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Antimyeloma activity of bromodomain inhibitors on the human myeloma cell line U266 by downregulation of MYCL.

ABSTRACT: Bromodomain and extraterminal protein (BET) inhibitors suppress the expression of c-MYC. U266, a human myeloma cell line, expresses the MYCL gene, but not the c-MYC gene. Our aim was to analyse the antimyeloma activity of BET inhibitors on U266 cells. Two BET inhibitors, I-BET151 and JQ1, were tested. U266 cell proliferation decreased to 61.5 and 54.0% of the control after incubation with 500 nmol/l I-BET151 for 72 and 96 h and to 53.5 and 56.4% of control after incubation with 500 nmol/l JQ1 for 72 and 96 h by MTS tetrazolium, respectively. BET inhibitors induced cell cycle arrest at the G1 phase in U266 cells, but did not induce apoptosis by flow cytometry. According to Gene Set Enrichment Analysis, MYC-related genes were significantly downregulated in U266 cells treated with I-BET151 similar to KMS11 cells that expressed c-MYC. The MYCL1 was expressed in U266 cells, whereas c-MYC and MYCN were not by quantitative real-time reverse-transcription-PCR. Incubation with I-BET151 induced downregulation of MYCL1 in U266 cells. BET inhibitors decreased the cell proliferation in U266 cells with overexpression of MYCL less than those without overexpression of MYCL. BET inhibitors induce G1 arrest without apoptosis and interfere with the proliferation of U266 myeloma cells, which express MYCL, but not c-MYC. BET inhibitors might be active in cancers that express MYCL, but not c-MYC.

SUBMITTER: Suzuki K

PROVIDER: S-EPMC4969056 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Antimyeloma activity of bromodomain inhibitors on the human myeloma cell line U266 by downregulation of MYCL.

Suzuki Kazuhito K Yamamoto Kouhei K Arakawa Yasuhiro Y Yamada Hisashi H Aiba Keisuke K Kitagawa Masanobu M

Anti-cancer drugs 20160901 8

Bromodomain and extraterminal protein (BET) inhibitors suppress the expression of c-MYC. U266, a human myeloma cell line, expresses the MYCL gene, but not the c-MYC gene. Our aim was to analyse the antimyeloma activity of BET inhibitors on U266 cells. Two BET inhibitors, I-BET151 and JQ1, were tested. U266 cell proliferation decreased to 61.5 and 54.0% of the control after incubation with 500 nmol/l I-BET151 for 72 and 96 h and to 53.5 and 56.4% of control after incubation with 500 nmol/l JQ1 fo ...[more]

PMID: 27276402

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Project description:Multiple Myeloma (MM) remains incurable, and new drugs with novel mechanisms of action are still needed. In this report, we have analyzed the action of Zalypsis, an alkaloid analogous to certain natural marine compounds, in MM. Zalypsis turned out to be the most potent antimyeloma agent we have tested so far, with IC50s from picomolar to low nanomolar ranges. It also showed remarkable ex vivo potency in plasma cells from patients and in MM cells in vivo xenografted in mice. Besides the induction of apoptosis and cell cycle arrest, Zalypsis provoked DNA double-strand-breaks (DSB), evidenced by an increase in phospho-Histone-H2AX and phospho-CHK2, followed by a striking overexpression of p53 in p53-wild type cell lines. In addition, in those cell lines in which p53 was mutated, Zalypsis also provoked DSB and induced cell death, although higher concentrations were required. Immunohistochemical studies in tumours also demonstrated Histone-H2AX phosphorylation and p53 overexpression. Gene expression profile studies were concordant with these results, revealing an important deregulation of genes involved in DNA-damage response. The potent in vitro and in vivo antimyeloma activity of Zalypsis uncovers the high sensitivity of tumour plasma cells to DSB, and strongly supports the use of this compound in MM patients. Experiment Overall Design: MM cells treated in vitro with Zalypsis (5 nM) were harvested at the beginning of induction of cell death (15-20% cell death as assessed by Annexin V-FITC staining). Total RNA was extracted using Trizol reagent (Life Technologies, MD, USA) and purified with RNAeasy Mini Kit (Qiagen, CA, USA). RNA integrity was verified with the Agilent 2100 Bioanalyzer (Agilent, CA, USA). Double-stranded cDNA and biotinylated cRNA were synthesized with T7-polyT primer and the BioArray RNA labeling kit (Enzo, NY, USA), respectively. The labelled RNA was then fragmented and hybridized to HG-U133 Plus 2.0 oligonucleotide arrays (Affymetrix, CA, USA), which were scanned in a Gene Array Scanner and analyzed using the DNA-Chip Analyzer software (DChip). Changes in gene expression greater than two-fold were considered significant.

Dataset Information

Antimyeloma activity of bromodomain inhibitors on the human myeloma cell line U266 by downregulation of MYCL.

Publications

Antimyeloma activity of bromodomain inhibitors on the human myeloma cell line U266 by downregulation of MYCL.

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