Specific Immunotherapy in Hymenoptera Venom Allergy and Concomitant Malignancy: A Retrospective Follow-up Focusing on Effectiveness and Safety.

BACKGROUND: Malignancies are often considered a contraindication for allergen-specific immunotherapy. Consequently, patients with severe Hymenoptera venom allergy and cancer require specific care. The aim of this retrospective study was to assess patients with Hymenoptera venom allergy and cancer undergoing venom immunotherapy (VIT). METHODS: The study population comprised all patients referred for evaluation of Hymenoptera venom allergy or for a routine check-up during VIT from January 1, 2004 to December 31, 2008. RESULTS: Of the patients assessed, 2% (51 of 2594) had a documented Hymenoptera venom allergy and cancer (25 female, 26 male; mean age 58 years). Of these, 42 patients received VIT (82%): 25 patients had a previously diagnosed malignancy, 16 were diagnosed with malignancy during VIT, and 1 patient was diagnosed with cancer after completion of VIT. The most frequent type of tumor was breast cancer in female patients (60%) and prostate cancer in male patients (39%). Systemic allergic reactions during VIT were recorded in 7% of patients. A total of 19 patients experienced a field sting or underwent a sting challenge test during VIT: 95% tolerated the sting well. VIT was halted definitively in 9 patients (new diagnosis of cancer in 7 patients, reactivation of cancer in 1, and progressive polyneuropathy in 1). CONCLUSIONS: The effectiveness and adverse effects of VIT in patients with Hymenoptera venom allergy and cancer in remission are comparable to those of patients without malignancy. Our findings show that patients with Hymenoptera venom allergy and cancer are eligible for VIT.

Influence of total and specific IgE, serum tryptase, and age on severity of allergic reactions to Hymenoptera stings.

BACKGROUND: The aim of this study was to analyze the influence of total serum IgE and other potential risk factors on severity of systemic allergic Hymenoptera sting reactions. METHODS: In a retrospective analysis of one thousand and two patients referred for insect allergy over 5 years, 865 reported systemic allergic sting reactions, most often by honey bees and wasps. In 758, total IgE, venom-specific IgE, and baseline tryptase levels were available and analyzed together with atopy state, age, and sex in relation to severity of sting reactions according to H. L. Mueller. RESULTS: In a binary logistic regression model considering, besides IgE, also other risk factors for severity, an influence of total and specific IgE on severity of systemic allergic sting reactions could not be shown, while high severity of systemic allergic sting reactions was significantly more often reported in patients with a baseline tryptase of ≥11.4 µg/l (P < 0.0001) and higher age (P = 0.026). In a bivariate analysis, however, in patients with grade IV reactions total IgE (P = 0.003) and honey bee venom-specific IgE (P = 0.001) were significantly lower than in lower severity grades. Bee venom-specific mean IgE rank was significantly higher in bee than in Vespula venom allergic patients (P = 0.0001). CONCLUSIONS: Connection of high severity sting reactions with lower IgE is mainly because of older age, which is associated with lower total IgE, and moreover with cardiovascular disease and elevated baseline serum tryptase, which are both risk factors for severe reactions.

Hymenoptera venom allergy: analysis of double positivity to honey bee and Vespula venom by estimation of IgE antibodies to species-specific major allergens Api m1 and Ves v5.

BACKGROUND: In patients with hymenoptera venom allergy diagnostic tests are often positive with honey bee and Vespula venom causing problems in selection of venoms for immunotherapy. METHODS: 100 patients each with allergic reactions to Vespula or honey bee stings and positive i.e. skin tests to the respective venom, were analysed for serum IgE to bee venom, Vespula venom and crossreacting carbohydrate determinants (CCDs) by UNICAP (CAP) and ADVIA Centaur (ADVIA). IgE-antibodies to species specific recombinant major allergens (SSMA) Api m1 for bee venom and Ves v5 for Vespula venom, were determined by ADVIA. 30 history and skin test negative patients served as controls. RESULTS: By CAP sensitivity was 1.0 for bee and 0.91 for Vespula venom, by ADVIA 0.99 for bee and 0.91 for Vespula venom. None of the controls were positive with either test. Double positivity was observed in 59% of allergic patients by CAP, in 32% by ADVIA. slgE to Api m1 was detected in 97% of bee and 17% of Vespula venom allergic patients, slgE to Ves v5 in 87% of Vespula and 17% of bee venom allergic patients. slgE to CCDs were present in 37% of all allergic patients and in 56% of those with double positivity and were more frequent in bee than in Vespula venom allergic patients. CONCLUSIONS: Double positivity of IgE to bee and Vespula venom is often caused by crossreactions, especially to CCDs. IgE to both Api m1 and Ves v5 indicates true double sensitization and immunotherapy with both venoms.

[Hymenoptera venom anaphylaxis and cardiovascular disease]. / Hymenopterengiftanaphylaxie und Herz-Kreislauf-Erkrankungen.

Preexisting cardiovascular disease may worsen the course of anaphylaxis. This is illustrated based on the example of Hymenoptera venom allergy. Fatal sting anaphylaxis is most often observed in elderly patients. During autopsy preexisting cardiovascular disease is frequently found. Preexisting cardiovascular disease in patients with anaphylaxis may also cause lasting morbidity, e.g. cerebral or myocardial infarction. Heart medications, notably beta-blockers und ACE-inhibitors may worsen the course of anaphylactic reactions due to their pharmacologic effects. Since cardiovascular diseases are much more frequent than anaphylaxis and these medications are very effective, these drugs cannot be substituted in patients with both diseases without a careful risk analysis. Epinephrine is the drug of first choice for treatment of anaphylaxis. It may however, especially following rapid intravenous administration, cause severe arrhythmias or myocardial infarction. Adrenaline should therefore preferably be given intramuscularly, or by slow intravenous infusion.

Discriminating two classes of toxicants through expression analysis of HepG2 cells with DNA arrays.

Microarray technology provides a rapid and cost-effective method to associate specific cellular responses with unique gene expression patterns. If characteristic expression patterns of a small number of genes could be associated with drug toxicity, this association may be used for toxicity prediction, and thereby to reduce the need for traditional toxicity testing. To test this hypothesis, we have designed an array composed of 92 known human genes of toxicological interest (including seven housekeeping genes) and eight bacterial controls. HepG2 cells were treated with either ethanol or one of two quinone containing anticancer drugs, mitomycin C or doxorubicin. RNA was isolated from treated and untreated cells, differentially labeled with fluorescent dyes, and then hybridized to the array. Our results show that the expression patterns induced by ethanol and the anticancer drugs are different. Both of the anticancer drugs, but not ethanol had a differential effect on the regulation of several genes, including CYP4F2/3, CYP3A3, TNFRSF6 and CHES1, demonstrating that the two drugs might function through a similar mechanism, which differs from that of ethanol. These results suggest that microarray-based expression analysis may offer a rapid and efficient means for assessing drug toxicity.

Novel surface and multicolor charge coupled device-based fluorescent imaging system for DNA microarrays.

We report a novel support, concomitant attachment chemistry, and a fluorescent imaging system for DNA microarrays. The support consists of soda lime glass coated with a layer of chromium, which eliminates any autofluorescence from the underlying glass substrate and reduces nonspecific probe binding. Attachment of DNA fragments exceeding 300 nucleotides in length is achieved without chemical modifications of either the chrome surface or the DNA itself. The charge coupled device (CCD)-based imaging system employs a 175 W xenon arc lamp as the light source, allowing the use of many different fluorophors. A 14 mm x 9 mm sample area is imaged with a single exposure, which takes between 5 and 20 s for each color plane in typical genomic comparative genomic hybridization type assays. The spatial resolution is limited only by the pixel size of the CCD chip (9 microm). The oblique illumination geometry combined with effective background reduction afforded by the chromium surface enables the system to achieve a detection limit of <5 x 10(7) fluorophors/cm(2) with 10 s integration. In a model system with arrayed lambda DNA targets a dose response was observed over four orders of magnitude in response to hybridizations with increasing amounts of the fluorescent labeled lambda probe.

New developments in the diagnosis and treatment of hymenoptera venom allergy.

According to most textbooks, diagnostic tests with Hymenoptera venoms are reliable, and immunotherapy with these venoms in Hymenoptera-venom-allergic patients leads in near to 100% to full protection. Careful analysis of the literature shows however that the specificity of diagnostic tests is far from perfect and that both efficacy and tolerance, especially in patients receiving honeybee venom immunotherapy, are still suboptimal. The major allergens of honeybee and vespid venoms are now available in recombinant form. Preliminary trials analyzing diagnostic tests with recombinant allergens in honeybee venom allergy are promising: the specificity is clearly increased in both skin testing and in determining venom-specific IgE antibodies when compared to natural venom allergens. An important recent finding is the frequent association of severe Hymenoptera venom allergy and elevated basal serum levels of the mast-cell-specific enzyme tryptase. Elevated levels are found in up to 30% of the patients with a history of severe shock reactions following Hymenoptera stings. The current findings indicate that basal tryptase levels indicating an increased mast cell load are much more frequent than previously thought and are a risk factor for severe or even fatal sting reactions. Premedication with antihistamines in the initial phase of venom immunotherapy reduced both local and systemic allergic side effects in several controlled studies. In a retrospective analysis of one of these trials it was found that reexposure during immunotherapy resulted in significantly more systemic allergic reactions in patients on placebo than on antihistamine premedication, suggesting that initial antihistamine premedication might increase the efficacy of venom immunotherapy. Different ways of allergen modification for venom immunotherapy have been proposed. While the results with chemical modifications were not convincing, recent studies with T-cell epitope peptides from the major bee venom allergen phospholipase A(2) look promising. Patient-tailored cocktails of recombinant venom allergens or isoforms thereof may be another possibility in the future. A number of prospective studies analyzing the duration of venom immunotherapy required for long-term protection have been published in the last decade. While most patients are still fully protected 1 year after discontinuation of therapy, relapses may occur in up to 20% of patients reexposed many years after treatment. Various risk factors for such relapses have been identified.

Are anaphylactic reactions to snake bites immunoglobulin E-mediated?

BACKGROUND: Bites by poisonous European snakes of the genus Vipera lead to local tissue damage and systemic symptoms such as generalized oedema, hypotension, gastrointestinal symptoms, haemolysis and renal dysfunction. Not rarely anaphylactic symptoms like urticaria, localized angioedema and asthma are observed. OBJECTIVE: To look for snake venom-specific immunoglobulin (Ig) E antibodies in patients with a history of bites by European vipers and for cross-reactions with Hymenoptera venoms, that have a similar composition. METHOD: Ten patients with a history of bites by Vipera aspis or Vipera berus were investigated. Three patients had been bitten only once, and two of these had developed only local reactions. Four reported previous allergic reactions to Hymenoptera stings. All patients, 10 Hymenoptera venom-allergic and five nonallergic individuals who served as controls underwent i.c. skin test endpoint titration with snake (V. aspis, V. berus) and Hymenoptera venoms (honey bee, Vespula spp.) and were investigated for specific serum IgE antibodies to the same venoms. RESULTS: Seven of the eight patients with systemic snake bite reactions had both positive skin tests and serum IgE antibodies to snake venoms, while these tests were negative in the two patients with only local reactions to snake bites and all controls. Seven of the eight patients with systemic snake bite reaction also had positive skin tests and specific IgE with one or both Hymenoptera venoms. By RAST-inhibition with sera of four patients with high IgE to both Vipera and Hymenoptera venoms, partial cross-reactivity could be demonstrated in one. CONCLUSIONS: Anaphylactic reactions following snake bites may be IgE-mediated, especially in patients with repeated bites.

Adaptation of capillary isoelectric focusing to microchannels on a glass chip.

As a first step toward adaptation of capillary isoelectric focusing (cIEF) to microchannels on a glass chip, we have compared the three most common mobilization methods: chemical, hydrodynamic, and electroosmotic flow (EOF)-driven mobilization. Using a commercial cIEF apparatus with coated or uncoated fused-silica capillaries, both chemical and hydrodynamic mobilization gave superior separation efficiency and reproducibility. However, EOF-driven mobilization, which occurs simultaneously with focusing, proved most suitable for miniaturization because of high speed, EOF compatibility and low instrumentation requirements. When this method was tested in a 200-micron-wide, 10-micron-deep, and 7-cm-long channel etched into planar glass, a mixture of Cy5-labeled peptides could be focused in less than 30 s, with plate heights of 0.4 micron (410 plates/s) upon optimization. For a total analysis time of less than 5 min, we estimate a maximum peak capacity of approximately 30-40. Interestingly, the order of migration was found to be reversed compared to capillary-based focusing.

Bee sting allergy in beekeepers.

BACKGROUND: Beekeepers are strongly exposed to honey bee stings and therefore at an increased risk to develop IgE-mediated allergy to bee venom. OBJECTIVE: We wondered whether bee venom-allergic beekeepers were different from normally exposed bee venom-allergic patients with regard to clinical and immunological parameters as well as their response to venom immunotherapy. METHOD: Among the 459 bee venom-allergic patients seen over the 5 year period 1987-91, 62 (14%) were beekeepers and 44 (10%) family members of beekeepers. These two groups were compared with 101 normally exposed bee venom-allergic patients matched with the allergic beekeepers for age and sex, regarding clinical parameters, skin sensitivity, specific IgE and IgG antibodies to bee venom as well as safety and efficacy of venom immunotherapy. RESULTS: As expected, allergic beekeepers had been stung most frequently before the first allergic reaction. The three groups showed a similar severity of allergic symptoms following bee stings and had an equal incidence of atopic diseases. Allergic beekeepers showed higher levels of bee venom-specific serum IgG, lower skin sensitivity and lower levels of bee venom specific serum IgE than bee venom-allergic control patients. A negative correlation between number of stings and skin sensitivity as well as specific IgE was found in allergic beekeepers and their family members, while the number of stings was positively correlated with specific IgG in these two groups. Venom immunotherapy was equally effective in the three groups, but better tolerated by allergic beekeepers than the two other groups. The majority of allergic beekeepers continued bee-keeping successfully under the protection of venom immunotherapy. CONCLUSION: The lower level of sensitivity in diagnostic tests and the better tolerance of immunotherapy in allergic beekeepers is most likely related to the high level of specific IgG in this group.

Simultaneous multiple analyte detection using fluorescent peptides and capillary isoelectric focusing.

Analyte-specific detection based on the isoelectric point of the detection moiety is a new concept that is under investigation at Vysis. We have developed methods for the synthesis of of fluorescent synthetic peptides that can be conjugated to bioanalytes such as nucleic acids and antibodies, processed in a hybridization or binding assay, and then chemically released prior to detection by capillary isoelectric focusing (cIEF)-laser-induced fluorescence (LIF) detection. A two-step cIEF method in coated capillaries using salt mobilization has been used that produces high peak efficiencies and good assay reproducibility. The concentration by focusing aspect of cIEF, which allows for the entire capillary to be filled with sample, enables detection limits in the pM as opposed to sub-nM level for conventional capillary electrophoresis (CE)-LIF. The simultaneous multiple detection of eleven different focusing entities has been achieved.

Long-term protection after stopping venom immunotherapy: results of re-stings in 200 patients.

BACKGROUND: Venom immunotherapy (VIT) protects most patients allergic to Hymenoptera stings while booster injections are continued. Few data on long-term protection after discontinuation of treatment are available. OBJECTIVE: We sought to investigate protection from re-stings over a prolonged period after stopping VIT. METHODS: Re-sting data were obtained from 200 of 322 patients in whom VIT had been stopped between 1988 and 1992 after a duration of at least 3 years. The 25 (12.5%) patients who again developed systemic allergic reactions were compared with 50 matched patients without re-sting reactions. Clinical data and diagnostic parameters (i.e., skin sensitivity and specific IgE and IgG) were studied. RESULTS: Of the 25 patients who had re-sting reactions, 19 had been treated with bee venom (relapse rate, 15.8%), and six had been treated with Vespula venom (relapse rate, 7.5%). About half of the re-sting reactions occurred on the first resting after stopping VIT. Most of these reactions were mild, whereas the majority of reactions occurring after repeated re-stings were severe. When re-sting reactions were related to the total re-stings per year, an accumulation of sting reactions was observed in years 3 to 5 after stopping VIT. Patients with re-sting reactions had been receiving VIT for a significantly shorter duration (43.35 months) than those with continued protection (54.65 months) (p < 0.01). Of the diagnostic parameters, only a negative intracutaneous skin test at 10(-3) gm/L predicted long-term protection reliably. CONCLUSION: Venom immunotherapy of 3 to 5 years duration induces long-term protection in most patients. In rare occasions severe re-sting reactions may, however, occur, especially after repeated re-stings.