aryl groups tested, and excellent binding activity
(IC50 = 7.0 nM).
The 1-MeO-4-CF3O-phen-2-yl group was then selected for the
C-3 position when we investigated the aryl substituents at the
C-6 position of indazole. As shown in Table 4, C-6 position seems