HuBC1-IL12: A very potent immunocytokine which targets the oncofetal fibronectin in the extracellular matrix of tumor vasculature

Proc Amer Assoc Cancer Res, Volume 45, 2004 4686 IL-12 is a heterodimeric cytokine with potent anti-tumor activity, but its use in the clinic has been hampered by its severe systemic toxicity. Here we describe a fusion protein, huBC1-IL12, which specifically targets IL-12 to the tumor vasculature and therefore has therapeutic potential. HuBC1 is a humanized antibody that targets a cryptic sequence of the human ED-B-containing fibronectin isoform, B-FN, which is present in the subendothelial extracellular matrix (ECM) of most aggressive tumors. B-FN is one of the best tumor markers because it is oncofetal and angiogenesis-associated, and is undetectable in most normal adult tissues. The original murine BC1 antibody has been used successfully for immunoscintigraphy to image brain tumor mass in glioblastoma patients. In huBC1-IL12, each of the immunoglobulin heavy chains is genetically fused to the N-terminus of the IL-12 p35 subunit, which in turn is disulfide-bonded to the p40 subunit, resulting in a hexameric molecule of MW of ∼300 kilodaltons. Because of the interstitial pressure of a solid tumor, the ECM of the tumor neovasculature should be more accessible than the tumor cells themselves to an intravenously administered drug, making B-FN an ideal target for immunotherapy with a complex molecule such as huBC1-IL12. Since human IL12 has no biological activity in mice, we first produced huBC1-muIL12 as a surrogate molecule for animal tumor models. Despite the poor PK profile of this molecule in mice, one cycle of treatment with huBC1-muIL12 showed very potent anti-tumor activity in several xenogeneic models, such as PC3mm2, A431, and HT29 subcutaneous tumor models and PC3mm2 lung metastasis model, in SCID mice which lack T and B cells. A fusion protein that contains human IL12 (huBC1-huIL12), which is therefore a suitable molecule for investigation as a therapeutic, has also been produced. This protein has been shown to have a longer serum half-life than huBC1-muIL12 in mice, and retains both antigen binding and IL-12 activity in in vitro assays.