A requirement for calcium in the caspase-independent killing of Burkitt lymphoma cell lines by Rituximab

The therapeutic monoclonal antibody rituximab has previously been shown to kill B cells in a caspase-independent manner. The signalling pathways underpinning this novel death pathway are unknown. The present study showed that rituximab treatment of Burkitt lymphoma cell lines induced a slow rise in intracellular calcium (Ca²⁺]_i). This rise was only witnessed in cell lines that were killed by antibody, suggesting a critical role for Ca²⁺ in mediating rituximab-driven caspase-independent cell death. Inhibition of the two main intracellular store-located Ca²⁺ channels, i.e. the ryanodine and inositol-1,4,5-triphosphate receptor channels by dantrolene and xestospongen-c respectively did not prevent the rise in Ca²⁺ seen with rituximab or protect cells from subsequent death. In sharp contrast, inhibition of Ca²⁺ entry via plasma membrane channels with (2-aminoethoxy) diphenylborate or SKF-96365 or complete chelation of extracellular Ca²⁺ with ethyleneglycol bis (aminoethylether) tetra-acetate inhibited the rise in Ca²⁺]_i and increased cell viability. Together, these data suggest that ligation of the CD20 receptor with rituximab allows a slow sustained influx of Ca²⁺ from the external environment that under certain conditions can lead to cell death.