Arsenic trioxide inhibits human cancer cell growth and tumor development in mice by blocking Hedgehog/GLI pathway
Elspeth M. Beauchamp, Lymor Ringer, Gülay Bulut, Kamal P. Sajwan, Michael D. Hall, Yi-Chien Lee, Daniel Peaceman, Metin Özdemirli, Olga Rodriguez, Tobey J. Macdonald, Chris Albanese, Jeffrey A. Toretsky, Aykut Üren
Elspeth M. Beauchamp, Lymor Ringer, Gülay Bulut, Kamal P. Sajwan, Michael D. Hall, Yi-Chien Lee, Daniel Peaceman, Metin Özdemirli, Olga Rodriguez, Tobey J. Macdonald, Chris Albanese, Jeffrey A. Toretsky, Aykut Üren
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Research Article

Arsenic trioxide inhibits human cancer cell growth and tumor development in mice by blocking Hedgehog/GLI pathway

Abstract

The Hedgehog (Hh) pathway is activated in some human cancers, including medulloblastoma. The glioma-associated oncogene homolog (GLI) transcription factors are critical mediators of the activated Hh pathway, and their expression may be elevated in some tumors independent of upstream Hh signaling. Thus, therapies targeting GLI transcription factors may benefit a wide spectrum of patients with mutations at different nodal points of the Hh pathway. In this study, we present evidence that arsenic trioxide (ATO) suppresses human cancer cell growth and tumor development in mice by inhibiting GLI1. Mechanistically, ATO directly bound to GLI1 protein, inhibited its transcriptional activity, and decreased expression of endogenous GLI target genes. Consistent with this, ATO inhibited the growth of human cancer cell lines that depended on upregulated GLI expression in vitro and in vivo in a xenograft model of Ewing sarcoma. Furthermore, ATO improved survival of a clinically relevant spontaneous mouse model of medulloblastoma with activated Hh pathway signaling. Our results establish ATO as a Hh pathway inhibitor acting at the level of GLI1 both in vitro and in vivo. These results warrant the clinical investigation of ATO for tumors with activated Hh/GLI signaling, in particular patients who develop resistance to current therapies targeting the Hh pathway upstream of GLI.

Authors

Elspeth M. Beauchamp, Lymor Ringer, Gülay Bulut, Kamal P. Sajwan, Michael D. Hall, Yi-Chien Lee, Daniel Peaceman, Metin Özdemirli, Olga Rodriguez, Tobey J. Macdonald, Chris Albanese, Jeffrey A. Toretsky, Aykut Üren

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Figure 4

ATO directly binds to GLI1.

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ATO directly binds to GLI1. (A) Coomassie-stained gel shows induction of...
(A) Coomassie-stained gel shows induction of full-length GLI1 protein expression in bacteria. Protein in the inclusion bodies was bound to Ni+ NTA column, refolded on the column, and eluted with an imidazole gradient. IPTG, isopropyl-D-1-thiogalactopyranoside. (B) 40 nM full-length GLI1 protein was preincubated with either BAL (50 μM for 10 minutes) or ATO (200, 100, 50, and 25 μM for 2 hours). 200 nM FlAsH-EDT was then added for 20 minutes. EWS-FLI1 with 200 nM FlAsH was used as negative control. Fluorescence intensity was measured with excitation at 508 nm and emission at 528 nm; relative fluorescence intensity is expressed as the fold change over the control (FlAsH alone). *P = 0.02, 2-tailed Student’s t test. (C) COS7 cells were transfected with EGFP alone, EGFP-DVL3, and EGFP-GLI1 and then treated with 2.5 μM ReAsH-EDT2 for 1 hour. Colocalization was examined by confocal fluorescence microscopy (all images are at ×600 magnification).