DOT1L and EZH2 cooperatively repress PRC2 targets to maintain the GC B cell identity in DLBCL [RNA-seq]

Previous published data suggested that DOT1L indirectly represses PRC2 target genes in murine B cell model. To address whether this relationship persists in human GCB-DLBCL, by integration of transcriptomic and epigenetic analysis, we defined the candidate direct targets of PRC2. GSEA of this genes list showed that DOT1L inhibition was able to significantly derepress candidate PRC2-target genes in Oci-Ly7 and Oci-Ly8. This effect was also observed upon combination treatment of DOT1Li and EZH2i, suggesting that DOT1L indirectly regulates the repression of PRC2 target genes. Moreover, it has been established that in the absence of Dot1l or Ezh2, GC B cells cannot maintain their identity and instead differentiate into a dysfunctional plasma cell-like state. We therefore, investigated whether the combined inhibition of DOT1L and EZH2 enhances plasma cell differentiation. We first established differentiation-state specific signatures unique for NBC, CB, MBC, PB, and BMPC using the GenomicScape database. Unsupervised clustering of the GSVA scores revealed that all the cell lines treated with the combination treatment were clustered together and gained a BMPC signature at the expense of a CB-signature. This crosstalk between DOT1L and EZH2 appeared to be conserved among different GCB-DLBCL cell lines and was independent of their responsiveness to the treatments. To investigate the effect of DOT1L and EZH2 inhibition on cell identity, we treated for 6 days four GCB-DLBCL cell lines with DOT1L inhibitor (EPZ-5676) and EZH2 inhibitor (GSK343) as a single agent or in combination. We then performed an unsupervised clustering anlysis using data obtained from RNA-seq analysis of four GCB-DLBCL cell lines treated for 6 days with the above mentioned inhibitors.