American Society of Hematology

Figure 3.

$Figure 3. Increased transfer of engineered CD4+ T cells enhances tumor-free survival of mice. (A) Groups of 10 scid mice were given intravenous injections of 5 × 106 MDA-MB-435-erbB2 cells at day 0 prior to intravenous injection of scFv-CD28-ζ–transduced T cells at day 5. Mice treated with a 1:1 ratio of CD4+-transduced (5 × 106) and CD8+-transduced (5 × 106) T cells (▪) demonstrated 100% survival compared to 30% survival of mice receiving unfractionated transduced T cells (□; *P < .005, Mann-Whitney test). No survival of mice was observed for those receiving CD4+-transduced T cells alone (▪), CD8+-transduced T cells alone (○), control transduced T cells (▵), or no T cells (▴). (B) Enhanced survival of mice was due to increased transfer of antigen-specific CD4+ T cells. All scid mice bearing 5-day MDA-MB-435-erbB2 tumor that received a 1:1 ratio of CD8+-transduced (5 × 106) and CD4+-transduced (5 × 106) T cells (▪) survived compared with mice receiving either a 1:1 combination of transduced CD8+ T cells (5 × 106) and naïve CD4+ T cells (5 × 106; □), 1:1 combination of transduced CD8+ (5 × 106) T cells and CD4+ T cells (5 × 106) transduced with an irrelevant scFv-anti-CEA-γ receptor (▵) or no T-cell treatment (▴; *P < .001, Mann-Whitney test). (C) Transfer of a greater number of transduced CD8+ T cells alone does not rescue all mice from disease. Mice were given intravenous injections of 5 × 106 MDA-MB-435-erbB2 cells at day 0 prior to intravenous injection of scFv-CD28-ζ–transduced T cells. Mice treated at day 5 with a single dose of CD4+-transduced (5 × 106) and CD8+-transduced (5 × 106) T cells (▪) in a 1:1 ratio demonstrated 100% survival compared to about 30% survival for mice treated at days 5 and 6 with transduced CD8+ T cells alone (107/injection; ○)or about 80% survival for mice treated at days 5 and 6 with transduced unfractionated T cells (107/injection; □). Mice receiving control T cells (▵) did not survive. The difference between groups of mice receiving a 1:1 ratio of transduced CD8+ and CD4+ T cells and transduced CD8+ T cells alone was significant (*P < .01, Mann-Whitney test). (D) To determine the optimal ratio of transduced CD4+ and CD8+ T cells required to achieve 100% survival of mice bearing 5-day established MDA-MB-435-erbB2 lung metastases, mice were treated with transduced CD8+ and CD4+ T cells at the following ratios: 1:1 (5 × 106 CD8+,5 × 106 CD4+ T cells; □), 9:1 (9 × 106 CD8+, 1 × 106 CD4+ T cells; ▴), 3:1 (7.5 × 106 CD8+, 2.5 × 106 CD4+ T cells; ▵), 1:3 (2.5 × 106 CD8+, 7.5 × 106 CD4+ T cells; ▪), or 1:9 (1 × 106 CD8+, 9 × 106 CD4+ T cells; ○). Control mice were treated with mock-transduced CD4+ and CD8+ T cells at a 1:1 ratio (▪; *P < .05, **P < .001, Mann-Whitney test). All results are calculated as the percentage of each group surviving and are representative of 2 experiments performed. Arrows depict days of T-cell transfer.$

Increased transfer of engineered CD4⁺ T cells enhances tumor-free survival of mice. (A) Groups of 10 scid mice were given intravenous injections of 5 × 106 MDA-MB-435-erbB2 cells at day 0 prior to intravenous injection of scFv-CD28-ζ–transduced T cells at day 5. Mice treated with a 1:1 ratio of CD4⁺-transduced (5 × 106) and CD8⁺-transduced (5 × 106) T cells (▪) demonstrated 100% survival compared to 30% survival of mice receiving unfractionated transduced T cells (□; *P < .005, Mann-Whitney test). No survival of mice was observed for those receiving CD4⁺-transduced T cells alone (▪), CD8⁺-transduced T cells alone (○), control transduced T cells (▵), or no T cells (▴). (B) Enhanced survival of mice was due to increased transfer of antigen-specific CD4⁺ T cells. All scid mice bearing 5-day MDA-MB-435-erbB2 tumor that received a 1:1 ratio of CD8⁺-transduced (5 × 106) and CD4⁺-transduced (5 × 106) T cells (▪) survived compared with mice receiving either a 1:1 combination of transduced CD8⁺ T cells (5 × 106) and naïve CD4⁺ T cells (5 × 106; □), 1:1 combination of transduced CD8⁺ (5 × 106) T cells and CD4⁺ T cells (5 × 106) transduced with an irrelevant scFv-anti-CEA-γ receptor (▵) or no T-cell treatment (▴; *P < .001, Mann-Whitney test). (C) Transfer of a greater number of transduced CD8⁺ T cells alone does not rescue all mice from disease. Mice were given intravenous injections of 5 × 106 MDA-MB-435-erbB2 cells at day 0 prior to intravenous injection of scFv-CD28-ζ–transduced T cells. Mice treated at day 5 with a single dose of CD4⁺-transduced (5 × 106) and CD8⁺-transduced (5 × 106) T cells (▪) in a 1:1 ratio demonstrated 100% survival compared to about 30% survival for mice treated at days 5 and 6 with transduced CD8⁺ T cells alone (107/injection; ○)or about 80% survival for mice treated at days 5 and 6 with transduced unfractionated T cells (107/injection; □). Mice receiving control T cells (▵) did not survive. The difference between groups of mice receiving a 1:1 ratio of transduced CD8⁺ and CD4⁺ T cells and transduced CD8⁺ T cells alone was significant (*P < .01, Mann-Whitney test). (D) To determine the optimal ratio of transduced CD4⁺ and CD8⁺ T cells required to achieve 100% survival of mice bearing 5-day established MDA-MB-435-erbB2 lung metastases, mice were treated with transduced CD8⁺ and CD4⁺ T cells at the following ratios: 1:1 (5 × 106 CD8⁺,5 × 106 CD4⁺ T cells; □), 9:1 (9 × 106 CD8⁺, 1 × 106 CD4⁺ T cells; ▴), 3:1 (7.5 × 106 CD8⁺, 2.5 × 106 CD4⁺ T cells; ▵), 1:3 (2.5 × 106 CD8⁺, 7.5 × 106 CD4⁺ T cells; ▪), or 1:9 (1 × 106 CD8⁺, 9 × 106 CD4⁺ T cells; ○). Control mice were treated with mock-transduced CD4⁺ and CD8⁺ T cells at a 1:1 ratio (▪; *P < .05, **P < .001, Mann-Whitney test). All results are calculated as the percentage of each group surviving and are representative of 2 experiments performed. Arrows depict days of T-cell transfer.

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