We performed comprehensive genome-wide gene expression profiling (GEP) of extranodal nasal-type natural killer/T-cell lymphoma (NKTL) using formalin-fixed, paraffin-embedded tissue (n = 9) and NK cell lines (n = 5) in comparison with normal NK cells, with the objective of understanding the oncogenic pathways involved in the pathogenesis of NKTL and to identify potential therapeutic targets. Pathway and network analysis of genes differentially expressed between NKTL and normal NK cells revealed significant enrichment for cell cycle-related genes and pathways, such as PLK1, CDK1, and Aurora-A. Furthermore, our results demonstrated a pro-proliferative and anti-apoptotic phenotype in NKTL characterized by activation of Myc and nuclear factor kappa B (NF-κB), and deregulation of p53. In corroboration with GEP findings, a significant percentage of NKTLs (n = 33) overexpressed c-Myc (45.4%), p53 (87.9%), and NF-κB p50 (67.7%) on immunohistochemistry using a tissue microarray containing 33 NKTL samples. Notably, overexpression of survivin was observed in 97% of cases. Based on our findings, we propose a model of NKTL pathogenesis where deregulation of p53 together with activation of Myc and NF-κB, possibly driven by EBV LMP-1, results in the cumulative up-regulation of survivin. Down-regulation of survivin with Terameprocol (EM-1421, a survivin inhibitor) results in reduced cell viability and increased apoptosis in tumour cells, suggesting that targeting survivin may be a potential novel therapeutic strategy in NKTL.

Key Points• This study has uncovered an oncogenic role of EZH2 independent of its methyltransferase activity in NKTL.• This study suggests that targeting EZH2 may have therapeutic usefulness in NKTL.The role of enhancer of zeste homolog 2 (EZH2) in cancer is complex and may vary depending on the cellular context. We found that EZH2 is aberrantly overexpressed in the majority of natural killer/T-cell lymphoma (NKTL), an aggressive lymphoid malignancy with very poor prognosis. We show that EZH2 upregulation is mediated by MYC-induced repression of its regulatory micro RNAs and EZH2 exerts oncogenic properties in NKTL. Ectopic expression of EZH2 in both primary NK cells and NKTL cell lines leads to a significant growth advantage. Conversely, knock-down of EZH2 in NKTL cell lines results in cell growth inhibition. Intriguingly, ectopic EZH2 mutant deficient for histone methyltransferase activity is also able to confer growth advantage and rescue growth inhibition on endogenous EZH2 depletion in NKTL cells, indicating an oncogenic role of EZH2 independent of its gene-silencing activity. Mechanistically, we show that EZH2 directly promotes the transcription of cyclin D1 and this effect is independent of its enzymatic activity. Furthermore, depletion of EZH2 using a PRC2 inhibitor 3-deazaneplanocin A significantly inhibits growth of NK tumor cells. Therefore, our study uncovers an oncogenic role of EZH2 independent of its methyltransferase activity in NKTL and suggests that targeting EZH2 may have therapeutic usefulness in this lymphoma. (Blood. 2013;121(22):4512-4520) Introduction Nasal-type natural killer/T-cell lymphoma (NKTL) is an aggressive lymphoid malignancy associated with very poor survival outcomes.1 A better understanding of the molecular abnormalities underlying this disease will provide important insights into the biology of this tumor; however, studies on NKTL are often limited by the lack of adequate tissue in small nasal biopsies and the presence of necrosis in biopsy specimens. Although more effective therapy is now available, treatment is still completely reliant on radiotherapy and combinations of chemotherapy. 2,3 We and others have recently performed whole-genome gene expression studies and identify a number of genes that are differentially expressed in NKTL as well as pathways that are activated in NKTL. Enhancer of zeste homolog 2 (EZH2), one of the genes identified in our study to be aberrantly overexpressed in NKTL, 4 is a H3K27-specific histone methyltransferase and a component of the polycomb repressive complex 2 (PRC2), which plays a key role in the epigenetic maintenance of repressive chromatin mark. EZH2 protein contains a catalytic domain (SET domain) at the COOH-terminus that provides the methyltransferase activity. The catalytic domain must partner with other noncatalytic proteins, such as EED and SUZ12, to form the PRC2 in order to attain robust histone methyltransferase activity. Genome-wide approaches have demonstrated the importance of the PRC2 complex in the transcriptional re...

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Key Points• JAK3-mediated phosphorylation of EZH2 resulted in EZH2 oncogenic function independent of its enzymatic activity.• Targeted inhibition of JAK3 may be a promising treatment in NK/TL through suppressing noncanonical EZH2 activity.The best-understood mechanism by which EZH2 exerts its oncogenic function is through polycomb repressive complex 2 (PRC2)-mediated gene repression, which requires its histone methyltransferase activity. However, small-molecule inhibitors of EZH2 that selectively target its enzymatic activity turn out to be potent only for lymphoma cells with EZH2-activating mutation. Intriguingly, recent discoveries, including ours, have placed EZH2 into the category of transcriptional coactivators and thus raised the possibility of noncanonical signaling pathways. However, it remains unclear how EZH2 switches to this catalytic independent function. In the current study, using natural killer/T-cell lymphoma (NKTL) as a disease model, we found that phosphorylation of EZH2 by JAK3 promotes the dissociation of the PRC2 complex leading to decreased global H3K27me3 levels, while it switches EZH2 to a transcriptional activator, conferring higher proliferative capacity of the affected cells. Gene expression data analysis also suggests that the noncanonical function of EZH2 as a transcriptional activator upregulates a set of genes involved in DNA replication, cell cycle, biosynthesis, stemness, and invasiveness. Consistently, JAK3 inhibitor was able to significantly reduce the growth of NKTL cells, in an EZH2 phosphorylation-dependent manner, whereas various compounds recently developed to inhibit EZH2 methyltransferase activity have no such effect. Thus, pharmacological inhibition of JAK3 activity may provide a promising treatment option for NKTL through the novel mechanism of suppressing noncanonical EZH2 activity. (Blood. 2016;128(7):948-958)