Helicase CHD4 is an epigenetic coregulator of PAX3-FOXO1 in alveolar rhabdomyosarcoma
Maria Böhm, Marco Wachtel, Joana G. Marques, Natalie Streiff, Dominik Laubscher, Paolo Nanni, Kamel Mamchaoui, Raffaella Santoro, Beat W. Schäfer
Maria Böhm, Marco Wachtel, Joana G. Marques, Natalie Streiff, Dominik Laubscher, Paolo Nanni, Kamel Mamchaoui, Raffaella Santoro, Beat W. Schäfer
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Research Article Oncology

Helicase CHD4 is an epigenetic coregulator of PAX3-FOXO1 in alveolar rhabdomyosarcoma

Abstract

A vast number of cancer genes are transcription factors that drive tumorigenesis as oncogenic fusion proteins. Although the direct targeting of transcription factors remains challenging, therapies aimed at oncogenic fusion proteins are attractive as potential treatments for cancer. There is particular interest in targeting the oncogenic PAX3-FOXO1 fusion transcription factor, which induces alveolar rhabdomyosarcoma (aRMS), an aggressive cancer of skeletal muscle cells for which patient outcomes remain dismal. In this work, we have defined the interactome of PAX3-FOXO1 and screened 60 candidate interactors using siRNA-mediated depletion to identify candidates that affect fusion protein activity in aRMS cells. We report that chromodomain helicase DNA binding protein 4 (CHD4), an ATP-dependent chromatin remodeler, acts as crucial coregulator of PAX3-FOXO1 activity. CHD4 interacts with PAX3-FOXO1 via short DNA fragments. Together, they bind to regulatory regions of PAX3-FOXO1 target genes. Gene expression analysis suggested that CHD4 coregulatory activity is essential for a subset of PAX3-FOXO1 target genes. Depletion of CHD4 reduced cell viability of fusion-positive but not of fusion-negative RMS in vitro, which resembled loss of PAX3-FOXO1. It also caused specific regression of fusion-positive xenograft tumors in vivo. Therefore, this work identifies CHD4 as an epigenetic coregulator of PAX3-FOXO1 activity, providing rational evidence for CHD4 as a potential therapeutic target in aRMS.

Authors

Maria Böhm, Marco Wachtel, Joana G. Marques, Natalie Streiff, Dominik Laubscher, Paolo Nanni, Kamel Mamchaoui, Raffaella Santoro, Beat W. Schäfer

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Figure 7

CHD4 inhibition causes tumor regression in mouse xenografts.

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CHD4 inhibition causes tumor regression in mouse xenografts. In vivo tre...
In vivo treatment of NOD/SCID mice engrafted with RH4 cells containing stably integrated doxycycline-inducible shCHD4#1 or shscr expression vectors. Mice bearing palpable tumors were treated i.p. for 2 days with either vehicle control or doxycycline at a dose of 53.3 mg/kg. Additionally, they were fed with doxycycline-supplemented or control food. (A) Absolute tumor volumes of FP-RMS xenografts as measured by caliper. Vehicle-treated control groups consisted of 6 mice, and doxycycline-treated groups consisted of 5 (shscr) and 7 (shCHD4#1) mice. (B) CHD4 mRNA expression level in indicated tumors of doxycycline- or vehicle control–treated mice analyzed by quantitative real-time PCR. Tumors were isolated at the end of treatment (black; tumor set shown in A) or after 4 days of treatment (red; separate tumor set). Fold changes of mRNA expression are normalized to the mean of CHD4 expression in shscr vehicle control–treated tumors. (C) Immunohistochemical staining of tumors with indicated antibodies. Tumors were isolated 4 days after the start of treatment.