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Original article
FISH assay development for the detection of p16/CDKN2A deletion in malignant pleural mesothelioma
  1. Catherine T-S Chung1,2,
  2. Gilda Da Cunha Santos2,3,
  3. David M Hwang2,3,
  4. Olga Ludkovski4,
  5. Melania Pintilie5,
  6. Jeremy A Squire2,3,4,6,
  7. Ming-Sound Tsao2,3,4,6
  1. 1Division of Pathology, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
  2. 2Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
  3. 3Department of Pathology, University Health Network, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada
  4. 4Division of Applied Molecular Oncology, University Health Network, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada
  5. 5Division of Biostatistics, University Health Network, Ontario Cancer Institute/Princess Margaret Hospital, Toronto, Ontario, Canada
  6. 6Department of Medical Biophysics, University of Toronto, Toronto, Ontario, Canada
  1. Correspondence to Ming-Sound Tsao, Department of Pathology, University Health Network, 200 Elizabeth Street, 11th floor, Toronto, Ontario M5G 2C4, Canada; ming.tsao{at}uhn.on.ca

Abstract

Aims To develop a fluorescence in-situ hybridisation (FISH) assay for detecting p16/CDKN2A deletion on paraffin tissue sections for use as an ancillary test to distinguish reactive from malignant mesothelial proliferations.

Method Dual-colour FISH for p16/CDKN2A and chromosome 9 (CEP-9) was performed on 11 benign mesothelial proliferations and 54 malignant pleural mesothelioma (MPM) cases to establish cut-off values for p16/CDKN2A deletion. A third MYC probe was used to verify cases showing homozygous deletion. Eight equivocal biopsies were used for assay testing.

Results Cut-off values for p16/CDKN2A deletion were calculated based on FISH signalling patterns obtained from the benign controls (mean percent nuclei plus three standard deviations). Hemizygous deletion was defined as >44% of nuclei showing the hemizygous (one p16/CDKN2A, two CEP-9 signals) or >15% of nuclei showing the monosomy (one p16/CDKN2A, one CEP-9 signal) deletion patterns. None of the benign cases showed a homozygous deletion pattern (no p16/CDKN2A, at least one CEP-9 signal). In the malignant cases, the percentage of nuclei showing homozygous deletion ranged from 1% to 87%. Therefore, the cut-off value for homozygous deletion was defined as >10%. P16/CDKN2A deletion was detected in 61% (33/54) of MPM cases. Among the equivocal biopsies, four showed homozygous and one showed hemizygous p16/CDKN2A deletion. Age over 60 years, asbestos exposure and p16/CDKN2A deletion were associated with a worse prognosis.

Conclusion Distinction between benign and malignant mesothelial proliferations can be diagnostically challenging. FISH for p16/CDKN2A deletion is a useful test for confirming the diagnosis of MPM.

  • Mesothelioma
  • p16
  • CDKN2A
  • gene deletion
  • fluorescence in situ hybridisation
  • FISH
  • malignant tumours
  • pleura

This is an open-access article distributed under the terms of the Creative Commons Attribution Non-commercial License, which permits use, distribution, and reproduction in any medium, provided the original work is properly cited, the use is non commercial and is otherwise in compliance with the license. See: http://creativecommons.org/licenses/by-nc/2.0/ and http://creativecommons.org/licenses/by-nc/2.0/legalcode.

https://doi.org/10.1136/jcp.2010.076794

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    Footnotes

    • Funding This project was supported by grants from the Physicians' Services Incorporated Foundation (PSI) and the Canadian Cancer Society/National Cancer Institute of Canada. The funding sources had no involvement in the study design, collection, analysis and interpretation of data or in the writing of the report or decision to submit the paper for publication.

    • Competing interests None.

    • Ethics approval This study was conducted with the approval of the University Health Network.

    • Provenance and peer review Not commissioned; externally peer reviewed.