ABSTRACT
Platelet-derived growth factor receptor α (PDGFRα) is a receptor tyrosine kinase that promotes cell survival and is expressed in both the tumor and the stromal components of human cancers. We have developed a fully human monoclonal antibody, MEDI-575, that selectively binds to human PDGFRα with high affinity, with no observable affinity for murine PDGFRα. To more fully characterize the role of PDGFRα in the regulation of tumor stroma, we evaluated the in vivo antitumor effects of MEDI-575 in tumor-bearing severe combined immunodeficient (SCID) mice and in genetically altered SCID mice expressing human PDGFRα in place of murine PDGFRα. We used the Calu-6 non-small cell lung cancer model because it lacks an in vitro proliferative response to PDGFRα activation. Antitumor activity was observed when the study was performed in mice expressing the human receptor, but no activity was observed in the mice expressing the murine receptor. Immunohistologic analysis of the tumors from mice expressing human PDGFRα showed a highly significant reduction in stromal fibroblast content and only minor changes in tumor proliferative index in tumors exposed to MEDI-575 compared with the results seen in vehicle-treated tumors or in tumors from mice expressing murine PDGFRα. Additional in vitro studies indicated that exposure of primary cancer-associated fibroblasts to MEDI-575 can directly affect proliferation and key signaling pathways in these cells. These results highlight the potential for observing antitumor activity with MEDI-575 through modulation of the stromal component of tumors and confirm that the PDGFRα pathway can play a role in maintaining a tumor microenvironment conducive to tumor growth.
Subject(s)
Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/pharmacology , Carcinoma, Non-Small-Cell Lung/pathology , Fibroblasts/drug effects , Lung Neoplasms/pathology , Receptor, Platelet-Derived Growth Factor alpha/antagonists & inhibitors , Animals , Antibodies, Monoclonal/therapeutic use , Antineoplastic Agents/therapeutic use , Carcinoma, Non-Small-Cell Lung/drug therapy , Carcinoma, Non-Small-Cell Lung/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Fibroblasts/metabolism , Fibroblasts/pathology , Humans , Lung Neoplasms/drug therapy , Lung Neoplasms/metabolism , Mice , Mice, Nude , Mice, SCID , NIH 3T3 Cells , Neoplasm Transplantation , Phosphorylation , Receptor, Platelet-Derived Growth Factor alpha/genetics , Stromal Cells/drug effects , Stromal Cells/metabolism , Stromal Cells/pathology , Transplantation, HeterologousABSTRACT
Localized angiopoietin-2 (Ang2) expression has been shown to function as a key regulator of blood vessel remodeling and tumor angiogenesis, making it an attractive candidate for antiangiogenic therapy. A fully human monoclonal antibody (3.19.3) was developed, which may have significant pharmaceutical advantages over synthetic peptide-based approaches in terms of reduced immunogenicity and increased half-life to block Ang2 function. The 3.19.3 antibody potently binds Ang2 with an equilibrium dissociation constant of 86 pmol/L, leading to inhibition of Tie2 receptor phosphorylation in cell-based assays. In preclinical models, 3.19.3 treatment blocked blood vessel formation in Matrigel plug assays and in human tumor xenografts. In vivo studies with 3.19.3 consistently showed broad antitumor activity as a single agent across a panel of diverse subcutaneous and orthotopic xenograft models. Combination studies of 3.19.3 with cytotoxic drugs or anti-vascular endothelial growth factor agents showed significant improvements in antitumor activity over single-agent treatments alone with no apparent evidence of increased toxicity. Initial pharmacokinetic profiling studies in mice and nonhuman primates suggested that 3.19.3 has a predicted human half-life of 10 to 14 days. These studies provide preclinical data for 3.19.3 as a potential new antiangiogenic therapy as a single agent or in combination with chemotherapy or vascular endothelial growth factor inhibitors for the treatment of cancer.
Subject(s)
Angiopoietin-2/immunology , Antibodies, Monoclonal/pharmacology , Antineoplastic Agents/pharmacology , Antineoplastic Combined Chemotherapy Protocols/pharmacology , Vascular Endothelial Growth Factors/antagonists & inhibitors , Xenograft Model Antitumor Assays , Animals , Antibodies, Monoclonal/administration & dosage , Antibodies, Monoclonal/pharmacokinetics , Antibodies, Monoclonal/therapeutic use , Antibody Specificity/drug effects , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/pharmacokinetics , Antineoplastic Agents/therapeutic use , Cell Death/drug effects , Collagen/metabolism , Drug Combinations , Humans , Laminin/metabolism , Mice , Neovascularization, Pathologic/drug therapy , Phosphorylation/drug effects , Primates , Protein Binding/drug effects , Proteoglycans/metabolism , Receptor, TIE-2/metabolismABSTRACT
Fusion proteins containing immunoglobulin Fc domains attached to bioactive moieties have been developed as therapeutic agents against several diseases. Here, we describe the development and characteristics of a novel fusion protein (FLSC R/T-IgG1) that targets CCR5, the major coreceptor for HIV-1 during primary infection. FLSC R/T-IgG1 was expressed from a synthetic gene that linked a single chain gp120-CD4 complex containing an R5 gp120 sequence with the hinge-CH2-CH3 portion of human immunoglobulin gamma subtype 1. Purified FLSC R/T-IgG1 exhibited a molecular mass of 189 kDa under reducing conditions, which matched the expected size of one polypeptide chain. Chemically crosslinked or untreated FLSC R/T-IgG1 exhibited a mass of a 360-kDa polypeptide under reducing and nonreducing conditions, which indicated that the molecule adopts a disulfide-linked bivalent structure. The chimeric molecule bound specifically to CCR5-expressing cells and to peptides derived from the CCR5 N-terminus. Such binding was more efficient than what was obtained with a monomeric single chain gp120-CD4 complex. FLSC R/T-IgG1 binding to CCR5 was blocked by preincubation of coreceptor-expressing cells with CCR5 ligands and by antibody to the coreceptor binding domain of gp120. Conversely, FLSC R/T-IgG1 blocked the binding of chemokine to CCR5. However, FLSC R/T-IgG1 did not trigger intracellular Ca2+ mobilization in peripheral blood mononuclear cells. FLSC R/T-IgG1 potently neutralized primary R5 HIV-1 in both a PBMC-based assay and cell line-based assays but did not affect the replication of X4 viruses. These findings suggest that FLSC R/T-IgG1 might be used as a possible therapeutic agent against HIV.