Abstract 1485: E7011, a novel anti-angiogenesis inhibitor targeting Notch4 receptor signal pathway

Notch4 is selectively expressed on tumor endothelial cells and hypothesized to contribute to cancer malignancy through mechanisms including tumor angiogenesis regulation and promoting drug resistance. We developed E7011, a humanized IgG2κ antibody targeting Notch4 receptor signaling. To obtain anti-human Notch4 monoclonal antibody, human Notch4 3EGFR (epidermal growth factor repeat) domain - NRR (negative regulatory region) - SEAP (secreted alkaline phosphatase) - His protein was immunized to mice and 6698 hybridomas were obtained. By screening these clones with the Notch4 binding assay and Notch4-specific gal4-fusion luciferase reporter assay, mouse anti-human Notch4 mAb clone (6-3-A6) was selected. The CDRs (complementarity determining regions) of 6-3-A6 were grafted into the variable regions of human IgG2κ, and E7011, novel humanized anti-human Notch4 monoclonal antibody, was developed by screening in assays above. The biomolecular interaction analysis showed that E7011 is able to bind to human and mouse Notch4-NRR domain-SEAP-His, and inhibit human Notch4 receptor signaling in an antibody-dose dependent manner as well as 6-3-A6. IHC analysis showed that Notch4 was expressed on CD31 positive endothelial cells in Calu6 human non-small cell lung carcinoma xenograft tumor. Intravenous administration of E7011 at 1, 3, 10 mg/kg in twice a week significantly decreased tumor growth compared with control IgG treatment without severe body weight loss. We also investigated the functional tumor angiogenesis by hoechest dye-perfusion and staining with anti-CD31 antibody in the Calu6 xenograft model. E7011 treatment significantly decreased hoechst-positive perfusion area compared with vehicle treatment. In conclusion, E7011 is a novel antitumor agent that is a specific signal inhibitor targeting the Notch4 receptor and suppresses tumor growth through decreasing tumor vessel function in based on preclinical studies. Citation Format: Yusuke Adachi, Yoshimasa Sakamoto, Youya Nakazawa, Sho Tachino, Ken Ito, Takanori Abe, Yukinori Minoshima, Kana Hoshino-Negishi, Hideaki Ogasawara, Tomomi Kawakatsu, Miyuki Nishimura, Masashi Shimizu, Kazuhiro Tahara, Toshitaka Sato, Katsuhisa Suzuki, Kishan Agarwala, Masao Iwata, Yasuhiro Funahashi, Kenichi Nomoto, Yoichi Ozawa, Junji Matsui, Toshio Imai, Yu Kato. E7011, a novel anti-angiogenesis inhibitor targeting Notch4 receptor signal pathway [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Res