Identification of 7 Nicotinic Acetylcholine Receptor Agonists for their Assessment in Improving Cognition in Schizophrenia - Accounts in Drug Discovery Case Studies in Medicinal Chemistry - page 335

O

O

DPPA or DPPC
TEA, CH 2 Cl 2

R

+ RNH 2

OH

H

R

R

Figure 15.2 Coupling conditions for acid and amine fragments (left) and data from
the evaluation of the compounds in the FLIPR assay (right). Adapted
with permission of the American Chemical Society from J. Med. Chem.,
2005, 48, 905.

profile fully, we used a binding assay and a hippocampal neuron assay to
validate the chimera assay as a primary screen predictive of native activity. In
the a7 nAChR binding assay, PNU-282987 displaced the a7 nAChR-selective
antagonist methyllycaconitine (MLA) from rat brain homogenate (K i ¼ 27
nM), and when applied to cultured rat hippocampal neurons it evoked a
rapidly desensitizing inward whole-cell current that was concentration depen-
dent and blocked by MLA, consistent with the opening of the a7 nAChR. 18
Another critical aspect of the screen was selectivity, starting with 5-HT 3
selectivity. PNU-282987 was selective against the neuromuscular junction form
of the receptor (a1b1gd) and the predominant ganglionic nAChR (a3b4), which
was critical because activation of these receptors causes many of the undesir-
able effects of nonspecific nAChR agonists such as epibatidine and nicotine. 19
In addition to its selectivity profile, PNU-282987 had very desirable PK
properties, with robust metabolic stability in rat and human liver microsomes
(HLM), low in vitro clearance, excellent brain penetration, and high oral
bioavailability in rats. 17,20 A key safety issue identified in the early profiling of
this compound was hERG (human ether-a` -go-go) potassium channel activity.
Significant hERG activity with this compound was an obstacle for further
development and would be a critical criterion in identifying additional chemical
matter, but 14-day rodent in vivo toxicology studies strengthened our con-
fidence in this template as a safe chemotype for further SAR studies.
One of the most significant milestones for PNU-282987 was its early in vivo
proof of pharmacology, which was established through tests in a rat model of
impaired sensory gating (P50) that was also validated with GTS-21. 8 P50 was
measured in a rat model of auditory sensory gating in which normal auditory
gating was disrupted by systemic administration of d-amphetamine. 21 Systemic
administration of d-amphetamine (0.3 or 1mg kg 1 iv) significantly disrupted
auditory gating in anesthetized rats owing to simultaneous decreases of con-
ditioning responses and increases in test responses. 22 Subsequent administration