High-throughput screening for N-type calcium channel blockers using a scintillation proximity assay.
N-type calcium channels located on presynaptic nerve terminals regulate neurotransmitter release, including that from the spinal terminations of primary afferent nociceptors. Accordingly, N-type calcium channel blockers may have clinical utility as analgesic drugs. A selective N-type calcium channel inhibitor, ziconotide (Prialt), is a neuroactive peptide recently marketed as a novel nonopioid treatment for severe chronic pain. To develop a small-molecule N-type calcium channel blocker, the authors developed a 96-well plate high-throughput screening scintillation proximity assay (SPA) for N-type calcium channel blockers using [125I]-labeled omega-conotoxin GVIA as a channel-specific ligand. Assay reagents were handled using Caliper's Allegro automation system, and bound ligands were detected using a PerkinElmer TopCount. Using this assay, more than 150,000 compounds were screened at 10 microM and approximately 340 compounds were identified as hits, exhibiting at least 40% inhibition of [125I]GVIA binding. This is the 1st demonstration of the use of [125I]-labeled peptides with SPA beads to provide a binding assay for the evaluation of ligand binding to calcium channels. This assay could be a useful tool for drug discovery.
Zhang
Sui-Po
SP
Drug Discovery, Johnson & Johnson Pharmaceutical Research and Development, LLC, Spring House, Pennsylvania 19477, USA. spzhang@prdus.jnj.com
Kauffman
Jack
J
Yagel
Susan K
SK
Codd
Ellen E
EE
D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000032:analysis* / Q000737:chemistry; D020864:Calcium Channels, N-Type / Q000378:metabolism*; D004353:Drug Evaluation, Preclinical; D051381:Rats; D012588:Scintillation Counting
D002121:Calcium Channel Blockers; D020864:Calcium Channels, N-Type