Encapsulation of a TRPM8 Agonist, WS12, in Lipid Nanocapsules Potentiates PC3 Prostate Cancer Cell Migration Inhibition through Channel Activation.
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ABSTRACT:
SUBMITTER: Grolez GP
PROVIDER: S-EPMC6538610 | biostudies-literature | 2019 May
REPOSITORIES: biostudies-literature
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Project description:In prostate carcinogenesis, androgens are known to control the expression of the transient receptor potential melastatin 8 (TRPM8) protein via activation of androgen receptor (AR). Overexpression and/or activity of TRPM8 channel was shown to suppress prostate cancer (PCa) cell migration. Here we report that at certain concentrations androgens facilitate PCa cell migration. We show that underlying mechanism is inhibition of TRPM8 by activated AR which interacts with the channel within lipid rafts microdomains of the plasma membrane. Thus, our study has identified an additional nongenomic mechanism of the TRPM8 channel regulation by androgens that should be taken into account upon the development of novel therapeutic strategies.
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Project description:Transient receptor potential channels are a family of cation channels involved in diverse cellular functions. Most of these channels are expressed in the nervous system and play a key role in sensory physiology. TRPM8 (transient receptor potential melastatine 8), a member of this family, is activated by cold, cooling substances such menthol and icilin and voltage. Although TRPM8 is a thermosensitive channel highly expressed in cold sensory neurons, the mechanisms underlying its temperature sensitivity are still poorly understood. Here we show that, in sensory neurons, TRPM8 channel is localized in cholesterol-rich specialized membrane domains known as lipid rafts. We also show that, in heterologous expression systems, lipid raft segregation of TRPM8 is favored by glycosylation at the Asn(934) residue of the polypeptide. In electrophysiological and imaging experiments, using cold and menthol as agonists, we also demonstrate that lipid raft association modulates TRPM8 channel activity. We found that menthol- and cold-mediated responses of TRPM8 are potentiated when the lipid raft association of the channel is prevented. In addition, lipid raft disruption shifts the threshold for TRPM8 activation to a warmer temperature. In view of these data, we suggest a role for lipid rafts in the activity and temperature sensitivity of TRPM8. We propose a model wherein different lipid membrane environments affect the cold sensing properties of TRPM8, modulating the response of cold thermoreceptors.
| S-EPMC2666574 | biostudies-literature
Project description:Cancer is a serious health problem due to the complexity of establishing an effective treatment. The purpose of this work was to evaluate the activity of a triazaspirane as a migration and invasion inhibitor in PC3 prostatic tumor cells through a possible negative regulation of the FAK/Src signal transduction pathway and decreased secretion of metalloproteinases 2 and 9. Molecular docking analysis was performed using Moe 2008.10 software. Migration (wound-healing assay) and invasion (Boyden chamber assay) assays were performed. In addition, the Western blot technique was used to quantify protein expression, and the zymography technique was used to observe the secretion of metalloproteinases. Molecular docking showed interactions in regions of interest of the FAK and Src proteins. Moreover, the biological activity assays demonstrated an inhibitory effect on cell migration and invasion, an important suppression of metalloproteinase secretion, and a decrease in the expression of p-FAK and p-Src proteins in treated PC3 cells. Triazaspirane-type molecules have important inhibitory effects on the mechanisms associated with metastasis in PC3 tumor cells.
| S-EPMC10254162 | biostudies-literature
Project description:The cold and menthol receptor TRPM8 is highly expressed in prostate and prostate cancer (PC). Recently, we identified that TRPM8 is as an ionotropic testosterone receptor. The TRPM8 mRNA is expressed in early prostate tumors with high androgen levels, while anti-androgen therapy greatly reduces its expression. Here, from the chromatin-immunoprecipitation (ChIP) analysis, we found that an androgen response element (ARE) mediates androgen regulation of trpm8. Furthermore, using immunofluorescence, calcium-imaging and planar lipid bilayers, we identified that TRPM8 channel is functionally regulated by androgens in the prostate. Although TRPM8 mRNA is expressed at high levels, we found that the TRPM8 protein undergoes ubiquitination and degradation in PC cells. The mass-spectrometry analysis of TRPM8, immunoprecipitated from LNCaP cells identified ubiquitin-like modifier-activating enzyme 1 (UBA1). PYR-41, a potent inhibitor of initial enzyme in the ubiquitination cascade, UBA1, increased TRPM8 activity on the plasma membrane (PM) of LNCaP cells. Furthermore, PYR-41-mediated PMTRPM8 activity was accompanied by enhanced activation of p53 and Caspase-9. Interestingly, we found that the trpm8 promoter possesses putative binding sites for p53 and that the overexpression of p53 increased the TRPM8 mRNA levels. In addition to the genomic regulation of TRPM8 by AR and p53, our findings indicate that the testosterone-induced PMTRPM8 activity elicits Ca2+ uptake, subsequently causing apoptotic cell death. These findings support the strategy of rescuing PMTRPM8 expression as a new therapeutic application through the regulation of PC cell growth and proliferation.
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Project description:Transient receptor potential melastatin member 8 (TRPM8) is a calcium ion (Ca2+)-permeable cation channel that serves as the primary cold and menthol sensor in humans. Activation of TRPM8 by cooling compounds relies on allosteric actions of agonist and membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2), but lack of structural information has thus far precluded a mechanistic understanding of ligand and lipid sensing by TRPM8. Using cryo-electron microscopy, we determined the structures of TRPM8 in complex with the synthetic cooling compound icilin, PIP2, and Ca2+, as well as in complex with the menthol analog WS-12 and PIP2 Our structures reveal the binding sites for cooling agonists and PIP2 in TRPM8. Notably, PIP2 binds to TRPM8 in two different modes, which illustrate the mechanism of allosteric coupling between PIP2 and agonists. This study provides a platform for understanding the molecular mechanism of TRPM8 activation by cooling agents.
| S-EPMC6478609 | biostudies-literature
Project description:The transient receptor potential melastatin 8 (TRPM8) channel is the primary molecular transducer responsible for the cool sensation elicited by menthol and cold in mammals. TRPM8 activation is controlled by cooling compounds together with the membrane lipid phosphatidylinositol 4,5-bisphosphate (PIP2). Our knowledge of cold sensation and the therapeutic potential of TRPM8 for neuroinflammatory diseases and pain will be enhanced by understanding the structural basis of cooling agonist- and PIP2-dependent TRPM8 activation. We present cryo-electron microscopy structures of mouse TRPM8 in closed, intermediate, and open states along the ligand- and PIP2-dependent gating pathway. Our results uncover two discrete agonist sites, state-dependent rearrangements in the gate positions, and a disordered-to-ordered transition of the gate-forming S6-elucidating the molecular basis of chemically induced cool sensation in mammals.
| S-EPMC9795508 | biostudies-literature
Project description:Prostate cancer (PCa) is the second leading cause of cancer?related mortality among American males. Studies suggest that cigarette smoking is associated with the progression of PCa; however, the molecular mechanisms underlying this process have not been extensively investigated. PCa progression is characterized by increased cell migration and alterations in extracellular matrix (ECM)? and cell adhesion molecule (CAM)?related gene expression. In the present study, the influence of cigarette smoke medium (SM) on cell migration and on the expression of ECM? and CAM?related genes in PC3 prostate adenocarcinoma cells was investigated. According to a wound?healing assay, SM treatment promoted PC3 cell migration. RNA expression levels from SM?treated and control cells were analyzed using a polymerase chain reaction (PCR) array. Of 84 genes analyzed, 27.38% (23/84) exhibited a ?2?fold change in threshold cycle in PC3 cells following 0.5% SM treatment. Functional gene grouping analysis demonstrated that SM treatment modulated the RNA transcription of approximately 18.4% of CAMs and 33.93% of ECM?related genes. Quantitative PCR analysis showed that SM treatment led to a significant decrease in transcription levels of the following genes: Collagen 5 ??1(V), connective tissue growth factor, integrin ??2, kallmann syndrome 1, laminin ? 3, matrix metallopeptidase 7 (MMP7), MMP13, secreted protein acidic cysteine?rich, thrombospondin?2 and versican; and that SM significantly increased the transcription levels of MMP2 and MMP12. Furthermore, MMP2 knockdown significantly reduced the migration of SM?treated PC3 cells. The present study provides novel insights into the association of cigarette smoking with PCa progression, via the alteration of ECM/CAM interactions.
| S-EPMC4626126 | biostudies-literature
Project description:Endothelial cell adhesion and migration are critical steps of the angiogenic process, whose dysfunction is associated with tumor growth and metastasis. The TRPM8 channel has recently been proposed to play a protective role in prostate cancer by impairing cell motility. However, the mechanisms by which it could influence vascular behavior are unknown. Here, we reveal a novel non-channel function for TRPM8 that unexpectedly acts as a Rap1 GTPase inhibitor, thereby inhibiting endothelial cell motility, independently of pore function. TRPM8 retains Rap1 intracellularly through direct protein-protein interaction, thus preventing its cytoplasm-plasma membrane trafficking. In turn, this mechanism impairs the activation of a major inside-out signaling pathway that triggers the conformational activation of integrin and, consequently, cell adhesion, migration, in vitro endothelial tube formation, and spheroid sprouting. Our results bring to light a novel, pore-independent molecular mechanism by which endogenous TRPM8 expression inhibits Rap1 GTPase and thus plays a critical role in the behavior of vascular endothelial cells by inhibiting migration.
| S-EPMC5496606 | biostudies-literature
Project description:BackgroundPhysical cooling of the eye surface relieves ocular discomfort, but translating this event to drug treatment of dry eye discomfort not been studied. Here, we synthesized a water-soluble TRPM8 receptor agonist called cryosim-3 (C3, 1-diisopropylphosphorylnonane) which selectively activates TRPM8 (linked to cooling) but not TRPV1 or TRPA1 (linked to nociception) and tested C3 in subjects with mild forms of dry eye disease.MethodsA set of 1-dialkylphosphoryalkanes were tested for activation of TRPM8, TRPV1 and TRPA1 receptors in transfected cells. The bioactivity profiles were compared by perioral, topical, and intravenous delivery to anesthetized rats. The selected lead candidate C3 or vehicle (water) was applied with a cotton gauze pad to upper eyelids of patients with dry eye disease (n = 30). Cooling sensation, tear film break-up time (TBUT), basal tear secretion, and corneal staining were evaluated. C3 was then applied four times daily for 2 weeks to patients using a pre-loaded single unit applicator containing 2 mg/mL of C3 in water (n = 20) or water only. TBUT, basal tear secretion, and corneal staining, and three questionnaires surveys of ocular discomfort (VAS scale, OSDI, and CVS symptoms) were analyzed before and at 1 and 2 weeks thereafter.ResultsC3 was a selective and potent TRPM8 agonist without TRPV1 or TRPA1 activity. In test animals, the absence of shaking behavior after C3 perioral administration made it the first choice for further study. C3 increased tear secretion in an animal model of dry eye disease and did not irritate when wiped on eyes of volunteers. C3 singly applied (2 mg/ml) produced significant cooling in <5 min, an effecting lasting 46 min with an increase in tear secretion for 60 min. C3 applied for 2 weeks also significantly increased basal tear secretion with questionnaire surveys of ocular discomfort indices clearly showing improvement of symptoms at 1 and 2 weeks. No complaints of irritation or pain were reported by any subject.ConclusionsC3 is a promising candidate for study of TRPM8 function on the eye surface and for relief of dry eye discomfort.Trial registrationISRCTN24802609 and ISRCTN13359367 . Registered 23 March 2015 and 2 September 2015.
| S-EPMC5485613 | biostudies-literature
Project description:BackgroundThe transient receptor potential channel (TRP) family includes more than 30 proteins; they participate in various Ca(2+) dependent processes. TRPs are functionally diverse involving thermal, chemical and mechanical transducers which modulate the concentration of intracellular Ca(2+) ([Ca(2+)]i). Ca(2+) triggers and/or regulates principal sperm functions during fertilization such as motility, capacitation and the acrosome reaction. Nevertheless, the presence of the TRPM subfamily in sperm has not been explored.Principal findingsHere we document with RT-PCR, western blot and immunocitochemistry analysis the presence of TRPM8 in human sperm. We also examined the participation of this channel in sperm function using specific agonists (menthol and temperature) and antagonists (BCTC and capsazepine). Computer-aided sperm analysis revealed that menthol did not significantly alter human sperm motility. In contrast, menthol induced the acrosome reaction in human sperm. This induction was inhibited about 70% by capsazepine (20 microM) and 80% by BCTC (1.6 microM). Activation of TRPM8 either by temperature or menthol induced [Ca(2+)]i increases in human sperm measured by fluorescence in populations or individual sperm cells, effect that was also inhibited by capsazepine (20 microM) and BCTC (1.6 microM). However, the progesterone and ZP3-induced acrosome reaction was not inhibited by capsazepine or BCTC, suggesting that TRPM8 activation triggers this process by a different signaling pathway.ConclusionsThis is the first report dealing with the presence of a thermo sensitive channel (TRPM8) in human sperm. This channel could be involved in cell signaling events such as thermotaxis or chemotaxis.
| S-EPMC2705237 | biostudies-literature