RESUMO
BACKGROUND: The tumor suppressors p14(ARF) (ARF) and p16(INK4A) (p16) are encoded by overlapping reading frames at the CDKN2A/INK4A locus on chromosome 9p21. In human melanoma, the accumulated evidence has suggested that the predominant tumor suppressor at 9p21 is p16, not ARF. However, recent observations from melanoma-prone families and murine melanoma models suggest a p16-independent tumor suppressor role for ARF. We analyzed a group of melanoma metastases and cell lines to investigate directly whether somatic alterations to the ARF gene support its role as a p16-independent tumor suppressor in human melanoma, assuming that two alterations (genetic and/or epigenetic) would be required to inactivate a gene. METHODS: We examined the p16/ARF locus in 60 melanoma metastases from 58 patients and in 9 human melanoma cell lines using multiplex ligation-dependent probe amplification and multiplex polymerase chain reaction (PCR) to detect deletions, methylation-specific PCR to detect promoter methylation, direct sequencing to detect mutations affecting ARF and p16, and, in a subset of 20 tumors, immunohistochemistry to determine the effect of these alterations on p16 protein expression. All statistical tests were two-sided. RESULTS: We observed two or more alterations to the ARF gene in 26/60 (43%) metastases. The p16 gene sustained two or more alterations in 13/60 (22%) metastases (P = .03). Inactivation of ARF in the presence of wild-type p16 was seen in 18/60 (30%) metastases. CONCLUSION: Genetic and epigenetic analyses of the human 9p21 locus indicate that modifications of ARF occur independently of p16 inactivation in human melanoma and suggest that ARF is more frequently inactivated than p16.
Assuntos
Inativação Gênica , Genes p16 , Melanoma/genética , Melanoma/patologia , Proteína Supressora de Tumor p14ARF/genética , Animais , Linhagem Celular Tumoral , Cromossomos Humanos Par 9 , Inibidor p16 de Quinase Dependente de Ciclina , Metilação de DNA , Deleção de Genes , Humanos , Immunoblotting , Imuno-Histoquímica , Melanoma Experimental/genética , Melanoma Experimental/patologia , Camundongos , Repetições de Microssatélites , Mutação , Proteínas de Neoplasias/genética , Reação em Cadeia da Polimerase/métodos , Regiões Promotoras GenéticasRESUMO
A recent study on an Asian population reported a six-nucleotide insertion-deletion polymorphism (-652 6N del) in the CASP8 promoter region to be strongly associated with a decreased risk of multiple types of cancer, including breast cancer (BC). Here, we investigate the effect of this deletion in four independent large European BC case-control studies, including data from a total of 7,753 cases and 7,921 controls. The combined per allele odds ratio (OR) was 0.97 (95% confidence interval (CI), 95% CI = 0.93-1.02). The present result indicates that the CASP8 -652 6N del variant has no significant effect on BC risk in Europeans.