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Identification of a mast-cell-specific receptor crucial for pseudo-allergic drug reactions.

McNeil BD, Pundir P, Meeker S, Han L, Undem BJ, Kulka M, Dong X - Nature (2014)

Bottom Line: Although they are classically activated by immunoglobulin (Ig)E antibodies, a unique property of mast cells is their antibody-independent responsiveness to a range of cationic substances, collectively called basic secretagogues, including inflammatory peptides and drugs associated with allergic-type reactions.The pathogenic roles of these substances have prompted a decades-long search for their receptor(s).Finally, we determine that Mrgprb2 and MRGPRX2 are targets of many small-molecule drugs associated with systemic pseudo-allergic, or anaphylactoid, reactions; we show that drug-induced symptoms of anaphylactoid responses are significantly reduced in knockout mice; and we identify a common chemical motif in several of these molecules that may help predict side effects of other compounds.

View Article: PubMed Central - PubMed

Affiliation: The Solomon H. Snyder Department of Neuroscience, Department of Neurosurgery, Center for Sensory Biology, Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA.

ABSTRACT
Mast cells are primary effectors in allergic reactions, and may have important roles in disease by secreting histamine and various inflammatory and immunomodulatory substances. Although they are classically activated by immunoglobulin (Ig)E antibodies, a unique property of mast cells is their antibody-independent responsiveness to a range of cationic substances, collectively called basic secretagogues, including inflammatory peptides and drugs associated with allergic-type reactions. The pathogenic roles of these substances have prompted a decades-long search for their receptor(s). Here we report that basic secretagogues activate mouse mast cells in vitro and in vivo through a single receptor, Mrgprb2, the orthologue of the human G-protein-coupled receptor MRGPRX2. Secretagogue-induced histamine release, inflammation and airway contraction are abolished in Mrgprb2- mutant mice. Furthermore, we show that most classes of US Food and Drug Administration (FDA)-approved peptidergic drugs associated with allergic-type injection-site reactions also activate Mrgprb2 and MRGPRX2, and that injection-site inflammation is absent in mutant mice. Finally, we determine that Mrgprb2 and MRGPRX2 are targets of many small-molecule drugs associated with systemic pseudo-allergic, or anaphylactoid, reactions; we show that drug-induced symptoms of anaphylactoid responses are significantly reduced in knockout mice; and we identify a common chemical motif in several of these molecules that may help predict side effects of other compounds. These discoveries introduce a mouse model to study mast cell activation by basic secretagogues and identify MRGPRX2 as a potential therapeutic target to reduce a subset of drug-induced adverse effects.

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Human mast cells are activated by basic secretagogues and drugs associated with pseudo-allergic reactions in an MrgprX2-dependent manner.a. Human LAD2 mast cells were treated with different concentrations of compound 48/80, mastoparan, icatibant, atracurium, and ciprofloxacin. The activation of mast cells in response to these substances was characterized by the release of β-hexosaminidase, TNF, PGD2, and histamine. In addition, 0.1 μg/ml streptavidin stimulation of biotin-conjugated human IgE sensitized LAD2 cells caused a robust release of β-hexosaminidase (71.3±1.8% release), compared to untreated cells (4.1±0.3% release). Group data are expressed as mean ± standard error of the mean.b. Knockdown of human MrgprX2 significantly reduced mast cell activation evoked by basic secretagogues and drugs associated with pseudo-allergic reactions, but not by IgE. Human LAD2 mast cells were first transfected with MrgprX2 siRNA or control siRNA. Two days after the transfection, the cells were treated with compound 48/80 (0.1 μg/ml), mastoparan (5 μg/ml), icatibant (10 μg/ml), atracurium (25 μg/ml), and ciprofloxacin (75 μg/ml). The activation of mast cells in response to these substances characterized by the release of β-hexosaminidase was significantly reduced in MrgprX2 siRNA treated cells, compared to release in the control group. IgE-mediated mast cell degranulation was unaffected by MrgprX2 siRNA knockdown. Group data are expressed as mean ± standard error of the mean. Two-tailed unpaired Student's t test was used to determine significance in statistical comparisons, and differences were considered significant at * p < 0.05; ** p<0.01; *** p< 0.005 (the experiments were repeated three times).
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Figure 14: Human mast cells are activated by basic secretagogues and drugs associated with pseudo-allergic reactions in an MrgprX2-dependent manner.a. Human LAD2 mast cells were treated with different concentrations of compound 48/80, mastoparan, icatibant, atracurium, and ciprofloxacin. The activation of mast cells in response to these substances was characterized by the release of β-hexosaminidase, TNF, PGD2, and histamine. In addition, 0.1 μg/ml streptavidin stimulation of biotin-conjugated human IgE sensitized LAD2 cells caused a robust release of β-hexosaminidase (71.3±1.8% release), compared to untreated cells (4.1±0.3% release). Group data are expressed as mean ± standard error of the mean.b. Knockdown of human MrgprX2 significantly reduced mast cell activation evoked by basic secretagogues and drugs associated with pseudo-allergic reactions, but not by IgE. Human LAD2 mast cells were first transfected with MrgprX2 siRNA or control siRNA. Two days after the transfection, the cells were treated with compound 48/80 (0.1 μg/ml), mastoparan (5 μg/ml), icatibant (10 μg/ml), atracurium (25 μg/ml), and ciprofloxacin (75 μg/ml). The activation of mast cells in response to these substances characterized by the release of β-hexosaminidase was significantly reduced in MrgprX2 siRNA treated cells, compared to release in the control group. IgE-mediated mast cell degranulation was unaffected by MrgprX2 siRNA knockdown. Group data are expressed as mean ± standard error of the mean. Two-tailed unpaired Student's t test was used to determine significance in statistical comparisons, and differences were considered significant at * p < 0.05; ** p<0.01; *** p< 0.005 (the experiments were repeated three times).

Mentions: Finally, we determined whether drugs associated with pseudo-allergies activate human mast cells through MrgprX2. We found that representative members of each examined drug class evoked release of histamine, TNF, PGD2, and β-hexosaminidase from LAD2 cells (Extended Data Fig. 10a). 48/80 and mastoparan were used as positive controls. Importantly, MrgprX2 siRNA-treated LAD2 cells exhibited significantly less β-hexosaminidase release evoked by these substances, compared to responses in control siRNA-treated cells, while IgE-mediated release was comparable (Extended Data Fig, 10b). The remaining release observed in MrgprX2 siRNA-treated cells are likely due to incomplete mRNA and/or protein knockdown.


Identification of a mast-cell-specific receptor crucial for pseudo-allergic drug reactions.

McNeil BD, Pundir P, Meeker S, Han L, Undem BJ, Kulka M, Dong X - Nature (2014)

Human mast cells are activated by basic secretagogues and drugs associated with pseudo-allergic reactions in an MrgprX2-dependent manner.a. Human LAD2 mast cells were treated with different concentrations of compound 48/80, mastoparan, icatibant, atracurium, and ciprofloxacin. The activation of mast cells in response to these substances was characterized by the release of β-hexosaminidase, TNF, PGD2, and histamine. In addition, 0.1 μg/ml streptavidin stimulation of biotin-conjugated human IgE sensitized LAD2 cells caused a robust release of β-hexosaminidase (71.3±1.8% release), compared to untreated cells (4.1±0.3% release). Group data are expressed as mean ± standard error of the mean.b. Knockdown of human MrgprX2 significantly reduced mast cell activation evoked by basic secretagogues and drugs associated with pseudo-allergic reactions, but not by IgE. Human LAD2 mast cells were first transfected with MrgprX2 siRNA or control siRNA. Two days after the transfection, the cells were treated with compound 48/80 (0.1 μg/ml), mastoparan (5 μg/ml), icatibant (10 μg/ml), atracurium (25 μg/ml), and ciprofloxacin (75 μg/ml). The activation of mast cells in response to these substances characterized by the release of β-hexosaminidase was significantly reduced in MrgprX2 siRNA treated cells, compared to release in the control group. IgE-mediated mast cell degranulation was unaffected by MrgprX2 siRNA knockdown. Group data are expressed as mean ± standard error of the mean. Two-tailed unpaired Student's t test was used to determine significance in statistical comparisons, and differences were considered significant at * p < 0.05; ** p<0.01; *** p< 0.005 (the experiments were repeated three times).
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Figure 14: Human mast cells are activated by basic secretagogues and drugs associated with pseudo-allergic reactions in an MrgprX2-dependent manner.a. Human LAD2 mast cells were treated with different concentrations of compound 48/80, mastoparan, icatibant, atracurium, and ciprofloxacin. The activation of mast cells in response to these substances was characterized by the release of β-hexosaminidase, TNF, PGD2, and histamine. In addition, 0.1 μg/ml streptavidin stimulation of biotin-conjugated human IgE sensitized LAD2 cells caused a robust release of β-hexosaminidase (71.3±1.8% release), compared to untreated cells (4.1±0.3% release). Group data are expressed as mean ± standard error of the mean.b. Knockdown of human MrgprX2 significantly reduced mast cell activation evoked by basic secretagogues and drugs associated with pseudo-allergic reactions, but not by IgE. Human LAD2 mast cells were first transfected with MrgprX2 siRNA or control siRNA. Two days after the transfection, the cells were treated with compound 48/80 (0.1 μg/ml), mastoparan (5 μg/ml), icatibant (10 μg/ml), atracurium (25 μg/ml), and ciprofloxacin (75 μg/ml). The activation of mast cells in response to these substances characterized by the release of β-hexosaminidase was significantly reduced in MrgprX2 siRNA treated cells, compared to release in the control group. IgE-mediated mast cell degranulation was unaffected by MrgprX2 siRNA knockdown. Group data are expressed as mean ± standard error of the mean. Two-tailed unpaired Student's t test was used to determine significance in statistical comparisons, and differences were considered significant at * p < 0.05; ** p<0.01; *** p< 0.005 (the experiments were repeated three times).
Mentions: Finally, we determined whether drugs associated with pseudo-allergies activate human mast cells through MrgprX2. We found that representative members of each examined drug class evoked release of histamine, TNF, PGD2, and β-hexosaminidase from LAD2 cells (Extended Data Fig. 10a). 48/80 and mastoparan were used as positive controls. Importantly, MrgprX2 siRNA-treated LAD2 cells exhibited significantly less β-hexosaminidase release evoked by these substances, compared to responses in control siRNA-treated cells, while IgE-mediated release was comparable (Extended Data Fig, 10b). The remaining release observed in MrgprX2 siRNA-treated cells are likely due to incomplete mRNA and/or protein knockdown.

Bottom Line: Although they are classically activated by immunoglobulin (Ig)E antibodies, a unique property of mast cells is their antibody-independent responsiveness to a range of cationic substances, collectively called basic secretagogues, including inflammatory peptides and drugs associated with allergic-type reactions.The pathogenic roles of these substances have prompted a decades-long search for their receptor(s).Finally, we determine that Mrgprb2 and MRGPRX2 are targets of many small-molecule drugs associated with systemic pseudo-allergic, or anaphylactoid, reactions; we show that drug-induced symptoms of anaphylactoid responses are significantly reduced in knockout mice; and we identify a common chemical motif in several of these molecules that may help predict side effects of other compounds.

View Article: PubMed Central - PubMed

Affiliation: The Solomon H. Snyder Department of Neuroscience, Department of Neurosurgery, Center for Sensory Biology, Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205, USA.

ABSTRACT
Mast cells are primary effectors in allergic reactions, and may have important roles in disease by secreting histamine and various inflammatory and immunomodulatory substances. Although they are classically activated by immunoglobulin (Ig)E antibodies, a unique property of mast cells is their antibody-independent responsiveness to a range of cationic substances, collectively called basic secretagogues, including inflammatory peptides and drugs associated with allergic-type reactions. The pathogenic roles of these substances have prompted a decades-long search for their receptor(s). Here we report that basic secretagogues activate mouse mast cells in vitro and in vivo through a single receptor, Mrgprb2, the orthologue of the human G-protein-coupled receptor MRGPRX2. Secretagogue-induced histamine release, inflammation and airway contraction are abolished in Mrgprb2- mutant mice. Furthermore, we show that most classes of US Food and Drug Administration (FDA)-approved peptidergic drugs associated with allergic-type injection-site reactions also activate Mrgprb2 and MRGPRX2, and that injection-site inflammation is absent in mutant mice. Finally, we determine that Mrgprb2 and MRGPRX2 are targets of many small-molecule drugs associated with systemic pseudo-allergic, or anaphylactoid, reactions; we show that drug-induced symptoms of anaphylactoid responses are significantly reduced in knockout mice; and we identify a common chemical motif in several of these molecules that may help predict side effects of other compounds. These discoveries introduce a mouse model to study mast cell activation by basic secretagogues and identify MRGPRX2 as a potential therapeutic target to reduce a subset of drug-induced adverse effects.

Show MeSH
Related in: MedlinePlus