Phosphorylation-mediated EZH2 inactivation promotes drug resistance in multiple myeloma
Jiro Kikuchi, … , Bjarne Bogen, Yusuke Furukawa
Jiro Kikuchi, … , Bjarne Bogen, Yusuke Furukawa
Published October 26, 2015
Citation Information: J Clin Invest. 2015;125(12):4375-4390. https://doi.org/10.1172/JCI80325.
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Research Article Oncology

Phosphorylation-mediated EZH2 inactivation promotes drug resistance in multiple myeloma

Abstract

Alterations in chromatin modifications, such as histone methylation, have been suggested as mediating chemotherapy resistance in several cancer types; therefore, elucidation of the epigenetic mechanisms that underlie drug resistance may greatly contribute to the advancement of cancer therapies. In the present study, we identified histone H3–lysine 27 (H3K27) as a critical residue for epigenetic modification in multiple myeloma. We determined that abrogation of drug-induced H3K27 hypermethylation is associated with cell adhesion–mediated drug resistance (CAM-DR), which is the most important form of drug resistance, using a coculture system to evaluate stroma cell adhesion–dependent alterations in multiple myeloma cells. Cell adhesion counteracted anticancer drug–induced hypermethylation of H3K27 via inactivating phosphorylation of the transcription regulator EZH2 at serine 21, leading to the sustained expression of antiapoptotic genes, including IGF1, B cell CLL/lymphoma 2 (BCL2), and hypoxia inducible factor 1, α subunit (HIF1A). Pharmacological and genetic inhibition of the IGF-1R/PI3K/AKT pathway reversed CAM-DR by promoting EZH2 dephosphorylation and H3K27 hypermethylation both in vitro and in refractory murine myeloma models. Together, our findings identify and characterize an epigenetic mechanism that underlies CAM-DR and suggest that kinase inhibitors to counteract EZH2 phosphorylation should be included in combination chemotherapy to increase therapeutic index.

Authors

Jiro Kikuchi, Daisuke Koyama, Taeko Wada, Tohru Izumi, Peter O. Hofgaard, Bjarne Bogen, Yusuke Furukawa

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Figure 3

H3K27 hypermethylation is correlated with drug sensitivity but is not a consequence of cell death.

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H3K27 hypermethylation is correlated with drug sensitivity but is not a ...
(A) KMS12-BM cells were harvested after 48 hours of culture described in Figure 2A and stained with Alexa Fluor 488–conjugated anti-H3K27me3 and 7-AAD for flow cytometric analysis. (B) The y axis shows the proportion of cells positive for both H3K27me3 and 7-AAD. P values were calculated by 1-way ANOVA with the Student-Newman-Keuls multiple comparisons test (n = 3). (C) Whole cell lysates were prepared from RPMI8226 cells cultured with 0.4 μM ADM at the indicated time points and subjected to immunoblot analyses for H3K27 trimethylation and caspase-9 activation. (D) RPMI8226 cells were cultured in the absence (–) or presence (+) of 0.4 μM ADM with or without a peptide inhibitor for caspase-9 (z-LEHD-fmk). Left panel: after 72 hours of culture, cell proliferation was determined by the MTT reduction assay and expressed as a percentage of the values of corresponding untreated cells. Mean ± SD of 3 independent experiments are shown. P values were calculated by 1-way ANOVA with the Student-Newman-Keuls multiple comparisons test. *P <0.05. Right panel: whole cell lysates were prepared after 48 hours for immunoblotting.