Depletion of the C3 Component of Complement Enhances the Ability of Rituximab-Coated Target Cells to Activate Human NK Cells and Improves the Efficacy of Monoclonal Antibody Therapy in An in Vivo Model

Abstract

Antibody-dependent cellular cytotoxicity (ADCC) is felt to play an important role in mediating the anti-tumor effects of Rituximab (R). We previously reported that C3b deposition on R-coated target cells interferes with the binding of the Fc portion of R to NK cell CD16. This interaction inhibits the activation of NK cells and NK cell-mediated ADCC of 51Cr-labeled R-coated target cells. The current studies were designed to determine whether C3 depletion enhances the ability of mAb-coated targets to activate NK cells in vitro and improves mAb therapy in vivo. Normal human serum inhibited the ability of R-coated lymphoma cells to activate NK cells as previously reported. NK activation was increased when serum was pre-incubated with cobra venom factor (CVF) to deplete C3. Similar results were found when non-malignant ascites or transudative pleural fluid, as a surrogate for extravascular fluid, was used as the source of complement. For in vivo analysis, we utilized a syngeneic, immunocompetent murine model in which ADCC has been previously demonstrated to be a key mechanism of action. CVF or a human C3 derivative with CVF-like functions (HC3-1496) was used to deplete C3 in vivo. In this model, C3H/HeN mice were inoculated with murine 38C13 lymphoma cells (day 0) and treated with 2 doses of CVF or HC3-1496 (day 3 and day 5). Four hours after the initial dose of CVF or HC3-1496, mice were treated with a single dose of an anti-lymphoma mAb directed against the 38C13 idiotype (MS11G6). Untreated mice all developed tumor and died with a median survival of 28 days. All mice treated with mAb alone eventually developed tumor and died with a median survival of 42 days. Survival following treatment with CVF plus mAb was superior to that of mAb alone (Fig 1, p=0.0312) with 50% of mice remaining tumor free. Survival following treatment with HC3-1496 plus mAb was also superior to that of mAb alone (Fig 2, p=0.0002) with 80% of mice remaining tumor free. In summary, depletion of the C3 component of complement enhanced the ability of R-coated target cells to activate human NK cells, and improved the efficacy of mAb therapy in an in vivo model of lymphoma. Furthermore, these studies suggest the inhibitory effects of complement on NK activation and ADCC may be seen in the extravascular compartment such as within involved lymph nodes. We conclude that depletion of complement through use of agents such as CVF or HC3-1496 could be considered as an approach to enhancing the efficacy of R therapy. [Display omitted] [Display omitted]