title: Efficient Payload Delivery by a Bispecific Antibody-Drug Conjugate Targeting HER2 and CD63.
abstract: Antibody-drug conjugates (ADC) are designed to be stable in circulation and to release potent cytotoxic drugs intracellularly following antigen-specific binding, uptake, and degradation in tumor cells. Efficient internalization and routing to lysosomes where proteolysis can take place is therefore essential. For many cell surface proteins and carbohydrate structures on tumor cells, however, the magnitude of these processes is insufficient to allow for an effective ADC approach. We hypothesized that we could overcome this limitation by enhancing lysosomal ADC delivery via a bispecific antibody (bsAb) approach, in which one binding domain would provide tumor specificity, whereas the other binding domain would facilitate targeting to the lysosomal compartment. We therefore designed a bsAb in which one binding arm specifically targeted CD63, a protein that is described to shuttle between the plasma membrane and intracellular compartments, and combined it in a bsAb with a HER2 binding arm, which was selected as model antigen for tumor-specific binding. The resulting bsHER2xCD63hisdemonstrated strong binding, internalization and lysosomal accumulation in HER2-positive tumor cells, and minimal internalization into HER2-negative cells. By conjugating bsHER2xCD63histo the microtubule-disrupting agent duostatin-3, we were able to demonstrate potent cytotoxicity of bsHER2xCD63his-ADC against HER2-positive tumors, which was not observed with monovalent HER2- and CD63-specific ADCs. Our data demonstrate, for the first time, that intracellular trafficking of ADCs can be improved using a bsAb approach that targets the lysosomal membrane protein CD63 and provide a rationale for the development of novel bsADCs that combine tumor-specific targeting with targeting of rapidly internalizing antigens. Mol Cancer Ther; 15(11); 2688-97. ©2016 AACR.
authors: Genmab, Utrecht, the Netherlands. BGO@genmab.com.
authors: Bart E C G
authors: BE
authors: de Goeij
authors: Genmab, Utrecht, the Netherlands.
authors: Tom
authors: T
authors: Vink
authors: Genmab, Utrecht, the Netherlands.
authors: Hendrik
authors: H
authors: Ten Napel
authors: Genmab, Utrecht, the Netherlands.
authors: Esther C W
authors: EC
authors: Breij
authors: Genmab, Utrecht, the Netherlands.
authors: David
authors: D
authors: Satijn
authors: Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, the Netherlands.
authors: Richard
authors: R
authors: Wubbolts
authors: Concortis Biosystems Corp., San Diego, California.
authors: David
authors: D
authors: Miao
authors: Genmab, Utrecht, the Netherlands.|Department of Cancer and Inflammation Research, Institute of Molecular Medicine, University of Southern Denmark, Odense, Denmark.|Department of Immunohematology and Blood Transfusion, Leiden University Medical Center, Leiden, the Netherlands.
authors: Paul W H I
authors: PW
authors: Parren
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000008:administration & dosage* / Q000276:immunology / Q000493:pharmacokinetics; D000915:Antibody Affinity / Q000276:immunology; D000970:Antineoplastic Agents / Q000008:administration & dosage* / Q000493:pharmacokinetics; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004195:Disease Models, Animal; D065546:Drug Liberation; D005260:Female; D006801:Humans; D018796:Immunoconjugates / Q000008:administration & dosage* / Q000493:pharmacokinetics; D007140:Immunoglobulin Fab Fragments / Q000276:immunology / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D058990:Molecular Targeted Therapy; D011485:Protein Binding; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors*; D060149:Tetraspanin 30 / Q000037:antagonists & inhibitors*; D047368:Tumor Burden / Q000187:drug effects; D023041:Xenograft Model Antitumor Assays
chemical_list: D018033:Antibodies, Bispecific; D000970:Antineoplastic Agents; D018796:Immunoconjugates; D007140:Immunoglobulin Fab Fragments; D018719:Erb-b2 Receptor Tyrosine Kinases; D060149:Tetraspanin 30