IL-2 administered in vivo after complex with anti-IL-2 mAb effectively expanded CD8⁺ T lymphocytes and augmented antitumor immunity induced by vaccination with tumor cells secreting gp96-Ig in lymphoma-bearing syngeneic HSCT recipients. Transplants, tumor inoculation, and treatments were performed as in Figure 4, and mice received IL-2_S4B6 1 day following each vaccination as indicated. (A) IL-2_S4B6 treatment induced robust expansion of CD8⁺ T cells in vaccinated HSCT recipients with lymphoma during the first 3 weeks after HSCT. CD8⁺ T-cell frequency in the peripheral blood; n = 20 from a pool of 4 experiments; x, no vaccine; ♦, E.G7; ●, E.G7-gp96-Ig; ■, E.G7-gp96-Ig + IL-2_S4B6. (B) Tumor-reactive CD8⁺ T lymphocytes markedly expanded at the vaccine site and other lymphoid tissues of lymphoma-bearing HSCT recipients 5 days following 3 treatments with IL-2_S4B6 and gp96-Ig vaccine. Tumor-reactive CD8⁺ T-cell number in the peritoneal cavity, spleen, and draining LNs; n = 2. (C) Combination therapy with vaccination and IL-2_S4B6 increased MST and overall survival of syngeneic HSCT recipients with lymphoma. Noncomplexed, unbound IL-2 in combination with vaccination failed to enhance survival of lymphoma-bearing HSCT recipients; n = 5; ●, E.G7-gp96-Ig; ♦, E.G7-gp96-Ig + IL-2; ■, E.G7-gp96-Ig + IL-2_S4B6. (D) IL-2_S4B6 in the absence of vaccination marginally enhanced survival of lymphoma bearing HSCT recipients; n = 5; x, IL-2_S4B6; ●, E.G7-gp96-Ig; ■, E.G7-gp96-Ig + IL-2_S4B6. (E) MST and overall survival of T cell-replete syngeneic HSCT recipients with lymphoma following vaccination and IL-2_S4B6 was independent of transgenic antigen-specific CD8⁺ T-cell presence; n = 5; ●, E.G7-gp96-Ig; ○, E.G7-gp96-Ig (no OT-I); ■, E.G7-gp96-Ig + IL-2_S4B6; □, E.G7-gp96-Ig + IL-2_S4B6 (no OT-I). (F) CD8⁺ effector T-cell numbers at the vaccine site were markedly increased only following vaccine and IL-2_S4B6 treatment. CD8⁺ T_eff (CD62L⁻ CD44⁺), including endogenous non-TCR transgenic DimerX−K^b−OVA_257-264⁺, cell numbers in the peritoneal cavity; n = 5. (G) Combined EL4-gp96-Ig and IL-2_S4B6 vaccination strategy prolonged MST in EL4-bearing HSCT recipients; n = 5; x, no vaccine; ♦, IL-2_S4B6; ●, EL4-gp96-Ig; ■, EL4-gp96-Ig + IL-2_S4B6.