title: Antitumor activity of a dual cytokine/single-chain antibody fusion protein for simultaneous delivery of GM-CSF and IL-2 to Ep-CAM expressing tumor cells.
abstract: Cytokine targeting to tumor-associated antigens via antibody cytokine fusion proteins has demonstrated potent antitumor activity in numerous animal models and has led to the clinical development of 2 antibody-interleukin-2 (IL-2) fusion proteins. We previously reported on the construction and in vitro properties of a "dual" cytokine fusion protein for simultaneous targeted delivery of human granulocyte macrophage-colony stimulating factor (GM-CSF) and IL-2 to human tumors. The fusion protein is based on a heterodimerized core structure formed by human CH1 and Ckappa domains (heterominibody) with C-terminally fused human cytokines and N-terminally fused single-chain antibody fragments specific for the tumor-associated surface antigen epithelial cell adhesion molecule (Ep-CAM). For testing the antitumor activity in syngeneic mouse xenograft models, we developed "dual cytokine heterominibodies" with murine cytokines (mDCH). mDCH fusion proteins and, as controls, "single cytokine heterominibodies" (SCH) carrying either murine GM-CSF (mGM-CSF) or murine IL-2 (mIL-2) were constructed, of which all retained the specific activities of cytokines and binding to the Ep-CAM antigen on human Ep-CAM transfected mouse colon carcinoma CT26-KSA cells. Over a 5-day treatment course, DCH fusion proteins induced significant inhibition of established pulmonary CT26-KSA metastases in immune-competent Balb/c mice at low daily doses of 1 mug of fusion protein per mouse. However, with the tested dosing schemes, antitumor activity of mDCH was largely independent of cytokine targeting to tumors as demonstrated by a control protein with mutated Ep-CAM binding sites. Single cytokine fusion proteins mSCH-GM-CSF and mSCH-IL-2 showed similar antitumor activity as the dual cytokine fusion protein mDCH, indicating that GM-CSF and IL-2 in one molecule did not significantly synergize in tumor rejection under our experimental conditions. Our results seem to contradict the notion that IL-2 and GM-CSF can synergize in antitumor activity and that with conventional dose regimens, their specific targeting to tumors, as tested here with 2 antibodies of different affinities, enhances their antitumor activity.
authors.affiliation: Micromet AG, Staffelseestrasse 2, 81477 Munich, Germany.
authors.forename: Juergen M
authors.initials: JM
authors.lastname: Schanzer
authors.forename: Iduna
authors.initials: I
authors.lastname: Fichtner
authors.forename: Patrick A
authors.initials: PA
authors.lastname: Baeuerle
authors.forename: Peter
authors.initials: P
authors.lastname: Kufer
mesh_terms: D000818:Animals; D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm / Q000096:biosynthesis* / Q000235:genetics; D000970:Antineoplastic Agents / Q000008:administration & dosage / Q000627:therapeutic use*; D015815:Cell Adhesion Molecules / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D006224:Cricetinae; D016503:Drug Delivery Systems; D000071858:Epithelial Cell Adhesion Molecule; D005260:Female; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000008:administration & dosage / Q000235:genetics*; D006801:Humans; D016233:Immunotherapy, Active; D007376:Interleukin-2 / Q000008:administration & dosage / Q000235:genetics*; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000276:immunology / Q000556:secondary; D051379:Mice; D008807:Mice, Inbred BALB C; D009368:Neoplasm Transplantation; D011993:Recombinant Fusion Proteins / Q000008:administration & dosage / Q000235:genetics / Q000627:therapeutic use*; D011994:Recombinant Proteins; D014183:Transplantation, Heterologous; D023041:Xenograft Model Antitumor Assays
chemical_list: D000912:Antibodies, Neoplasm; D000951:Antigens, Neoplasm; D000970:Antineoplastic Agents; D015815:Cell Adhesion Molecules; D000071858:Epithelial Cell Adhesion Molecule; D007376:Interleukin-2; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor