Fig. 4

Agonist immunotherapy treatment rescues inhibition of mouse T-cell function induced by MEK inhibition. Purified CD8⁺ or CD4⁺ murine T cells were stimulated with α-CD3 (1 µg/ml) and α-CD28 (0.5 µg/ml) antibodies and treated with vehicle (DMSO), 2A3 isotype, α-4-1BB or α-OX-40 antibody, trametinib alone, or combination of trametinib and agonist antibody. a, b Cell proliferation was measured by 3H-thymidine incorporation (added at 48 h) after 72 h of incubation with treatments. Proliferation in counts per minute (CPM) was measured for CD4⁺ and CD8⁺ T cells for a α-4-1BB antibody and b α-OX-40 antibody combinations. c, d IFNγ cytokine production (pg/ml) was measured from 72 h cultured supernatants via CBA analysis of c α-4-1BB antibody and d α-OX-40 antibody combinations in both CD8⁺ and CD4⁺ T cells, respectively. Experiments were performed in quadruplicate and is representative of 2–3 independent repeats. Controls groups are duplicated between panels as these experiments were performed concurrently. Data are presented as mean ± SEM. P-values represent one-way ANOVA and post hoc Fisher’s LSD tests. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001