Icilin

Icilin is an agonist of TRPM8 and hENaCδ [1, 2]

Icilin is a synthetic supercooling compound. It is reported that icilin can activate TRPM8 to some small degree in the absence of extra-cellular Ca2+. It further enhances the potency of icilin and the subsequent activation of TRPM8 channel. Icilin is also an agonist of hENaCδ. In the homomeric hENaCδ-expressing oocytes, 100μM icilin induces an inward current significantly. This effect can be reduced when the external Na+ is removed. In addition, icilin shows anti-proliferation efficacy in PC-3 cells. It induces G1 arrest via modulating the expression of cell cycle regulators including cyclin A, cyclin D1, CDK1 and CDK2. Icilin plays this role without affecting TRPM8 but through activating NK and p38 kinase pathways [1, 2 and 3]

References:
[1] Chuang H, Neuhausser W M, Julius D. The super-cooling agent icilin reveals a mechanism of coincidence detection by a temperature-sensitive TRP channel. Neuron, 2004, 43(6): 859-869.
[2] Yamamura H, Ugawa S, Ueda T, et al. Icilin activates the δ-subunit of the human epithelial Na+ channel. Molecular pharmacology, 2005, 68(4): 1142-1147.
[3] Kim S H, Kim S Y, Park E J, et al. Icilin induces G1 arrest through activating JNK and p38 kinase in a TRPM8-independent manner. Biochemical and biophysical research communications, 2011, 406(1): 30-35.

Icilin reduces voltage-gated calcium channel currents in naive and injured DRG neurons in the rat spinal nerve ligation model.[Pubmed:24560602]

Brain Res. 2014 Apr 4;1557:171-9.

Recently, the transient receptor potential (TRP) channels TRPM8 and TRPA1 have been identified as molecular sensors for cold, and it has been suggested that they play a crucial role in allodynia by modulating voltage-gated calcium channel currents (ICa(V)). The aim of this study was to analyze the modulation of ICa(V) by the TRPM8-agonist Icilin in vitro and to investigate the analgesic effect of Icilin in a neuropathic pain model in vivo. Whole cell patch-clamp recordings were performed on isolated naive and injured rat dorsal root ganglia (DRG) neurons, and the analgesic efficacy of Icilin applied topically to the paws or intrathecally was tested in rats after spinal nerve ligation (SNL). ICa(V) (depolarization from -80 to 0mV) in naive DRG neurons was reduced dose dependently (0.002-200microM) by Icilin (18-80%). Subtype isolation of calcium channels show a marked reduction of L-type channel currents compared to N-type channel currents. The effects of Icilin on ICa(V) were not significantly different in non-injured and SNL-injured DRG neurons. In vivo, neither topical (10-200microM) nor intrathecal application of Icilin (0.1nM to 1microM) affected tactile allodynia or thermal hyperalgesia after SNL, but it increases cold allodynia 6h after application. We conclude that the Icilin-induced modulation of ICa(V) in DRG neurons is unlikely to mediate analgesic effects or contribute directly to the pathogenesis of cold allodynia in the rat SNL model, but it is a potential mechanism for the analgesic effects of Icilin in other pain models.

Icilin inhibits E2F1-mediated cell cycle regulatory programs in prostate cancer.[Pubmed:24239550]

Biochem Biophys Res Commun. 2013 Nov 29;441(4):1005-10.

Aberrant expression of cell cycle regulators have been implicated in prostate cancer development and progression. Therefore, understanding transcriptional networks controlling the cell cycle remain a challenge in the development of prostate cancer treatment. In this study, we found that Icilin, a super-cooling agent, down-regulated the expression of cell cycle signature genes and caused G1 arrest in PC-3 prostate cancer cells. With reverse-engineering and an unbiased interrogation of a prostate cancer-specific regulatory network, master regulator analysis discovered that Icilin affected cell cycle-related transcriptional modules and identified E2F1 transcription factor as a target master regulator of Icilin. Experimental analyses confirmed that Icilin reduced the activity and expression levels of E2F1. These results demonstrated that Icilin inactivates a small regulatory module controlling the cell cycle in prostate cancer cells. Our study might provide insight into the development of cell cycle-targeted cancer therapeutics.

Camphor activates and sensitizes transient receptor potential melastatin 8 (TRPM8) to cooling and icilin.[Pubmed:23828908]

Chem Senses. 2013 Sep;38(7):563-75.

Camphor is known to potentiate both heat and cold sensations. Although the sensitization to heat could be explained by the activation of heat-sensitive transient receptor potential (TRP) channels TRPV1 and TRPV3, the camphor-induced sensitization to cooling remains unexplained. In this study, we present evidence for the activation of the cold- and menthol-sensitive channel transient receptor potential melastatin 8 (TRPM8) by camphor. Calcium transients evoked by camphor in HEK293 cells expressing human and rat TRPM8 are inhibited by the TRPM8 antagonists 4-(3-chloro-2-pyridinyl)-N-[4-(1,1-dimethylethyl)phenyl]-1-piperazinecarboxamide and 2-aminoethyl diphenylborinate. Camphor also sensitized the cold-induced calcium transients and evoked desensitizing outward-rectifying currents in TRPM8-expressing HEK293 cells. In the presence of ruthenium red (a blocker of TRPV1, TRPV3, and TRPA1), the camphor sensitivity of cultured rat dorsal root ganglion neurons was highest in a subpopulation of cold- and Icilin-sensitive neurons, strongly suggesting that camphor activates native TRPM8. Camphor has a dual action on TRPM8: it not only activates the channel but also inhibits its response to menthol. The Icilin-insensitive chicken TRPM8 was also camphor insensitive. However, camphor was able to activate an Icilin-insensitive human TRPM8 mutant channel. The activation and sensitization to cold of mammalian TRPM8 are likely to be responsible for the psychophysical enhancement of innocuous cold and "stinging/burning" cold sensations by camphor.

Structure-activity relationships of the prototypical TRPM8 agonist icilin.[Pubmed:25935641]

Bioorg Med Chem Lett. 2015 Jun 1;25(11):2285-90.

A series of structural analogues of the TRPM8 agonist Icilin was prepared. The compounds were examined for their ability to exert agonist or antagonist effects in HEK-293 cells expressing the TRPM8 receptor. Most structural modifications of the Icilin structure largely met with diminished TRPM8 agonist activity. Cinnamamide 'open-chain' analogs of Icilin, however, demonstrated significant antagonistic actions at the TRPM8 receptor. Optimal potency (IC50=73 nM) was observed in the 3-iodo derivative 18l.

Characterization of the mouse cold-menthol receptor TRPM8 and vanilloid receptor type-1 VR1 using a fluorometric imaging plate reader (FLIPR) assay.[Pubmed:14757700]

Br J Pharmacol. 2004 Feb;141(4):737-45.

1. TRPM8 (CMR1) is a Ca(2+)-permeable channel, which can be activated by low temperatures, menthol, eucalyptol and Icilin. It belongs to the transient receptor potential (TRP) family, and therefore is related to vanilloid receptor type-1 (VR1, TRPV1). We tested whether substances which are structurally related to menthol, or which produce a cooling sensation, could activate TRPM8, and compared the responses of TRPM8 and VR1 to these ligands. 2. The effects of 70 odorants and menthol-related substances on recombinant mouse TRPM8 (mTRPM8), expressed in HEK293 cells, were examined using a FLIPR assay. In all, 10 substances (linalool, geraniol, hydroxycitronellal, WS-3, WS-23, FrescolatMGA, FrescolatML, PMD38, CoolactP and Cooling Agent 10) were found to be agonists. 3. The EC(50) values of the agonists defined their relative potencies: Icilin (0.2+/-0.1 microM)>FrescolatML (3.3+/-1.5 microM) > WS-3 (3.7+/-1.7 microM) >(-)menthol (4.1+/-1.3 microM) >frescolatMAG (4.8+/-1.1 microM) > cooling agent 10 (6+/-2.2 microM) >(+)menthol (14.4+/-1.3 microM) > PMD38 (31+/-1.1 microM) > WS-23 (44+/-7.3 microM) > Coolact P (66+/-20 microM) > geraniol (5.9+/-1.6 mM) > linalool (6.7+/-2.0 mM) > eucalyptol (7.7+/-2.0 mM) > hydroxycitronellal (19.6+/-2.2 mM). 4. Known VR1 antagonists (BCTC, thio-BCTC and capsazepine) were also able to block the response of TRPM8 to menthol (IC(50): 0.8+/-1.0, 3.5+/-1.1 and 18+/-1.1 microM, respectively). 5. The Ca(2+) response of hVR1-transfected HEK293 cells to the endogenous VR1 agonist N-arachidonoyl-dopamine was potentiated by low pH. In contrast, menthol- and Icilin-activated TRPM8 currents were suppressed by low pH. 6. In conclusion, in the present study, we identified 10 new agonists and three antagonists of TRPM8. We found that, in contrast to VR1, TRPM8 is inhibited rather than potentiated by protons.

ANKTM1, a TRP-like channel expressed in nociceptive neurons, is activated by cold temperatures.[Pubmed:12654248]

Cell. 2003 Mar 21;112(6):819-29.

Mammals detect temperature with specialized neurons in the peripheral nervous system. Four TRPV-class channels have been implicated in sensing heat, and one TRPM-class channel in sensing cold. The combined range of temperatures that activate these channels covers a majority of the relevant physiological spectrum sensed by most mammals, with a significant gap in the noxious cold range. Here, we describe the characterization of ANKTM1, a cold-activated channel with a lower activation temperature compared to the cold and menthol receptor, TRPM8. ANKTM1 is a distant family member of TRP channels with very little amino acid similarity to TRPM8. It is found in a subset of nociceptive sensory neurons where it is coexpressed with TRPV1/VR1 (the capsaicin/heat receptor) but not TRPM8. Consistent with the expression of ANKTM1, we identify noxious cold-sensitive sensory neurons that also respond to capsaicin but not to menthol.

Identification of a cold receptor reveals a general role for TRP channels in thermosensation.[Pubmed:11882888]

Nature. 2002 Mar 7;416(6876):52-8.

The cellular and molecular mechanisms that enable us to sense cold are not well understood. Insights into this process have come from the use of pharmacological agents, such as menthol, that elicit a cooling sensation. Here we have characterized and cloned a menthol receptor from trigeminal sensory neurons that is also activated by thermal stimuli in the cool to cold range. This cold- and menthol-sensitive receptor, CMR1, is a member of the TRP family of excitatory ion channels, and we propose that it functions as a transducer of cold stimuli in the somatosensory system. These findings, together with our previous identification of the heat-sensitive channels VR1 and VRL-1, demonstrate that TRP channels detect temperatures over a wide range and are the principal sensors of thermal stimuli in the mammalian peripheral nervous system.