title: 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.
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 (FcɛRII). 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 k(D)≈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.
authors.affiliation: Department of Systems Immunology, College of Biomedical Science, Kangwon National University, Chuncheon, Gangwon, South Korea.
authors.forename: Hyo-jung
authors.initials: HJ
authors.lastname: Choi
authors.forename: Suk-yoon
authors.initials: SY
authors.lastname: Song
authors.forename: Jae-bong
authors.initials: JB
authors.lastname: Yoon
authors.forename: Li-kun
authors.initials: LK
authors.lastname: Liu
authors.forename: Kristine
authors.initials: K
authors.lastname: Kim
authors.forename: Sang-hoon
authors.initials: SH
authors.lastname: Cha
mesh_terms: D000915:Antibody Affinity / Q000276:immunology*; D000918:Antibody Specificity / Q000276:immunology; D045744:Cell Line, Tumor; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology* / Q000302:isolation & purification*; D007147:Immunoglobulin Light Chains / Q000276:immunology / Q000378:metabolism; D016393:Lymphoma, B-Cell / Q000378:metabolism*; D019151:Peptide Library; D011485:Protein Binding / Q000276:immunology; D017455:Receptors, IgE / Q000276:immunology* / Q000378:metabolism*; D020298:U937 Cells
chemical_list: D007140:Immunoglobulin Fab Fragments; D007147:Immunoglobulin Light Chains; D019151:Peptide Library; D017455:Receptors, IgE