title: A new Tri-Fab bispecific antibody for pretargeting Trop-2-expressing epithelial cancers.
abstract: RS7 is an internalizing anti-Trop-2 pancarcinoma antibody capable of targeting most epithelial cancers. Because pretargeting strategies could improve the tumor localization of radionuclides, a new anti-Trop-2 × antihapten bispecific antibody for pretargeting, based on humanized RS7, was prepared and evaluated with a radiolabeled hapten-peptide in vitro and in vivo to determine whether its internalization properties would interfere with pretargeting.
  
  
  METHODS
  The anti-Trop-2 × antihapten bispecific antibody, TF12, was prepared using the modular dock-and-lock method. TF12 and humanized RS7 binding was assessed by cell binding assays and fluorescence-activated cell sorting analysis in a variety of human carcinoma cell lines. The internalization of TF12 was evaluated in vitro using a fluorescent TF12 conjugate or hapten-peptide and (111)In-labeled TF12 and RS7. The biodistribution of TF12 and its use as a pretargeting agent with an (111)In-labeled hapten-peptide were assessed in several human epithelial cancer xenografts. Dose optimization was examined in 2 tumor models.
  
  
  RESULTS
  TF12 internalizes, but a substantial fraction remained accessible on the tumor surface. Fluorescence-activated cell sorting analysis showed only a minor change in fluorescent signal when the tumor was probed with a fluorescent hapten-peptide over 4 h, and microscopy showed substantial membrane staining when reassessed at 24 h after TF12 exposure. Only 40.1% of (111)In-TF12 was internalized after 24 h. In vivo, excellent tumor localization of the (111)In-labeled peptide was observed in several tumor models.
  
  
  CONCLUSIONS
  TF12 was retained sufficiently on the cell surface in several epithelial cancers, thereby making it suitable for pretargeted imaging and therapy of various Trop-2-expressing carcinomas.
authors.affiliation: Garden State Cancer Center, Center for Molecular Medicine and Immunology, Morris Plains, New Jersey 07950, USA. rmsharkey@gscancer.org
authors.forename: Robert M
authors.initials: RM
authors.lastname: Sharkey
authors.forename: Catharina M
authors.initials: CM
authors.lastname: van Rij
authors.forename: Habibe
authors.initials: H
authors.lastname: Karacay
authors.forename: Edmund A
authors.initials: EA
authors.lastname: Rossi
authors.forename: Cathelijne
authors.initials: C
authors.lastname: Frielink
authors.forename: Celeste
authors.initials: C
authors.lastname: Regino
authors.forename: Thomas M
authors.initials: TM
authors.lastname: Cardillo
authors.forename: William J
authors.initials: WJ
authors.lastname: McBride
authors.forename: Chien-Hsing
authors.initials: CH
authors.lastname: Chang
authors.forename: Otto C
authors.initials: OC
authors.lastname: Boerman
authors.forename: David M
authors.initials: DM
authors.lastname: Goldenberg
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000276:immunology* / Q000378:metabolism / Q000493:pharmacokinetics; D000951:Antigens, Neoplasm / Q000276:immunology* / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000276:immunology* / Q000378:metabolism*; D045744:Cell Line, Tumor; D015972:Gene Expression Regulation, Neoplastic; D006241:Haptens / Q000378:metabolism; D006801:Humans; D007553:Isotope Labeling; D051379:Mice; D009375:Neoplasms, Glandular and Epithelial / Q000378:metabolism / Q000473:pathology*; D021381:Protein Transport
chemical_list: D018033:Antibodies, Bispecific; D000951:Antigens, Neoplasm; D015815:Cell Adhesion Molecules; D006241:Haptens; C499380:TACSTD2 protein, human