title: Visualization of effective tumor targeting by CD8+ natural killer T cells redirected with bispecific antibody F(ab')(2)HER2xCD3.
abstract: HER2 is an attractive immunotherapeutic target for neoplastic disease because this cell surface molecule is overexpressed on a large fraction of malignant tumor cells. To directly assess therapeutic responses to targeted therapy by noninvasive in vivo imaging in small animals, human HER2-expressing ovarian carcinoma cells were genetically modified with a firefly luciferase gene, and light emission was used for visualization of tumor growth and response to therapy. This imaging approach was able to demonstrate in real-time tumor regression in a HER2 xenograft mouse model by adoptive transfer of in vitro induced and expanded cytotoxic CD8+ natural killer T (NKT) cells retargeted with a humanized bispecific antibody F(ab')(2)HER2xCD3. Immunotherapy with effector cells alone or a humanized monoclonal antibody anti-p185(HER2) (4D5-8) resulted in significant but slower reduction in tumor burden. Long-term survival of tumor xenografts correlated inversely with visible residual tumor burden. In vitro, F(ab')(2)HER2xCD3 substantially augmented cytotoxic activity of CD8+ NKT cells. By flow-sorting, CD8+ NKT cells coexpressing CD56 were found to have the highest redirected killing ability. Treatment with concanamycin A or EGTA abrogated CD8+ NKT cytotoxicity indicating that perforin is a major pathway of tumor cell lysis. In contrast, when CD8+ NKT cell were cross-linked with F(ab')(2)HER2xCD3 neither the immunosuppressants cyclosporine A and FK506, nor the increase of intracellular cyclic AMP by dibutyryl cyclic AMP were able to inhibit cytotoxicity demonstrating that signaling via the CD3 antigen changes the biological activity of non-MHC-restricted effector cells. These studies have demonstrated that CD8+ NKT cells can be successfully redirected to tumor cells using bispecific antibodies and offer a promising strategy for adoptive immunotherapy of neoplastic diseases.
authors.affiliation: Division of Bone Marrow Transplantation, Stanford University School of Medicine, California 94305, USA.
authors.forename: Christian
authors.initials: C
authors.lastname: Scheffold
authors.forename: Martin
authors.initials: M
authors.lastname: Kornacker
authors.forename: Yolanda C
authors.initials: YC
authors.lastname: Scheffold
authors.forename: Christopher H
authors.initials: CH
authors.lastname: Contag
authors.forename: Robert S
authors.initials: RS
authors.lastname: Negrin
mesh_terms: D000818:Animals; D018033:Antibodies, Bispecific / Q000276:immunology*; D017252:CD3 Complex / Q000276:immunology*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology*; D000242:Cyclic AMP / Q000378:metabolism; D016572:Cyclosporine / Q000494:pharmacology; D003602:Cytotoxicity, Immunologic; D005260:Female; D006801:Humans; D016219:Immunotherapy, Adoptive / Q000379:methods*; D018799:Intercellular Adhesion Molecule-1 / Q000276:immunology; D007694:Killer Cells, Natural / Q000276:immunology*; D008163:Luminescent Measurements; D016169:Lymphocyte Function-Associated Antigen-1 / Q000276:immunology; D016393:Lymphoma, B-Cell / Q000276:immunology / Q000628:therapy; D008562:Membrane Glycoproteins / Q000276:immunology; D051379:Mice; D010051:Ovarian Neoplasms / Q000276:immunology / Q000628:therapy; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000276:immunology*; D016559:Tacrolimus / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
chemical_list: D018033:Antibodies, Bispecific; D017252:CD3 Complex; D000242:Cyclic AMP; D016572:Cyclosporine; D018799:Intercellular Adhesion Molecule-1; D016169:Lymphocyte Function-Associated Antigen-1; D008562:Membrane Glycoproteins; D054353:Perforin; D052899:Pore Forming Cytotoxic Proteins; D018719:Erb-b2 Receptor Tyrosine Kinases; D016559:Tacrolimus