Recent data suggest that monocytes are the dominant effector cells during immunotherapy using the anti-CD20 monoclonal antibody rituximab, depleting B cells through FcγR-dependent pathways. Because monocyte colony-stimulating factor (M-CSF) enhances the antibody-dependent cellular cytotoxicity (ADCC) of monocytes, the clinical efficacy of rituximab might be improved by the addition of M-CSF. We have studied the effect of M-CSF in the enhancement of rituximab-mediated ADCC against B-cell lymphoma.
Monocytes were isolated by negative selection of PBMCs from healthy individuals for the absence of T-cell, B-cell, and NK-cell markers. Cytotoxicity was determined by a flow cytometry using two fluorescent dyes, calcein-AM and ethidium homodimer to specifically stain living and dead cells respectively. Monocytes were cultured for 48 hours in the presence or absence of human recombinant M-CSF (66 ng/ml). The B-cell lymphoma cell line Daudi was used as target in the presence of rituximab (5 μg/ml) or human IgG1 as control for 30 min at room temperature. Effector cells and target cells were incubated at different ratios ranging from 1:1 to 15:1 for 4 hours at 37°C. The expression of FcγRl, FcγRII, and FcγRIII on monocytes was determined using a flow cytometry.
Monocytes treated with M-CSF showed a significant increase in rituximab-mediated cytotoxicity against B lymphoma cells: specific lysis at an E:T ratio of 15:1 was 39% ± 7% (mean ± SD) vs. 21% ± 5%, M-CSF-treated monocytes vs. non-treated monocytes. Lysis of lymphoma cells treated with rituximab alone was 8% ± 4%. Treatment with M-CSF led to a 1.5- to 2.0-fold increase of FcγRI and FcγRIII expression in monocytes, while FcγRII expression remained unchanged.
Pretreatment of monocytes with M-CSF enhances their rituximab-mediated ADCC against B-cell lymphoma, which may partly result from increasing expression of FcγRI and FcγRIII on monocytes via M-CSF stimulation. These in vitro results may provide a new approach to improve the therapeutic activity of rituximab.
Disclosure: No relevant conflicts of interest to declare.