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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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MrgprB2 mediates mast cell responsiveness and side effects of peptidergic therapeutic drugsa. Percentage of responding cells from WT and MrgprB2MUT peritoneal mast cells after drug application, assayed using Fluo-4 imaging. Concentrations of drugs (in μg/ml): icatibant, 50; cetrorelix, 20; leuprolide, 100; octreotide, 10; sermorelin, 60; insulin, 80. n=3/genotype; >150 cells counted/substance, except >100 cells counted for insulin. Difference between insulin responsiveness was not significant.b. Left, representative images of Evans Blue extravasation 15 minutes after intraplantar injection of icatibant (right, arrow, 10 mg/ml, 5 μl in saline) or saline (left). Right, quantification of Evans Blue leakage into the paw after 15 minutes. n=6/genotype. Difference after saline injection was not significant.c. Total histamine release from WT (red diamonds) and MrgprB2MUT (black squares) mice after incubation with named substances. Note: no significant difference between WT and MrgprB2MUT cells was found at any dose of anti-IgE antibody. Experiments were repeated >3 times.Data are presented as mean ± SEM. Two-tailed unpaired Student's t test: *, p < 0.05. **, p<0.01.
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Figure 3: MrgprB2 mediates mast cell responsiveness and side effects of peptidergic therapeutic drugsa. Percentage of responding cells from WT and MrgprB2MUT peritoneal mast cells after drug application, assayed using Fluo-4 imaging. Concentrations of drugs (in μg/ml): icatibant, 50; cetrorelix, 20; leuprolide, 100; octreotide, 10; sermorelin, 60; insulin, 80. n=3/genotype; >150 cells counted/substance, except >100 cells counted for insulin. Difference between insulin responsiveness was not significant.b. Left, representative images of Evans Blue extravasation 15 minutes after intraplantar injection of icatibant (right, arrow, 10 mg/ml, 5 μl in saline) or saline (left). Right, quantification of Evans Blue leakage into the paw after 15 minutes. n=6/genotype. Difference after saline injection was not significant.c. Total histamine release from WT (red diamonds) and MrgprB2MUT (black squares) mice after incubation with named substances. Note: no significant difference between WT and MrgprB2MUT cells was found at any dose of anti-IgE antibody. Experiments were repeated >3 times.Data are presented as mean ± SEM. Two-tailed unpaired Student's t test: *, p < 0.05. **, p<0.01.

Mentions: We next considered whether MrgprB2 factors in allergic-type reactions. We specifically addressed drug-induced reactions because many therapeutic drugs are cationic. Up to 15% of drug-induced adverse reactions appear to be allergic in nature; however, many are not well-correlated with IgE antibody titer, indicating that antibody-independent, or pseudo-allergic, mechanisms participate18. We focused first on peptidergic drugs because most are introduced subcutaneously or intramuscularly at millimolar concentrations (Supplementary Information), high enough for cationic peptides to activate mast cells. The most frequent allergic-type response described in FDA labels of these drugs is an injection-site reaction (ISR), a local swelling and/or flare of variable size which can be accompanied by pain or pruritus. In a survey of FDA-approved peptidergic drugs, we found that the vast majority associated with ISRs are cationic (Supplementary Information). We found that representative members of all common, commercially available classes of these cationic drugs activated mast cells in an MrgprB2-dependent manner, while the innocuous protein insulin had no effect (Fig. 3a; Extended Data Fig. 9b,c). Consistently, all of these peptides except insulin activate both MrgprB2-HEK and MrgprX2-HEK cells (Extended Data Fig. 2). We selected the drug icatibant for further study because it induces ISRs nearly in every patient19. Icatibant at the clinical concentration induced extensive extravasation and swelling, similar to human ISRs, in WT mice but not in MrgprB2MUT mice (Fig. 3b). Mice pretreated with the mast cell stabilizer ketotifen also showed no inflammation (without ketotifen: 40.7±2.1% increase in paw thickness; with ketotifen: 3.1±0.6% increase; n=4 each; p=2.2e-6), strongly indicating that mast cells mediated the inflammation. Furthermore, icatibant (as well as positive controls 48/80 and mastoparan) induced histamine release from WT peritoneal mast cells, while MrgprB2MUT mast cells released substantially less (Fig. 3c). However, IgE-mediated histamine release was unaffected by MrgprB2 deletion (Fig. 3c). These data anticipate that drug-induced ISRs may be alleviated by targeting MrgprX2 or by using peptides with less potent MrgprX2 agonist properties.


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)

MrgprB2 mediates mast cell responsiveness and side effects of peptidergic therapeutic drugsa. Percentage of responding cells from WT and MrgprB2MUT peritoneal mast cells after drug application, assayed using Fluo-4 imaging. Concentrations of drugs (in μg/ml): icatibant, 50; cetrorelix, 20; leuprolide, 100; octreotide, 10; sermorelin, 60; insulin, 80. n=3/genotype; >150 cells counted/substance, except >100 cells counted for insulin. Difference between insulin responsiveness was not significant.b. Left, representative images of Evans Blue extravasation 15 minutes after intraplantar injection of icatibant (right, arrow, 10 mg/ml, 5 μl in saline) or saline (left). Right, quantification of Evans Blue leakage into the paw after 15 minutes. n=6/genotype. Difference after saline injection was not significant.c. Total histamine release from WT (red diamonds) and MrgprB2MUT (black squares) mice after incubation with named substances. Note: no significant difference between WT and MrgprB2MUT cells was found at any dose of anti-IgE antibody. Experiments were repeated >3 times.Data are presented as mean ± SEM. Two-tailed unpaired Student's t test: *, p < 0.05. **, p<0.01.
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Figure 3: MrgprB2 mediates mast cell responsiveness and side effects of peptidergic therapeutic drugsa. Percentage of responding cells from WT and MrgprB2MUT peritoneal mast cells after drug application, assayed using Fluo-4 imaging. Concentrations of drugs (in μg/ml): icatibant, 50; cetrorelix, 20; leuprolide, 100; octreotide, 10; sermorelin, 60; insulin, 80. n=3/genotype; >150 cells counted/substance, except >100 cells counted for insulin. Difference between insulin responsiveness was not significant.b. Left, representative images of Evans Blue extravasation 15 minutes after intraplantar injection of icatibant (right, arrow, 10 mg/ml, 5 μl in saline) or saline (left). Right, quantification of Evans Blue leakage into the paw after 15 minutes. n=6/genotype. Difference after saline injection was not significant.c. Total histamine release from WT (red diamonds) and MrgprB2MUT (black squares) mice after incubation with named substances. Note: no significant difference between WT and MrgprB2MUT cells was found at any dose of anti-IgE antibody. Experiments were repeated >3 times.Data are presented as mean ± SEM. Two-tailed unpaired Student's t test: *, p < 0.05. **, p<0.01.
Mentions: We next considered whether MrgprB2 factors in allergic-type reactions. We specifically addressed drug-induced reactions because many therapeutic drugs are cationic. Up to 15% of drug-induced adverse reactions appear to be allergic in nature; however, many are not well-correlated with IgE antibody titer, indicating that antibody-independent, or pseudo-allergic, mechanisms participate18. We focused first on peptidergic drugs because most are introduced subcutaneously or intramuscularly at millimolar concentrations (Supplementary Information), high enough for cationic peptides to activate mast cells. The most frequent allergic-type response described in FDA labels of these drugs is an injection-site reaction (ISR), a local swelling and/or flare of variable size which can be accompanied by pain or pruritus. In a survey of FDA-approved peptidergic drugs, we found that the vast majority associated with ISRs are cationic (Supplementary Information). We found that representative members of all common, commercially available classes of these cationic drugs activated mast cells in an MrgprB2-dependent manner, while the innocuous protein insulin had no effect (Fig. 3a; Extended Data Fig. 9b,c). Consistently, all of these peptides except insulin activate both MrgprB2-HEK and MrgprX2-HEK cells (Extended Data Fig. 2). We selected the drug icatibant for further study because it induces ISRs nearly in every patient19. Icatibant at the clinical concentration induced extensive extravasation and swelling, similar to human ISRs, in WT mice but not in MrgprB2MUT mice (Fig. 3b). Mice pretreated with the mast cell stabilizer ketotifen also showed no inflammation (without ketotifen: 40.7±2.1% increase in paw thickness; with ketotifen: 3.1±0.6% increase; n=4 each; p=2.2e-6), strongly indicating that mast cells mediated the inflammation. Furthermore, icatibant (as well as positive controls 48/80 and mastoparan) induced histamine release from WT peritoneal mast cells, while MrgprB2MUT mast cells released substantially less (Fig. 3c). However, IgE-mediated histamine release was unaffected by MrgprB2 deletion (Fig. 3c). These data anticipate that drug-induced ISRs may be alleviated by targeting MrgprX2 or by using peptides with less potent MrgprX2 agonist properties.

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