Isolation of human Fab antibodies specific for the low-affinity IgE receptor (CD23) by selecting a hierarchical antibody library system against B lymphoblastic IM-9 cells

• Published in: Immunology letters 2011-05, Vol.136 (2), p.213-220
• Main Authors: Choi, Hyo-jung, Song, Suk-yoon, Yoon, Jae-bong, Liu, Li-kun, Kim, Kristine, Cha, Sang-hoon
• Format: Article
• Language: English
• Subjects: Allergic diseases; Allergy and Immunology; Antibody Affinity - immunology; antibody libraries; Antibody Specificity - immunology; CD23; CD23 antigen; Cell Line, Tumor; Cell panning; Chronic lymphatic leukemia; double prime B-cell lymphoma; Dual-vector system; Fab; Human antibody; Humans; IM-9; Immunoglobulin E; Immunoglobulin Fab Fragments - immunology; Immunoglobulin Fab Fragments - isolation & purification; Immunoglobulin Light Chains - immunology; Immunoglobulin Light Chains - metabolism; Interleukin 4; Lymphocytes B; Lymphoma, B-Cell - metabolism; Monoclonal antibodies; Panning; Peptide Library; Phage display; Protein Binding - immunology; Receptors, IgE - immunology; Receptors, IgE - metabolism; U937 Cells
• ISSN: 0165-2478; 1879-0542
• DOI: 10.1016/j.imlet.2011.01.012

Abstract The development of human antibodies specific for certain B cell markers is required to generate therapeutic antibody leads with improved therapeutic indices against B-cell lymphomas. To meet this demand, we selected a primary human antibody library, HuDVFab-8L, against human B lymphoblastic IM-9 cells via a ‘Biopanning and Rapid Analysis of Selective Interactive Ligands (BRASIL)’ cell panning approach. Six Fab clones that specifically bound to IM-9 cells were successfully isolated. Among these clones, two clones (IM-L6-E and IM-L8-G), were found to be specific for CD23 (FcεRII). Affinity maturation of these Fab clones was then performed in a hierarchical manner by constructing secondary antibody libraries through combining heavy (H) chains of two Fabs with the human kappa L chain sublibrary HuNL-D3 followed by biopanning against the CD23 antigen. Clone IM-L6-5, one of the affinity maturated Fab derivatives from IM-L6-E, has a binding affinity of kD ≈ 30 nM to soluble CD23. In addition, IM-L6-5 Fab is able to bind to an inducible form of CD23 expressed on U937 cells upon IL-4 stimulation, and inhibits binding of human IgE to CD23. Since the Fab IM-L6-5 is derived from a fully human naïve origin, we believe that IM-L6-5 can be utilized for the development of a therapeutic mAb which may have an improved therapeutic index over lumiliximab, a primatized anti-CD23 mAb, for the treatment of CLL or allergic diseases.