Figure 7.

Combined TMZ and TTFields efficacy correlates with treatment sequence and ARL13B⁺ cilia. (A, B) S7 parental or ARL13B KO (clone G12) cells were grown as spheres for 4 days and then treated with vehicle (veh) or 50 or 100 µM TMZ and exposed to an additional 3 days without (A) or with TTFields. Bar graphs show the normalized cell number (% of control) at the end of 7 days. (C, D) TMZ “pre”-experiment: S7 parental cells were treated with 100 µM TMZ for 3 days before a 24-h treatment of TTFields or control (con). Cells were counted immediately after this treatment (acute count), and then pooled, and 2.5 × 10⁴ cells/well were expanded in fresh media for 7 days in 24-well plates (n = 12 wells/group), and the fold expansion was calculated (fold count) (D). (E, F) TMZ “post”-experiment: TMZ (100 µM) was given for 3 days after a 24-h TTFields treatment with S7 cells being counted acutely (E) or expanded in fresh media for 7 days (F). (G, H) TMZ “with” experiment: S7 cells were grown for 6 days and then simultaneously treated with TMZ (100 µM) and TTFields for 24 h, counted acutely (G) or examined for fold expansion after 7 days (H). (I) Similar experiment as (F) but examining fold expansion of S7 ARL13B KO cells after 6 days. n.d., not determined; ns, not significant; ^* p < 0.05; ^** p < 0.01; ^*** p < 0.001; ^**** p < 0.0001 (ANOVA).