An Integrated Multiassay Approach to the Discovery of Small-Molecule N-Type Voltage-Gated Calcium Channel Antagonists

التفاصيل البيبلوغرافية
العنوان: An Integrated Multiassay Approach to the Discovery of Small-Molecule N-Type Voltage-Gated Calcium Channel Antagonists
المؤلفون: Yi Liu, Michael F.A. Finley, Mary Lou Lubin, Ning Qin, Michael P. Neeper, Christopher M. Flores, Edward J. Beck
المصدر: ASSAY and Drug Development Technologies. 8:685-694
بيانات النشر: Mary Ann Liebert Inc, 2010.
سنة النشر: 2010
مصطلحات موضوعية: Patch-Clamp Techniques, Calcitonin Gene-Related Peptide, Presynaptic Terminals, Pain, Pharmacology, omega-Conotoxins, Cell Line, Small Molecule Libraries, Calcium Channels, N-Type, Dorsal root ganglion, Ganglia, Spinal, Drug Discovery, Calcium flux, medicine, Animals, Humans, Patch clamp, Ion channel, Neurons, Analgesics, Ziconotide, Voltage-dependent calcium channel, Chemistry, Calcium channel, Calcium Channel Blockers, High-Throughput Screening Assays, Rats, Electrophysiology, HEK293 Cells, medicine.anatomical_structure, Spinal Cord, Luminescent Measurements, Molecular Medicine, Neuroscience, medicine.drug
الوصف: The N-type voltage-gated calcium channel (Cav2.2) has been intensively explored as a target for novel, small-molecule analgesic drugs because of its distribution in the pain pathway and its role in nociceptive processing. For example, Cav2.2 is localized at presynaptic terminals of pain fibers in the dorsal horn, and it serves as a downstream effector of μ-opioid receptors. Most importantly, antagonism of the channel by the highly specific and potent Cav2.2 blocker ω-conotoxin MVIIA (ziconotide) produces clinical efficacy in the treatment of severe, intractable pain. To identify novel small-molecule Cav2.2 inhibitors, we developed new tools and screening methods critical to enhance the efficiency and probability of success. First, we established and characterized a new cell line stably expressing the three subunits of the Cav2.2, including an α-subunit splice variant that is uniquely expressed by dorsal root ganglion neurons. Second, using this cell line, we validated and employed a fluorescence-based calcium flux assay. Third, we developed a new "medium-throughput" electrophysiology assay using QPatch-HT to provide faster turnaround on high-content electrophysiology data that are critical for studying ion channel targets. Lastly, we used a therapeutically relevant, ex vivo spinal cord calcitonin gene-related peptide-release assay to confirm activities in the other assays. Using this approach we have identified compounds exhibiting single-digit nM IC₅₀ values and with a positive correlation across assay methods. This integrated approach provides a more comprehensive evaluation of small-molecule N-type inhibitors that may lead to improved therapeutic pharmacology.
تدمد: 1557-8127
1540-658X
URL الوصول: https://explore.openaire.eu/search/publication?articleId=doi_dedup___::021e926842b8366d4a8742a010cc48f2
https://doi.org/10.1089/adt.2010.0311
رقم الأكسشن: edsair.doi.dedup.....021e926842b8366d4a8742a010cc48f2
قاعدة البيانات: OpenAIRE