Asparaginase Immune Complexes Detectable after Asparaginase-Induced Hypersensitivities Activate Basophils Via FcγRIII

Background: Asparaginase (ASNase) is an important chemotherapeutic agent for the treatment of acute lymphoblastic leukemia (ALL). However, the development of anti-ASNase IgG antibodies are frequent, can neutralize the enzyme activity of ASNase, and can increase the risk of ALL relapse. Nevertheless, the role of anti-ASNase IgG antibodies on the onset of ASNase-induced hypersensitivity reactions is unknown, but likely require the formation of ASNase immune complexes (ICs) to activate cells involved in hypersensitivities due to the low binding affinity of the Fcγ receptor for the IgG immunoglobulin.

Objective: To detect plasma ASNase ICs after the onset of ASNase hypersensitivities and to demonstrate that binding of ASNase ICs to peripheral blood cells, and the subsequent immune cell activation, is FcγRIII-dependent.

Methods: Mice were sensitized to native E.coli ASNase using aluminum hydroxide adjuvant. ICs prepared using the plasma of sensitized mice were precipitated using polyethylene glycol to remove free ASNase and anti-ASNase IgG. ICs were detected by flow cytometry using APC labeled ASNase and protein G beads, characterized by dynamic light scattering and TEM, and the binding properties of ASNase ICs or free ASNase were determined by flow cytometry using non-sensitized and sensitized peripheral murine blood cells and the anti-IgE and/or anti-FcγRIIβ/III blocking mAb. Activation of basophils, which can occur in an IgE- or FcγRIII-dependent manner, was determined ex vivo by measuring changes in CD200R1 or CD200R3 expression after incubating peripheral blood cells with ASNase or ASNase ICs.

Results: ASNase ICs were detected after the onset of ASNase hypersensitivities in mice. ASNase ICs formed ex vivo at high anti-ASNase IgG₁ concentrations, bound to naïve peripheral blood cells (e.g., neutrophils macrophage/monocytes, and basophils), and activated basophils in an FcγRIII-dependent manner. In contrast, basophil binding and activation by free ASNase was IgE- and not FcγRIII-dependent.

Conclusion: Our data indicate that ASNase sensitized mice form ICs after drug administration and can activate basophils via the FcγRIII receptor in mice, whereas the binding and activation by free ASNase occurs via cell-associated IgE. Our study suggests that ASNase ICs can contribute to the onset and severity of ASNase hypersensitivity.