Superior Efficacy of a Combined Epigenetic Therapy against Human Mantle Cell Lymphoma Cells

Abstract

A deregulated epigenome contributes to the transformed phenotype of Mantle Cell Lymphoma (MCL). This involves activity of the PRC (polycomb repressive complex) 2, containing three core proteins EZH2, SUZ12 and EED, in which the SET domain of EZH2 mediates the histone methyltransferase activity. This induces trimethylation (3Me) of lysine (K)-27 on histone H3 (3MeK27H3), regulates the expression of HOX genes and promotes cell proliferation and aggressiveness of the transformed cells. Here, we demonstrate that treatment with the S-adenosylhomocysteine hydrolase inhibitor 3-Deazaneplanocin A (DZNep) depletes EZH2, SUZ12 and 3MeK27H3 in the cultured human MCL cells. Treatment with DZNep increased the expression of p21, p27 and FBXO32, while depleting Cyclin D1 and Cyclin E1 levels in MCL cells. Additionally, DZNep treatment induced cell cycle arrest and apoptosis in cultured and primary MCL cells. Further, as compared to treatment with each agent alone, co-treatment with DZNep and the pan-histone deacetylase inhibitor panobinostat (PS) caused greater depletion of EZH2, SUZ12, 3MeK27H3 and Cyclin D1 levels, while inducing greater expression of FBXO32, p16, p21 and p27. Combined treatment with DZNep and PS also synergistically induced apoptosis of cultured and primary MCL cells while relatively sparing normal CD34+ cells. Co-treatment with DZNep and PS also caused significantly greater inhibition of tumor growth of JeKo-1 xenografts in NOD/SCID mice. These preclinical in vitro and in vivo findings demonstrate that the co-treatment with DZNep and PS is an active combined epigenetic therapy worthy of further in vivo testing against MCL.

The Polycomb group (PcG) protein EZH2 supports the survival of PAX3-FOXO1 alveolar rhabdomyosarcoma by repressing FBXO32 (Atrogin1/MAFbx)

“…33,34 Both pharmacological treatments phenocopied the effects of genetic inhibition of EZH2 in vitro enhancing the apoptotic rate of fusion-positive alveolar RMS cells, upregulating FBXO32 and downregulating Myogenin. Moreover, even if higher concentrations of DZNep compared with those used in this study can affect histone methyltransferases other than EZH2, 28 the decrease in H3K27me3, but not in H3K9me3 (another repressive transcriptional mark), observed after treatment with the two compounds suggests a selective function anti-EZH2.…”

“…15,[19][20][21][22][23][24] Notably, its molecular or pharmacological inhibition leads to the reversal of the malignant phenotype. [24][25][26][27][28][29] Collectively, these findings demonstrate a role of EZH2 as a potential prognostic marker and therapeutic target in cancer. We and others have recently reported that (i) as compared with the normal myoblasts and muscle tissues EZH2 is markedly overexpressed in RMS 30,31 and (ii) EZH2 downregulation in vitro leads to myogenic-like differentiation of an embryonal RMS cell line.…”

“…We then provide evidence that this phenomenon is due, at least in part, to the derepression of the tumor suppressor gene FBXO32, already shown to be induced by EZH2 knockdown in cells of solid and blood EZH2 supports PAX3-FOXO1 rhabdomyosarcoma survival R Ciarapica et al cancers. 23,28,40 Indeed, the simultaneous depletion of FBXO32 and EZH2 greatly reduced the apoptotic cell response, suggesting that EZH2 protects fusion-positive alveolar RMS cells from apoptosis in part by repressing FBXO32. Of note, FBXO32 derepression is associated with both a reduction in EZH2 recruitment and a decrease in H3K27me3 mark to the FBXO32 gene promoter, indicating that in our cell context EZH2 is recruited at this promoter and is enzymatically active to repress transcription, as already shown.…”

“…26,27,29 In the attempt to evaluate the effect of pharmacologic inhibition of EZH2, we have used DZNep, which has been already shown to induce FBXO32 expression in cancer cells 25,28,40,66 in parallel with an EZH2 catalytic inhibitor, MC1945. 33,34 Both pharmacological treatments phenocopied the effects of genetic inhibition of EZH2 in vitro enhancing the apoptotic rate of fusion-positive alveolar RMS cells, upregulating FBXO32 and downregulating Myogenin.…”

Acquired platinum resistance involves epithelial to mesenchymal transition through ubiquitin ligase FBXO32 dysregulation

“…Furthermore, Chou et al reported the potent ability of FBXO32 to resensitize cancer cells to CDDP, suggesting that the regulation of FBXO32 expression may be crucial for cancer progression (38). Recent studies on the epigenetic regulation of FBXO32 expression in malignancies revealed that efficient apoptosis induction by the chromatin modulator DZNep via trimethylation of H3K27 required FBXO32 (37,40,41).…”