thapsigargin and Dermatitis--Contact

There is a major clinical need for new therapies for the treatment of chronic itch. Many of the molecular components involved in itch neurotransmission are known, including the neuropeptide NPPB, a transmitter required for normal itch responses to multiple pruritogens in mice. Here, we investigated the potential for a novel strategy for the treatment of itch that involves the inhibition of the NPPB receptor NPR1 (natriuretic peptide receptor 1). Because there are no available effective human NPR1 (hNPR1) antagonists, we performed a high-throughput cell-based screen and identified 15 small-molecule hNPR1 inhibitors. Using in vitro assays, we demonstrated that these compounds specifically inhibit hNPR1 and murine NPR1 (mNPR1). In vivo, NPR1 antagonism attenuated behavioral responses to both acute itch- and chronic itch-challenged mice. Together, our results suggest that inhibiting NPR1 might be an effective strategy for treating acute and chronic itch.

Topics: Animals; Behavior, Animal; Cell-Free System; Dermatitis, Contact; Disease Models, Animal; Ganglia, Spinal; Humans; Mice, Inbred C57BL; Mice, Knockout; Neurons; Pruritus; Receptors, Atrial Natriuretic Factor; Reproducibility of Results; Signal Transduction; Small Molecule Libraries

In nonexcitable cells, Ca(2+) entry is mediated predominantly through the store depletion-dependent Ca(2+) channels called store-operated Ca(2+) (SOC) or Ca(2+) release-activated Ca(2+) channels. YM-58483, a pyrazole derivative, inhibited an anti-CD3 mAb-induced sustained Ca(2+) influx in acute T cell leukemia, Jurkat cells. But it did not affect an anti-CD3 mAb-induced transient intracellular Ca(2+) increase in Ca(2+)-free medium, nor anti-CD3 mAb-induced phosphorylation of phospholipase Cgamma1. It was suggested that YM-58483 inhibited Ca(2+) influx through SOC channels without affecting the TCR signal transduction cascade. Furthermore, YM-58483 inhibited thapsigargin-induced sustained Ca(2+) influx with an IC(50) value of 100 nM without affecting membrane potential. YM-58483 inhibited by 30-fold the Ca(2+) influx through SOC channels compared with voltage-operated Ca(2+) channels, while econazole inhibited both SOC channels and voltage-operated Ca(2+) channels with an equivalent range of IC(50) values. YM-58483 potently inhibited IL-2 production and NF-AT-driven promoter activity, but not AP-1-driven promoter activity in Jurkat cells. Moreover, this compound inhibited delayed-type hypersensitivity in mice with an ED(50) of 1.1 mg/kg. Therefore, we concluded that YM-58483 was a novel store-operated Ca(2+) entry blocker and a potent immunomodulator, and could be useful for the treatment of autoimmune diseases and chronic inflammation. Furthermore, YM-58483 would be a candidate for the study of capacitative Ca(2+) entry mechanisms through SOC/CRAC channels and for identification of putative Ca(2+) channel genes.

Topics: Anilides; Animals; Antibodies, Monoclonal; Calcium; Calcium Channel Blockers; Calcium Channels; Calcium Signaling; CD3 Complex; Dermatitis, Contact; DNA-Binding Proteins; Gene Expression Regulation; Genes, Reporter; Humans; Interleukin-2; Intracellular Fluid; Ion Channel Gating; Jurkat Cells; Male; Membrane Potentials; Mice; NFATC Transcription Factors; Nuclear Proteins; PC12 Cells; Phospholipase C gamma; Phosphorylation; Phytohemagglutinins; Picryl Chloride; Pyrazoles; Rats; T-Lymphocytes; Thapsigargin; Thiadiazoles; Transcription Factor AP-1; Transcription Factors; Type C Phospholipases; Tyrosine