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Reactivation of death receptor 4 (DR4) expression sensitizes medulloblastoma cell lines to TRAIL

  • laboratory Investigation - Human/Animal Tissue
  • Published:
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Abstract

Object Apoptosis, a key cellular response to therapeutic agents is often inactivated in tumor cells. In this study, we evaluated the expression of the tumor necrosis family of death receptors, DR4 and DR5, in medulloblastoma tumor samples and cell lines to determine if epigenetic modulation of gene expression could sensitize tumor cell lines to TRAIL-mediated apoptosis. Methods Human medulloblastoma samples and cell lines were analyzed for DR4 and DR5 expression by quantitative PCR and immunofluorescence assays. Cell lines with downregulated expression of one or both genes were treated with the histone deacetylase inhibitor, MS-275, and the expression of DR4 and DR5 measured by quantitative PCR, Western blotting, flow cytometry and chromatin immunoprecipitation assays. Induction of apoptosis in the presence of MS-275 was evaluated by TUNEL assay and its ability to augment TRAIL-mediated cytotoxicity was determined by MTT assays, Western blotting and flow cytometry. Results Compared to normal cerebellum, DR4, but not DR5 expression was consistently downregulated in medulloblastoma tumor samples and in Daoy and D283 cell lines. Interestingly, MS-275 decreased cell growth and induced apoptosis in Daoy and D283 cells. In Daoy cells, this coincided with increased histone H3 and H4 acetylation at the DR4 promoter and enhanced DR4 gene and protein expression as well as elevated Caspase-8 activity. The involvement of DR4 in the cellular response to MS-275 was further confirmed by the observation that knockdown of DR4 and FADD abrogated apoptosis. Further, addition of TRAIL to MS-275 treated cells resulted in an enhancement of apoptosis, suggesting that the upregulated death receptors were functional. Conclusion Our study provides an understanding of the role of DR4 in apoptosis of medulloblastoma cell lines and suggests a potential contribution of aberrant histone deacetylation to the resistance of medulloblastoma cells to therapeutic death.

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Abbreviations

FADD:

FAS-associated death domain

MTT:

3-(4, 5 dimethylthiazole-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-Tetrazolium

TRAIL:

TNF-related apoptosis-inducing ligand

MS-275:

N-(2-aminophenyl)-4-[(N-pyridine-3-ylmetoxycarbonyl) aminomethyl] Benzamide

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Acknowledgements

We would like to thank Drs. Peter Zage, Joya Chandra and Douglas Cardillo for their input on this manuscript. We also wish to thank Dr. Ashish Kamath for sharing xenografts of bladder cancer for our analyses.

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Author notes

  1. Neeta D. Sinnappah-Kang

    Present address: Betty Cowan Research and Innovation Centre, Christian Medical College and Hospital, Brown Road, Ludhiana, Punjab, 141008, India

Authors and Affiliations

  1. Department of Pediatrics, University of Texas, M.D. Anderson Cancer Center, 1515 Holcombe Blvd, Unit 853, Houston, TX, 77030, USA

    Dolly G. Aguilera, Chandra M. Das, Neeta D. Sinnappah-Kang, Celine Joyce, Pete H. Taylor, Johannes E. Wolff & Vidya Gopalakrishnan

  2. Department of Biostatistics, University of Texas at Houston, 1515 Holcombe Blvd, Houston, TX, 77030, USA

    Sijin Wen

  3. Institute of Neuropathology, University Hospital, 48129, Munster, Germany

    Martin Hasselblatt & Werner Paulus

  4. Department of Pathology, University of Texas, M.D. Anderson Cancer Center, 1515 Holcombe Blvd, Unit 853, Houston, TX, 77030, USA

    Greg Fuller

  5. Graduate School of Biomedical Sciences, University of Texas at Houston, 1515 Holcombe Blvd, Houston, TX, 77225, USA

    Johannes E. Wolff & Vidya Gopalakrishnan

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  1. Dolly G. Aguilera
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Correspondence to Vidya Gopalakrishnan.

Additional information

This work was supported in part by Award P50CA127001 from the National Cancer Institutes and Grants from the National Brain Tumor Foundation and the American Cancer Society-IRG to VG.

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Aguilera, D.G., Das, C.M., Sinnappah-Kang, N.D. et al. Reactivation of death receptor 4 (DR4) expression sensitizes medulloblastoma cell lines to TRAIL. J Neurooncol 93, 303–318 (2009). https://doi.org/10.1007/s11060-008-9788-x

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