ATRX mutant neuroblastoma is sensitive to EZH2 inhibition via modulation of neuronal differentiation

We report on the characterization of ATRX in-frame fusion neuroblastoma and identify that ATRX IFF proteins re-locate from H3K9me3 enriched regions to active chromatin, such as the promoter of neural repressor REST. We further identify that REST is upregulated in ATRX IFF NB and that several neurogenesis and REST target genes are transcriptionally downregulated. Through ChIP-seq analysis, we observe that REST is bound to ATRX IFF Down genes, which have higher levels of H3K27me3. We further show that ATRX in-frame fusion neuroblastoma cells are sensitive to EZH2 inhibitors through de-repression of H3K27me3 bound neuronal function genes, includiing a subset of REST targets. We examine the genomic occupancy of ATRX in one ATRX wild-type neuroblastoma cell line and two ATRX in-frame fusion neuroblastoma cell lines; REST in two ATRX in-frame fusion neuroblastoma cell lines; relevent histone modifications and the transcriptome of two ATRX wild-type neuroblastoma cell lines, two ATRX in-frame fusion neuroblastoma cell lines, and two ATRX in-frame fusion tumor sample. We further interrogate one ATRX wild-type and two ATRX in-frame fusion neuroblastoma cell lines for transcriptional changes upon EZH2 inhibition. Finally, we include here RNA-seq analysis of five ATRX wild-type neuroblastoma tumor samples, 5 MYCN-amplified neuroblastoma tumor samples, and 5 ATRX in-frame fusion and mutant tumor samples. We also utilized RNA-seq analysis of SK-N-FI cells expressing MS2-NLS-control and MS2-NLS-ATRX-helicase as well as CHLA-90 control and REST knockdown cells.