Design, syntheses, and SAR of 2,8-diazaspiro[4.5]decanones as T-type calcium channel antagonists

Article abstract of DOI:10.1016/j.bmcl.2010.09.098

It was hypothesized that an appropriately substituted 2,8-diazaspiro[4.5] decan-1-one could effectively approximate a 5-feature T-type pharmacophore model published in the literature. Compounds were designed and synthesized to test our hypothesis and were found to be potent T-type calcium channel inhibitors with modest selectivity over L-type calcium channels. The synthesis and SAR of the series is described.

Full text of DOI:10.1016/j.bmcl.2010.09.098

Bioorganic & Medicinal Chemistry Letters 20 (2010) 6375–6378
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Design, syntheses, and SAR of 2,8-diazaspiro[4.5]decanones as T-type
calcium channel antagonists
⇑
Paul C. Fritch , Jeffrey Krajewski
Icagen, Inc., Suite 390, 4222 Emperor Blvd., Durham, NC 27703, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
It was hypothesized that an appropriately substituted 2,8-diazaspiro[4.5]decan-1-one could effectively
approximate a 5-feature T-type pharmacophore model published in the literature. Compounds were
designed and synthesized to test our hypothesis and were found to be potent T-type calcium channel
inhibitors with modest selectivity over L-type calcium channels. The synthesis and SAR of the series is
described.
Received 11 August 2010
Revised 14 September 2010
Accepted 16 September 2010
Available online 19 September 2010
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
T-type calcium channel
CaV3.2
Diazaspirodecanones
Calcium is an important signaling molecule for many physiolog-
ical processes in the human body. These include electrical signaling
in the nervous system, as well as controlling heart and smooth
muscle contraction and hormone release. The entry of calcium into
cells is regulated by a diverse set of proteins called calcium chan-
nels. A fundamental role of Ca2+ channels is to translate an electri-
cal signal on the surface membrane into a chemical signal within
the cytoplasm, which, in turn, activates many important intracellu-
lar processes including contraction, secretion, neurotransmission,
regulation of enzymatic activities, and gene expression, as well
as cell death.1
Voltage-gated calcium channels are divided into two primary
groups based on electrophysiological characteristics: low voltage
activated (LVA or T-type) and high voltage activated (HVA).2 The
HVA group includes the L-, N-, P-, Q-, and R-types and requires a
relatively large membrane depolarization to open and displays
slow inactivation kinetics. In contrast, the LVA or T-type channels
require a smaller membrane depolarization for activation. These
channels are so named because they carry a transient current with
a low voltage of activation and rapid inactivation.
Rapid gating kinetics and smaller membrane depolarization
thresholds make T-type channels well suited to control neuronal
excitability.3,4 For example, the roles of T-type channels in regulat-
ing sleep rhythms and in certain forms of epilepsy have been
established.5 More recent evidence of a link between T-type chan-
nels and weight maintenance is intriguing and may suggest a po-
tential for treatment of obesity.6
The role of T-type channels in nociception is less established,
but evidence has been accumulating. It has been demonstrated
that T-type channel-dependent, low-threshold Ca2+ spikes regulate
excitability in DRG neurons. Furthermore, T-type activity is upreg-
ulated in neuropathic animals resulting in spontaneous burst firing
of action potentials.7
The analgesic nitrous oxide selectively blocks DRG T-type cur-
rents with no effect on HVA currents at clinically relevant concen-
trations. A number of anesthetics including isoflurane were found
to block T-type currents.8
The modestly selective T-type/L-type calcium channel antago-
nist mibefradil, briefly marketed as Posicor™ for the treatment of
hypertension and angina pectoris, was withdrawn from the market
less than a year after its introduction due to problems associated
with inhibition of cytochrome P450 enzymes.9 Nevertheless, mibe-
fradil has been a useful tool for investigating the role of T-type cal-
cium channels in in vitro and in vivo models.
The T-type channels are further subdivided into three subtypes
based on the amino acid sequences of their pore-forming a1 sub-
units.3 Three genes that encode these subunits have been identi-
fied: CACNA1G for CaV3.1 (
a
1G), CACNA1H for CaV3.2 (a1H), and
CACNA1I for CaV3.3 ( 1I). These three
a
a
1 subunits are differentially
Mibefradil and the T-type antagonist ethosuximide reduce in vivo
hyperalgesic responses to thermal or mechanical stimuli induced by
chemical agents or experimental nerve injury. Moreover, selective
T-type knock-out animals provide further evidence for the role of T-
type calcium channels in peripheral nerve pain signaling.10
The desire to find novel mechanisms of action and therapeutic
targets for the treatment of hypertension, epilepsy, insomnia, obes-
ity, and neuropathic pain has spurred considerable interest in the
and widely expressed in the central and peripheral nervous sys-
tems, cardiac, and vascular smooth muscle, kidneys, sperm, adre-
nal, and pituitary glands, as well the pancreas.3
⇑
Corresponding author. Tel.: +1 919 771 5580.
E-mail address: pfritch@icagen.com (P.C. Fritch).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.09.098

6376
P. C. Fritch, J. Krajewski / Bioorg. Med. Chem. Lett. 20 (2010) 6375–6378
Table 1
Percent Inhibition of
development of novel and selective antagonists to T-type chan-
nels.11,12 The 5-feature T-type calcium channel pharmacophore
model described by Pae and co-workers, consisting of three
hydrophobic regions, one hydrogen bond acceptor and one positive
ionizable region, captured our attention.11b It was recognized that
2,8-diazaspiro[4.5]decan-1-one, which contains a hydrogen bond
acceptor and a positive ionizable atom could serve as a semi-rigid
core from which to attach hydrophobic moieties. Furthermore, the
spatial arrangement of the attached hydrophobic moieties, hydro-
gen bond acceptor, and positive ionizable atom appeared to
approximate the model described by Pae and co-workers. It was
hypothesized that compounds derived from such a scaffold would
a1H and a1C at concentration in l
Ma
Compd
R
a1H% inhib @
a1C% inhib @
(l
M)
(lM)
1
2
3
4
5
6
7
8
9
NAb @ 3
58 1 @ 0.03
66 8 @ 1
65 3 @ 3
59 4 @ 0.1
67 6 @ 3
56 14 @ 0.3
40 4 @ 0.3
NA @ 1
31 5 @ 0.1
59 7 @ 3
4-FPhCH2
(4-FPh)2CHCH2
2-EtOPhCH2
3-PhOPhCH2
10
11
12
45 4 @ 3
30 11 @ 0.03
91 1 @ 0.3
NA @ 0.03
52 1 @ 0.03
28 2 @ 1
44 9 @ 1
76 3 @ 1
54 8 @ 3
be potent inhibitors of the a1H T-type calcium channel.
4-ClPhCH2CH2
3-Cl PhCH2
Herein we describe the design, syntheses, and structure–activity
relationship (SAR) of a series of novel 2,8-diazaspiro[4.5]decanone
analogs. To test our hypothesis, we coupled commercially available
reagents 2-(4-chlorobenzyl)-2,8-diazospiro[4.5]decan-1-one 1 and
4,4-difluorobenzhydryl chloride to form 2-(4-chlorobenzyl)-8-(4,4-
difluorobenzhydryl)-2,8-diazaspiro[4.5]decan-1-one 2 (Scheme 1).
13
14
15
16
17
18
19
20
21
22
3-MeOPhCH2
MeOCH2CH2
MeOCH2CH2CH2
iPrOCH2CH2CH2
67 14 @ 0.3
58 8 @ 10
56 5 @ 3
O(CH2CH2)2NCH2CH2CH2 35 2 @ 1
iPr
57 6 @ 3
44 13 @ 0.3
NA @ 3
52 3 @ 3
60 13 @ 0.3
Compound 2 was found to inhibit the a1H T-type calcium channel
t-BuCH2CH2
subtype 58 1% at a concentration of 30 nM in patch clamp electro-
physiology assays, thus validating our hypothesis (Table 1).13 Com-
pound 2 displayed modest (ꢀ10-fold) selectivity (T-type/L-type Ca
H
68 3 @ 1
59 4 @ 0.3
42 0 @ 0.1
44 2 @ 0.1
channels) as it inhibited the a1C L-type calcium channel 31 5% at
a
For assay conditions see references.
NA (not active) denotes <20% inhibition.
0.1 lM in our patch clamp assay. The potency and T-type/L-type
b
selectivity of 2 was similar to that of mibefradil.14 The unsubstituted
2-(4-chlorobenzyl)-2,8-diazospiro[4.5]decan-1-one, 1, was not ac-
tive at 3 lM concentration in the a1H T-type assay.
Next the amide carbonyl hydrogen bond acceptor was removed
by treating compound 2 with lithium aluminum hydride to pro-
and the benzhydryl moiety (compound 5) led to a modest decrease
in 1H potency (59 4% at 0.1 M).
Analogs 6 and 7 attempted to fill the third hydrophobic region
by attaching a moiety to the benzyl substituent; 1H potency in-
creased in relation to the size of the moiety off the benzyl group
(6, EtO, 67 6 at 3 M and 7, PhO, 56 14 at 0.3 M). A Strecker
a
l
duce the bis-amine 3. This compound inhibited
a1H (66 8% at
a
1
lM) but was far less potent than 2, indicating the importance
of the hydrogen bond acceptor for potent T-type activity. Reductive
amination of diazaspirodecanone 1 with aldehydes by treatment
with sodium triacetoxyborohydride in 1,2-dichloroethane afforded
analogs 4–7. Compound 4 demonstrated that removal of one of the
l
l
and Bruylants reaction sequence with diazaspirodecanone 1 affor-
ded analog 8,15 which replaced the two phenyl rings of the benzhy-
dryl moiety with two isopropyl groups; this analog inhibited
a1H
three hydrophobic regions led to a large decrease in a1H potency
(40 4% at 0.3 M) but was less potent than 2. Acylation and sulf-
l
(65 3% at 3
lM). Inserting a methylene spacer between the amine
onylation of 1 afforded compounds 9 and 10, which were inactive
and weakly active, respectively.
Compounds 11–20 were prepared to explore changes to the
hydrophobic region on the amide side of the spirocycle. Alkylation
of ethyl 1-tert-butoxycarbonylpiperidine-4-carboxylate with allyl
bromide followed by oxidative cleavage afforded the ethyl ester
aldehyde (Scheme 2). Reductive aminations and concomitant
cyclization afforded Boc-protected diazaspirodecanones. Acid
catalyzed deprotections and alkylations with 4,4-difluorobenzhyd-
ryl chloride afforded compounds 11–19. Insertion of a methylene
F
F
Cl
b
Cl
O
N
N
N
F
N
N
F
2
a
3
Cl
Cl
R
O
N
spacer into the 4-chlorobenzyl moiety (11, 30 11% at 0.03
lM)
O
N
c
NH
or moving the chloro from the ortho to the meta position (12,
N
91 1% at 0.3
lM) reduced
a1H potency, however the 3-methoxy-
4-7
d
1
Cl
e
O
N
F
f
O
Cl
Boc
O
Boc
N
N
b
8
N
a
N
F
O
N
EtO2C
EtO2C
CHO
9
R N
F
O O
N
Cl
S
O
11-20
c
O
N
F
NH
HCl
10
HN
Scheme 1. Reagents and conditions: (a) 4,4-difluorobenzhydryl chloride, Cs2CO3,
DMF, 70 °C, 16 h, 17%; (b) LiAlH4, THF, rt, 16 h, 7%; (c) RCHO, NaBH(OAc)3, DCE,
5–16 h, 77–99%; (d) (i) isobutyraldehyde, (CH3)3SiCN, CH3CN, rt, 16 h, 51%; (ii)
iPrMgBr, THF, rt, 16 h, 21%; (e) 4-fluorobenzoyl chloride, NaOH, THF, H2O, rt, 1 h,
81%; (f) 4-fluorophenylsulfonyl chloride, NaOH, THF, H2O, rt, 12 h, 69%.
Scheme 2. Reagents and conditions: (a) (i) ((CH3)3Si)2NLi, allyl bromide, THF, ꢁ70–
0 °C, 4 h, 97%; (ii) NaIO4, OsO4(cat), dioxane, H2O, rt, 16 h, 76%; (b) (i) RNH2,
NaBH(OAc)3, DCE, rt, 1 h then reflux, 16 h, 45–86%; (ii) CF3CO2H, DCM, rt, 1 h, 100%;
(iii) 4,4-difluorobenzhydryl chloride, K2CO3, DMF, 80 °C, 16 h, 11–21%; (c) 4,4-
difluorobenzhydryl chloride, K2CO3, DMF, 80 °C, 16 h, 10%.

P. C. Fritch, J. Krajewski / Bioorg. Med. Chem. Lett. 20 (2010) 6375–6378
6377
benzyl analog 13 (52 1% at 0.03
l
M) was nearly equipotent
21) and (59 4% at 0.3
Analysis against
position had little effect on the L-type calcium channel. Indeed
compounds 21 (42 0% at 0.1 M) and 22 (44 2% at 0.1 M) were
slightly more potent against 1C than compound 2.
Based on the 5-feature T-type calcium channel pharmacophore
model described by Pae and co-workers, a series of compounds de-
rived from a 2,8-diazaspiro[4.5]decan-1-one core was designed
that appeared to approximate the spatial arrangement of the phar-
macophore model. Our initial compound to test this hypothesis, 2,
l
M, 22) against the
a
1H T-type channel.
against 1H with compound 2. Like compound 2, compound 13
a
a1C showed that moving the amide carbonyl
was found to have modest (ꢀeightfold) T-type/L-type calcium
channel selectivity.
l
a
l
Compounds 14–17 incorporated alkyl ether moieties into the
hydrophobic region on the amide side of the pharmacophore. The
a
1H potency increased with the size of the moiety; methoxyethyl
(28 2% at 1 M, 14), methoxypropyl (44 9% at 1 M, 15), and
isopropoxypropyl (76 3% at 1 M, 16). Incorporation of a basic
amine into this region, morpholinopropyl 17, resulted in decreased
1H potency (35 2% at 1 M).
Alkyl analogs 18 (iPr, 57 6% at 3
44 13% at 0.3 M) also showed increasing
l
l
l
a
l
was a potent inhibitor of the
approximately 10-fold selective over the a
a
1H T-type calcium channel and was
1C L-type calcium chan-
lM) and 19 (tBuCH2CH2,
l
a1H potency with an
nel. Evaluation of the SAR indicated the importance of the amide
carbonyl hydrogen bond acceptor and its position in the 2,8-diaza-
increase in size of alkyl substituent but remained much less potent
that the benzyl analogs. The unsubstituted amide 20 was prepared
by alkylation of commercially available 2,8-diazaspiro[4.5]decan-
1-one hydrochloride with 4,4-difluorobenzhydryl chloride, the
spiro[4.5]decane core for optimal
a1H inhibition. The size of the
moiety attached to the 8-N appeared to correlate with
a1H inhibi-
tion. Large groups (such as benzhydryl) that could effectively fill
two hydrophobic regions contributed to increased potency. Moie-
ties that were less effective at interacting with two hydrophobic
regions gave less potent compounds. With respect to groups at-
tached to the 2-N (amide nitrogen), the best activity was obtained
from benzyl groups; as other groups, in general, afforded decreased
potency in relation to the size of the substituent. The 2,8-diazaspi-
ro[4.5]decanones disclosed herein appear to validate the 5-feature
T-type pharmacophore model and provide new insights for the
development of potent and selective T-type calcium channel
antagonists.
compound was not active at 3 lM concentration against a1H.
Additional SAR regarding the position of the amide carbonyl
hydrogen bond acceptor was generated through compounds 21
and 22. Compound 21 was prepared following the procedure of
Smith et al.16 Reaction of 1-tert-butoxycarbonyl-4-piperidone with
stabilized Wittig reagent (methoxycarbonylmethylene)triphenyl-
phosphorane afforded the unsaturated ester, which underwent Mi-
chael addition of nitromethane when refluxed in the presence of
1,1,3,3-tetramethylguanidine (Scheme 3). Subsequent reduction
with hydrogen and Raney nickel followed by concomitant cycliza-
tion afforded the Boc-protected 2,8-diazaspiro[4.5]decan-3-one.
Alkylation with 4-chlorobenzyl chloride and sodium hydride affor-
ded 21.
Acknowledgments
Compound 22 was prepared in a three-step sequence from
commercially available 2,8-diazaspiro[4.5]decane-2-carboxylic
acid tert-butyl ester. Alkylation of the amine with 4,4-dif-
luorobenzhydryl chloride followed by Boc deprotection and
acylation with 4-chlorobenzoyl chloride afforded 22 (Scheme 4).
Compounds 21 and 22 demonstrated the importance of the
The authors would like to thank Eva Prazak and Michelle Mese-
ke for contributing to this research.
References and notes
1. (a) Tsien, R. W.; Lipscombe, D.; Madison, D. V.; Bley, K. R.; Fox, A. P. Trends
Neurosci. 1988, 11, 431; (b) Tsien, R. W.; Wheeler, D. B. Voltage-gated Calcium
Channels. In Calcium as a Cellular Regulator; Carafoli, E., Klee, C., Eds.; Oxford
University Press: New York, 1999; pp 171–199; (c) Berridge, M. J.; Bootman, M.
D.; Lipp, P. Nature 1998, 395, 645; (d) Clapham, D. E. Cell 2007, 131, 1047.
2. (a) Catterall, W. A.; Perez-Reyes, E.; Snutch, T. P.; Striessnig, J. Pharmacol. Rev.
2005, 57, 411; (b) Ertel, S. I.; Ertel, E. A. Trends Pharmacol. Sci. 1997, 18, 37; (c)
Ertel, E. A.; Campbell, K. P.; Harpold, M. M.; Hofmann, F.; Mori, Y.; Perez-Reyes,
E.; Schwartz, A.; Snutch, T. P.; Tanabe, T.; Tsien, R. W.; Catterall, W. A. Neuron
2000, 25, 533.
position of the amide carbonyl for
a1H T-type inhibition. Moving
the amide carbonyl from the 1-position of the 2,8-diazaspi-
ro[4.5]decanone to the 3-position, compound 21, or shifting the
amide carbonyl outside of the 2,8-diazaspirodecane, compound
22, resulted in at least a 10-fold loss in potency (68 3% at 1 lM,
F
3. Perez-Reyes, E. Physiol. Rev. 2003, 83, 117. and references cited therein.
4. (a) Iftinca, M. C.; Zamponi, G. W. Trends Pharmacol. Sci. 2008, 30, 32; (b) Fox, A.
P. J. Physiol. 1987, 394, 149.
5. (a) Lee, J.; Kim, D.; Shin, H.-S. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 18195; (b)
Renger, J. J.; Koblan, K. S. PCT Int. Appl. WO 2004035000.; (c) Steriade, M. Trends
Neurosci. 2005, 28, 317; (d) Reger, T. S.; Yang, Z-Q.; Schlegel, K.-A. S.; Shu, Y.;
Cube, R. V.; Mattern, C.; Rittle, K. E.; Ngo, P. L.; Shipe, W. D.; Yang, V.; Lindsley,
C.; Barrow, J.; Coleman, P.; Hartman, G. D.; Tang, C.; Ballard, J.; Kuo, Y.;
Prueksaritanont, T.; Kane, S. A.; Urban, M. O.; Liang, A.; Sain, N. M.; Uebele, V.
N.; Nuss, C. E.; Doran, S. M.; Garson, S. L.; Fox, S. V.; Kraus, R. L.; Renger, J. J.
Abstracts of Papers, 237th National Meeting of the American Chemical Society,
Salt Lake City, UT, March 22–26, 2009; American Chemical Society:
Washington, DC, 2009; MEDI-292.; (e) Meldrum, B. S.; Rogawski, M. A.
Neurotherapeutics 2007, 4, 18; (f) Khosravani, H.; Zamponi, G. W. Physiol. Rev.
2006, 86, 941; (g) Nelson, M. T.; Todorovic, S. M.; Perez-Reyes, E. Curr. Pharm.
Des. 2006, 12, 2189.
Boc
Cl
Boc
N
N
a
b
N
HN
O
F
N
O
O
21
Scheme 3. Reagents and conditions: (a) (i) Ph3PCHCO2CH3, toluene, reflux, 16 h,
93%; (ii) CH3NO2, N,N,N0,N0-tetramethylguanidine, reflux, 16 h, 30%; (iii) H2, Raney
Ni, EtOH, 3 d, 75%; (b) (i) NaH, 4-ClBnCl, THF, rt then reflux, 12 h, 99%; (ii) CF3CO2H,
DCM, rt, 1 h, 94%; (iii) 4,4-difluorobenzhydryl chloride, K2CO3, DMF, 80 °C, 5 h, 6%.
6. Uebele, V. N.; Gotter, A. L.; Nuss, C. E.; Kraus, R. L.; Doran, S. M.; Garson, S. L.;
Reiss, D. R.; Li, Y.; Barrow, J. C.; Reger, T. S.; Yang, Z.-Q.; Ballard, J. E.; Tang, C.;
Metzger, J. M.; Wang, S.-P.; Koblan, K. S.; Renger, J. J. J. Clin. Invest. 2009, 119,
1659.
F
7. (a) Lovinger, D. M.; White, G. Neurosci. Lett. 1989, 102, 50; (b) White, G.;
Lovinger, D. M. Proc. Natl. Acad. Sci. U.S.A. 1989, 86, 6802; (c) Study, R. E.; Kral,
M. G. Pain 1996, 65, 235.
8. (a) Todorovic, S. M.; Lingle, C. J. Neurophysiol. 1998, 79, 240; (b) Todorovic, S.
M.; Jevtovic-Todorovic, V.; Perez-Reyes, E.; Zorumski, C. F. Mol. Pharmacol.
2001, 60, 603; (c) Todorovic, S. M.; Perez-Reyes, E.; Lingle, C. J. Mol. Pharmacol.
2000, 58, 98.
Cl
NH
a
N
Boc N
N
F
O
22
Scheme 4. Reagents and conditions: (a) (i) 4,4-difluorobenzhydryl chloride,
Cs2CO3, DMF, 70 °C, 16 h, 20%; (ii) CF3CO2H, DCM, rt, 1 h, 100%; (iii) 4-chlorobenzoyl
chloride, NaOH, THF, H2O, rt, 1 h, 98%.
9. SoRelle, R. Circulation 1998, 98, 831.
10. (a) Todorovic, S. M.; Jevtovic-Todorovic, V.; Meyenburg, A.; Mennerick, S.;
Perez-Reyes, E.; Romano, C.; Olney, J. W.; Zorumski, C. F. Neuron 2001, 31, 75;

6378
P. C. Fritch, J. Krajewski / Bioorg. Med. Chem. Lett. 20 (2010) 6375–6378
(b) Dogrul, A.; Gardell, L.; Ossipov, M.; Tulunay, F.; Lai, J.; Porreca, F. Pain 2003,
K.-A. S.; Shu, Y.; Rittle, K. E.; Bock, M. G.; Hartman, G. D.; Tang, C.; Ballard, J. E.;
Kuo, Y.; Adarayan, E. D.; Prueksaritanont, T.; Zrada, M. M.; Uebele, V. N.; Nuss,
C. E.; Connolly, T. M.; Doran, S. M.; Fox, S. V.; Kraus, R. L.; Marino, M. J.;
Graufields, V. K.; Vargas, H. M.; Bunting, P. B.; Hasbun-Manning, M.; Evans, R.
M.; Koblan, K. S.; Renger, J. L. J. Med. Chem. 2008, 51, 3692; (t) Yang, Z.-Q.;
Barrow, J. C.; Shipe, W. D.; Schlegel, K.-A. S.; Shu, Y.; Yang, F. V.; Lindsley, C. W.;
Rittle, K. E.; Bock, M. G.; Hartman, G. D.; Uebele, V. N.; Nuss, C. E.; Fox, S. V.;
Kraus, R. L.; Doran, S. M.; Connolly, T. M.; Tang, C.; Ballard, J. E.; Kuo, Y.;
Adarayan, E. D.; Prueksaritanont, T.; Zrada, M. M.; Marino, M. J.; Graufields, V.
K.; DiLella, A. G.; Reynolds, I. G.; Vargas, H. M.; Bunting, P. B.; Woltmann, R. F.;
Magee, M. M.; Koblan, K. S.; Renger, J. L. J. Med. Chem. 2008, 51, 6471; (u) Oh, Y.;
Kim, Y.; Seo, S. H.; Lee, J. K.; Rhim, H.; Pae, A. N.; Jeong, K.-S.; Choo, H.; Cho, Y. S.
Bull. Korean Chem. Soc. 2008, 29, 1881.
105, 159; (c) Todorovic, S. M.; Meyenburg, A.; Jevtovic-Todorovic, V. Pain 2004,
109, 328; (d) Flatters, S. J. L.; Bennett, G. J. Pain 2004, 109, 150; (e) Mathews, E.
A.; Dickenson, A. H. Eur. J. Pharmacol. 2001, 415, 141; (f) Flatters, S. J. L. Drugs
Future 2005, 30, 573; (g) Todorovic, S. M.; Meyenburg, A.; Jevtovic-Todorovic,
V. Brain Res. 2002, 951, 336; (h) Dogrul, A.; Yesilyurt, O.; Isimer, A.; Guzeldemir,
M. E. Pain 2001, 93, 61; (i) Bilici, D.; Akpinar, E.; Gursan, N.; Ozbakis Dengiz, G.;
Bilici, S.; Altas, S. Pharmacol. Res. 2001, 44, 527; (j) Kim, D.; Park, D.; Choi, S.;
Lee, S.; Sun, M.; Kim, C.; Shin, H.-S. Science 2003, 302, 117; (k) Kim, D.; Song, I.;
Keum, S.; Lee, T.; Jeong, M.-J.; Kim, S.-S.; McEnery, M. W.; Shin, H.-S. Neuron
2001, 31, 35; (l) Bourinet, E.; Alloui, A.; Monteil, A.; Barrere, C.; Couette, B.;
Poirot, O.; Pages, A.; McRory, J.; Snutch, T. P.; Eschalier, A.; Nargeot, N. EMBO J.
2005, 24, 315; (m) Choi, S.; Na, H. S.; Kim, J.; Lee, J.; Lee, S.; Kim, D.; Park, J.;
Chen, C.-C.; Campbell, K. P.; Shin, H.-S. Genes Brain Behav. 2007, 6, 425.
11. (a) Schenck, H. A.; Lenkowski, P. W.; Choudhury-Mukherjee, I.; Ko, S.-H.;
Stables, J. P.; Patel, M. K.; Brown, M. L. Bioorg. Med. Chem. 2004, 12, 979; (b)
Doddareddy, M. R.; Jung, H. K.; Lee, J. Y.; Lee, Y. S.; Cho, Y. S.; Koh, H. Y.; Pae, A.
N. Bioorg. Med. Chem. 2004, 12, 1605; (c) Doddareddy, M. R.; Jung, H. K.; Cha, J.
H.; Cho, Y. S.; Koh, H. Y.; Chang, M. H.; Pae, A. N. Bioorg. Med. Chem. 2004, 12,
1613; (d) Jung, H. K.; Doddareddy, M. R.; Cha, J. H.; Rhim, H.; Cho, Y. S.; Koh, H.
Y.; Jung, B. Y.; Pae, A. N. Bioorg. Med. Chem. 2004, 12, 3965; (e) Lee, Y. S.; Lee, B.
H.; Park, S. J.; Kang, S. B.; Rhim, H.; Park, J.-Y.; Lee, J.-H.; Jeong, S.-W.; Lee, J. Y.
Bioorg. Med. Chem. Lett. 2004, 14, 3379; (f) McCalmont, W. F.; Heady, T. N.;
Patterson, J. R.; Lindenmuth, M. A.; Haverstick, D. M.; Gray, L. S.; Macdonald, T.
L. Bioorg. Med. Chem. Lett. 2004, 14, 3691; (g) McCalmont, W. F.; Patterson, J. R.;
Lindenmuth, M. A.; Heady, T. N.; Haverstick, D. M.; Gray, L. S.; Macdonald, T. L.
Bioorg. Med. Chem. 2005, 13, 3821; (h) Rhim, H.; Lee, Y. S.; Park, S. J.; Chung, B.
Y.; Lee, J. Y. Bioorg. Med. Chem. Lett. 2005, 15, 283; (i) Ku, I. W.; Cho, S.;
Doddareddy, M. R.; Jang, M. S.; Keum, G.; Lee, J.-H.; Chung, B. Y.; Kim, Y.; Rhim,
H.; Kang, S. B. Bioorg. Med. Chem. Lett. 2006, 16, 5244; (j) Park, S. J.; Park, S. J.;
Lee, M. J.; Rhim, H.; Kim, Y.; Lee, J.-H.; Chung, B. Y.; Lee, J. Y. Bioorg. Med. Chem.
2006, 14, 3502; (k) Choi, J. Y.; Seo, H. N.; Lee, M. J.; Park, S. J.; Park, S. J.; Jeon, J.
Y.; Kang, J. H.; Pae, A. N.; Rhim, H.; Lee, J. Y. Bioorg. Med. Chem. Lett. 2007, 17,
471; (l) Kim, H. S.; Kim, Y.; Doddareddy, M. R.; Seo, S. H.; Rhim, H.; Tae, J.; Pae,
A. N.; Choo, H.; Cho, Y. S. Bioorg. Med. Chem. Lett. 2007, 17, 476; (m) Seo, H. N.;
Choi, J. Y.; Choe, Y. J.; Kim, Y.; Rhim, H.; Lee, S. H.; Kim, J.; Joo, D. J.; Lee, J. Y.
Bioorg. Med. Chem. Lett. 2007, 17, 5740; (n) Doddareddy, M. R.; Choo, H.; Yong,
S.; Rhim, H.; Koh, H. Y.; Lee, J.-H.; Jeong, S.-W.; Pae, A. N. Bioorg. Med. Chem.
2007, 15, 1091; (o) Park, J. H.; Choi, J. K.; Lee, E.; Lee, J. K.; Rhim, H.; Seo, S. H.;
Kim, Y.; Doddareddy, M. R.; Pae, A. N.; Kang, J.; Roh, E. J. Bioorg. Med. Chem.
2007, 15, 1409; (p) Hangeland, J. L.; Cheney, D. L.; Friends, T. J.; Swartz, S.;
Levesque, P. C.; Rich, A. J.; Sun, L.; Bridal, T. R.; Adam, L. P.; Normandin, D. E.;
Murugesan, N.; Ewing, W. R. Bioorg. Med. Chem. Lett. 2008, 18, 474; (q) Heo, J.
H.; Seo, H. N.; Choe, Y. J.; Kim, S.; Chun, R. O.; Kim, Y. D.; Rhim, H.; Choo, D. J.;
Kim, J.; Lee, J. Y. Bioorg. Med. Chem. Lett. 2008, 18, 3899; (r) Lee, H. K.; Lee, Y. S.;
Roh, E. J.; Rhim, H.; Lee, J. Y.; Shin, K. J. Bioorg. Med. Chem. Lett. 2008, 18, 4424;
(s) Shipe, W. D.; Barrow, J. C.; Yang, Z.-Q.; Lindsley, C. W.; Yang, F. V.; Schlegel,
12. (a) Pacofsky, G. J.; Suto, M. J.; Fritch, P. C. PCT Int. Appl. WO 2007075852.; (b)
Pacofsky, G. J.; Suto, M. J.; Fritch, P. C. PCT Int. Appl. WO 2007073497.
13.
a
1H (Cav3.2, T-type) and
standard whole-cell electrophysiology with cell lines prepared in house. The
subunits for the L-type channel are 1C/ 2d1/b2. 10 mM DMSO solutions of
a1C (Cav2.2, L-type) currents were recorded using
a
a
compounds were diluted with external solution. Test compounds were
superfused until steady-state inhibition was reached (typically within 1–
2 min). Data are reported as percent inhibition compared to control. Protocol
for a1H: external solution in mM NaCl (132), KCl (5.4), CaCl2 (1.8), MgCl2 (0.8),
4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid (HEPES) (10), glucose (5);
pH to 7.4 with NaOH. Internal solution in mM CsAsp (110), CsCl (20), ethylene
glycol-bis(2-aminoethylether)-N,N,N0,N0-tetraacetic acid (EGTA) (5), MgCl2 (1),
MgATP (5), Na-GTP (0.2), HEPES (10), pH to 7.3 with CsOH. The half-
inactivation potential was determined by the potential needed to generate
half-maximal current. The cell was held at the half-inactivation potential then
stepped to ꢁ20 mV for 200 ms at 15 s intervals. Protocol for
a1C: external
solution in mM NaCl (132), KCl (5.4), BaCl2 (10), MgCl2 (0.8), HEPES (10),
glucose (5); pH to 7.4 with NaOH. Internal solution in mM CsAsp (110), CsCl
(20), EGTA (5), MgCl2 (1), MgATP (5), Na-GTP (0.2), HEPES (10), pH to 7.3 with
CsOH. The half-inactivation potential was determined by the potential needed
to generate half-maximal current. For compound testing the cell was held at
ꢁ90 mV then stepped to the half-inactivation potential for 8 s. Current was
activated by stepping to +25 mV for 200 ms. The protocol was repeated every
20 s.
14. Five-point IC50’s for mibefradil were determined by standard whole-cell
electrophysiology. 1H (Cav3.2, T-type) = 32 nM and 1C (Cav2.2, L-
type) = 416 nM, 13-fold selective.
a
a
15. Briner, K.; Collado, I.; Fisher, M. J.; Garcia-Paredes, C.; Husain, S.; Kuklish, S. L.;
Mateo, A. I.; O’Brien, T. P.; Ornstein, P. L.; Zgombick, J.; De Frutos, O. Bioorg.
Med. Chem. Lett. 2006, 16, 3449.
16. Smith, P. W.; Cooper, A. W. J.; Bell, R.; Beresford, I. J. M.; Gore, P. M.; McElroy, A.
B.; Pritchard, J. M.; Saez, V.; Taylor, N. R.; Sheldrick, R. L. G.; Ward, P. J. Med.
Chem. 1995, 38, 3772.