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. 2024 Aug 28;5(9):101700. doi: 10.1016/j.xcrm.2024.101700

Figure 5.

Figure 5

Gemcitabine deactivates SHH signaling in SHH-ATRTs through loss of SIRT1, increasing SHH-ATRT-specific drug sensitivity

(A) t-SNE clustering of 49 individual tumor samples of patients with ATRT (dataset GSE: 70678) based on overall mRNA expression profiles (perplexity: 12). SHH subgroup is shown as a distinct group from other ATRT samples in upper-left corner as confirmed by overall high GLI2 expression (blue to red) versus low GLI2 expression in the non-SHH ATRT (yellow to green).

(B) mRNA expression levels of SIRT1 in SHH ATRT (n = 16) versus non-SHH ATRT (n = 33) (Wilcoxon rank-sum: p = 0.028).

(C) Immunofluorescent stainings of GLI2 (green), α-tubulin (red), and DAPI (blue) in VUMC-ATRT-03 cells treated with 20 nM gemcitabine for 24 h show the loss of nuclear localization of GLI2. Upper: an average depiction of all wells. Lower: zoomed depictions of the indicated area in the upper panels.

(D) Quantification of percentage nuclear (DAPI) overlap with GLI2 between DMSO and gemcitabine-treated VUMC-ATRT-01 (n = 20) and VUMC-ATRT-03 cells (n = 20) (one-way ANOVA: ∗∗∗∗p < 0.0001).

(E and F) Quantification of total GLI2-positive nuclei per well between DMSO and gemcitabine-treated VUMC-ATRT-01 (n = 20) and VUMC-ATRT-03 cells (n = 20) (one-way ANOVA: ∗∗∗∗p < 0.0001). (F) Illustration of the proposed mechanism through which gemcitabine causes tumor toxicity in ATRT, including the mechanisms that cause extra sensitivity in SHH-subgroup ATRT.