Fig. 3: RNA sequencing reveals significant changes in cell cycle and differentiation pathways following MEK inhibition.

A Venn diagram depicting the number of significantly differentially expressed genes that are unique to one timepoint (day 3 or day 7) or that overlap between both timepoints following trametinib treatment. B, C Enriched pathways and biological processes as determined by GSEA following 3 days (B) or 7 days (C) trametinib treatment of UI226 tumorspheres. GSEA results for Hallmark, Canonical Pathways (C2:CP), and Gene Ontology (C5) (q value < 0.01) were used to build the enrichment map. Node coloring is based on the normalized enrichment score (NES), with blue representing enrichment in vehicle and red representing enrichment in trametinib samples. D–G Heat maps depicting genes associated with E2F targets (D), MYC targets (E), G2M checkpoint (F), as well as neural stem cell markers (G) that are enriched in downregulated genes following trametinib treatment (negative NES in GSEA analysis). Significant enrichment was observed at day 3, and this was further enhanced at day 7. For Hallmark G2M_checkpoint Day 3, q = 0.0087. For all other categories and timepoints, q < 0.0001. H–J GSEA results for gene sets upregulated in GCNPs after SHH stimulation (H), markers for embryonal cerebral cortex astrocytes, (I) or ciliopathy markers (J) that are enriched in downregulated (H) or upregulated (I, J) genes following 7 days of trametinib treatment. padj < 0.05* for all signatures. For GCNP after SHH stimulation in (H): Day 7, q = 0.0002, NES = −1.84. For astrocyte signature in (I): Day 7, q = 0.047, NES = 1.62. For cilopathy signature in (J): Day 7, q < 0.0001, NES = 2.26.