A chimeric humanized single-chain antibody against the type I insulin-like growth factor (IGF) receptor renders breast cancer cells refractory to the mitogenic effects of IGF-I.
Insulin-like growth factors (IGFs) stimulate breast cancer proliferation, motility, and survival. The type I IGF receptor (IGF1R) mediates the effects of IGF-I. Thus, inhibition of IGF1R activation could inhibit IGF action in breast cancer cells. A single-chain antibody directed against IGF1R (IGF1R scFv-Fc) has been shown to partially inhibit xenograft growth of MCF-7 cells in athymic mice. In this study, we have examined the effects of scFv-Fc on IGF1R signaling in the estrogen receptor-positive (ER+) MCF-7 breast cancer cells in vitro and in vivo. The antibody stimulated IGF1R activation in vitro in MCF-7 cells and was unable to block IGF-I effects. The antibody also stimulated proliferation of MCF-7 cells in monolayer growth assays. To determine how scFv-Fc could stimulate in vitro growth yet inhibit in vivo tumor growth, we examined the effect of scFv-Fc on IGF1R expression. In MCF-7 cells, scFv-Fc down-regulated IGF1R levels after 2 h, and the levels were greatly reduced after 24 h. In contrast, IGF-I treatment over the same time period did not affect IGF1R levels. Twenty-four-h pretreatment of cells with scFv-Fc blocked IGF-I mediated phosphorylation of insulin receptor substrate-1 and subsequent extracellular signal-regulated kinase 1/extracellular signal-regulated kinase 2 and phosphatidylinositol 3'-kinase activation. In contrast, cells treated with 5 nM IGF-I for 24 h still retained the ability to further activate downstream signaling pathways in response to IGF-I. Moreover, pretreatment of MCF-7 cells with scFv-Fc rendered them refractory to further proliferation induced by additional IGF-I. Twenty-four-h pretreatment of cells with scFv-Fc also inhibited IGF-I stimulated anchorage-independent growth. scFv-Fc did not enhance antibody-dependent cell-mediated cytotoxicity. In vivo, treatment of mice bearing MCF-7 xenograft tumors with scFv-Fc resulted in near complete down-regulation of IGF1R. Our data show that scFv-Fc stimulates biochemical activation of IGF1R, then causes receptor down-regulation, making MCF-7 cells refractory to additional IGF-I exposure. These results indicate that such chimeric single-chain antibodies against IGF1R have future potential in breast cancer therapy by causing down-regulation of receptor.
Sachdev
Deepali
D
Department of Medicine, University of Minnesota Cancer Center, Minneapolis, Minnesota 55455, USA.
Li
Shu-Lian
SL
Hartell
Julie S
JS
Fujita-Yamaguchi
Yoko
Y
Miller
Jeffrey S
JS
Yee
Douglas
D
D016475:3T3 Cells; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology / Q000494:pharmacology; D001943:Breast Neoplasms / Q000276:immunology / Q000628:therapy; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D015536:Down-Regulation / Q000187:drug effects; D005260:Female; D007128:Immunoglobulin Fragments / Q000276:immunology / Q000494:pharmacology*; D007074:Immunoglobulin G / Q000276:immunology / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D007334:Insulin-Like Growth Factor I / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D051379:Mice; D008819:Mice, Nude; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D017526:Receptor, IGF Type 1 / Q000819:agonists* / Q000276:immunology* / Q000378:metabolism / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D014407:Tumor Cells, Cultured; D023041:Xenograft Model Antitumor Assays
D000911:Antibodies, Monoclonal; D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D055504:Insulin Receptor Substrate Proteins; C526295:Irs1 protein, mouse; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; D007334:Insulin-Like Growth Factor I; D019869:Phosphatidylinositol 3-Kinases; D017526:Receptor, IGF Type 1; D020928:Mitogen-Activated Protein Kinases