Elsevier

Human Pathology

Volume 35, Issue 1, January 2004, Pages 25-33
Human Pathology

Original contribution
Expression and localization of mutant p16 proteins in melanocytic lesions from familial melanoma patients

https://doi.org/10.1016/j.humpath.2003.08.017Get rights and content

Abstract

Little is known about the correlation between the loss of p16 expression and tumor progression in familial melanoma; no systematic study has been conducted on p16 expression in melanocytic tumors from patients carrying germline CDKN2A mutations. We analyzed 98 early primary lesions from familial patients, previously tested for germline CDKN2A status, by quantitative immunohistochemistry using 3 p16 antibodies. We found that p16 expression was inversely correlated with tumor progression and was significantly lower in melanomas, including in situ lesions, than in nevi. Of other features analyzed, tumor thickness showed the most significant correlation with p16 levels. Lesions from mutation-negative patients displayed combined nuclear and cytoplasmic staining. However, some mutation-positive lesions (ie, G101W, 113insR, M53I, R24P, and 33ins24), including benign nevi, showed nuclear mislocalization, confirming previous studies suggesting that subcellular distribution indicates functional impairment of p16.

Section snippets

Tissue samples

A total of 98 formalin-fixed, paraffin-embedded lesions were collected from patients belonging to melanoma kindreds. Samples came from pathology units in Italy, Sweden, and Australia, and all assays were performed in the same laboratory in Italy. In detail, 49 lesions came from the Department of Pathology at San Martino Hospital and 9 came from the Department of Pathology at the National Institute for Cancer Research, both in Genova. One lesion was provided by the San Gallicano Hospital in

Mutational analysis of CDKN2A in familial melanoma patients

Of the 37 families from Italy, Sweden, and Australia analyzed for CDKN2A status, 25 were found to be mutation-positive. Twelve of the 24 Italian families had no mutations, and 12 carried the G101W mutation (2 of which had not been described previously). The 9 Australian families carried a variety of mutations13 in either exon 1 (9del24, 33ins24, L16P, R24P, L32P, and G35A) or exon 2 (M53I), and the 4 families from Sweden carried the 113insR mutation12 (Table 1).

Specificity and establishment of reproducible conditions for all antibodies

Lesions were tested with at

Discussion

Because the progression stages of melanoma have been well characterized, genetic studies are essential for the development of a model of melanoma pathogenesis.18 The INK4a/ARF locus encodes 2 cell cycle regulatory proteins, the CDK inhibitor p16INK4a and the p53 activator p14ARF. Alterations affecting this locus occur in familial melanoma kindreds and cause functional and quantitative defects in p16 and p14 expression.5 Although the role of CDKN2A has been widely studied, few efforts have been

Acknowledgements

The authors thank L. Luzzatto for advice and critical reading of the manuscript, and P. Grammatico and A. Amantea for providing a lesion from a mutated patient. Special thanks to S. Gargiulo for helping to prepare sections from Italian patients, M. Mantelli and L. Pastorino for providing technical assistance, and L. V. Battistuzzi for assisting in the preparation of the manuscript.

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    Supported by grants FIRB post genoma RBNE014975 2001, Ministry of Health ICS 030.1/RF99.32, and AIRC 2002 (to G.B.S.); grants from the Cancer Society of Stockholm, the King Gustav V Jubilee Fund, the Karolinska Institute Research Funds, and the Swedish Radiation Protection Institute (to J.H. and A.P.); and funding from the National Health and Medical Research Council of Australia (to N.H.)

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