Journal: Nature Communications

Article Title: Canonical PRC1 controls sequence-independent propagation of Polycomb-mediated gene silencing

doi: 10.1038/s41467-019-09628-6

Figure Lengend Snippet: cPRC1 and H3K27me3 are required for maintenance of reporter gene silencing. a Flow cytometry histograms before and after 6 days of Dox treatment of CRISPR mutant clones isolated from sgRNA-treated cPRC1-mESCs. Percentage indicates fraction of GFP-negative reporter cells. b Percentage of GFP- and BFP-negative cells before and after 6 days of Dox treatment in response to increasing concentrations of Ezh2 inhibition by GSK126. Data are mean ± SD (error bars) of two independent experiments

Article Snippet: 4 × 10 3 cPRC1-TetO mESCs were treated for three days on 96 well plates in both absence and presence of 1 µg/ml doxycycline (Sigma, D9891) with following chemical inhibitors: Ezh2 inhibitor GSK126 (Axora, BV-2282), increasing concentrations of negative control compound UNC4219, Cbx4/7 antagonist UNC3866 alone or in combination with 4 µM GSK126 .

Techniques: Flow Cytometry, CRISPR, Mutagenesis, Clone Assay, Isolation, Inhibition

Journal: Nature Communications

Article Title: Canonical PRC1 controls sequence-independent propagation of Polycomb-mediated gene silencing

doi: 10.1038/s41467-019-09628-6

Figure Lengend Snippet: Interaction of Cbx7 with H3K27me3 is essential for cPRC1-dependent inheritance. a ChIP qPCR analyses compares relative enrichments of PcG proteins and histone modifications in reporter cells expressing wild-type and mutant FLAG-TetR-Cbx7. ChIP enrichments for H2AK119ub1 and H3K27me3 are normalized to negative control locus (IAP). Data are mean ± SD (error bars) of at least two independent experiments. Source data are provided as a Source Data file. b Flow cytometry histograms compare GFP expression before and after three days of Dox treatment of wild-type (gray—upper panels) and Cbx7 KO dual reporter cells expressing TetR-Cbx7 W35A (gray—lower panels). Percentages indicate fraction of silenced reporter cells expressing either mutant or wild-type TetR fusion (in brackets). c Percentage of GFP-negative cells before and after 6 days of Dox treatment in response to increasing concentrations of Cbx7 inhibitor (UNC3866) alone, in combination with 4 μM GSK126 or control compound (UNC4219). Data are mean ± SD (error bars) of two independent experiments. d GFP histograms before and after 6 days of Dox addition to TetR-Cbx7 reporter cells with overexpression of Bap1 and Asxl1 (PR-DUB OE), components of the human PR-DUB complex specific for H2AK119ub1. Percentages indicate fraction of silenced cells in PR-DUB OE and wild-type reporter cell lines (in brackets)

Article Snippet: 4 × 10 3 cPRC1-TetO mESCs were treated for three days on 96 well plates in both absence and presence of 1 µg/ml doxycycline (Sigma, D9891) with following chemical inhibitors: Ezh2 inhibitor GSK126 (Axora, BV-2282), increasing concentrations of negative control compound UNC4219, Cbx4/7 antagonist UNC3866 alone or in combination with 4 µM GSK126 .

Techniques: Expressing, Mutagenesis, Negative Control, Flow Cytometry, Over Expression

Journal: Journal of Cellular and Molecular Medicine

Article Title: Targeting EZH 2 as a novel therapeutic strategy for sorafenib‐resistant thyroid carcinoma

doi: 10.1111/jcmm.14365

Figure Lengend Snippet: Enhancer of zeste homolog 2 ( EZH 2) was modulated by miR‐124‐3p and miR‐506‐3p in sorafenib resistant cells. A, The expression levels of miR‐124‐3p and miR‐506‐3p were decreased in TC ‐13 cells compared with the TC ‐07 cells. The mRNA levels of miR‐124‐3p and miR‐506‐3p were determined by a qRT ‐ PCR . The results represent the mean ± SD from three independent experiments. *** P < 0.001. B, The protein levels of EZH 2 and H3K27me3 was up‐regulated, whereas, the H3K27Ac was down‐regulated in TC ‐13 cells compared with the TC ‐07 cells. A Western blot of cell lysates of TC ‐13 and TC ‐07 cell was performed to examine the protein levels of EZH 2, H3K27me3 and H3K27Ac. H3 was used as the loading control. C, The relative mRNA levels of EZH 2 and H3K27me3 were increased and H3K27Ac was decreased in the TC ‐13 cells compared with the TC ‐07 cells. The mRNA levels of EZH 2, H3K27me3 and H3K27Ac were tested by a qRT ‐ PCR . The results represent the mean ± SD from three independent experiments. *** P < 0.001, ** P < 0.05

Article Snippet: The EZH2 inhibitor EPZ‐6438 was obtained from Adooq Bioscience (Shanghai, China).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: Targeting EZH 2 as a novel therapeutic strategy for sorafenib‐resistant thyroid carcinoma

doi: 10.1111/jcmm.14365

Figure Lengend Snippet: Targeting enhancer of zeste homolog 2 by overexpression of miR‐124/506 or EPZ ‐6438 treatment inhibits the proliferation ability of sorafenib resistant thyroid tumor cells via epigenetic regulation. A, The proliferation ability of TC ‐13 cells was inhibited when cells were overexpressed with miR‐124/506 or treated with 1 μmol/L EPZ ‐6438. The cell viability was examined for 5 d by CCK ‐8 kit and the absorbance results at 450 nm were represented as mean ± SD from three independent repeats. B, Overexpression of miR‐124/506 or EPZ ‐6438 treatment promotes the protein expression of H3K27me3 and suppresses the protein level of H3K27Ac in thyroid tumour cells. A Western blot was performed to determine the protein levels of H3K27me3 and H3K27Ac with indicated antibodies. H3 was tested as a loading control

Article Snippet: The EZH2 inhibitor EPZ‐6438 was obtained from Adooq Bioscience (Shanghai, China).

Techniques: Over Expression, CCK-8 Assay, Expressing, Western Blot, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: Targeting EZH 2 as a novel therapeutic strategy for sorafenib‐resistant thyroid carcinoma

doi: 10.1111/jcmm.14365

Figure Lengend Snippet: Combination of sorafenib with miR‐124/506 overexpression or enhancer of zeste homolog 2 inhibitor improves the survival in mice model. Survival curve of the mice were injected with TC ‐07 or TC ‐13 cells (1 × 10 6 cells/mouse) by subaxillary inoculation. The sorafenib was given every day for 20 d by 20 mg/kg with intravenous injection or a combination with miR‐124/506 mimics or the amount of EPZ ‐6438 by 200 mg/kg. Each group has 20 mice

Article Snippet: The EZH2 inhibitor EPZ‐6438 was obtained from Adooq Bioscience (Shanghai, China).

Techniques: Over Expression, Injection

Journal: NPJ Precision Oncology

Article Title: Relapse-free survival in a pediatric patient with recurrent EZH2 -mutant melanoma treated with adjuvant tazemetostat

doi: 10.1038/s41698-025-00826-8

Figure Lengend Snippet: A First relapse; a 1.4 cm tumor was identified in the left external auditory canal three months after initial debulking surgery and prior to any systemic treatment. B Second relapse; after one year of adjuvant nivolumab, the patient experienced a second relapse when a new 4.0 cm mass was identified along the floor of the left middle cranial fossa with mass effect on the left temporal lobe. C Post-operative imaging; the patient underwent intracranial tumor resection after two cycles of neoadjuvant ipilimumab/ nivolumab. D Representative surveillance imaging while on tazemetostat; eighteen months post-operatively and fifteen months into tazemetostat therapy, the patient continued to demonstrate no evidence of relapsed disease. Blue arrows indicate the location of the tumor or resection site.

Article Snippet: After discussion at the Johns Hopkins Molecular Tumor Board and careful consideration, she was started on the EZH2 inhibitor tazemetostat based upon the presence of the pathogenic EZH2 A692V mutation identified on NGS .

Techniques: Adjuvant, Imaging

Journal: NPJ Precision Oncology

Article Title: Relapse-free survival in a pediatric patient with recurrent EZH2 -mutant melanoma treated with adjuvant tazemetostat

doi: 10.1038/s41698-025-00826-8

Figure Lengend Snippet: Next-generation sequencing (NGS) comparing first and second relapse

Article Snippet: After discussion at the Johns Hopkins Molecular Tumor Board and careful consideration, she was started on the EZH2 inhibitor tazemetostat based upon the presence of the pathogenic EZH2 A692V mutation identified on NGS .

Techniques: Next-Generation Sequencing

Journal: Journal for Immunotherapy of Cancer

Article Title: Targeting disialoganglioside GD2 with chimeric antigen receptor-redirected T cells in lung cancer

doi: 10.1136/jitc-2021-003897

Figure Lengend Snippet: Upregulation of GD2 expression by EZH2 inhibition in GD2 low/neg lung cancer cell lines makes them sensitive to GD2.CAR-T cell therapy. (A) GD2 expression on the cell surface of the SCLC cell line H748 and NSCLC cell line H1792 cultured with tazemetostat at 1 or 10 µM, respectively, or equivalent volumes of DMSO for 7 days (upper panel) and for 21 days (lower panel). The Ewing sarcoma cell line RD-ES was used as a positive control. The results are shown as percentages of positive cells and MFI of GD2 (GD2 MFI/isotype control MFI). (B) Lung cancer cell lines pretreated with tazemetostat or DMSO for 21 days were cocultured with either CD19.CAR-T cells or GD2.CAR-T cells at the T-cell to tumor cell ratio of 1 to 2. On day 5, tumor cells (CD276 + ) and T cells (CD3 + ) were enumerated by flow cytometry. The Ewing’s sarcoma cell line RD-ES was used as a positive control. Quantification of residual tumor cells are illustrated. Data represent mean±SD (n=3, *p<0.05; ***p<0.001, tazemetostat vs DMSO pretreatment). (C, D) Summary of IFNγ (C) and IL-2 (D) released by GD2.CAR-T cells in the culture supernatant after 24 hours of coculture with the tumor cell lines as measured by ELISA. Data represent mean±SD (n=3, *p<0.05; **p<0.01; ****p<0.0001, tazemetostat vs DMSO pretreatment). (E) Representative CFSE dilution of CFSE-labeled GD2.CAR-T cells cocultured with tumor cell lines pretreated with tazemetostat or DMSO, for 4 days at 1 to 1 ratio analyzed by flow cytometry. (F) The percentages of CFSE dilution were measured relative to CFSE-labeled NT. Summary of CFSE-dilution assays. data represent mean±SD (n=3, *p<0.05; **p<0.01, tazemetostat vs DMSO pretreatment). CAR, chimeric antigen receptor; CFSE, carboxyfluorescein diacetate succinimidyl ester; DMSO, dimethyl sulfoxide; EZH2, Enhancer of zeste homolog 2; FSC, forward scatter; MFI, mean fluorescent intensity; NSCLC, non-small cell lung cancer; NT, non-transduced T cells; SCLC, small cell lung cancer.

Article Snippet: Tumor cells were treated with the EZH2 (Enhancer of zeste homolog 2) inhibitor tazemetostat (EPZ-6438, KareBayBiochem, USA) dissolved in dimethyl sulfoxide (DMSO) or DMSO alone added at a concentration of 1 or 10 μM.

Techniques: Expressing, Inhibition, Cell Culture, Positive Control, Control, Flow Cytometry, Enzyme-linked Immunosorbent Assay, Labeling

Journal: The American Journal of Pathology

Article Title: Epigenetic Silencing of miRNA-34a in Human Cholangiocarcinoma via EZH2 and DNA Methylation

doi: 10.1016/j.ajpath.2017.06.014

Figure Lengend Snippet: EZH2 represses miR-34a expression through H3K27 trimethylation in cholangiocarcinoma (CCA) cells. A: Representative immunohistochemistry for EZH2 in human CCA tissue. The brown color indicates positive signals; nuclei were counterstained as blue. The boxed area in the left panel is shown at higher magnification in the right panel. B: Western blot analysis for EZH2 in nonmalignant human cholangiocyte cell (H69) and CCA cells (CCLP1, SG231, HUCCT1, and TFK1). C: The levels of miR-34a in CCA cells with/without GSK126 treatment for 72 hours, as determined by quantitative RT-PCR (RT-qPCR). D: Chromatin immunoprecipitation (ChIP) assay. The chromatin extracted from CCLP1 and SG231 cells treated with or without GSK126 was subjected to immunoprecipitation with H3K27me3 antibody, and the precipitated DNA was subjected to RT-qPCR analysis using two sets of primers to amplify the miR-34a promoter region, as shown in the schematic diagram. Normal mouse IgG was used as the negative control. E: RT-qPCR analysis for miR-34a in SG231 cells transfected with two individual EZH2 shRNA or control vector (pSMP). F: ChIP assay with EZH2 antibody followed by RT-qPCR analysis in SG231 cells with or without GSK126 treatment. Data are expressed as means ± SD (D and F); data are expressed as means ± SEM (C and E). ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001. Original magnification: ×100 (A, left panel); ×200 (A, right panel). TSS, transcription start site.

Article Snippet: EZH2 inhibitor, GSK126, was purchased from Cellagen Technology (San Diego, CA) and Sellekchem (Houston, TX).

Techniques: Expressing, Immunohistochemistry, Western Blot, Quantitative RT-PCR, Chromatin Immunoprecipitation, Immunoprecipitation, Negative Control, Transfection, shRNA, Control, Plasmid Preparation

Journal: The American Journal of Pathology

Article Title: Epigenetic Silencing of miRNA-34a in Human Cholangiocarcinoma via EZH2 and DNA Methylation

doi: 10.1016/j.ajpath.2017.06.014

Figure Lengend Snippet: Effect of EZH2 inhibitor, GSK126, on cholangiocarcinoma cell growth in vivo and in vitro. A: Experimental design of tumor xenograft and GSK126 intraperitoneal injection in SCID mice. CCLP1 cells (1.0 × 106) were mixed in Matrigel solution and inoculated into severe combined immunodeficiency (SCID) mice (initially started with six mice per group) via intrahepatic injection and the mice were treated with vehicle (20% Captisol) or 150 mg/kg GSK126 twice a week for 5 weeks. B: Tumor volume of vehicle and GSK126-treated mice. After injection of CCLP1 cells into the left liver lobe in the mice, intrahepatic metastasis was determined at 8 weeks by observation of the tumor nodule on the uninjected liver lobes. All of the vehicle-treated mice (five mice) had intrahepatic metastasis, whereas one of four GSK126-treated mice had intrahepatic metastasis (P < 0.05, Fisher exact test). C: Body weight changes of the mice. D: The level of trimethylated H3K27 in liver tumor tissues from mice treated with vehicle or GSK126 (only three liver tumor samples were available for Western blot because of the very small tumor size in the GSK126-treated group). E: The growth of CCLP1 and SG231 cells was determined by WST1 assay after treatment of GSK126 or vehicle for 72 hours. Colony-forming assay in CCLP1 and SG231 cells with or without GSK126 treatment for 10 days. Data are expressed as means ± SD. ∗∗∗P < 0.001. Veh, vehicle.

Article Snippet: EZH2 inhibitor, GSK126, was purchased from Cellagen Technology (San Diego, CA) and Sellekchem (Houston, TX).

Techniques: In Vivo, In Vitro, Injection, Western Blot

Journal: The American Journal of Pathology

Article Title: Epigenetic Silencing of miRNA-34a in Human Cholangiocarcinoma via EZH2 and DNA Methylation

doi: 10.1016/j.ajpath.2017.06.014

Figure Lengend Snippet: EZH2-mediated H3K27 trimethylation and DNA methylation independently silence miR-34a expression. A: Relative miR-34a expression in cholangiocarcinoma (CCA) cells treated with 3 μmol/L 5-Aza-2′-deoxycytidine (5-Aza-CdR) or vehicle for 72 hours, as determined by quantitative RT-PCR (RT-qPCR) analysis. B: Methylation-specific PCR (MSP) of genomic DNA from CCLP1 and SG231 cells treated with or without 3 μmol/L 5-Aza-CdR treatment. C: MSP analysis in CCLP1 and SG231 cells treated with 2.5 or 10 μmol/L GSK126. D: Western blot analysis for DNMT1, H3K27 trimethylation (H3K27me3), and EZH2 in CCLP1 and SG231 cells with or without GSK126 treatment. E: Chromatin immunoprecipitation (ChIP) assay. The chromatin extracted from SG231 cells treated with 5-Aza-CdR or vehicle was subjected to immunoprecipitation with H3K27me3 antibody, and the precipitated DNA was subjected to RT-qPCR analysis using two sets of primers to amplify the miR-34a promoter region. Data are expressed as means ± SEM. ∗P < 0.05, ∗∗P < 0.01. 5Aza, 5-Aza-CdR; M, methylation-specific primer to the miR-34a promoter sequence; U, unmethylation-specific primer to the miR-34a promoter sequence.

Article Snippet: EZH2 inhibitor, GSK126, was purchased from Cellagen Technology (San Diego, CA) and Sellekchem (Houston, TX).

Techniques: DNA Methylation Assay, Expressing, Quantitative RT-PCR, Methylation, Western Blot, Chromatin Immunoprecipitation, Immunoprecipitation, Sequencing

Journal: Cancers

Article Title: BET and CDK Inhibition Reveal Differences in the Proliferation Control of Sympathetic Ganglion Neuroblasts and Adrenal Chromaffin Cells

doi: 10.3390/cancers14112755

Figure Lengend Snippet: Effect of IGF, EZH2, ALK, and WNT inhibitors on the proliferation of chromaffin cells, neuroblasts, and NESTIN-expressing cells (NECs). Dose–response curves are shown for IGFR inhibitor (PPP) ( A ), EZH2 inhibitor (EPZ6438) ( B ), Alk inhibitor (Alectinib), ( C ) and Wnt inhibitor (ICG001) ( D ). Data represent the mean ± s. e. m. of at least three independent experiments.

Article Snippet: The inhibitors that were used were the BET inhibitors JQ1 (Tocris Biotechne, Wiesbaden, Germany; 4499) and GSK1324726A (iBET 726) (Selleckchem Biozol, Eching, Germany), the CDK-7 inhibitors THZ1 (Medchem Express Biotrend, Köln, Germany) and YKL-5-125 (Selleckchem), the CDK12/13 inhibitor THZ 531 (Selleckchem), the IGF1-R inhibitor picropodophyllin (PPP) (Tocris 2956), the EZH2 inhibitor EPZ 6438 (Axon Medchem, Groningen, NL; 2227), the WNT inhibitor ICG001 (Axon Medchem 1766), and the ALK inhibitor Alectinib (Selleckchem S276).

Techniques: Expressing