Figure 1.

Improved tumor control and lymphocyte infiltration by addition of MEKi and PI3Ki to BRAFi. (A) C57BL/6 mice were injected subcutaneously with 3 × 10⁵ D4M.3A mouse melanoma cells and targeted therapies (14 d) were started after 7 d. BRAFi [B] PLX4720 was provided in chow; MEKi [M] trametinib was dosed by daily oral gavage at 15 µg (on average 0.75 mg/kg); PI3Ki [P] BKM120 daily oral gavage at 400 µg (on average 20 mg/kg); and mTORi [mT] everolimus daily oral gavage at 100 µg (on average 5 mg/kg). Shown are tumor growth curves from mice treated with the indicated combinations (mean ± SEM and n = 9–11). (B) Mean tumor size ± SD at day 28 (no mice removed from experiments) is depicted in a dot plot (Mann–Whitney U-test). (C) D4M.3A tumor-bearing C57BL/6 mice (each group n = 5) were treated for 3 d with MAPK and/or PI3K pathway inhibitors, and tumors were analyzed by immunohistochemistry for CD3⁺ cell infiltration and analyzed by flow cytometry for (D) proportion of IFNγ producing CD8⁺ T cells; (E) PD-L1 expression of CD45⁻ cells; and (F) expression of PD-1 on CD8⁺ T cells.