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Background Recent data suggest that Bruton's tyrosine kinase ( BTK) is an emerging therapeutic target in IgE receptor (Ig ER)-cross-linked basophils. Methods We examined the effects of four BTK inhibitors (ibrutinib, dasatinib, AVL-292, and CNX-774) on IgE-dependent activation and histamine release in blood basophils obtained from allergic patients (n=11) and nonallergic donors (n=5). In addition, we examined the effects of these drugs on the growth of the human basophil cell line KU812 and the human mast cell line HMC-1. Results All four BTK blockers were found to inhibit anti-IgE-induced histamine release from basophils in nonallergic subjects and allergen-induced histamine liberation from basophils in allergic donors. Drug effects on allergen-induced histamine release were dose dependent, with IC50 values ranging between 0.001 and 0.5 μmol/L, and the following rank order of potency: ibrutinib> AVL-292>dasatinib> CNX-774. The basophil-targeting effect of ibrutinib was confirmed by demonstrating that IgE-dependent histamine release in ex vivo blood basophils is largely suppressed in a leukemia patient treated with ibrutinib. Dasatinib and ibrutinib were also found to counteract anti-IgE-induced and allergen-induced upregulation of CD13, CD63, CD164, and CD203c on basophils, whereas AVL-292 and CNX-774 showed no significant effects. Whereas dasatinib and CNX-774 were found to inhibit the growth of HMC-1 cells and KU812 cells, no substantial effects were seen with ibrutinib or AVL-292. Conclusions BTK-targeting drugs are potent inhibitors of IgE-dependent histamine release in human basophils. The clinical value of BTK inhibition in the context of allergic diseases remains to be determined. [ABSTRACT FROM AUTHOR]

The induction of blocking Ig G antibodies that compete with Ig E for allergen binding is one important mechanism of allergen-specific immunotherapy. The application of blocking antibodies may be an alternative treatment strategy. A synthetic gene coding for a single-chain fragment ( Sc Fv) specific for the major timothy grass pollen allergen Phl p 2 was inserted into plasmid pCANTAB 5 E, and the recombinant Sc Fv was expressed in Escherichia coli and purified by affinity chromatography. The Sc Fv was tested for allergen binding by ELISA, and its association and dissociation were measured by surface plasmon resonance (Biacore) technology. The ability of the Sc Fv to inhibit allergic patients' Ig E binding to Phl p 2 and Phl p 2-induced basophil degranulation was studied by ELISA competition and basophil activation ( CD203c) assays. We report the expression, purification, biochemical and immunological characterization of a monomeric single-chain fragment ( Sc Fv) of human origin specific for the major timothy grass pollen allergen, Phl p 2. The Phl p 2- Sc Fv showed high affinity binding to the allergen and blocked the binding of allergic patients' polyclonal Ig E to Phl p 2 up to 98%. Furthermore, it inhibited allergen-induced basophil activation. The Phl p 2- Sc Fv inhibited allergic patients' Ig E binding to Phl p 2 as well as Phl p 2-induced basophil activation and might be useful for passive immunotherapy of grass pollen allergy. [ABSTRACT FROM AUTHOR]

Background Recent data obtained in mouse models have initiated a controversy whether basophils are the key antigen-presenting cells ( APCs) in allergy. Here, we investigate whether basophils are of importance for the presentation of allergen and the induction of T cell proliferation in allergic patients. Methods T cells, basophils, and APCs depleted of basophils were purified from allergic patients. Co-culture systems based on purified major allergens were established to study allergen-specific T cell responses using proliferation assays. Results Only co-cultures of T cells with APCs depleted of basophils but not with basophils proliferated in response to allergen. Even addition of IL-3 to T cell-basophil co-cultures failed to induce allergen-specific T cell proliferation. Conclusions Our data demonstrate by classical in vitro proliferation assays that basophils are not key antigen-presenting cells that promote T cell proliferation in secondary immune responses to allergen in allergic patients. [ABSTRACT FROM AUTHOR]