Doxorubicin plus tumor necrosis factor α combination treatments in EL4-lymphoma-bearing C57BL/6 mice

The therapeutic efficacy of a total of 42 single-agent or combination protocols involving doxorubicin (Adriamycin, ADM) and tumor necrosis factor alpha (TNF alpha) were evaluated in the syngeneic murine lymphoma model, C57BL/6-EL4. Combination treatments were the most effective and the therapeutic effects were schedule-dependent; e.g. it was generally advantageous for ADM to precede TNF alpha administration. Two protocols selected for further study were 4 mg/kg ADM i.v. on days 1 and 8 plus TNF alpha i.v., at either 16000 U (7 mu g)/injection, on days 1 and 8 or 4000 U (1.7 mu g)/injection, on days 11-15. Survival of mice bearing one of four EL4 sublines having different in vitro drug sensitivities was assessed. These sublines were E10 (ADM-sensitive/TNF alpha-resistant), E16 (sensitive/sensitive), ER2 (ADM-resistant/TNF alpha-sensitive) and ER13 (resistant/resistant). Between 80% and 100% long-term survivors (i.e. tumor free on day 60) were obtained with the two treatments in mice bearing ADM-sensitive sublines, even though one of these sublines, E10, was resistant to TNF alpha in vitro. Induction of long-term survival appeared, therefore, to correlate with in vitro defined sensitivity/resistance to ADM, but not to TNF alpha. Treatment-induced modulations of tumoricidal immune effector functions were also examined. Taken together, the results indicated that induction of longterm survival involved complex interactions of: (1) ADM-induced tumor modifications, including, but not limited to, tumor debulking, (2) combination-treatment-induced modifications of splenic cytolytic T cell and macrophage activities, and (3) the restoration of thymus cellularity. Finally, when long-term survivors resulting from treatment of E10- or E16-bearing mice were implanted with ER2 on day 120, the majority survived, indicating that long-term immune memory, capable of recognizing drug resistant variants, had been established.