Rituximab (RTX), an anti-CD20 antibody, revolutionized treatment for B-cell malignancies, but it is not without its own shortcomings, most notably tumor relapse. Recent research has provided evidence supporting the increased efficacy of RTX when combined with interleukin-15 (IL-15). IL-15 enhances antibody-dependent cellular cytotoxicity (ADCC), an important mechanism of RTX, by increasing the proliferation and activation of natural killer (NK) cells, as well as monocytes and macrophages. However, the majority of this evidence has been obtained through in vitro experiments and in vivo models using xenografts in immuno-deficient mice. Given the complexity of the immune system, we used an immuno-competent, syngeneic mouse model of human B-cell lymphoma to further investigate the effect of combining IL-15 with RTX to enhance ADCC. Wild-type (WT) C57BL/6 mice (n = 40) were distributed into treatment groups of 10 mice each, and inoculated intravenously with EL4-CD20 cells, a mouse lymphoma line transfected with human CD20. IL-15 was given five times per week for 4 weeks (5μg/mouse), starting on day 3 after tumor inoculation, and RTX was given once per week for 4 weeks (100 μg/mouse), starting on day 5. While IL-15 and RTX individually prolonged survival of the mice when compared with the control (p<0.05), the combination of IL-15 and RTX showed an additive effect, prolonging mouse survival significantly when compared with the monotherapies alone (p<0.01). At day 75 after tumor inoculation, 90% of the mice in the combination group were still alive. In comparison, only 30% of the mice in each of the IL-15 and RTX groups and none of the mice in the control group were alive. In contrast, treatment of EL4-CD20-bearing FcRγ-/- mice with RTX, under the same dose and dosing schedule as those used in WT mice, did not show a therapeutic effect when compared with the control. We further confirmed the role of ADCC by performing an ex vivo experiment using NK cells purified from splenocytes of either WT or FcRγ-/- mice treated with IL-15 five days before extraction. NK cells were cultured in triplicate with 51Cr-labeled EL4-CD20 cells with or without RTX and lysis was evaluated using a chromium-51 release assay. NK cells from IL-15-treated WT mice combined with RTX resulted in a three-fold increase in ADCC when compared with the monotherapies alone (16% combination vs. 6% IL-15 and 5% RTX, at a 10:1 E:T ratio). In contrast, NK cells from FcRγ-/- mice showed no difference in ADCC with or without RTX treatment. In agreement with previously published research, our study gives further evidence that IL-15 increases the efficacy of RTX, primarily through enhancing ADCC. Confirming this synergistic effect in an immuno-competent mouse model better supports a future study of RTX with IL-15 in human clinical trials for indolent B-cell lymphomas.

Citation Format: Bernard Wen, Meili Zhang, David Dilillo, Jeffrey V. Ravetch, Thomas A. Waldmann. Interleukin-15 enhances rituximab-dependent cytotoxicity ex vivo and in vivo against a mouse lymphoma expressing human CD20. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1332. doi:10.1158/1538-7445.AM2015-1332