title: Effector cell recruitment with novel Fv-based dual-affinity re-targeting protein leads to potent tumor cytolysis and in vivo B-cell depletion.
abstract: Bispecific antibodies capable of redirecting the lytic potential of immune effector cells to kill tumor targets have long been recognized as a potentially potent biological therapeutic intervention. Unfortunately, efforts to produce such molecules have been limited owing to inefficient production and poor stability properties. Here, we describe a novel Fv-derived strategy based on a covalently linked bispecific diabody structure that we term dual-affinity re-targeting (DART). As a model system, we linked an Fv specific for human CD16 (FcgammaRIII) on effector cells to an Fv specific for mouse or human CD32B (FcgammaRIIB), a normal B-cell and tumor target antigen. DART proteins were produced at high levels in mammalian cells, retained the binding activity of the respective parental Fv domains as well as bispecific binding, and showed extended storage and serum stability. Functionally, the DART molecules demonstrated extremely potent, dose-dependent cytotoxicity in retargeting human PBMC against B-lymphoma cell lines as well as in mediating autologous B-cell depletion in culture. In vivo studies in mice demonstrated effective B-cell depletion that was dependent on the transgenic expression of both CD16A on the effector cells and CD32B on the B-cell targets. Furthermore, DART proteins showed potent in vivo protective activity in a human Burkitt's lymphoma cell xenograft model. Thus, DART represents a biologically potent format that provides a versatile platform for generating bispecific antibody fragments for redirected killing and, with the selection of appropriate binding partners, applications outside of tumor cell cytotoxicity.
authors.affiliation: MacroGenics, Inc., 1500 East Gude Drive, Rockville, MD 20850, USA. johnsons@macrogenics.com
authors.forename: Syd
authors.initials: S
authors.lastname: Johnson
authors.forename: Stephen
authors.initials: S
authors.lastname: Burke
authors.forename: Ling
authors.initials: L
authors.lastname: Huang
authors.forename: Sergey
authors.initials: S
authors.lastname: Gorlatov
authors.forename: Hua
authors.initials: H
authors.lastname: Li
authors.forename: Weili
authors.initials: W
authors.lastname: Wang
authors.forename: Wenjun
authors.initials: W
authors.lastname: Zhang
authors.forename: Nadine
authors.initials: N
authors.lastname: Tuaillon
authors.forename: Jonah
authors.initials: J
authors.lastname: Rainey
authors.forename: Bhaswati
authors.initials: B
authors.lastname: Barat
authors.forename: Yinhua
authors.initials: Y
authors.lastname: Yang
authors.forename: Linda
authors.initials: L
authors.lastname: Jin
authors.forename: Valentina
authors.initials: V
authors.lastname: Ciccarone
authors.forename: Paul A
authors.initials: PA
authors.lastname: Moore
authors.forename: Scott
authors.initials: S
authors.lastname: Koenig
authors.forename: Ezio
authors.initials: E
authors.lastname: Bonvini
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000235:genetics / Q000276:immunology* / Q000493:pharmacokinetics; D001402:B-Lymphocytes / Q000276:immunology / Q000473:pathology*; D002051:Burkitt Lymphoma / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D045744:Cell Line, Tumor; D003602:Cytotoxicity, Immunologic; D005260:Female; D058851:GPI-Linked Proteins; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000276:immunology*; D007963:Leukocytes, Mononuclear / Q000276:immunology*; D016393:Lymphoma, B-Cell / Q000276:immunology / Q000473:pathology*; D051379:Mice; D008819:Mice, Nude; D008822:Mice, Transgenic; D009368:Neoplasm Transplantation; D055550:Protein Stability; D017452:Receptors, IgG / Q000235:genetics / Q000276:immunology*; D014183:Transplantation, Heterologous
chemical_list: D018033:Antibodies, Bispecific; C428200:FCGR3B protein, human; C080571:Fc gamma receptor IIB; D058851:GPI-Linked Proteins; D007135:Immunoglobulin Variable Region; D017452:Receptors, IgG