Daratumumab (DARA) is a human CD38 monoclonal antibody with broad-spectrum killing activity. DARA is in clinical development for multiple myeloma (MM) and has potential in other hematological tumors on which CD38 is expressed. In these clinical studies the adverse events have been manageable and marked reductions in paraprotein and bone marrow plasma cells have been observed. The killing activity of DARA on CD38-expressing tumor cells has previously been mainly attributed to complement-dependent cytotoxicity (CDC) and antibody-dependent cell-mediated cytotoxicity (ADCC).
In this study we explored whether DARA could also kill CD38-positive cells via induction of apoptosis after immobilization of DARA via Fc receptors (FcRs) in vitro. To this end, target cells were co-incubated in vitro with hFcgRI-expressing cells, lacking ADCC activity, in the absence or presence of DARA. Apoptosis was measured in flow cytometry for annexin-V/7AAD positivity. Treatment of the Burkitt's lymphoma cell line Ramos with DARA in combination with hFcgRI-expressing cells, led to a significant enhancement of annexin–V/7AAD positive cells (p<0.05 Dunn's multiple comparison test). Similar results were obtained with patient-derived multiple myeloma cell lines L363 and UM9, which are transduced with CD38 to obtain comparable expression levels to primary myeloma cells. This indicates that FcR-mediated crosslinking of DARA induces apoptosis which might contribute to the mechanism of action. We are currently performing experiments to investigate the contribution of DARA-mediated apoptosis in vivo in tumor growth inhibtion. Therefore, DARA efficacy is studied in a peritoneal syngeneic mouse model with EL4-CD38 cells, a mouse lymphoma cell line transfected with human CD38. The EL4-CD38 cells showed efficient apoptosis induction after FcR-mediated crosslinking of DARA in vitro. For this mouse model we will make use of NOTAM mice, which have a functionally inactive FcR associated gamma chain leading to cell membrane expression of all FcRs, but without signaling capacity (de Haij et al. Cancer Research 2010). Leucocytes of these mice are capable of antibody crosslinking via FcRs, but incapable of inducing cytotoxicity via ADCC.
In conclusion, in vitro data suggest that FcR-mediated crosslinking of DARA induces apoptosis of CD38 expressing tumor cells, and can thus be considered an additional mechanism of action for DARA.
Jansen:Genmab BV: Research Funding. Overdijk:Genmab BV: Employment. Lammerts van Bueren:Genmab BV: Employment. Parren:Genmab BV: Employment.
Asterisk with author names denotes non-ASH members.