Improving antibody-based immunotherapy protocols in vitro : Study on optimization of antitumor immunity by antibody-dependent phagocytosis with the monoclonal antibody rituximab

Sammanfattning: The monoclonal antibody Rituximab (RTX) has been widely and successfully used to treat non-Burkitt B cell lymphoma in human patients. The combination of immunotherapy with low-dose chemotherapy has increased the success of the treatment and translated into increased patients’ survival. On the other hand, many patients react differently to the chemotherapy, the antibody treatment, and the combination of the two, translating into a need for studies that evaluate strategies to improve response and enhance antibody-dependent phagocytosis by the patient’s immune cells. In the present study, I investigated separately four different lymphoma B cell lines grown in 3D tumor spheroid models which better represent cancer in vivo than the conventional in vitro 2D cell suspensions. These tumor models were treated with the monoclonal antibody Rituximab in the FDA approved IgG1 isotype form, with other non-approved isotypes and with a combination of them. The RTX-opsonized lymphoma models were then co-cultured with IFNγ activated monocytes, followed by an evaluation of the tumor cell killing efficacy by flow cytometry. The use of the chemotherapeutic agent staurosporine (RTX) in combination with RTX was also investigated. RTX isotype IgG3 proved to induce the highest percentage of phagocytosis of all the investigated tumor cell lines, both in 2D suspension and in 3D spheroid models. The phagocytosis efficacy increased when in combination with previous chemotherapeutic treatment with STR for all cell lines, with slight variation between them. We identified CD47 as a cell surface receptor that might have a role in this effect. CD47 is part of the CD47-SIRPalfa axis, used by somatic cells and cancer cells as a “don’t eat me” signal to prevent being engulfed by the body’s immune cells. We found a positive correlation between the expression of this surface receptor and the change in phagocytosis percentage when combined with apoptosis inducing agents. Our data matches previous works that studied the role of CD47 and how it becomes non-active, or hidden, on apoptotic cells. We suggest that apoptosis inducing agents should be paired with RTX immunotherapy for the treatment of tumors expressing large amounts of CD47 on their surface. The usage of CD47 as a prognostic biomarker in B-cell non-Hodgkin lymphomas (NHLs) has the potential to dramatically change how we use CD20-targeting antibodies and needs to be further investigated in future studies.