Figure 3. Adoptively transferred T cells are not fully functional in response to established tumors.

(A) FACS analysis comparing the accumulation of SIN-reactive, OT-I T cells in the mediastinal DLN and the inguinal peripheral lymph nodes (PLN) three days after adoptive transfer of naïve OT-I cells into Lenti-x or Lenti-LucOS tumor-bearing mice 10 weeks after tumor initiation (similar results observed four weeks after tumor initiation). Percent of total CD8⁺ cells that are anti-SIN/K^b+ is indicated. n≥ 3 mice for Lenti-LucOS or ≥1 mouse for Lent-x tumor-bearing mice in each of two independent experiments.

(B) Histogram plots of CD8⁺anti-SIN/K^b+ gated cells for CFSE dilution or surface expression of CD44, CD62L, or CD69 from the DLN or PLN of Lenti-LucOS (open histogram) or Lenti-x (filled histogram) tumor-bearing mice. Undivided (CFSE^hi) CD8⁺anti-SIN/K^b+ cells were examined in CD69 plots to detect initial T cell activation. n≥ 3 mice for Lenti-LucOS or ≥1 mouse for Lent-x tumor-bearing mice in each of two independent experiments.

(C) FACS analysis for the presence of OT-I T cells in the lungs of Lenti-LucOS tumor-bearing mice 12 days after adoptive transfer of naïve CD45.1⁺ OT-I T cells (tumor-bearing mice were CD45.2⁺). n≥ 3 mice in at least three independent experiments.

(D) Percent of CD8⁺CD45.1⁺ gated OT-I T cells producing IFNγ⁺TNFα⁺ or IFNγ⁺TNFα^neg from the DLN or lung after transfer of naïve OT-I T cells. WSN-SIN represents analysis of OT-I T cells 7 days after WSN-SIN infection. Lenti-LucOS represents analysis of OT-I T cells 12 days after transfer into Lenti-LucOS tumor-bearing mice between 12 and 16 weeks after tumor initiation. n= 6–7 mice per group.

(E) Percent of CD8⁺CD45.1⁺ gated OT-I T cells producing IFNγ⁺TNFα⁺ or IFNγ⁺TNFα^neg from the DLN or lung after transfer of in vitro-activated OT-I T cells. Lenti-LucOS represents analysis one day (lung only, n= 6 mice) and 12 days after transfer (n= 3 mice) into Lenti-LucOS tumor-bearing mice between 10 and 14 weeks after tumor initiation. WSN-SIN in the DLN represents analysis of naïve OT-I T cells 14 days after WSN-SIN infection (n= 3 mice).

Data are mean ± SEM. See also Figure S3.