A single mouse monoclonal antibody, E58 modulates multiple IgE epitopes on group 1 cedar pollen allergens

Abstract

We recently described a dominant role for conformational epitopes on the group 1 allergen of the mountain cedar (Juniperus ashei, Cupressaceae), Jun a 1, in pollen hypersensitivity in South Central U.S.A. Since these epitopes are surface exposed and are likely to be flexible, they may be susceptible to molecular or physical perturbations. This may make Jun a 1 a potential target for new forms of therapy for cedar pollinosis. Here, we describe a mouse monoclonal antibody, termed E58, which binds to the group 1 allergens of the cedar pollens from three highly populated regions of the world (central U.S.A., France and Japan). Upon binding to these allergens, E58 strongly reduces the binding of patient's IgE antibodies to these dominant allergens. This characteristic of E58, and potentially other similar antibodies, suggests an opportunity to develop preventative or therapeutic agents that may inhibit cedar pollen sensitization or prevent their allergic reactions.

Keywords: Allergen structure; Cedar pollen allergy; Conformational epitope; IgE epitope; Jun a 1; Monoclonal antibody.

Fig 1. E58 inhibits the binding of other mAbs in ELISAs

(A) Denaturation of Jun a 1 with 6M guanidine did not significantly reduced the reactivity of E58 to Jun a 1. N = 3, mean and SE were shown. N.S.: not significant. (B) However, preincubating Jun a 1 with mAb E58 inhibits the subsequent binding of 4 other mAbs (G1–4) to their conformational epitopes of Jun a 1. N = 3, mean and SE were shown. *: p<0.05 compared to negative control, +: p<0.05 compared to inhibition by E58. (C) The same mAbs G1–4 do not inhibit the binding of E58 to Jun a 1. *: p<0.05 compared to negative control, +: p<0.05 compared to inhibition by E58. (D) ■: E58 inhibition of E58 binding to each allergen, ━: mean ± SD. E58 also inhibits the binding of IgE antibodies from sera of patients’ sensitive to other group 1 cedar allergens, Cup s 1 and Cry j 1. N.S.: not significant, *: p<0.05 between indicated groups.

Fig 2. rE58 scFv inhibits degranulation of human serum IgE-sensitized mast cells

rE58 scFv (a univalent Ab) inhibits Jun a 1-induced degranulation of mast cells sensitized with three different mountain cedar patients’ sera. N = 3, mean ±SD were shown. *: p<0.05 compared to no antibody control.

Fig. 3. Potential epitope of E58

Amino acid sequences of Jun a 1, Cup s 1 and Cry j 1 were aligned. Surface exposed amino acids are highlighted with gray and prediction of linear B cell epitopes were underlined. There were only two amino acids 40 and 46, which were surface exposed and predicted as linear B cell epitopes that differ between these three allergens. This region may also be part of the E58 epitopes.

Fig 4. Biacore analyses

Surface plasmon resonance analysis of the binding of: (A) native E58, (B) E58 scAb, (C) E58 Fab to Jun a 1. Graphs display Response Units (RU) as a function of the molar concentration of each analyte. The line through the points is the resulting fit from the steady state analysis procedure (part of the Biacore T100 analysis software).