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hyperalgesia and chronic pain.
36-38
However, it was also found that capsaze-
pine blocks receptors other than TRPV1, such as voltage-gated Ca
21
chan-
nels
39
and nicotinic acetylcholine receptors,
40
and does not act as an antagonist
when the TRPV1 channel is activated by heat or acid. Other evidence for the
role of TRPV1 in pain sensation has come from studies with TRPV1 knock-out
mice.
41,42
These TRPV1-deficient mice have a decreased sensitivity to painful
heat in inflammatory pain models.
Based on these considerations, we set out to discover and develop TRPV1
antagonists that block all modes of activation. We hypothesized that this type
of agent may offer a rapid onset of analgesic action by blockade of the pain-
signaling pathway with potentially fewer side effects.
43,44
13.2 Lead Identification
To generate and identify new leads we conducted a high-throughput screen
(HTS) of our corporate library consisting of approximately 1 million com-
pounds. For the purposes of our study, we employed rat or human TRPV1
channel recombinantly expressed in Chinese hamster ovary (CHO) cells in a
cell-based assay, where an increase in intracellular calcium resulted in light
emission through transfected aequorin. We then confirmed antagonism of the
primary hits in assays measuring both capsaicin- and pH-mediated influx of
45
Ca
21
.
45
As a result of the screen we identified a number of hits from various
structural classes (e.g., compounds 3-5; Figure 13.2).
Through traditional medicinal chemistry investigations, several potent ben-
zimidazole,
46
thiazole amide,
47
imidazole,
48
and cinnamide
49
TRPV1 antago-
nists were generated from these original hits and their structure-activity
O
Cl
H
N
S
HN
H
N
O
Cl
3
4
hTRPV1(Cap) IC
50
= 5,800 nM
hTRPV1(Cap) IC
50
= 370 nM
Cl
N
Cl
O
5
hTRPV1(Cap) IC
50
= 1,200 nM
Figure 13.2
Examples of initial hits obtained from TRPV1 high-throughput screen-
ing efforts.
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