EZH2 upregulates the PI3K/AKT pathway through IGF1R and MYC in clinically aggressive chronic lymphocytic leukemia

EZH2 is overexpressed in poor-prognostic chronic lymphocytic leukemia (CLL) cases, acting as an oncogene; however, thus far, the EZH2 target genes in CLL have not been disclosed. In this study, using ChIP-sequencing, we identified EZH2 and H3K27me3 target genes in two prognostic subgroups of CLL with distinct prognosis and outcome, i.e. cases with unmutated (U-CLL, n=6) or mutated IGHV genes (M-CLL, n=6). While the majority of oncogenic pathways were equally enriched for EZH2 target genes in both prognostic subgroups, PI3K pathway genes were differentially bound by EZH2 in U-CLL versus M-CLL. The occupancy of EZH2 for selected PI3K pathway target genes was validated in additional CLL samples (n=16) and CLL cell lines using siRNA-mediated EZH2 downregulation and ChIP assays. Intriguingly, we found that EZH2 directly binds to the <i>IGF1R</i> promoter along with MYC and upregulates IGF1R expression in U-CLL, leading to downstream PI3K activation. By investigating an independent CLL cohort (n=96), a positive correlation was observed between <i>EZH2</i> and <i>IGF1R</i> expression with higher levels in U-CLL compared to M-CLL. Accordingly, siRNA-mediated downregulation of either <i>EZH2, MYC</i> or <i>IGF1R</i> and treatment with EZH2 and MYC pharmacological inhibitors in the HG3 CLL cell line induced significant reduction in PI3K pathway activation. In conclusion, we characterize for the first time EZH2 target genes in CLL revealing a hitherto unknown implication of EZH2 in modulating the PI3K pathway in a non-cannonical, PRC2-independent way, with potential therapeutic implications considering that PI3K inhibitors are effective therapeutic agents for CLL.