Frequent upregulation of G9a promotes RelB-dependent proliferation and survival in multiple myeloma

Abstract

Background: Multiple myeloma is an incurable hematological malignancy characterized by a heterogeneous genetic and epigenetic landscape. Although a number of genetic aberrations associated with myeloma pathogenesis, progression and prognosis have been well characterized, the role of many epigenetic aberrations in multiple myeloma remain elusive. G9a, a histone methyltransferase, has been found to promote disease progression, proliferation and metastasis via diverse mechanisms in several cancers. A role for G9a in multiple myeloma, however, has not been previously explored. Methods: Expression levels of G9a/EHMT2 of multiple myeloma cell lines and control cells Peripheral Blood Mononuclear Cells (PBMCs) were analyzed. Correlation of G9a expression and overall survival of multiple myeloma patients were analyzed using patient sample database. To further study the function of G9a in multiple myeloma, G9a depleted multiple myeloma cells were built by lentiviral transduction, of which proliferation, colony formation assays as well as tumorigenesis were measured. RNA-seq of G9a depleted multiple myeloma with controls were performed to explore the downstream mechanism of G9a regulation in multiple myeloma. Results: G9a is upregulated in a range of multiple myeloma cell lines. G9a expression portends poorer survival outcomes in a cohort of multiple myeloma patients. Depletion of G9a inhibited proliferation and tumorigenesis in multiple myeloma. RelB was significantly downregulated by G9a depletion or small molecule inhibition of G9a/GLP inhibitor UNC0642, inducing transcription of proapoptotic genes Bim and BMF. Rescuing RelB eliminated the inhibition in proliferation and tumorigenesis by G9a depletion. Conclusions: In this study, we demonstrated that G9a is upregulated in most multiple myeloma cell lines. Furthermore, G9a loss-of-function analysis provided evidence that G9a contributes to multiple myeloma cell survival and proliferation. This study found that G9a interacts with NF-κB pathway as a key regulator of RelB in multiple myeloma and regulates RelB-dependent multiple myeloma survival. G9a therefore is a promising therapeutic target for multiple myeloma.

Dual EZH2 and G9a inhibition suppresses multiple myeloma cell proliferation by regulating the interferon signal and IRF4-MYC axis

“…Several studies have shown the antitumor effects of G9a inhibition in various human malignancies, including breast cancer, acute myeloid leukemia, and non-small cell lung cancer 20 – 22 . We and others also recently reported the antimyeloma effects of G9a inhibitors 10 , 19 . Although H3K27 and H3K9 methylation similarly control cellular processes through gene silencing, their functions have been investigated separately in most earlier studies 23 , 24 .…”

“…5F ). Recent studies reported that elevated expression of EZH2 and EHMT2 is significantly associated with a poor prognosis in MM patients, which indicates that both EZH2 and G9a may be associated with progression of MM 8 , 19 .…”

G9a/GLP targeting in MM promotes autophagy-associated apoptosis and boosts proteasome inhibitor–mediated cell death

“…3 G9a overexpression in HMCLs was also recently reported in MM, and patients with high G9a levels had a significantly worse outcome. 8 In the present study, we validated the prognostic value of G9a in 2 independent cohorts of newly diagnosed patients and 1 cohort of relapsed patients.…”

“…3,7 G9a is overexpressed in many cancer types, including MM, resulting in the repression of tumor suppressor genes, like p53, CDH1, RUNX3, and E-cadherin. 3,8 High G9a expression is often associated with poor prognosis; G9a targeting, using short hairpin RNA (shRNA) or specific small molecule inhibitors, inhibits proliferation, migration, and metastasis of cancer cells. 3,4,9,10 A recent study showed that simultaneous targeting of DNMTs and G9a in acute myeloid leukemia, acute lymphoblastic leukemia, and diffuse large B-cell lymphoma cell lines using the dual G9a/DNMT inhibitor CM-272 inhibits proliferation and promotes apoptosis by inducing interferon-stimulated genes and immunogenic cell death.…”

“…12 Moreover, only very recently was G9a shown to be overexpressed in HMCLs, and G9a shRNA inhibited proliferation in vitro and tumorigenesis in xenograft mice. 8 However, the mechanisms of action were not thoroughly investigated nor was the capacity to boost the antimyeloma activity of proteasome inhibitors (PIs). Moreover, to the best of our knowledge, no study has investigated the expression or prognostic value of GLP in MM patients or validated the therapeutic value of G9a/GLP targeting using primary human MM samples and syngeneic immunocompetent murine myeloma models.…”