Cell - 8 September 2016

Figure S6. PDA-InfiltratinggdT Cells InhibitabT Cells but Exhaustion Ligand Blockade Is Tumor Protective and Activates CD8+T Cells ingdT
Cell-Competent Hosts, Related toFigures 5and 7
(A–E) SplenicabT cells from untreated WT mice were cultured in 96 well plates either unstimulated, or stimulated withaCD3/aCD28 alone or in co-culture with
PDA-infiltratinggdT cells (5:1 ratio) orgdT cell conditioned media. The fraction of CD4+and CD8+T cells (A and B) adopting a CD62L–CD44+phenotype and (C and
D) expressing IFN-gwas determined at 72h by flow cytometry. (E) Cytokine expression was also determined by analysis of cell culture supernatant. Experiments
were repeated twice (n = 4/group).
(F and G) WT and Tcrd–/–mice were orthotopically implanted with KPC-derived tumor cells and serially treated withaPD-L1 oraGalectin-9 neutralizing mAbs or
respective isotype controls. Pancreatic tumors were harvested at 3 weeks, and CD8+T lymphocytes from each cohort were analyzed by flow cytometry for
expression of (F) T-bet and (G) TNF-a.
(H) Cohorts of 6 week old KC;TCRd+/+and KC;TCRd–/–mice were serially treated withaPD-L1 or isotype control for 8 weeks and pancreata were harvested at
14 weeks. Comparative tumor weights are shown (n = 5/group). Controls were previously shown inFigure 3B.
(I and J) Cohorts of WT mice were orthotopically implanted with KPC-derived tumor cells. In parallel, Tcrd–/–mice were similarly treated but tumor cells were co-
injected with FACS-sorted PDA-infiltratinggdT cells that were treated ex-vivo with either (I) Rat IgG2b isotype, or (J)aPD-L1. PDA tumors were measured at
21 days (n = 4/group).
(K–M) Tcrd–/–mice were subcutaneously implanted with KPC-derived tumor cells engineered to express OVA. On day 10, tumors were directly inoculated with
PBS, FACS-sorted PDA-infiltratinggdT cells treatedex-vivowith Rat IgG2b, or PDA-infiltratinggdT cells treated withaPD-L1. On day 15 (K) tumor volume (scale
bar = 1cm), (L) the fraction of CD8+OVA Pentamer+T cells among all CD8+T cells, and (M) OVA Pentamer+T cell expression of CD107a were recorded (n =
5/group;p < 0.05, p < 0.01, p < 0.001).