Histone methyltransferases EHMT1 and EHMT2 (GLP/G9A) maintain PARP inhibitor resistance in high-grade serous ovarian carcinoma
| Autor: | Miriam D. Post, Kian Behbakht, Connor J. Hughes, Alexandra McMellen, Hyunmin Kim, Tomomi M. Yamamoto, Benjamin G. Bitler, Zachary L. Watson, Lindsay J. Wheeler |
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| Jazyk: | angličtina |
| Rok vydání: | 2019 |
| Předmět: |
Resistance
H3K9me2 Mass Spectrometry Piperazines Histones Mice 0302 clinical medicine Histocompatibility Antigens Genetics (clinical) Ovarian Neoplasms 0303 health sciences EHMT1 EHMT2 Chemistry Cell cycle 3. Good health 030220 oncology & carcinogenesis Histone methyltransferase PARP inhibitor Disease Progression Female DNA damage DNA repair Poly(ADP-ribose) Polymerase Inhibitors 03 medical and health sciences Histone H3 Ovarian cancer Cell Line Tumor Genetics Animals Humans HGSOC Epigenetics Molecular Biology 030304 developmental biology Sequence Analysis RNA Research Gene Expression Profiling Histone-Lysine N-Methyltransferase Xenograft Model Antitumor Assays Cystadenocarcinoma Serous Drug Resistance Neoplasm Cancer research Phthalazines Neoplasm Grading Developmental Biology |
| Zdroj: | Clinical Epigenetics |
| ISSN: | 1868-7083 1868-7075 |
| Popis: | BackgroundEuchromatic histone-lysine-N-methyltransferases 1 and 2 (EHMT1/2, aka GLP/G9A) catalyze dimethylation of histone H3 lysine 9 (H3K9me2) and have roles in epigenetic silencing of gene expression. EHMT1/2 also have direct roles in DNA repair and are implicated in chemoresistance in several cancers. Resistance to chemotherapy and PARP inhibitors (PARPi) is a major cause of mortality in high-grade serous ovarian carcinoma (HGSOC), but the contribution of the epigenetic landscape is unknown.ResultsTo identify epigenetic mechanisms of PARPi resistance in HGSOC, we utilized unbiased exploratory techniques, including RNA-Seq and mass spectrometry profiling of histone modifications. Compared to sensitive cells, PARPi-resistant HGSOC cells display a global increase of H3K9me2 accompanied by overexpression of EHMT1/2. EHMT1/2 overexpression was also observed in a PARPi-resistant in vivo patient-derived xenograft (PDX) model. Genetic or pharmacologic disruption of EHMT1/2 sensitizes HGSOC cells to PARPi. Cell death assays demonstrate that EHMT1/2 disruption does not increase PARPi-induced apoptosis. Functional DNA repair assays show that disruption of EHMT1/2 ablates homologous recombination (HR) and non-homologous end joining (NHEJ), while immunofluorescent staining of phosphorylated histone H2AX shows large increases in DNA damage. Propidium iodide staining and flow cytometry analysis of cell cycle show that PARPi treatment increases the proportion of PARPi-resistant cells in S and G2 phases, while cells treated with an EHMT1/2 inhibitor remain in G1. Co-treatment with PARPi and EHMT1/2 inhibitor produces an intermediate phenotype. Immunoblot of cell cycle regulators shows that combined EHMT1/2 and PARP inhibition reduces expression of specific cyclins and phosphorylation of mitotic markers. These data suggest DNA damage and altered cell cycle regulation as mechanisms of sensitization. RNA-Seq of PARPi-resistant cells treated with EHMT1/2 inhibitor showed significant gene expression changes enriched in pro-survival pathways that remain unexplored in the context of PARPi resistance, including PI3K, AKT, and mTOR.ConclusionsThis study demonstrates that disrupting EHMT1/2 sensitizes HGSOC cells to PARPi, and suggests a potential mechanism through DNA damage and cell cycle dysregulation. RNA-Seq identifies several unexplored pathways that may alter PARPi resistance. Further study of EHMT1/2 and regulated genes will facilitate development of novel therapeutic strategies to successfully treat HGSOC. |
| Databáze: | OpenAIRE |
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