Characterization and pharmacologic targeting of EZH2, a fetal retinal protein and epigenetic regulator, in human retinoblastoma

Abstract

Retinoblastoma (RB) is the most common primary intraocular cancer in children, a nd the third most common cancer overall in infants. No molecular-targeted therapy for this lethal tumor exists. Since the tumor suppressor RB1, whose genetic inactivation underlies RB, is upstream of the epigenetic regulator EZH2, a pharmacologic target for many solid tumors, we reasoned that EZH2 might regulate human RB tumorigenesis. Histologic and immunohistochemical analyses were performed using an EZH2 antibody in sections from 43 samples of primary, formalin-fixed, paraffin embedded human RB tissue, cryopreserved mouse retina; and in whole cell lysates from human RB cell lines (Y79 and WERI-Rb1), primary human fetal RPE and fetal and adult retina, mouse retina and embryonic stem (ES) cells. While enriched during fetal human retinal development, EZH2 protein was not present in the normal postnatal retina. However, EZH2 was detected in all 43 analyzed human RB specimens, indicating that EZH2 is a fetal protein expressed in postnatal human RB. EZH2 expression marked single RB cell invasion into the optic nerve, a site of invasion whose involvement may influence the decision for systemic chemotherapy. To assess the role of EZH2 in RB cell survival, human RB and primary RPE cells were treated with two EZH2 inhibitors (EZH2i), GSK126 and SAH-EZH2 (SAH). EZH2i inhibitors impaired intracellular ATP production, an indicator of cell viability, in a time and dose-dependent manner, but did not affect primary human fetal RPE. Thus, aberrant expression of a histone methyltransferase protein is a feature of human RB. This is the first time this mechanism has been implicated for an eye, adnexal, or orbital tumor. The specificity of EZH2i toward human RB cells, but not RPE, warrants further in vivo testing in animal models of RB, especially those EZH2i currently in clinical trials for solid tumors and lymphoma.

Postnatal onset of retinal degeneration by loss of embryonic Ezh2 repression of Six1

“…Among the two HMTs that regulate H3K27 trimethylation, Ezh1 and Ezh2 , and the demethylase Jmjd3 , Ezh2 and Jmjd3 are widely expressed in retinal progenitors during development; Ezh1 is expressed only weakly in the postnatal retina 29 . The transient expression of Ezh2 during development, which is completely repressed postnatally in the retina, was also confirmed during human retinal development 11 . A cell type specific effect was also observed with Jmjd3 deficiency, which resulted in impaired differentiation of subsets of bipolar cells, despite strong Jmjd3 expression in the RGCs 9 .…”

“…Ezh2 trimethylates histone H3 at lysine 27 (H3K27me3), a repressive histone mark that is known to persist into adulthood 7 8 . Previous reports have shown that Ezh2 / EZH2 is abundant during embryonic development but is absent during the late postnatal period and adulthood in mice and humans 9 10 11 . Conditional loss-of-function experiments in which an early (E9.5) retinal Cre driver, such as Dkk3 - 12 or Pax6-αCre , was used to delete retinal Ezh2, has demonstrated a role for this HMT in the proliferation and differentiation of retinal progenitor cells (RPCs) 13 14 .…”

Implementation of the Chick Chorioallantoic Membrane (CAM) Model in Radiation Biology and Experimental Radiation Oncology Research

“…The above in vitro models are complemented by classical xenograft or orthotopic murine tumor models with established human or syngeneic murine cancer cell lines, genetically engineered murine tumor models, or patient-derived xenograft tumors grown in immunodeficient mice [173,174,175,176,177,178,179,180,181,182,183,184].…”

<p>Diosgenin and GSK126 Produce Synergistic Effects on Epithelial–Mesenchymal Transition in Gastric Cancer Cells by Mediating EZH2 via the Rho/ROCK Signaling Pathway</p>

“…21 Several studies conducted with solid tumor cell lines have suggested that GSK126 suppresses cell migration and angiogenesis, but has a limited therapeutic effect when used alone. [22][23][24] Therefore, we hypothesized that a combination of GSK126 and diosgenin might be more potent for attenuating GC metastasis when compared to using GSK126 or diosgenin alone.…”