Doxorubicin enhances ¹³¹I-rituximab induced apoptotic cell death in Raji cells

Non-Hodgkin's lymphoma (NHL) is a group of hematological malignancies of which >90% is caused due to B cells. Although there are several therapeutic agents for NHL, they have their limitations for complete treatment. The objective of the study was to evaluate the enhancement in cell death when combining radioimmunotherapy with chemotherapy at optimum conditions thereby mitigating the limitations of each therapeutic modality. Raji cells which are CD20 positive Burkitt lymphoma, were treated with doxorubicin in combination with ¹³¹I-rituximab (anti CD20) and cell toxicity and underlying signaling pathways were elucidated. ¹³¹I-rituximab was prepared by iodogen method and purified by PD-10 column. Raji cells were grown in RPMI1640 complete media and seeded in 24 well plates at a density of 1 × 106 cells per well. Doxorubicin (0.5-10 μg/mL) was added in cultured cells and incubated for 4 h followed by addition of 1.85 MBq of ¹³¹I-rituximab for 2 h. Cells were harvested after 6 h, washed with PBS and further incubated for 12 h. Harvested cells were counted for cell death in trypan blue dye. Extent of apoptosis was evaluated by ELISA method and cell death signaling pathways were established. It was found that cellular toxicity was highest in Raji cells treated with doxorubicin (10 μg/mL) in combination with ¹³¹I-rituximab. However, apoptosis study showed that DNA fragmentation and PARP cleavage were highest in Raji cells treated with doxorubicin (2 μg/ mL) in combination with ¹³¹I-rituximab. Western blotting showed that downregulation of bcl-xl and cleavage of PARP protein was highest in the Raji cells treated with doxorubicin in combination with ¹³¹I-rituximab. It is concluded that doxorubicin has the potential to sensitize Raji cells by inducing cellular toxicity and apoptotic cell death induced by ¹³¹I-rituximab. (author)