NK cells activated by monoclonal antibody-coated target cells enhance bispecific antibody-induced T cell proliferation, activation and cytotoxicity

Background: We previously demonstrated that T cell help can enhance the viability and cytotoxic potential of Natural Killer (NK) cells activated by monoclonal antibody (mAb)-coated target cells. Here we evaluated whether the reverse is also true, and that NK cells coated by mAb can enhance proliferation, activation, and the cytotoxic potential of bispecific antibody (bsAb) retargeted T cells. Methods: Raji lymphoma cells and normal donor peripheral blood mononuclear cells depleted of NK cells were mixed at an effector to target ratio of 5:1. Autologous NK cells were added back in controlled concentrations (0%, 5% or 20% of total effector cells). Co-cultures were treated with anti-CD3×CD19 bispecific antibody (blinatumomab), anti-CD20 monoclonal antibody (rituximab) or both. Media and antibodies were refreshed on days 2 and 4. After 5 days, T cell proliferation (Ki67) and production of cytotoxic molecules (including granzyme B) was assessed as was the number of remaining Raji cells as a measure of cytotoxicity. Results: Treatment with the combination of blinatumomab and rituximab enhanced T cell proliferation and activation when larger numbers of NK cells were present. The presence of NK cells also enhanced elimination of Raji cells. The impact of NK cells on T cell proliferation and activation in response to single agent blinatumomab or rituximab was less pronounced. Similar results were seen with other mAb and bsAb combinations. Conclusion: NK cells, activated by mAb-coated tumor cells, promote the proliferation, activation and cytotoxic capacity of bsAb-retargeted T cells. These data provide evidence that NK cells activated by mAb and T cells activated by bsAb are synergistic and provide cross help to each other. This provides additional rationale for evaluating therapeutic strategies involving concurrent administration of mAb and bsAb that allows for simultaneous co-engagement of NK cells (via mAbs) and T cells (via bsAbs). Number T cells (×10 3 ) No Ab Rituximab Blinatumomab Rituximab & Blinatumomab 0% NK 54±13 57±16 56±62 77±74 5% NK 56±20 68±34 60±64 153±137 20% NK 67±39 92±34 81±86 212±147 T cell Granzyme B 0% NK 96±204 128±351 2328±1534 3660±1277 5% NK 265±500 1412±834 4508±2754 11409±5077 20% NK 1043±1491 6882±2830 10876±3252 27083±9834 Remaining Raji cells (×10 3 ) 0% NK 314±72 230±82 184±26 145±27 5% NK 259±84 93±80 174±20 50±79 20% NK 241±68 21±35 124±11 9±15