Fig 2: Integrated screen establishes SSRP1 as the top drug candidate and a prognostic marker.A Flowchart of bioinformatics analysis. B Genes with inhibitor were chosen for further validation. C Box plots of SSRP1 expression levels in different subgroups of medulloblastoma and normal cerebellum (CB) from the Pomeroy dataset. D Box plots of SSRP1 expression levels in medulloblastoma and normal cerebellum from the u133p2 dataset. E Box plots of SSRP1 expression levels in various subgroups of medulloblastoma from the Cavalli dataset. F, G Kaplan–Meier plots that show the OS rate in Cavalli dataset of all MB patients and group three MB patients. (*P < 0.05, **P < 0.01, and ***P < 0.001).

Fig 3: CBL0137 decreases the transcription of NEUROD1 and OTX2-associated genes.A, B Volcano plot of the normalized expression of genes in MYC-amplified medulloblastoma cells after CBL0137 treatment. Red points represent genes with P-value <0.05. C NEUROD1 expression in the MYC-amplified medulloblastoma cells which were treated with CBL0137 or DMSO was measured using RT-qPCR. D–G Enrichment plots of genes in shOTX2_48 h and shOTX2_96 h gene sets in DMSO-treated cells and CBL0137-treated cells. H ChIP-qPCR analysis of SSRP1 levels at MYC in MB cells treated with DMSO or CBL0137. The seven sites at the promoter region of NEUROD1 ware arranged up the histogram. (*P < 0.05, **P < 0.01, and ***P < 0.001).

Fig 4: CPF suppresses transcription of FACT and c‐MYC genes. A, Quiescent A549 cells were transfected with either empty vector (control), or FACT plasmid (SPT16+ or SSRP1+), or treated with CPF at GI90, or a combination of FACT plasmid and CPF at GI90 (rescue) upon cell cycle re‐entry for 36h. The cells were harvested for cell cycle analysis. *P < .05 vs control; B, ChIP was performed in the presence of CPF at GI90 or DMSO for 36 h following release from 3‐day contact inhibition in A549 cells. PCR primers were designed based on FACT and c‐MYC promoter or control region. *P < .05 vs control. C, Knockdown of c‐MYC with three sets of siRNA and its impact on FACT mRNA and protein levels by RT‐PCR, immunoblotting and quantification. *P < .05 vs NC control

Fig 5: Targeted knockdown of SSRP1 using CRISPR-Cas9 or RNA interference attenuates the growth of medulloblastoma.A Boxplot of CRISPR gene effect scores in the MB cell lines. B, C RT-qPCR and western blot analyses were performed to assess the efficiency of shRNA SSRP1 knockdown. D Cell growth of the medulloblastoma cells infected with shRNA lentivirus was measured at days 0, 2, and 4. (*P < 0.05, **P < 0.01, ***P < 0.001).