Genetic regulation of the RUNX transcription factor family has antitumor effects
Ken Morita, … , Hiroshi Sugiyama, Yasuhiko Kamikubo
Ken Morita, … , Hiroshi Sugiyama, Yasuhiko Kamikubo
Published May 22, 2017
Citation Information: J Clin Invest. 2017;127(7):2815-2828. https://doi.org/10.1172/JCI91788.
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Research Article Hematology Oncology Article has an altmetric score of 12

Genetic regulation of the RUNX transcription factor family has antitumor effects

Abstract

Runt-related transcription factor 1 (RUNX1) is generally considered to function as a tumor suppressor in the development of leukemia, but a growing body of evidence suggests that it has pro-oncogenic properties in acute myeloid leukemia (AML). Here we have demonstrated that the antileukemic effect mediated by RUNX1 depletion is highly dependent on a functional p53-mediated cell death pathway. Increased expression of other RUNX family members, including RUNX2 and RUNX3, compensated for the antitumor effect elicited by RUNX1 silencing, and simultaneous attenuation of all RUNX family members as a cluster led to a much stronger antitumor effect relative to suppression of individual RUNX members. Switching off the RUNX cluster using alkylating agent–conjugated pyrrole-imidazole (PI) polyamides, which were designed to specifically bind to consensus RUNX-binding sequences, was highly effective against AML cells and against several poor-prognosis solid tumors in a xenograft mouse model of AML without notable adverse events. Taken together, these results identify a crucial role for the RUNX cluster in the maintenance and progression of cancer cells and suggest that modulation of the RUNX cluster using the PI polyamide gene-switch technology is a potential strategy to control malignancies.

Authors

Ken Morita, Kensho Suzuki, Shintaro Maeda, Akihiko Matsuo, Yoshihide Mitsuda, Chieko Tokushige, Gengo Kashiwazaki, Junichi Taniguchi, Rina Maeda, Mina Noura, Masahiro Hirata, Tatsuki Kataoka, Ayaka Yano, Yoshimi Yamada, Hiroki Kiyose, Mayu Tokumasu, Hidemasa Matsuo, Sunao Tanaka, Yasushi Okuno, Manabu Muto, Kazuhito Naka, Kosei Ito, Toshio Kitamura, Yasufumi Kaneda, Paul P. Liu, Toshikazu Bando, Souichi Adachi, Hiroshi Sugiyama, Yasuhiko Kamikubo

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Figure 1

p53-dependent antitumor effect of RUNX1 depletion.

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p53-dependent antitumor effect of RUNX1 depletion. (A) Growth curves of ...
(A) Growth curves of MOLM-13 and MV4-11 cells transduced with control (sh_Luc) or with RUNX1 shRNAs (sh_Rx1 #1 and sh_Rx1 #2) in the presence of 3 μM doxycycline (n = 3). (B) RUNX1 depletion–mediated increase in number of cells with G0/G1 DNA content. Nondepleted and RUNX1-depleted MV4-11 cells were cultured in the presence of 3 μM doxycycline. Forty-eight hours after treatment, cells were harvested and analyzed by flow cytometry (n = 3). (C) Early apoptotic cell death induced by RUNX1 silencing. Nondepleted and RUNX1-depleted MV4-11 cells were treated as in B, and the early apoptotic cells (annexin V+ DAPI–) were scored by flow cytometric analysis (n = 3). (D) RUNX1 depletion–mediated stimulation of p53-dependent cell death pathway. Nondepleted and RUNX1-depleted MV4-11 cells were treated as in B. Cell lysates were analyzed by immunoblotting with the indicated antibodies. (E) Induction of p53-target genes in RUNX1-depleted MV4-11 cells. Nondepleted and RUNX1-depleted MV4-11 cells were treated as in B. Total RNA was prepared and then analyzed by real-time RT-PCR for the indicated p53-target genes. Values are normalized to that of control vector–transduced cells (n = 3). (F) Increase in half-life of p53 in RUNX1-silenced MV4-11 cells. MV4-11 cells were treated with 3 μM doxycycline as in B. Forty-eight hours after treatment, cells were exposed to cycloheximide (CHX). Cell lysates were then prepared at the indicated time points, and analyzed for p53 by immunoblotting. Results are normalized to that of control vector–transduced cells (n = 3). Data are the mean ± SEM values. *P < 0.05, **P < 0.01, ***P < 0.001, NS, not significant, by 2-tailed Student’s t test.