Simultaneous antagonism of interleukin-5, granulocyte-macrophage colony- stimulating factor, and interleukin-3 stimulation of human eosinophils by targetting the common cytokine binding site of their receptors

Human interleukin-5 (IL-5), granulocyte-macrophage colony-stimulating factor (GM-CSF), and IL-3 are eosinophilopoietic cytokines implicated in allergy in general and in the inflammation of the airways specifically as seen in asthma. All 3 cytokines function through cell surface receptors that comprise a ligand-specific α chain and a shared subunit (β(c)). Although binding of IL-5, GM-CSF, and IL-3 to their respective receptor α chains is the first step in receptor activation, it is the recruitment of β(c) that allows high-affinity binding and signal transduction to proceed. Thus, β(c) is a valid yet untested target for antiasthma drugs with the added advantage of potentially allowing antagonism of all 3 eosinophil-acting cytokines with a single compound. We show here the first development of such an agent in the form of a monoclonal antibody (MoAb), BION-1, raised against the isolated membrane proximal domain of β(c). BION-1 blocked eosinophil production, survival, and activation stimulated by IL-5 as well as by GM-CSF and IL-3. Studies of the mechanism of this antagonism showed that BION-1 prevented the high-affinity binding of ¹²⁵I-IL-5, ¹²⁵I-GM-CSF, and ¹²⁵I-IL-3 to purified human eosinophils and that it bound to the major cytokine binding site of β(c). Interestingly, epitope analysis using several β(c) mutants showed that BION-1 interacted with residues different from those used by IL- 5, GM-CSF, and IL-3. Furthermore, coimmunoprecipitation experiments showed that BION-1 prevented ligand-induced receptor dimerization and phosphorylation of β(c), suggesting that ligand contact with β(c) is a prerequisite for recruitment of β(c), receptor dimerization, and consequent activation. These results demonstrate the feasibility of simultaneously inhibiting IL-5, GM-CSF, and IL-3 function with a single agent and that BION- 1 represents a new tool and lead compound with which to identify and generate further agents for the treatment of eosinophil-dependent diseases such as asthma.