513 PHST001, a humanized anti-CD24 antibody, induces phagocytosis of human tumor cells in vitro and tumor clearance in vivo

Background

CD24 is a small and highly glycosylated tumor antigen with a restricted expression profile that acts as an important macrophage 'don't eat me' signal. CD24 is highly expressed by many human cancers, including ovarian and breast, and it interacts with the macrophage receptor Siglec10 on M2 macrophages to protect cancer cells from being engulfed by tumor-associated macrophages. PHST001 is a humanized anti-CD24 monoclonal antibody that binds to CD24 on the surface of cells with high affinity and specificity, blocking the interaction of CD24 with Siglec10 on macrophages.

Methods

In vitro phagocytosis: tumor cells engineered to express GFP were co-cultured with M2 macrophages for 2 hours. Phagocytosis was measured by flow cytometry, as the number of CD11b+/GFP+ macrophages, quantified as a percentage of the total CD11b+ macrophages. In vitro cytokine release: whole blood from 10 healthy human donors were treated with PHST001 or positive control antibodies (anti-CD3, anti-CD28 or anti-CD52). All antibodies were tested in soluble (5ug/mL) and plate-bound (100 ug/mL) forms. In vivo efficacy: Immunocompromised mice were implanted subcutaneously with luciferase-expressing MFM223 triple negative breast cancer or BT474 Her2+ breast cancer cells. Once tumors were detected, mice were treated with isotype control or PHST001 intraperitoneally at different concentrations, three times a week for 18 doses. Tumor volume was assessed once a week by bioluminescence.

Results

PHST001 is an optimally humanized antibody that binds specifically to human CD24 (figure 1). PHST001 binds CD24 on the surface of cancer cell lines from multiple indications, including Breast, Ovarian, Colorectal, Pancreatic, and Lung with high affinity (figure 2). PHST001 promotes macrophage-induced phagocytosis of Breast, Ovarian, Colorectal and Pancreatic cancer cell lines in vitro (figure 3). PHST001 treatment significantly inhibited the growth of MFM223 triple negative breast cancer and BT474 Her2+ breast cancer in vivo (figures 4 and 5). PHST001 does not induce release of IL6, TNFa, IL2 or IFNg by human PBMCs in vitro (figure 6). PHST001 detects CD24 in normal and cancer tissues by IHC.

Conclusions

PHST001, a humanized anti-CD24 monoclonal antibody, binds with high affinity to CD24 on the surface of cancer cells from multiple indications, inducing in vitro phagocytosis of cancer cells by macrophages and tumor growth control in vivo. PHST001 does not induce cytokine release in vitro and detects CD24 in human tissues. A clinical study to investigate the safety and efficacy of PHST001 in cancer patients is being developed.