gsk-2816126 and Melanoma

Hepatic metastasis develops in ~ 50% of uveal melanoma (UM) patients with no effective treatments. Although GNAQ/GNA11 mutations are believed to confer pathogenesis of UM, the underlying mechanism of liver metastasis remains poorly understood. Given that profound epigenetic evolution may occur in the long journey of circulating tumor cells (CTCs) to distant organs, we hypothesized that EZH2 endowed tumor cells with enhanced malignant features (e.g., stemness and motility) during hepatic metastasis in UM. We aimed to test this hypothesis and explore whether EZH2 was a therapeutic target for hepatic metastatic UM patients.. Expression of EZH2 in UM was detected by qRT-PCR, Western blotting and immunohistochemistry staining. Proliferation, apoptosis, cancer stem-like cells (CSCs) properties, migration and invasion were evaluated under circumstances of treatment with either EZH2 shRNA or EZH2 inhibitor GSK126. Antitumor activity and frequency of CSCs were determined by xenografted and PDX models with NOD/SCID mice. Hepatic metastasis was evaluated with NOG mice.. We found that EZH2 overexpressed in UM promoted the growth of UM; EZH2 increased the percentage and self-renewal of CSCs by miR-29c-DVL2-β-catenin signaling; EZH2 facilitates migration and invasion of UM cells via RhoGDIγ-Rac1 axis. Targeting EZH2 either by genetics or small molecule inhibitor GSK126 decreased CSCs and motility and abrogated the liver metastasis of UM.. These findings validate EZH2 as a druggable target in metastatic UM patients, and may shed light on the understanding and interfering the complicated metastatic process.

Topics: Animals; Apoptosis; Biomarkers, Tumor; Cell Proliferation; Enhancer of Zeste Homolog 2 Protein; Gene Expression Regulation, Neoplastic; Humans; Indoles; Melanoma; Mice; Mice, Inbred NOD; Mice, SCID; Neoplastic Stem Cells; Pyridones; Tumor Cells, Cultured; Tumor Suppressor Proteins; Ubiquitin Thiolesterase; Uveal Neoplasms; Xenograft Model Antitumor Assays

Metabolic reprogramming is a major factor in transformation, and particular metabolic phenotypes correlate with oncogenotype, tumor progression, and metastasis. By profiling metabolites in 17 patient-derived xenograft melanoma models, we identified durable metabolomic signatures that correlate with biological features of the tumors. BRAF mutant tumors had metabolomic and metabolic flux features of enhanced glycolysis compared to BRAF wild-type tumors. Tumors that metastasized efficiently from their primary sites had elevated levels of metabolites related to protein methylation, including trimethyllysine (TML). TML levels correlated with histone H3 trimethylation at Lys

Topics: Animals; Cell Movement; Down-Regulation; Enhancer of Zeste Homolog 2 Protein; Histone-Lysine N-Methyltransferase; Histones; Humans; Indoles; Melanoma; Metabolome; Metabolomics; Methylation; Mice; Mice, Inbred NOD; Neoplasm Metastasis; Principal Component Analysis; Protein Methyltransferases; Proto-Oncogene Proteins B-raf; Pyridones; RNA Interference; RNA, Small Interfering; Transplantation, Heterologous

The epigenetic modifier EZH2 is in the center of a repressive complex controlling differentiation of normal cells. In cancer EZH2 has been implicated in silencing tumor suppressor genes. Its role in melanoma as well as target genes affected by EZH2 are poorly understood. In view of this we have used an integrated systems biology approach to analyze 471 cases of skin cutaneous melanoma (SKCM) in The Cancer Genome Atlas (TCGA) for mutations and amplifications of EZH2. Identified changes in target genes were validated by interrogation of microarray data from melanoma cells treated with the EZH2 inhibitor GSK126. We found that EZH2 activation by mutations, gene amplification and increased transcription occurred in about 20% of the cohort. These alterations were associated with significant hypermethylation of DNA and significant downregulation of 11% of transcripts in patient RNASeq data. GSK126 treatment of melanoma lines containing EZH2 activation reversed such transcriptional repression in 98 candidate target genes. Gene enrichment analysis revealed genes associated with tumor suppression, cell differentiation, cell cycle inhibition and repression of metastases as well as antigen processing and presentation pathways. The identified changes in EZH2 were associated with an adverse prognosis in the TCGA dataset. These results suggest that inhibiting of EZH2 is a promising therapeutic avenue for a substantial fraction of melanoma patients.

Topics: DNA Copy Number Variations; DNA Methylation; Enhancer of Zeste Homolog 2 Protein; Epigenesis, Genetic; Gene Expression Regulation, Neoplastic; Genes, Tumor Suppressor; Humans; Immunity, Innate; Indoles; Melanoma; Microarray Analysis; Mutation; Polymorphism, Single Nucleotide; Pyridones

The epigenetic modifier EZH2 is part of the polycomb repressive complex that suppresses gene expression via histone methylation. Activating mutations in EZH2 are found in a subset of melanoma that contributes to disease progression by inactivating tumor suppressor genes. In this study we have targeted EZH2 with a specific inhibitor (GSK126) or depleted EZH2 protein by stable shRNA knockdown. We show that inhibition of EZH2 has potent effects on the growth of both wild-type and EZH2 mutant human melanoma in vitro particularly in cell lines harboring the EZH2Y646 activating mutation. This was associated with cell cycle arrest, reduced proliferative capacity in both 2D and 3D culture systems, and induction of apoptosis. The latter was caspase independent and mediated by the release of apoptosis inducing factor (AIFM1) from mitochondria. Gene expression arrays showed that several well characterized tumor suppressor genes were reactivated by EZH2 inhibition. This included activating transcription factor 3 (ATF3) that was validated as an EZH2 target gene by ChIP-qPCR. These results emphasize a critical role for EZH2 in the proliferation and viability of melanoma and highlight the potential for targeted therapy against EZH2 in treatment of patients with melanoma.

Topics: Activating Transcription Factor 3; Apoptosis; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Cell Survival; Disease Progression; Enhancer of Zeste Homolog 2 Protein; Epigenomics; Gene Expression Profiling; Gene Expression Regulation, Neoplastic; Gene Silencing; Genes, Tumor Suppressor; Genotype; Histones; Humans; Indoles; Inhibitory Concentration 50; Melanoma; Mutation; Polycomb Repressive Complex 2; Pyridones; Skin Neoplasms; Up-Regulation