FISH assay development for the detection of p16/CDKN2A deletion in malignant pleural mesothelioma.
OBJECTIVE
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.


METHODS
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.


CONCLUSIONS
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.
Chung
Catherine T-S
CT
Division of Pathology, Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada.
Santos
Gilda Da Cunha
Gda C
Hwang
David M
DM
Ludkovski
Olga
O
Pintilie
Melania
M
Squire
Jeremy A
JA
Tsao
Ming-Sound
MS
D001194:Asbestos / Q000009:adverse effects; D001706:Biopsy; D005260:Female; D017353:Gene Deletion; D019942:Genes, p16; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008297:Male; D008654:Mesothelioma / Q000175:diagnosis* / Q000209:etiology / Q000235:genetics / Q000473:pathology; D008875:Middle Aged; D010997:Pleural Neoplasms / Q000175:diagnosis* / Q000209:etiology / Q000235:genetics / Q000473:pathology; D011379:Prognosis; D011446:Prospective Studies; D012907:Smoking / Q000009:adverse effects; D016019:Survival Analysis
D001194:Asbestos