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Heat shock protein expression analysis in canine osteosarcoma reveals HSP60 as a potentially relevant therapeutic target

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Cell Stress and Chaperones Aims and scope

Abstract

Heat shock proteins (HSP) are highly conserved across eukaryotic and prokaryotic species. These proteins play a role in response to cellular stressors, protecting cells from damage and facilitating recovery. In tumor cells, HSPs can have cytoprotective effects and interfere with apoptotic cascades. This study was performed to assess the prognostic and predictive values of the gene expression of HSP family members in canine osteosarcoma (OS) and their potential for targeted therapy. Gene expressions for HSP were assessed using quantitative PCR (qPCR) on 58 snap-frozen primary canine OS tumors and related to clinic-pathological parameters. A significant increased expression of HSP60 was found in relation to shorter overall survival and an osteoblastic phenotype. Therefore, the function of HSP60 was investigated in more detail. Immunohistochemical analysis revealed heterogeneous staining for HSP60 in tumors. The highest immunoreactivity was found in tumors of short surviving dogs. Next HSP expression was shown in a variety of canine and human OS cell lines by qPCR and Western blot. In two highly metastatic cell lines HSP60 expression was silenced using siRNA resulting in decreased cell proliferation and induction of apoptosis in both cell lines. It is concluded that overexpression of HSP60 is associated with a poor prognosis of OS and should be evaluated as a new target for therapy.

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Acknowledgments

The authors would like to acknowledge the following individuals for their various contributions to the project: Malaysian Ministry of Higher Education (MOHE) for funding the PhD candidate working on this project; Peter van Kooten (Department of Infectious Diseases and Immunology, Utrecht University) for providing the LK-1 HSP60 antibody; Monique E. Van Wolferen for assisting in RNA isolation and cDNA synthesis; Annette van Drie (Department of Pathobiology, Utrecht University) for assistance with preparation of histology slides; Frank Riemers for valuable input on cell transfection protocols, siGlo transfection optimizations, western blotting and QPCR analyses; Ellen Martens for help maintaining cells in culture; Dr. Chand Khanna, Christina Mazcko (National Cancer Intsitute, NIH, USA) and Prof. Farese (University of Florida, USA) for kindly providing the canine OS cell lines utilized in this study. We also thank Jantine Posthuma de Boer from the University of Amsterdam and Dr. A.M.Cleton-Jansen from the Department of Pathobiology at the University of Leiden for providing the human OS cell lines and paraffin-embedded human OS tissue sections, respectively, for the comparative analysis.

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Authors and Affiliations

  1. Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, 3584 CM, Utrecht, The Netherlands

    Gayathri T. Selvarajah, Floor A. S. Bonestroo, Jolle Kirpensteijn, Elpetra P. M. Timmermans-Sprang, Adri Slob & Jan A. Mol

  2. Department of Veterinary Clinical Studies, Faculty of Veterinary Medicine, University Putra Malaysia, 43400 UPM, Serdang, Malaysia

    Gayathri T. Selvarajah

  3. Department of Pathobiology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht, The Netherlands

    Marja J. L. Kik

  4. Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584 CL, Utrecht, The Netherlands

    Ruurd van der Zee & Willem van Eden

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  1. Gayathri T. Selvarajah
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  2. Floor A. S. Bonestroo
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  8. Adri Slob
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  9. Jan A. Mol
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Correspondence to Jan A. Mol.

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Selvarajah, G.T., Bonestroo, F.A.S., Kirpensteijn, J. et al. Heat shock protein expression analysis in canine osteosarcoma reveals HSP60 as a potentially relevant therapeutic target. Cell Stress and Chaperones 18, 607–622 (2013). https://doi.org/10.1007/s12192-013-0414-2

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  • DOI: https://doi.org/10.1007/s12192-013-0414-2

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