Long-lasting anti-tumor immunity induced by the rejection of donor hematopoietic cells in mixed bone marrow chimeras

BACKGROUND Surprisingly, some patients who reject donor marrow grafts following nonmyeloablative HCT sustain remissions of advanced hematological malignancies (Br J Haematol2005, 128:351). In murine mixed chimeras prepared with nonmyeloablative conditioning, we previously showed that recipient leukocyte infusions (RLI) induced loss of donor chimerism and anti-tumor responses against host-type tumors (Blood2003, 102:2300) and that T cells in the RLI are necessary to achieve maximum anti-tumor effects (J Immunol2005, 175:665). We now examined whether or not this anti-tumor response demonstrates long-term memory. We attempted to enhance anti-tumor effects by adding exogenous IL-15, which promotes CD8 memory T cell survival. METHODS Mixed chimerism was achieved in BALB/c (H-2 d ) mice conditioned with depleting anti-CD4 and CD8 mAbs on Day -5, cyclophosphamide 200 mg/kg on Day -1 and 7 Gy thymic irradiation on Day 0 prior to transplantation of 25x10 6 B10.BR (H-2 k ) or B10.RIII (H-2 r ) bone marrow cells. Some groups received RLI (3x10 7 BALB/c spleen cells) seven weeks post-BMT. A20 cells (5x10 5 ) were given i.v. one week after RLI. Some groups received a secondary A20 challenge (10 5 ) 100 days after the first challenge. Some groups received 5 μg of IL-15 intraperitoneally, starting one week after RLI injection and every 12 h thereafter for a total of 10 doses. Spleen cells were isolated and anti-tumor cytotoxicity was evaluated after 5 days’ coculture with irradiated A20 cells. RESULTS We rechallenged RLI-treated long-term tumor survivors with a second lethal dose of A20 tumor cells 100 days after the first tumor challenge. One third of long-term survivors (n=20) rejected the lethal dose of A20 (median survival time [MST] 72 days). In contrast, all mice in the control group that did not previously receive RLI and received their first tumor challenge concurrently with the rechallenge group died, with MST 56 days (n=23) (p=0.02). Thus, 1/3 of long-term tumor survivors retained sufficiently strong anti-tumor immunity to reject secondary tumor. We have previously demonstrated in vitro tumor-specific cytotoxic responses 11 weeks after first tumor challenge. We now examined anti-tumor cytotoxicity at a later time point and observed such responses even 11 months after first tumor challenge in some of the tumor survivors. From these results, we speculated that memory CTL may play an important role in the anti-tumor effect of RLI. We hypothesized that exogenous IL-15 administration might enhance the anti-tumor effect of RLI, since it promotes generation and maintenance of memory T cells. Indeed, we observed increased tumor-free survival in mice that received RLI and IL-15 (n=14) compared to recipients of RLI alone (n=14) (MST 64 versus 48 days respectively, p=0.03). CONCLUSION Together, these data suggest that RLI therapy can evoke long-term anti-tumor immunity and that the anti-tumor response may be enhanced by administration of exogenous IL-15, reinforcing the potential of RLI therapy to provide a new HCT strategy lacking the risk of graft-versus-host disease.