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Abstract Agonists of TRPA1 such as mustard oil and its key component AITC cause pain and neurogenic inflammation in humans and pain behavior in rodents. TRPA1 is activated by numerous reactive compounds making it a sensor for reactive compounds in the body. Failure of AITC, formalin and other reactive compounds to trigger pain behavior in TRPA1 knockout mice, as well as the ability of TRPA1 antisense to alleviate cold hyperalgesia after spinal nerve ligation, suggest that TRPA1 is a potential target for novel analgesic agents. Here, we have characterized CHO cells expressing human and rat TRPA1 driven by an inducible promoter. As reported previously, both human and rat TRPA1 are activated by AITC and inhibited by ruthenium red. We have also characterized noxious cold response of these cell lines and show that noxious cold activates both human and rat TRPA1. Further, we have used CHO cells expressing human TRPA1 to screen a small molecule compound library and discovered that 'trichloro(sulfanyl)ethyl benzamides' (AMG2504, AMG5445, AMG7160 and AMG9090) act as potent antagonists of human TRPA1 activated by AITC and noxious cold. However, trichloro(sulfanyl)ethyl benzamides' (TCEB compounds) displayed differential pharmacology at rat TRPA1. AMG2504 and AMG7160 marginally inhibited rat TRPA1 activation by AITC, whereas AMG5445 and AMG9090 acted as partial agonists. In summary, we conclude that both human and rat TRPA1 channels show similar AITC and noxious cold activation profiles, but TCEB compounds display species-specific differential pharmacology at TRPA1.

The macronucleus of the ciliate Tetrahymena thermophila contains a fragmented somatic genome consisting of several hundred identifiable chromosome pieces. These pieces are generated by site-specific fragmentation of the germline chromosomes and most of them are represented at an average of 45 copies per macronucleus. In the course of successive divisions of an initially heterozygous macronucleus, the random distribution of alleles of loci carried on these copies eventually generates macronuclei that are pure for one allele or the other. This phenomenon is called phenotypic assortment. We have previously reported the existence of loci that assort together (coassort) and hypothesized that these loci reside on the same macronuclear piece. The work reported here provides new, rigorous genetic support for the hypothesis that macronuclear autonomously replicating chromosome pieces are the physical basis of coassortment groups. Thus, coassortment allows the mapping of the somatic genome by purely genetic means. The data also strongly suggest that the random distribution of alleles in the Tetrahymena macronucleus is due to the random distribution of the MAC chromosome pieces that carry them.

The vanilloid receptor TRPV1 (transient receptor potential vanilloid 1) is a cation channel that serves as a polymodal detector of pain-producing stimuli such as capsaicin, protons (pH < 5.7), and heat. TRPV1 antagonists block pain behaviors in rodent models of inflammatory, neuropathic, and cancer pain, suggesting their utility as analgesics. Here, we report that TRPV1 antagonists representing various chemotypes cause an increase in body temperature (hyperthermia), identifying a potential issue for their clinical development. Peripheral restriction of antagonists did not eliminate hyperthermia, suggesting that the site of action is predominantly outside of the blood- brain barrier. Antagonists that are ineffective against proton activation also caused hyperthermia, indicating that blocking capsaicin and heat activation of TRPV1 is sufficient to produce hyperthermia. All TRPV1 antagonists evaluated here caused hyperthermia, suggesting that TRPV1 is tonically activated in vivo and that TRPV1 antagonism and hyperthermia are not separable. TRPV1 antagonists caused hyperthermia in multiple species (rats, dogs, and monkeys), demonstrating that TRPV1 function in thermoregulation is conserved from rodents to primates. Together, these results indicate that tonic TRPV1 activation regulates body temperature. [ABSTRACT FROM AUTHOR]

Agonists of TRPA1 such as mustard oil and its key component AITC cause pain and neurogenic inflammation in humans and pain behavior in rodents. TRPA1 is activated by numerous reactive compounds making it a sensor for reactive compounds in the body. Failure of AITC, formalin and other reactive compounds to trigger pain behavior in TRPA1 knockout mice, as well as the ability of TRPA1 antisense to alleviate cold hyperalgesia after spinal nerve ligation, suggest that TRPA1 a potential target for novel analgesic agents. Here, we have characterized CHO cells expressing human and rat TRPA1 driven by an inducible promoter. As reported previously, both human and rat TRPA1 are activated by AITC and inhibited by ruthenium red. We have also characterized noxious cold response of these cell lines and show that noxious cold activates both human and rat TRPA1. Further, we have used CHO cells expressing human TRPA1 to screen a small molecule compound library and discovered that 'trichloro(sulfanyl)ethyl benzamides' (AMG2504, AMG5445, AMG7160 and AMG9090) act as potent antagonists of human TRPA1 activated by AITC and noxious cold. However, trichloro(sulfanyl)ethyl benzamides' (TCEB compounds) displayed differential pharmacology at rat TRPA1. AMG2504 and AMG7160 marginally inhibited rat TRPA1 activation by AITC, whereas AMG5445 and AMG9090 acted as partial agonists. In summary, we conclude that both human and rat TRPA1 channels show similar AITC and noxious cold activation profiles, but TCEB compounds display species-specific differential pharmacology at TRPA1. [ABSTRACT FROM AUTHOR]

The vanilloid receptor-1 (VR1 or TRPV1) is a membrane-bound, nonselective cation channel that is predominantly expressed by peripheral neurons sensing painful stimuli. TRPV1 antagonists produce antihyperalgesic effects in animal models of inflammatory and neuropathic pain. Herein, we describe the synthesis and the structure-activity relationships of a series of 2-(4-pyridin-2-ylpiperazin-1-yl)-1H-benzo[d]imidazoles as novel TRPV1 antagonists. Compound 46ad was among the most potent analogues in this series. This compound was orally bioavailable in rats and was efficacious in blocking capsaicin-induced flinch in rats in a dose-dependent manner. Compound 46ad also reversed thermal hyperalgesia in a model of inflammatory pain, which was induced by complete Freund's adjuvant (CFA).

The vanilloid receptor-1 (TRPV1 or VR1) is a member of the transient receptor potential (TRP) family of ion channels and plays a role in regulating the function of sensory nerves. A growing body of evidence demonstrates the therapeutic potential of TRPV1 modulators, particularly in the management of pain. As a result of our screening efforts, we identified (E)-3-(4-tert-butylphenyl)-N-(2,3-dihydrobenzo[b][1,4]dioxin-6-yl)acrylamide (1), an antagonist that blocks the capsaicin-induced and pH-induced uptake of (45)Ca(2+) in TRPV1-expressing Chinese hamster ovary cells with IC(50) values of 17 +/- 5 and 150 +/- 80 nM, respectively. In this report, we describe the synthesis and structure-activity relationship of a series of N-aryl cinnamides, the most potent of which (49a and 49b) exhibit good oral bioavailability in rats (F(oral) = 39% and 17%, respectively).