A Potent Human Anti-Eotaxin1 Antibody, CAT-213: Isolation by Phage Display and in Vitro and in Vivo Efficacy

Abstract

The CC chemokine, eotaxin1 (CCL11) is an important regulator of eosinophil function. A marked accumulation of eosinophils in tissues has been correlated with the up-regulation of eotaxin1 expression in several diseases. The potential therapeutic value of neutralizing the effects of eotaxin1 in inflammatory conditions (including asthma) is under investigation. A human singlechain fragment variable antibody that neutralizes human eotaxin1 (CAT-212) was produced using antibody phage display and converted to whole antibody IgG4 format (CAT-213). A novel approach to lead optimization in which the length of the variable heavy chain complementarity-determining region 3 was reduced by one amino acid resulted in an increase in potency of Ͼ1000-fold compared with the parent anti-eotaxin1 antibody. The optimized antibody binds eotaxin1 with high affinity (80.4 pM) and specificity. CAT-213 and CAT-212 do not bind or neutralize a range of other human proteins including human monocyte chemoattractant protein-1, a structurally similar chemokine. CAT-213 neutralizes the ability of eotaxin1 to cause an increase in intracellular calcium signaling (with an IC 50 value of 2.86 nM), migration of CCR3-expressing L1.2 cells (with an IC 50 value of 0.48 nM), and inhibition of the eotaxin1-evoked shape change of human eosinophils in vitro (with an IC 50 of 0.71 nM). Local administration of CAT-213 to mice (1-100 g kg Ϫ1 ) attenuates dermal eosinophilia induced by human eotaxin1, achieving Ͼ90% inhibition of eosinophil influx. CAT-213 may therefore be of therapeutic value in inhibiting diseases in which eotaxin1 and eosinophils play a major role, for example, severe asthma.

Primary eosinophilic disorders of the gastrointestinal tract

“…Mepolizumab, a humanised anti-IL5 monoclonal antibody, shown benefit in five patients without serious side effects 64 65. Specific eotaxin inhibitors are also under development 66. Medications successfully used in eosinophilic gastroenteritis such as cromolyn and ketotifen (mast cell stabilising medications), and suplatast tosilate (a selective Th2 IL4 and IL5 inhibitor) have not been studied in adults with eosinophilic oesophagitis.…”

Mucosal Eosinophils

“…These results provide substantial evidence that the biological activities attributed to eotaxins in animals are conserved in humans. This has prompted the development of a humanized anti-eotaxin-1 antibody (CAT-213) that has been shown to lower eosinophils in the respiratory tract (Main et al, 2006) and is now being developed for a variety of eosinophil-associated disorders including inflammatory bowel disease. In a human allergic GI disorder, EoE, human eotaxin-3 is the most upregulated gene, and may account for esophageal epithelium eosinophil infiltration (Blanchard et al, 2006).…”

Non-IBD and noninfectious colitis

“…53,64 At present, a humanized anti-eotaxin 1 antibody (CAT-213) is undergoing clinical trials. 65 CCR-3 (eotaxin-1 receptor) antagonists 66 and IL-5 blockade (mepolizumab) might also prove useful for treating eosinophilic colitis. 67 Studies have shown that the disease can be caused by one or more somatic mutation in the plateletderived growth factor receptor α (PDGFRα) gene, 68 which confer constitutive activation to the protein and thereby provide a rationale for treatment with tyrosine kinase inhibitors (e.g.…”