Figure 4. Regulation of BTN2A1 and Vγ9Vδ2 TCR-mediated killing of cancer cells.

(a, b) (a) Vγ9Vδ2 TCR tetramer staining MFI and (b) surface BTN2A1 MFI for Daudi-Cas9 KOs treated with 80 μM C991 (DMSO), DMSO (vehicle), 0.5 mM AICAR (aqueous), or water (vehicle) for 72 hours. (c) Survival of eGFP+ Daudi cells treated for 3 days with AICAR or water prior to co-culture (E:T 1:1) with primary Vγ9Vδ2 T cells, normalized to Daudi cells cultured without T cells. Measured by real-time quantitative live-cell imaging. (d) TEG-mediated killing of patient-derived cancer organoids and Daudi cells after 4 days of treatment with 10 μM pamidronate and AICAR, C991, or DMSO. Cancer cells were co-cultured with TEG001 or mock TEG-LM1 and 10 μM pamidronate for 2 days. (a) n=3 per condition, representative data from one of four independent experiments. Two-tailed unpaired Student’s t test. (b) n=3, representative data from one of two independent experiments. Two-tailed unpaired Student’s t test. (c) n=4 per condition, representative data from one of two independent experiments. Repeated measure two-way ANOVA with Geisser-Greenhouse correction. (d) n=4 per condition, data combined from two independent experiments (R4 organoid: n=2 per condition, data from one experiment, no statistical significance tests provided). One-way ANOVA comparison to the DMSO treatment condition with Dunnett’s multiple comparisons test. (a-d) Mean ± SD. p<0.0001 (****), p<0.001 (***), p<0.01 (**), p<0.05 (*).