W.S. Son, et al.
Bioorganic & Medicinal Chemistry Letters 29 (2019) 1168–1172
Fig. 1. Structural hybridization of pyrrolidine-based T-type calcium channel
inhibitors.
Scheme 2. Reagent and conditions: (i) 1.0 M LiAlH
89–96%; (ii) TsCl, TEA, DCM, rt, 64–87%; (iii) 3,3-dimethylbutanal or nicoti-
naldehyde, NaBH(OAc) , DCM, rt, 8–20%; (iv) 3,3-dimethylbutanoyl chloride,
TEA, THF, rt, 40%; (v) K CO , MeCN, reflux, 87–99%.
4
in THF, THF, 0 °C to rt,
profiles of pyrrolidine-based T-type calcium channel inhibitors. Newly
obtained compounds were tested in terms of their potency and ADME
properties. Furthermore, in vivo efficacy of representative compound
3
2
3
1
7h was evaluated in the SNL model.
Previously, in order to rationally design T-type calcium channel
inhibitors, we employed the common feature hypothesis generation
approach (HipHop) implemented in CATALYST program to generate a
were either purchased or accessed by reduction of commercially
available homobenzylic acid 14e–h using lithium aluminum hydride.
2
5
3
D ligand-based pharmacophore which resulted in a series of com-
Tosylation of 15d–h followed by S 2 reaction with either 12 or 13 with
N
pounds such as compound 1. We decided to further explore hydro-
phobic groups other than the 5-isobutyl-1-phenyl-1H-pyrazole moiety
in order to modulate potency and ADME properties (Fig. 1). The 3-
isopropylisoxazole moiety that exhibited consistent activity in our
potassium carbonate smoothly provided desired compounds 17d–m in
high yields.
Next, we evaluated in vitro T-type channel blocking activities of 12,
2
7
1
3, 17a–m employing a fluorescence-based FDSS6000 assay against
2
6
previous T-type calcium blockers and the 3-nitrophenyl-1,4-dihy-
dropyridine group extracted from the FDA-approved T-type channel
inhibitor nicardipine were selected to quickly characterize pyrrolidine-
based T-type calcium blockers.
Ca
v
3.1 and Ca 3.2 at 10 μM of compound concentration measuring %
2+
v
inhibitions of Ca
current (Table 1). As for compounds having 3-iso-
propylisoxazole, free pyrrolidine 12 did not inhibit T-type calcium
channels at 10 μM. As exemplified in the study of compound 1, in-
stallation of hydrophobic groups would be expected to increase po-
tency. However, 3,3-dimethylbutyl or 3,3-dimethylbutanoyl group was
barely effective in enhancing in vitro activity. Incorporation of differ-
ently substituted N-phenethyl groups to the pyrrolidine ring finally
conferred high T-type calcium channel inhibitory activities: most of N-
phenethylpyrrolidines harboring several different substituents such as
methyl-, fluoro-, chloro-, trifluoromethyl- and trifluoromethoxy groups
on the phenyl ring 17d-h inhibited greater than 40% of activation of
We envisaged that the designed compounds would be accessed by
various derivatizations of free pyrrolidine 12 or 13 serving as common
synthetic intermediates (Scheme 1). Their synthesis commenced with
amide formation between known tert-butyl (R)-3-(aminomethyl)pyrro-
2
4
lidine-1-carboxylate 8 and either 3-isopropyl-isoxazole-5-carboxylic
2
6
acid 7 came from 3 to isopropylisoxazole-5-carbaldehyde 6
using
Jones reagent or commercially available 9. Boc deprotection smoothly
produced free pyrrolidines 12 or 13 in high yield. Derivatizations of 12
or 13 were carried out by three different ways: acylation and alkylation
both Ca
v
3.1 and Ca 3.2 channels. In case of compounds possessing the
v
by either reductive alkylation or S 2 reaction (Scheme 2). N-(3,3-di-
N
nicardipine moiety, similar patterns of in vitro activity were observed. A
couple of N-phenethylpyrrolidines 17k and 17l showed great inhibitory
methybutyl)pyrrolidine 17a was obtained by reductive amination be-
tween 3,3-dimethylbutanal and 12. Next, acylation reaction of 12 with
activities against both Ca
v
3.1 and Ca
v
3.2 channels suppressing more
3
,3-dimethylbutanoyl chloride furnished 17b. Most of N-alkylated
2
+
than 65% of Ca
currents.
pyrrolidines (17d–m) were synthesized from alkylation of free pyrro-
lidines with tosylates (16d–h). First, homobenzylic alcohols 15d–h
Compounds exhibiting reasonable T-type calcium blocking activities
were subjected to microsomal stability test exploring remaining com-
pound percentage at 30 min after incubation in human liver microsome
(
Table 1). It was found that the in vitro metabolic stability of 17d-17h
was greatly improved as compared to the series of compounds having 5-
isobutyl-1-phenyl-1H-pyrazole moiety such as compound 1. Whereas
approximately 15% of compound 1 was left after 30 min of incubation
in human liver microsome, more than 35% of compounds stayed intact
after 30 min of incubation in most cases. However, it was observed that
compounds 17i–m were metabolized very fast being left less than 5% of
tested compounds after 30 min. Next, IC50 values against hERG chan-
2
8
nels were evaluated employing the whole-cell patch-clamp assay to
check potential cardiac side effects (Table 1). Although these series of
compounds were observed to potently inhibit hERG channels, several
compounds (17e–f, 17h, 17i, 17k and 17m) exhibited comparable
hERG activity to Mibefradil that shows approximately IC50 of 1.34 μM
Scheme 1. Reagent and conditions: (i) Jones reagent, acetone, 0 °C, 78%; (ii) 8,
CDI, THF, rt, 87–99%; (iii) TFA, DCM, rt, 79–99%.
29
against hERG.
1169
Son, Woo Seung
