2
R. M. McKinnell et al. / Bioorg. Med. Chem. Lett. xxx (2014) xxx–xxx
OH
H
N
OH
H
N
O
HO
O
HO
HO
HN
O
1
2
H
Cl
OH
OH
O
H
N
H
N
O
OH
H
N
O
O
HN
Cl
HO
HO
HO
O
HN
OH
3
4
5
O
Figure 1. Structures of the b2-agonists salmeterol (1), formoterol (2), olodaterol (3), indacaterol (4) and vilanterol (5).
molecule would enhance solubility and therefore facilitate formu-
lation development.
the 72 h time point in the guinea pig model. Following nebulized
dosing, both 15 and 16 were shown to possess excellent broncho-
protective activity over this extended time frame. Although neither
compound showed a statistically significant improvement over 7, a
promising trend towards improved efficacy was apparent (Fig. 3).
All three compounds were superior to salmeterol in this model.
During additional preclinical evaluation of compounds 15–16,
we observed significant oxidation (potentially catalyzed by mono-
amine oxidase12) of the primary amine functionality to the respec-
tive aldehyde in the plasma and/or hepatocytes of various species
(Scheme 2). These aldehyde metabolites were subsequently found
to have in vitro activity similar to parent molecules at the b2-
receptor (data not shown). The presence of circulating active
metabolites bearing a potentially reactive functional group was
seen as a risk for the development of these compounds, and further
evaluation was suspended.
To explore the impact of amine-substituted secondary binding
groups on b2 activity, we initially prepared a series of derivatives
of 7 in which a primary aminomethyl moiety was appended to
the ortho-, meta- and para-positions of the distal phenyl ring of
the secondary binding group (Scheme 1). Synthesis of these sec-
ondary binding groups was accomplished by Suzuki coupling of
bromide 8 with the appropriate benzonitrile boronic acids. Reduc-
tion of the nitrile substituent using borane in THF followed by
hydrogenation of the nitro group furnished intermediates 9–11.
The headgroup-linker fragment 13 was constructed by alkylation
of 4-bromophenethylamine with protected headgroup 12.10 Buch-
wald coupling of intermediates 9–11 to bromide 13 followed by
deprotection of the headgroup afforded the desired compounds
14–16.
These compounds were profiled in our in vitro screens and
compared to standards (Table 1). Binding affinities for the b1-
and b2-adrenoceptors and b2/b1 selectivity were measured using
competition binding assays with [3H]dihydroalprenolol as tracer
and plasma membranes from cells expressing the human recombi-
nant b1- and b2-adrenoceptors. Agonist potencies were measured
in cAMP accumulation assays using recombinant cell lines heterol-
ogously expressing the human b1-, b2-, and b3-adrenoceptors, and
the BEAS-2B lung epithelial cell line with endogenous expression
of the b2-adrenoceptor. BEAS-2B cells were also used to measure
intrinsic agonist activity (IA) relative to isoproterenol. Compounds
14–16 illustrate that, relative to 7, the addition of a basic moiety
was well tolerated in terms of b2-receptor affinity and functional
potency. However, b2/b1 selectivity of 14 was too low for progres-
sion, since selectivity similar to or greater than that of formoterol
(2) was targeted. Based on their attractive in vitro profile,
compounds 15 and 16 were advanced to the guinea pig in vivo
bronchoprotection model.11
Based on the encouraging efficacy of compounds 15–16, we
sought an alternative strategy by which to incorporate metaboli-
cally stable amines into this series. The large number of synthetic
steps required to produce 15–16 also led us to consider structural
simplification of the scaffold. The terminal alkyl ether group of 7
was identified as a convenient synthetic handle with which to rap-
idly explore a variety of basic amines in the same general region of
the molecule. In order to simplify the synthesis and reduce
structural similarity to 7, the distal phenyl ring of 7 was excised.
Synthesis of the parent compound 19 was accomplished by cou-
pling BOC-protected 4-bromophenethylamine with 4-ethoxyani-
line followed by deprotection to provided intermediate 17.
Treatment of 17 with epoxide 1813 followed by debenzylation
afforded 19 (Scheme 3). Synthesis of the amine-based compounds
was accomplished by nucleophilic aromatic substitution of p-fluo-
ronitrobenzene with the alkoxide of the requisite aminoalcohol.
Hydrogenation of the nitro group afforded aniline intermediates
which were then coupled to intermediate 13 using Buchwald
chemistry. Deprotection of the headgroup afforded compounds
20–30 (Scheme 4).
At this stage of our b2-agonist program, our criteria for advanc-
ing compounds required significant bronchoprotective efficacy at
'Secondary
Binding Group'
'Secondary
'Headgroup'
OH
'Linker'
'Headgroup'
OH
'Linker'
Binding Group'
H
N
H
N
O
HO
N
H
HO
N
H
HN
O
HN
H
OH
(7)
TD-5471
(6)
milveterol
O
Figure 2. Structure of milveterol (6) and TD-5471 (7) indicating the b2- agonist ‘headgroup’ as well as the regions we have termed the ‘linker’ and the ‘secondary binding
group’.