Cellular internalization and mechanism of cytotoxicity of 131 I-Rituximab in raji cells

摘要

Rituximab labeled with radioiodine (131I-rituximab) has a large potential to be employed for targeted therapy of non-Hodgkin's lymphoma. Studies of parameters such as cellular internalization, stability of 131I-rituximab bound to CD20 receptor of tumor cells, and the mechanism underlying cytotoxicity induced by 131I-rituximab will be useful for better clinical application. In this article we describe the efficacy of 131I-rituximab in CD20-expressing Raji cells. Rituximab labeled with 131I was purified on a PD-10 column and characterized using high-performance liquid chromatography and paper electrophoresis. Raji cells treated with 131I-rituximab (1.85 MBq for 2 hours) were washed then incubated. The culture medium collected from treated cells showed increased radioactivity over a longer period (6 hours), probably due to the deiodination/degradation of 131I-rituximab. The tumor cells treated with 131I-rituximab showed time-dependent internalization of radioactivity, and at 12 hours the radioactivity was almost equally distributed in the membrane and cytoplasm. At 24 hours ~70% of the radioactivity was internalized. Cellular toxicity after 131I-rituximab treatment showed a time-dependent increase in toxicity as estimated by lactate dehydrogenase. Tumor cells treated with 131I-rituximab showed signifcantly higher toxicity and apoptosis compared with the those treated with the same concentration of unlabeled rituximab. The increased apoptotic death in cells treated with 131I-rituximab was associated with cleavage of poly ADP ribose polymerase and upregulation of p53 protein. This study provides a deeper understanding about the cellular internalization/stability of 131I-rituximab bound to the CD20 receptor and its efficacy in killing Raji cells.