Rituximab-Mediated Antibody-Dependent Cellular Cytotoxicity Is Enhanced by Interleukin-15 or CpG Oligodeoxynucleotides.

Introduction

Antibody-dependent cellular cytotoxicity (ADCC) is an important mechanism in the clinical activity of rituximab for the treatment of B-cell malignancies. The NK cells, through the activating receptor FcγRIIIa, play a major role in the rituximab-mediated ADCC. We have studied the effect of NK stimulators such IL-15 and CpG oligodeoxynucleotides (ODN) in the enhancement of rituximab-mediated ADCC against B-cell lymphoma.

Methods

ADCC was performed by standard ⁵¹Cr release assays using peripheral blood mononuclear cells (PBMC) purified from 4 normal volunteers by Ficoll-Hypaque gradient centrifugation. Cells were cultured for 18 hours in the presence of human recombinant interleukin 15 (rhIL-15; 10 ng/ml) or CpG ODN (2,5 μg/ml). The B-lymphoma cell line Raji was used as target in the presence of rituximab (10 μg/ml) or human IgG1 as control (10 μg/ml) for 30 min at room temperature. Target cells and effector cells were incubated at different ratios ranging from 1:1 to 30:1, for 4 hours at 37°C. The polymorphism at amino acid 158 of the activating FcγRIIIa receptor (FcγRIIIa V→F) was analyzed in effector cells from normal donors with a polymerase chain reaction (PCR)-based allele-specific restriction analysis assay.

Results

Lysis of B lymphoma cells treated with rituximab alone was 40% ± 5% (mean ± SD) at the highest ratio, as compared to 12% ± 7% (p<0.001) in the presence of human IgG1 control. When the effector cells were treated with IL-15, a dramatic increase in the lysis of lymphoma cells was observed (70% ± 8% vs. 40% ± 5%, IL-15 plus rituximab vs. rituximab alone) (p<0.001). Effector cells treated with CpG ODN showed a significant increase in rituximab-mediated cytotoxicity (84% ± 2% vs. 53% ± 16%) (CpG ODN vs. control, respectively; p=0.03). The enhancement of rituximab-mediated ADCC when the effector cells were treated with IL-15 or CpG ODN was similar for both FcγRIIIa-V/V and FcγRIIIa-V/F genotypes on effector cells.

Conclusion

PBMC cells activated in the presence of IL-15 or CpG ODN become strong effectors in rituximab-mediated ADCC against B cell lymphoma. These results may be useful to improve the therapeutic activity of rituximab.