A plant-derived TRPV3 inhibitor suppresses pain and itch

PUBLISHED IN:- JOURNAL OF BRITISH PHARMACOLOGY. DOI: 10.1111/BPH.15390

Abstract

Background and Purpose: Itching is the most frequent pathology in dermatology that has significant impacts on people’s mental health and social life. Transient receptor potential vanilloid 3 (TRPV3) channel is a promising target for treating pruritus. However, few selecetive and potent antagonists have been reported. This study was designed to identify selective TRPV3 antagonist and elucidate its anti-pruritus pharmacology.

Experimental Approach: FlexStation and calcium fluorescence imaging were conducted to track the functional compounds. Whole-cell patch clamp was used to record itch-related ion channel currents. Homologous recombination and site-directed mutagenesis were employed to construct TRPV3 channel chimeras and point mutations for exploring pharmacological mechanism. Mouse models were used for in vivo anti-pruritus assay.

Key Results: An acridone alkaloid (citrusinine-II) was purified and characterized from Atalantia monophylla. It directly interacts with Y564 within S4 helix of TRPV3 to selectively inhibit the channel with a half maximal inhibitory concentration (IC50) of 12.43 μM. Citrusinine-II showed potential efficacy to attenuate both chronic and acute itch. Intradermal administration of citrusinine-II (143 ng/skin site) nearly completely inhibited itch behaviours. It also shows significant analgesic effects. Little side effects of the compound are observed.

Conclusion and Implications: By acting as a selective and potent inhibitor of TRPV3 channel, citrusinine-II shows valuable therapeutic effects in pruritus animal models and is a promising candidate drug and/or lead molecule for the development of anti- pruritus drugs.

KEYWORDS: citrusinine-II, Inhibition, Itch, Pain, TRPV3

Lancet. 2021 Oct 30;398(10311):1581-1592. doi: 10.1016/S0140-6736(21)01256-3.Epub 2021 Oct 28.

Efficacy and safety of maralixibat treatment in patients with Alagille syndrome and cholestatic pruritus (ICONIC): a randomised phase 2 study

Emmanuel Gonzales ¹, Winita Hardikar ², Michael Stormon ³, Alastair Baker ⁴, Loreto Hierro ⁵, Dorota Gliwicz ⁶, Florence Lacaille ⁷, Alain Lachaux ⁸, Ekkehard Sturm ⁹, Kenneth D R Setchell ¹⁰, Ciara Kennedy ¹¹, Alejandro Dorenbaum ¹², Jana Steinmetz ¹³, Nirav K Desai ¹⁴, Andrew J Wardle ¹⁵, Will Garner ¹⁵, Pamela Vig ¹⁵, Thomas Jaecklin ¹⁶, Etienne M Sokal ¹⁷, Emmanuel Jacquemin ¹⁸Affiliations expand

• PMID: 34755627
• DOI: 10.1016/S0140-6736(21)01256-3

Abstract

Background: Alagille syndrome is a rare genetic disease that often presents with severe cholestasis and pruritus. There are no approved drugs for management. Maralixibat, an apical, sodium-dependent, bile acid transport inhibitor, prevents enterohepatic bile acid recirculation. We evaluated the safety and efficacy of maralixibat for children with cholestasis in Alagille syndrome.

Methods: ICONIC was a placebo-controlled, randomised withdrawal period (RWD), phase 2b study with open-label extension in children (aged 1-18 years) with Alagille syndrome (NCT02160782). Eligible participants had more than three times the normal serum bile acid (sBA) levels and intractable pruritus. After 18 weeks of maralixibat 380 μg/kg once per day, participants were randomly assigned (1:1) to continue maralixibat or receive placebo for 4 weeks. Subsequently, all participants received open-label maralixibat until week 48. During the long-term extension (204 weeks reported), doses were increased up to 380 μg/kg twice per day. The primary endpoint was the mean sBA change during the RWD in participants with at least 50% sBA reduction by week 18. Cholestastic pruritus was assessed using observer-rated, patient-rated, and clinician-rated 0-4 scales. The safety population was defined as all participants who had received at least one dose of maralixibat. This trial was registered with ClinicalTrials.gov, NCT02160782, and is closed to recruitment.

Findings: Between Oct 28, 2014, and Aug 14, 2015, 31 participants (mean age 5·4 years [SD 4·25]) were enrolled and 28 analysed at week 48. Of the 29 participants who entered the randomised drug withdrawal period, ten (34%) were female and 19 (66%) were male. In the RWD, participants switched to placebo had significant increases in sBA (94 μmol/L, 95% CI 23 to 164) and pruritus (1·7 points, 95% CI 1·2 to 2·2), whereas participants who continued maralixibat maintained treatment effect. This study met the primary endpoint (least square mean difference -117 μmol/L, 95% CI -232 to -2). From baseline to week 48, sBA (-96 μmol/L, -162 to -31) and pruritus (-1·6 pts, -2·1 to -1·1) improved. In participants who continued to week 204 (n=15) all improvements were maintained. Maralixibat was generally safe and well tolerated throughout. The most frequent adverse events were gastrointestinal related. Most adverse events were self-limiting in nature and mild-to-moderate in severity.

Interpretation: In children with Alagille syndrome, maralixibat is, to our knowledge, the first agent to show durable and clinically meaningful improvements in cholestasis. Maralixibat might represent a new treatment paradigm for chronic cholestasis in Alagille syndrome.

Funding: Mirum Pharmaceuticals.

Copyright © 2021 Elsevier Ltd. All rights reserved.

presentation by Jong hyun

Type I hypersensitivity promotes Aedes aegypti blood feeding

Michael J. Conway, published in Scientific reports

Abstract:

Mosquitoes play a major role in human disease by serving as vectors of pathogenic microorganisms. Mosquitoes inject saliva into host skin during the probing process. Mosquito saliva contains a number of proteins that facilitate blood feeding by preventing hemostasis. Mosquito saliva also contains potent allergens that induce type I hypersensitivity reactions in some individuals. Type I hypersensitivity reactions in skin involve IgE-mediated degranulation of mast cells, which leads to vasodilation and an itch sensation. We hypothesized that hypersensitivity to mosquito saliva influences blood feeding. To test this hypothesis, we recruited human subjects who consented to Aedes aegypti bites. We measured their first sensation of itch, the strength of their itch sensation, the number of times mosquitoes attempted to feed, the number of times mosquitoes probed their skin, feeding time, engorgement status, and wheal diameter. Here we show that hypersensitive subjects had a stronger itch sensation, and that the time to first itch sensation was inversely correlated with wheal diameter; however, mosquitoes tended to probe less and engorge more on these subjects. Follow-up experiments testing the impact of oral antihistamine treatment on mosquito feeding parameters failed to reveal a statistically significant result. Histamine also failed to promote blood feeding on an artificial membrane feeder. This study suggests that mosquito saliva-induced type
I hypersensitivity promotes blood feeding but that this may be independent from histamine or histamine signaling.

IL-33 Signaling in Sensory Neurons Promotes Dry Skin Itch

Abstract

Background

Chronic pruritus, or itch, is common and debilitating, but the neuro-immune mechanisms that drive chronic itch are only starting to be elucidated. Recent studies demonstrate that the IL-33 receptor (IL-33R) is expressed by sensory neurons. However, whether sensory neuron-restricted activity of IL-33 is necessary for chronic itch remains poorly understood.

Objectives

We sought to determine if IL-33 signaling in sensory neurons is critical for the development of chronic itch in two divergent pruritic disease models.

Methods

Plasma levels of IL-33 were assessed in patients with atopic dermatitis (AD) and chronic pruritus of unknown origin (CPUO). Mice were generated to conditionally delete IL-33R from sensory neurons. The contribution of neuronal IL-33R signaling to chronic itch development was tested in mouse models that recapitulate key pathologic features of AD and CPUO, respectively.

Results

IL-33 was elevated in both AD and CPUO as well as their respective mouse models. While neuron-restricted IL-33R signaling was dispensable for itch in AD-like disease, it was required for the development of dry skin itch in a mouse model that mirrors key aspects of CPUO pathology.

Conclusion

These data highlight how IL-33 may be a predominant mediator of itch in certain contexts, depending on the tissue microenvironment. Further, this study provides insight for future therapeutic strategies targeting the IL-33 pathway for chronic itch.

Key Words

• Atopic dermatitis
• chronic pruritus of unknown origin
• dry skin
• IL-33
• itch
• neuroimmunology
• pruriceptor
• pruritogen

Abbreviations:

Ab ((antibody)), AD ((atopic dermatitis)), AEW ((acetone/ether plus water)), bp ((base pair)), Cap ((Capsaicin)), CPUO ((chronic pruritus of unknown origin)), CQ ((chloroquine)), DRG ((dorsal root ganglia)), DT ((diphtheria toxin)), epidermis ((Epi)), EtOH ((ethanol)), HC ((healthy control)), His ((histamine)), i.d ((intradermal)), ILC2s ((group 2 innate lymphoid cells)), IL-33R ((IL-33 receptor)), i.p ((intraperitoneal)), KCl ((potassium chloride)), LM ((littermate)), loxP ((locus of X-over P1)), mAb ((monoclonal antibody)), MACS ((magnetic-activated cell sorting)), MasTRECK ((mast cell-specific enhancer-mediated toxin receptor-mediated conditional cell knockout)), NS ((no significance)), rh ((recombinant human)), rm ((recombinant mouse)), RNA-seq ((RNA sequencing)), stratum corneum ((SC)), Veh ((Vehicle)), WT ((wild-type))

TRPC3 Antagonizes Pruritus in A Mouse Contact Dermatitis Model

Katherine Beattie , Haowu Jiang , Mayank Gautam , Mary K MacVittie , Barbara Miller , Minghong Ma , Qin Liu , Wenqin Luo 

Abstract

Contact dermatitis (CD), including allergic and irritant CD, are common dermatological diseases and characterized by an erythematous rash and severe itch. In this study, we investigated the function of TRPC3, a canonical TRP channel highly expressed in type 1 non-peptidergic (NP1) nociceptive primary afferents and other cell types, in a mouse CD model. Though TrpC3 null mice had little deficits in acute somatosensation, they showed significantly increased scratching with CD. In addition, TrpC3 null mice displayed no differences in mechanic and thermal hypersensitivity in an inflammatory pain model, suggesting that this channel preferentially functions to antagonize CD-induced itch. Using dorsal root ganglia (DRG) and pan-immune-specific TrpC3 conditional KO (CKO) mice, we determined that TrpC3 in DRG neurons, but not in immune cells, is required for this phenotype. Furthermore, the number of MRGPRD⁺ NP1 afferents in CD-affected DRGs is significantly reduced in TrpC3 mutant mice. Taken together, our results suggest that TrpC3 plays a critical role in NP1 afferents to cope with CD-induced excitotoxicity, and that degeneration of NP1 fibers may lead to an increased itch of CD. Our study identified a role of TrpC3 and NP1 afferents in CD pathology.

Keywords: Contact dermatitis; DRG; MRGPRD; TRPC3; non-peptidergic nociceptors; pruritus.

Reduced spontaneous itch in mouse models of cholestasis

Published in:- Scientific Reports (2021.03.17) IF~4.379

Jacqueline Langedijk, Ruth Bolier, Dagmar Tolenaars, Lysbeth ten Bloemendaal, Suzanne Duijst, Dirk de Waart, Ulrich Beuers, Piter Bosma & Ronald Oude Elferink

Abstract

Pruritus is one of the most distressing symptoms in cholestatic patients. Plasma autotaxin (ATX) activity correlates with the severity of pruritus in cholestatic patients, but the pathophysiology is unclear. To study pruritus in mice, we measured scratch activity in cholestatic Atp8b1 mutant mice, a model for Progressive Familial Intrahepatic Cholestasis type 1, and wild type mice (WT) with alphanaphthylisothiocyanate (ANIT)-induced cholestasis. To induce cholestasis, Atp8b1 mutant mice received a diet containing 0.1% cholic acid (CA) and WT mice were treated with ANIT. In these mice ATX was also overexpressed by transduction with AAV-ATX. Scratch activity was measured using an unbiased, electronic assay. Marked cholestasis was accomplished in both Atp8b1 mutant mice on a CA-supplemented diet and in ANIT-treatment in WT mice, but scratch activity was decreased rather than increased while plasma ATX activity was increased. Plasma ATX activity was further increased up to fivefold with AAV-ATX, but this did not induce scratch activity. In contrast to several reports two cholestatic mouse models did not display increased scratch activity as a measure of itch perception. Increasing plasma ATX activity by overexpression also did not lead to increased scratch activity in mice. This questions whether mice are suitable to study cholestatic itch.

TRPV1 and TRPA1 Channels Are Both Involved Downstream of Histamine-Induced Itch 

by Jenny Wilzopolski ^1,2,3,*, Manfred Kietzmann ¹, Santosh K. Mishra ², Holger Stark ⁴, Wolfgang Bäumer ^2,3 and Kristine Rossbach¹

¹Department of Pharmacology, Toxicology and Pharmacy, University of Veterinary Medicine Hannover, Foundation, 30559 Hannover, Germany

²Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27607, USA

³Department of Veterinary Medicine, Institute of Pharmacology and Toxicology, Freie Universität Berlin, 14195 Berlin, Germany

⁴Institute of Pharmaceutical and Medical Chemistry, Heinrich Heine University Düsseldorf, 40225 Duesseldorf, Germany

^*Author to whom correspondence should be addressed. Academic Editors: Emanuela Masini, Laura Lucarini and Alessandro AlaimoBiomolecules2021, 11(8), 1166; https://doi.org/10.3390/biom11081166Received: 13 July 2021 / Revised: 31 July 2021 / Accepted: 4 August 2021 / Published: 6 August 2021(This article belongs to the Special Issue New Developments in Histamine Research)

Abstract: Two histamine receptor subtypes (HR), namely H1R and H4R, are involved in the trans- mission of histamine-induced itch as key components. Although exact downstream signaling mechanisms are still elusive, transient receptor potential (TRP) ion channels play important roles in the sensation of histaminergic and non-histaminergic itch. The aim of this study was to investigate the involvement of TRPV1 and TRPA1 channels in the transmission of histaminergic itch. The potential of TRPV1 and TRPA1 inhibitors to modulate H1R- and H4R-induced signal transmission was tested in a scratching assay in mice in vivo as well as via Ca2+ imaging of murine sensory dorsal root ganglia (DRG) neurons in vitro. TRPV1 inhibition led to a reduction of H1R- and H4R- induced itch, whereas TRPA1 inhibition reduced H4R- but not H1R-induced itch. TRPV1 and TRPA1 inhibition resulted in a reduced Ca2+ influx into sensory neurons in vitro. In conclusion, these results indicate that both channels, TRPV1 and TRPA1, are involved in the transmission of histamine-induced pruritus.

Keywords: histamine; histamine H1 receptor; histamine H4 receptor; itch; signal transduction; TRPV1; TRPA1; dorsal root ganglion neurons (DRG); Ca2+-imaging

Dissecting the precise nature of itch-evoked scratching

Nivanthika K Wimalasena ¹, George Milner ², Ricardo Silva ³, Cliff Vuong ³, Zihe Zhang ¹, Diana M Bautista ⁴, Clifford J Woolf ⁵

Abstract

Itch is a discrete and irritating sensation tightly coupled to a drive to scratch. Acute scratching developed evolutionarily as an adaptive defense against skin irritants, pathogens, or parasites. In contrast, the itch-scratch cycle in chronic itch is harmful, inducing escalating itch and skin damage. Clinically and preclinically, scratching incidence is currently evaluated as a unidimensional motor parameter and believed to reflect itch severity. We propose that scratching, when appreciated as a complex, multidimensional motor behavior, will yield greater insight into the nature of itch and the organization of neural circuits driving repetitive motor patterns. We outline the limitations of standard measurements of scratching in rodent models and present new approaches to observe and quantify itch-evoked scratching. We argue that accurate quantitative measurements of scratching are critical for dissecting the molecular, cellular, and circuit mechanisms underlying itch and for preclinical development of therapeutic interventions for acute and chronic itch disorders.

Keywords: high-speed recording; itch; pruritis; rodent models; scratching behavior.

Statin-boosted cellular uptake of penetratin due to reduced membrane dipole
potential

published in bioRxiv, 2020.08.04

Gyula Batta, Levente Kárpáti, Gabriela Fulaneto Henrique, Szabolcs Tarapcsák, Tamás Kovács, Florina Zákány, István M. Mándity, Peter Nagy

Abstract

Since cell penetrating peptides are promising tools for delivery of cargo into cells, factors limiting or facilitating their cellular uptake are intensely studied. Using labeling with pH-insensitive and pH-sensitive dyes we report that escape of penetratin from acidic endo-lysosomal compartments is retarded compared to its cellular uptake. The membrane dipole potential, known to alter transmembrane transport of charged molecules, is shown to be negatively correlated with the concentration of penetratin in the cytoplasmic compartment. Treatment of cells with therapeutically relevant concentrations of atorvastatin, an inhibitor of HMG-CoA reductase and cholesterol synthesis, significantly increased the release of penetratin from acidic endocytic compartments in two different cell types. This effect of atorvastatin correlated with its ability to decrease the membrane dipole potential. These results highlight the importance of the dipole potential in regulating cellular uptake of cell penetrating peptides and suggest a clinically relevant way of boosting this process.

Identification of the dog orthologue of human MAS-related G protein coupled receptor X2 (MRGPRX2) essential for drug-induced pseudo-allergic reactions

Published in scientifc reports, 30 september 2020

Eri Hamamura-Yasuno, Takuma Iguchi, Kazuyoshi Kumagai, Yoshimi tsuchiya&Kazuhiko Mori

Abstarct

MAS-related G protein coupled receptor-X2 (MRGPRX2), expressed in human mast cells, is associated with drug-induced pseudo-allergic reactions. Dogs are highly susceptible to drug-induced anaphylactoid reactions caused by various drugs; however, the distribution and physiological function of canine MRGPR family genes, including MRGPRX2, remain largely unknown. In the present study, we clarified the distribution of dog MRGPR family genes by real-time quantitative PCR and in situ hybridisation. We also investigated the stimulatory effects of various histamine-releasing agents, including fluoroquinolones, on HEK293 cells transiently transfected with dog MRGPR family genes to identify their physiological function. Dog MRGPRX2 and MRGPRG were distributed in a limited number of tissues, including the skin (from the eyelid, abdomen, and cheek), whereas MRGPRD and MRGPRF were extensively expressed in almost all tissues examined. Histochemical and in situ hybridisation analyses revealed that MRGPRX2 was expressed in dog connective tissue-type mast cells in the skin. Intracellular Ca²⁺ mobilisation assay revealed that HEK293 cells, expressing dog MRGPRX2 or human MRGPRX2, but not dog MRGPRD, MRGPRF, and MRGPRG, responded to histamine-releasing agents. Our results suggest that dog MRGPRX2 is the functional orthologue of human MRGPRX2 and plays an essential role in drug-induced anaphylactoid reactions in dogs.

Presented by Jong hyun Im