title: Monoclonal antibodies against the C(epsilon)mX domain of human membrane-bound IgE and their potential use for targeting IgE-expressing B cells.
abstract: BACKGROUND
  IgE mediates immediate-type hypersensitivity reactions responsible for various allergic symptoms. It is secreted by IgE-producing plasma cells, which differentiate from B cells expressing membrane-bound IgE (mIgE) on their surface. The epsilon-chain of human mIgE contains a membrane-anchoring peptide and an extra 52-amino-acid (a.a.)-long domain (referred to as C(epsilon)mX) between the membrane anchor and the CH4 domain.
  
  
  OBJECTIVE
  The study was designed to evaluate the effects of C(epsilon)mX-specific monoclonal antibodies (mAbs) to target IgE-expressing B cells and decrease IgE production.
  
  
  METHODS
  A C(epsilon)mX-containing IgG1.Fc fusion protein was produced in CHO cells and used to immunize mice; five hybridoma clones secreting C(epsilon)mX-specific mAbs were obtained.
  
  
  RESULTS
  Characterization of the mAbs using ELISA, immunoprecipitation, and immunoblotting methods showed that they could bind to both native and denatured forms of C(epsilon)mX. The mAbs exhibited mutual inhibition of binding to mIgE. Epitope mapping using synthetic peptides revealed that all five mAbs recognize the same epitope, RADWPGPP, located near the C-terminus of C(epsilon)mX. Binding of one of the mAbs to mIgE on SKO-007 cells induced the cross-linking of mIgE molecules on the cell surface, resulting in their patching and capping. In vitro functional analysis revealed that mAbs are able to cause complement-mediated cytotoxicity on transfectants expressing the Fc portion of mIgE.
  
  
  CONCLUSIONS
  We have prepared several human mIgE-specific mAbs. The potential of the mAbs on targeting mIgE+ B cells was demonstrated by CDC analysis.
authors.affiliation: Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan, ROC.
authors.forename: Huan Yuan
authors.initials: HY
authors.lastname: Chen
authors.forename: Fu-Tong
authors.initials: FT
authors.lastname: Liu
authors.forename: Charlie M H
authors.initials: CM
authors.lastname: Hou
authors.forename: Janice S W
authors.initials: JS
authors.lastname: Huang
authors.forename: Bhavya Bhavna
authors.initials: BB
authors.lastname: Sharma
authors.forename: Tse Wen
authors.initials: TW
authors.lastname: Chang
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology* / Q000627:therapeutic use; D001402:B-Lymphocytes / Q000187:drug effects / Q000276:immunology; D001667:Binding, Competitive; D016466:CHO Cells; D002460:Cell Line; D003167:Complement Activation; D006224:Cricetinae; D016503:Drug Delivery Systems; D018604:Epitope Mapping; D006825:Hybridomas; D006969:Hypersensitivity, Immediate / Q000628:therapy; D015151:Immunoblotting; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D007139:Immunoglobulin epsilon-Chains / Q000737:chemistry / Q000276:immunology*; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D011940:Receptor Aggregation; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000494:pharmacology
chemical_list: D000911:Antibodies, Monoclonal; D007141:Immunoglobulin Fc Fragments; D007139:Immunoglobulin epsilon-Chains; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins