EZH2 inhibition induces senescence via ERK1/2 signaling pathway in multiple myeloma

Abstract

Epigenetic modifications play an important role in cellular senescence, and enhancer of zeste homolog 2 (EZH2) is a key methyltransferase involved in epigenetic remodeling in multiple myeloma (MM) cells. We have previously demonstrated that GSK126, a specific EZH2 inhibitor, exhibits anti-MM therapeutic efficacy and safety in vivo and in vitro; however, its specific mechanism remains unclear. This study shows that GSK126 induces cellular senescence in MM, which is characterized by the accumulation of senescence-associated heterochromatin foci (SAHF) and p21, and increased senescence-associated β galactosidase activity. Furthermore, EZH2 is inhibited in ribonucleotide reductase regulatory subunit M2 (RRM2)-overexpressing OCI-MY5 and RPMI-8226 cells. RRM2 overexpression inhibits the methyltransferase function of EZH2 and promotes its degradation through the ubiquitin-proteasome pathway, thereby inducing cellular senescence. In this senescence model, Lamin B1, a key component of the nuclear envelope and a marker of senescence, does not decrease but instead undergoes aberrant accumulation. Meanwhile, phosphorylation of extracellular signal-regulated protein kinase (ERK1/2) is significantly increased. The inhibition of ERK1/2 phosphorylation in turn partially restores Lamin B1 level and alleviates senescence. These findings suggest that EZH2 inhibition increases Lamin B1 level and induces senescence by promoting ERK1/2 phosphorylation. These data indicate that EZH2 plays an important role in MM cellular senescence and provide insights into the relationships among Lamin B1, p-ERK1/2, and cellular senescence.

1. Introduction

Cellular senescence is a stable state of cell cycle arrest and an important mechanism for preventing tumor cell proliferation [1‒3]. Cellular senescence is associated with haematopoietic stem cell function and the progression of multiple myeloma (MM) [4‒6]. The senescence response has been reported to be a key factor affecting chemotherapy sensitivity and treatment outcomes in patients with haematological malignancies. For example, AKI603 overcomes imatinib resistance in chronic myeloid leukaemia cells by inducing senescence [7]. Thus, unravelling the mechanisms of cellular senescence in MM may be key to overcoming therapeutic barriers in patients with relapsed/refractory MM.

Recently, several studies have focused on oncogene-induced senescence (OIS) [8‒10]. OIS is a persistent antiproliferative response that acts as a barrier against malignant transformation. Senescence-associated heterochromatin foci (SAHF) are multilayered structures centered on a condensed heterochromatic core that can be detected as DNA/chromatin-dense foci and can be induced by various trigger factors. Usually, OIS is accompanied by SAHF, whereas other forms of DNA damage response-induced senescence exhibit less SAHF [11]. Mitochondria dysfunction is also a representative hallmark of senescence [11‒13]. Normal cristae are crucial for mitochondrial function [14], while mitochondria from old samples lose their typical crista structure [15].

Enhancer of zeste homolog 2 (EZH2), a catalytic subunit of polycomb repressive complex 2 (PRC2), plays a key role in the epigenetic remodeling of MM through the trimethylation of Lys27 in histone 3 (H3K27me3) [16‒18]. Recent studies have shown that epigenetic modification plays an important role in cellular senescence [11]. In addition, inhibitors targeting EZH2 have recently been demonstrated to have therapeutic efficacy against various tumors [19,20]. High expression of EZH2 is associated with poor outcome in hematologic malignancies (myeloma and chronic lymphocytic leukaemia). In a previous study, we demonstrated that GSK126, a specific EZH2 inhibitor, exhibits anti-MM therapeutic efficacy and safety in vivo and in vitro [21]; however, its specific mechanism remains unclear. EZH2 plays a critical role in normal B cell development, and its expression level influences differentiation decisions [22]. EZH2 is degraded via the ubiquitin-proteasome pathway [23]. Phosphorylated EZH2 acts as a tumor suppressor, resulting in the inhibition of its own methyltransferase function [24,25]. Several studies have shown that EZH2 is involved in maintaining cell stemness as well as cellular senescence in several cancers [26‒28]. Furthermore, EZH2 depletion activates p21 and induces senescence in melanoma [29,30]. However, the relationship between EZH2 and senescence in MM remains unclear.