Abstract 540: AO-176, a normal cell sparing humanized anti-CD47 antibody
Authors: Prabir Chakraborty, Myriam N. Bouchlaka, Benjamin J. Capoccia, Ronald R. Hiebsch, Michael J. Donio, Robyn J. Puro, Vicki Sung, Daniel S. Pereira
Journal: Cancer Research (2019)

Abstract AO-176 is a next generation humanized anti-CD47 IgG2 that binds human and cynomolgus monkey CD47 equivalently. AO-176, like other CD47 antibodies, blocks the interaction of CD47 with SIRPα, inducing phagocytosis of tumor cells by activated macrophages. Unique to AO-176 is its ability to directly kill tumor cells via programmed cell death type III and immunogenic cell death in a cell autonomous (non-ADCC) manner. AO-176 also exhibits preferential binding to tumor vs. normal cells, a notable characteristic, as CD47 is expressed on many normal cells including red blood cells (RBC), platelets, T cells and endothelial cells. While other CD47 antibodies induce hematological toxicities such as anemia and thrombocytopenia in both primate models and in patients, AO-176 negligibly binds RBCs and platelets and even at high doses, minimally impacts hematology in cynomolgus monkey toxicology studies. AO-176 is currently being evaluated in phase 1 clinical trials for the treatment of solid tumors. In recent head to head experiments conducted with other CD47 antibodies, AO-176 bound all normal cells tested to a significantly lower degree. For example, unlike anti-CD47 antibodies such as Hu-5F9-G4, we observed negligible and minimal ex vivo binding to healthy human RBCs and platelets respectively and significantly lower binding of AO-176 to other hematologic and non-hematologic cells such as T cells and endothelial cells. Although AO-176 binds cynomolgus monkey RBCs to a slightly greater degree than human RBCs ex vivo, AO-176 versus comparative published Hu5F9-G4 findings has demonstrated dramatically reduced receptor occupancy when evaluated in monkey toxicology studies. We have also developed a clinic-ready receptor occupancy assay to measure AO-176 binding to circulating PBMCs in patients treated with AO-176 and to demonstrate minimal binding to CD47 on normal human cells. We have begun to evaluate mechanisms that may underlie the normal cell sparing effects of AO-176. Published studies have demonstrated that in addition to CD47 protein expression level, clustering and mobility of CD47 at the cell surface may impact binding to ligands, downstream signaling and subsequent cellular responses such as apoptosis. For example, avidity for SIRPα and clearance by macrophages is modulated by CD47 clustering that can be modulated by protein glycosylation, association with lipid rafts, cytoskeleton, integrins or other cis-acting factors. Here, we compare the distribution of CD47 on tumor and RBCs to evaluate receptor association with other cellular membrane components that may impact distribution on the cell surface. In summary, AO-176 is a next generation anti-CD47 antibody that, aside from induction of phagocytosis, possesses additional attributes that include direct tumor cell killing and preferential binding to tumor versus normal cells. These attributes of AO-176 differentiate it from other CD47 axis targeting agents currently in clinical evaluation. Citation Format: Prabir Chakraborty, Myriam N. Bouchlaka, Benjamin J. Capoccia, Ronald R. Hiebsch, Michael J. Donio, Robyn J. Puro, Vicki Sung, Daniel S. Pereira. AO-176, a normal cell sparing humanized anti-CD47 antibody [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 540.