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Intranasal treatment with CpG oligonucleotides provides protection from anaphylaxis in a murine model of peanut allergy
$206
50
Abstracts
IntranasalTreatmentwith CpG OligonucleotidesProvidesProtection from Anaphylaxisin a Murine Model of PeanutAllergy
J. D. Kattan, K. D. Srivastava, X. Li, H. A. Sampson; Pediatrics, Mount Sinai School of Medicine, New York, NY. RATIONALE: Little is known about the use of CpG oligonucleotides as a treatment for food allergy. We investigated the use of CpG for the treatment of peanut (PN) allergy in a murine model of peanut-induced anaphylaxis. METHODS: C3H/HeJ mice were orally sensitized with peanut and cholera toxin over seven weeks. Mice were subsequently treated with CpG intranasally weekly for three weeks or left untreated. One week following the last treatment, mice were given oral PN challenge. Following challenge, symptom scores and core body temperatures were evaluated, and serum immunoglobulins were determined by ELISA. Cytokine production by cultured splenocytes was analyzed by ELISA. RESULTS: CpG-treated mice were completely protected from anaphylactic reactions in response to PN challenge, whereas 67% of the untreated mice displayed symptoms of anaphylaxis. Body temperatures following challenge in CpG-treated mice (35.15+0.6~ were higher than in the untreated group (32.3+2.0~ A reduction in PN-specific IgE levels was seen in CpG-treated mice (1553-+ 156ng/ml) as compared to the untreated mice (1738-+290ng/ml). PN-specific lgG2a levels in CpG-treated mice (234.5-+90.3ug/ml) were more than three fold higher than in untreated mice (68.4+_4. lug/ml). No significant difference was observed in PN-specific IgG1 levels. A reduction in IL-5 levels in splenocyte supernatants from CpG-treated mice was detected (145.5+-43.1pg/ml) as compared to the untreated mice (293.5+85.6pg/ml). CONCLUSIONS: Intranasal CpG treatment was shown to protect PN allergic-mice from anaphylactic reactions to oral PN challenge suggesting that CpG may be a potential therapeutic approach for peanut allergy in humans. Funding: Dugan Family Foundation
551
Allergen Immunotborapywith Heat-killed Listeria Monocytogenes(HKL)AbrogatesPeanutSensitivity/Anaphylaxisin Dogs
O. L. Frick< S. S. Teuber 2, B. B. Buchanan 3, S. Morigasaki 3, D. T. Umetsu4; IUniversity of California, San Francisco, CA, 2University of California, Davis, CA, 3University of California, Berkeley, CA, 4Stanford University, Stanford, CA. RATIONALE: HKL potently stimulates IFN-yproduction in CD4 and CD8 T cells, and when used as adjuvant for immunotherapy, reduces lgE production and asthma in a murine model. We asked if such treatment could decrease on-going peanut induced anaphylactic responses in highly food allergic dogs. METHODS: Four atopic colony dogs highly sensitive to peanut developed anaphylaxis or vomiting upon oral exposure to peanut. The 4 dogs had skin test end-point titration to peanut of 10~ to 1(~6. Peanut extract ( h i 0 0 w/v Miles) 0.1 ml mixed with HKL (109 bacteria) was emulsified in Incomplete Freunds Adjuvant (IFA) and injected subcutaneously once. Skin test endToint was repeated at intervals for 20 weeks after immunization. IgE antibody-immunoblots were done with pre and post-immunization sera from peanut-sensitive dogs. Four months post-treatment each dog was serially challenged orally with 0.020 to 20 g peanut. RESULTS: in all 4 dogs, skin test end-points to peanut improved by 10 to 10,000-fold over the 5 month course. IgE-immunoblots of preimmunization sera from the peanut-sensitive dogs showed significant Ara hi binding, similar to that of sera from peanut allergic humans, but 3 months after immunization, Ara hl binding almost disappeared in each of the dog sera. Upon oral peanut challenge, one dog vomited after 4 g while 3 dogs had no signs after 20 g. CONCLUSIONS: hnmunotherapy with one dose of peanut and HKL as adjuvant in 1FA abrogated peanut anaphylaxis in highly atopic dogs. Therefore, immunotherapy with HKL-peanut may provide effective therapy for peanut allergy in humans. Funding: Children's Health
J ALLERGY CLIN IMMUNOL FEBRUARY 2003
552 Systemic Occult Bone Marrow Mastocytosis Presenting as Recurrent Anaphylaxis C. Akin, D. D. Metcalfe; Laboratory of Allergic Diseases, NIAID, NIH, Bethesda, MD. Unexplained episodes of recurrent anaphylaxis can be variably observed in patients with mastocytosis. Clinical characteristics of patients with mastocytosis who experience recurrent anaphylaxis, however, have not been analyzed. We have thus studied thirty consecutive patients evaluated at the NIH Clinical Center with a diagnosis of systemic mastocytosis. Thirty percent of the patients (9 total: 6 males, 3 females) had a history of recurrent unexplained anaphylaxis. Seven of 9 patients (78%) with anaphylaxis lacked any cutaneous lesions of mastocytosis. This finding was significantly different than the group without anaphylaxis where only 4 of 21 (20%) lacked skin lesions (p<0.005, Fisher's exact probability). Furthermore, patients with anaphylaxis had significantly less bone marrow involvement with mast cells. Thus, six of 9 (67%) of the patients with anaphylaxis lacked the characteristic compact mast cell aggregates in bone marrow biopsy as opposed to only 3 of 15 (20%) patients without anaphylaxis (p<0.02). Five patients with anaphylaxis had neither skin lesions nor characteristic infiltrates of mast cells in bone marrow biopsy and had baseline serum tryptase levels of less than 20 ng/ml. These patients, however, had other diagnostic features of disease such as aberrant mast ceil morphology, expression of CD2 and CD25 by mast cells, and the presence of codon 816 c-kit mutations. Such patients are termed to have "occult bone marrow mastocytosis" which is detectable by sensitive diagnostic approaches. In conclusion, systemic mastocytosis may be an underlying pathology in unexplained recurrent anaphylaxis in more patients than previously recognized. Funding: NIAID Intramural Funds
553 ,." AcetylhydrolaseDeficiencyPredisposesto FatalAnaphyP. Vadas 1, M. Gold 2, G. Liss 3, C. Smith 2, ]. Yeung4, B. Perelman-~; IMedicine, St. Michael's Hospital, University of Toronto, Toronto, ON, CANADA, 2Hospital for Sick Children, Toronto, ON, CANADA, 3Government of Ontario, Ministry of Labour, Toronto, ON, CANADA, 4National Food Processors Association, Washington, DC, 5St. Michael's Hospital, University of Toronto, Toronto, ON, CANADA. Platelet activating factor (PAF) is a central mediator of anaphylaxis in animal models. Manifestations of anaphylaxis are reproduced by exogenous PAF; PAF receptor antagonists protect against fatal anaphylaxis and PAF receptor knock-out mice show diminished anaphylactic responses. We examined the relationship between severity of anaphylaxis and levels of PAF acetylhydrolase (PAF-AH), the enzyme that inactivates PAE PAFAH levels were measured in sera of individuals who had died of peanut anaphylactic reactions and compared to control groups. Results were: fatal peanut anaphylaxis (n=8, PAF-AH 15.7 + 2.6 nmol/min/ml); healthy adults (n=26, PAF-AH 35.2 -- I 1.1 ), healthy children (n=26, PAF-AH 27.7 + 8.5); peanut allergic children (n=63, PAF-AH 24.9 +_5.8): non-fatal anaphylaxis (n=24, PAF-AH 29.6 -+ 9.0): non-anaphylactic fatalities (n=lO, PAF-AH 26.4 -+ 7.1). The differences between mean serum PAFAH activity in the fatal peanut anaphylaxis group and the control groups were highly significant (p=0.0003). whereas serum PAF-AH levels were comparable in all control groups. Analysis of genomic DNA for 3 patients with the lowest serum PAF-AH activities revealed no mutations in the coding exons of the PAF-AH gene. PAF-AH circulates as a complex with low density lipoproteins and we found a linear correlation between PAFAH activity and apolipoprotein B concentrations in patients with fatal and non-fatal anaphylactic reactions to peanut (r=0.614, p<0.0001). These data clearly show that mean serum PAF-AH activity was significantly lower in patients with fatal peanut anaphylaxis as compared to other groups, including healthy controls. Low density lipoprotein levels may be the major determinant of serum PAF-AH activity. Funding: Peanut Foundation
50
Abstracts
IntranasalTreatmentwith CpG OligonucleotidesProvidesProtection from Anaphylaxisin a Murine Model of PeanutAllergy
J. D. Kattan, K. D. Srivastava, X. Li, H. A. Sampson; Pediatrics, Mount Sinai School of Medicine, New York, NY. RATIONALE: Little is known about the use of CpG oligonucleotides as a treatment for food allergy. We investigated the use of CpG for the treatment of peanut (PN) allergy in a murine model of peanut-induced anaphylaxis. METHODS: C3H/HeJ mice were orally sensitized with peanut and cholera toxin over seven weeks. Mice were subsequently treated with CpG intranasally weekly for three weeks or left untreated. One week following the last treatment, mice were given oral PN challenge. Following challenge, symptom scores and core body temperatures were evaluated, and serum immunoglobulins were determined by ELISA. Cytokine production by cultured splenocytes was analyzed by ELISA. RESULTS: CpG-treated mice were completely protected from anaphylactic reactions in response to PN challenge, whereas 67% of the untreated mice displayed symptoms of anaphylaxis. Body temperatures following challenge in CpG-treated mice (35.15+0.6~ were higher than in the untreated group (32.3+2.0~ A reduction in PN-specific IgE levels was seen in CpG-treated mice (1553-+ 156ng/ml) as compared to the untreated mice (1738-+290ng/ml). PN-specific lgG2a levels in CpG-treated mice (234.5-+90.3ug/ml) were more than three fold higher than in untreated mice (68.4+_4. lug/ml). No significant difference was observed in PN-specific IgG1 levels. A reduction in IL-5 levels in splenocyte supernatants from CpG-treated mice was detected (145.5+-43.1pg/ml) as compared to the untreated mice (293.5+85.6pg/ml). CONCLUSIONS: Intranasal CpG treatment was shown to protect PN allergic-mice from anaphylactic reactions to oral PN challenge suggesting that CpG may be a potential therapeutic approach for peanut allergy in humans. Funding: Dugan Family Foundation
551
Allergen Immunotborapywith Heat-killed Listeria Monocytogenes(HKL)AbrogatesPeanutSensitivity/Anaphylaxisin Dogs
O. L. Frick< S. S. Teuber 2, B. B. Buchanan 3, S. Morigasaki 3, D. T. Umetsu4; IUniversity of California, San Francisco, CA, 2University of California, Davis, CA, 3University of California, Berkeley, CA, 4Stanford University, Stanford, CA. RATIONALE: HKL potently stimulates IFN-yproduction in CD4 and CD8 T cells, and when used as adjuvant for immunotherapy, reduces lgE production and asthma in a murine model. We asked if such treatment could decrease on-going peanut induced anaphylactic responses in highly food allergic dogs. METHODS: Four atopic colony dogs highly sensitive to peanut developed anaphylaxis or vomiting upon oral exposure to peanut. The 4 dogs had skin test end-point titration to peanut of 10~ to 1(~6. Peanut extract ( h i 0 0 w/v Miles) 0.1 ml mixed with HKL (109 bacteria) was emulsified in Incomplete Freunds Adjuvant (IFA) and injected subcutaneously once. Skin test endToint was repeated at intervals for 20 weeks after immunization. IgE antibody-immunoblots were done with pre and post-immunization sera from peanut-sensitive dogs. Four months post-treatment each dog was serially challenged orally with 0.020 to 20 g peanut. RESULTS: in all 4 dogs, skin test end-points to peanut improved by 10 to 10,000-fold over the 5 month course. IgE-immunoblots of preimmunization sera from the peanut-sensitive dogs showed significant Ara hi binding, similar to that of sera from peanut allergic humans, but 3 months after immunization, Ara hl binding almost disappeared in each of the dog sera. Upon oral peanut challenge, one dog vomited after 4 g while 3 dogs had no signs after 20 g. CONCLUSIONS: hnmunotherapy with one dose of peanut and HKL as adjuvant in 1FA abrogated peanut anaphylaxis in highly atopic dogs. Therefore, immunotherapy with HKL-peanut may provide effective therapy for peanut allergy in humans. Funding: Children's Health
J ALLERGY CLIN IMMUNOL FEBRUARY 2003
552 Systemic Occult Bone Marrow Mastocytosis Presenting as Recurrent Anaphylaxis C. Akin, D. D. Metcalfe; Laboratory of Allergic Diseases, NIAID, NIH, Bethesda, MD. Unexplained episodes of recurrent anaphylaxis can be variably observed in patients with mastocytosis. Clinical characteristics of patients with mastocytosis who experience recurrent anaphylaxis, however, have not been analyzed. We have thus studied thirty consecutive patients evaluated at the NIH Clinical Center with a diagnosis of systemic mastocytosis. Thirty percent of the patients (9 total: 6 males, 3 females) had a history of recurrent unexplained anaphylaxis. Seven of 9 patients (78%) with anaphylaxis lacked any cutaneous lesions of mastocytosis. This finding was significantly different than the group without anaphylaxis where only 4 of 21 (20%) lacked skin lesions (p<0.005, Fisher's exact probability). Furthermore, patients with anaphylaxis had significantly less bone marrow involvement with mast cells. Thus, six of 9 (67%) of the patients with anaphylaxis lacked the characteristic compact mast cell aggregates in bone marrow biopsy as opposed to only 3 of 15 (20%) patients without anaphylaxis (p<0.02). Five patients with anaphylaxis had neither skin lesions nor characteristic infiltrates of mast cells in bone marrow biopsy and had baseline serum tryptase levels of less than 20 ng/ml. These patients, however, had other diagnostic features of disease such as aberrant mast ceil morphology, expression of CD2 and CD25 by mast cells, and the presence of codon 816 c-kit mutations. Such patients are termed to have "occult bone marrow mastocytosis" which is detectable by sensitive diagnostic approaches. In conclusion, systemic mastocytosis may be an underlying pathology in unexplained recurrent anaphylaxis in more patients than previously recognized. Funding: NIAID Intramural Funds
553 ,." AcetylhydrolaseDeficiencyPredisposesto FatalAnaphyP. Vadas 1, M. Gold 2, G. Liss 3, C. Smith 2, ]. Yeung4, B. Perelman-~; IMedicine, St. Michael's Hospital, University of Toronto, Toronto, ON, CANADA, 2Hospital for Sick Children, Toronto, ON, CANADA, 3Government of Ontario, Ministry of Labour, Toronto, ON, CANADA, 4National Food Processors Association, Washington, DC, 5St. Michael's Hospital, University of Toronto, Toronto, ON, CANADA. Platelet activating factor (PAF) is a central mediator of anaphylaxis in animal models. Manifestations of anaphylaxis are reproduced by exogenous PAF; PAF receptor antagonists protect against fatal anaphylaxis and PAF receptor knock-out mice show diminished anaphylactic responses. We examined the relationship between severity of anaphylaxis and levels of PAF acetylhydrolase (PAF-AH), the enzyme that inactivates PAE PAFAH levels were measured in sera of individuals who had died of peanut anaphylactic reactions and compared to control groups. Results were: fatal peanut anaphylaxis (n=8, PAF-AH 15.7 + 2.6 nmol/min/ml); healthy adults (n=26, PAF-AH 35.2 -- I 1.1 ), healthy children (n=26, PAF-AH 27.7 + 8.5); peanut allergic children (n=63, PAF-AH 24.9 +_5.8): non-fatal anaphylaxis (n=24, PAF-AH 29.6 -+ 9.0): non-anaphylactic fatalities (n=lO, PAF-AH 26.4 -+ 7.1). The differences between mean serum PAFAH activity in the fatal peanut anaphylaxis group and the control groups were highly significant (p=0.0003). whereas serum PAF-AH levels were comparable in all control groups. Analysis of genomic DNA for 3 patients with the lowest serum PAF-AH activities revealed no mutations in the coding exons of the PAF-AH gene. PAF-AH circulates as a complex with low density lipoproteins and we found a linear correlation between PAFAH activity and apolipoprotein B concentrations in patients with fatal and non-fatal anaphylactic reactions to peanut (r=0.614, p<0.0001). These data clearly show that mean serum PAF-AH activity was significantly lower in patients with fatal peanut anaphylaxis as compared to other groups, including healthy controls. Low density lipoprotein levels may be the major determinant of serum PAF-AH activity. Funding: Peanut Foundation