Genetic and epigenetic aberrations of p16 in feline primary neoplastic diseases and tumor cell lines of lymphoid and non-lymphoid origins

- The tumor suppressor p16 is frequently inactivated in human cancer but poorly investigated in feline cancer.
- The feline p16 gene was cloned, and its polymorphism was characterized in domestic cats.
- Epigenetic regulation of the p16 was demonstrated in feline lymphoma cell lines.
- Genetic and epigenetic alterations of p16 were detected in feline cancer cell lines and primary cancers.
- Frequent promoter methylation of the p16 gene was observed in feline B-cell lymphoid malignancies.

The p16 gene acts as a tumor suppressor by regulating the cell cycle and is frequently inactivated in human and canine cancers. The aim of this study was to characterize genetic and epigenetic alterations of the p16 in feline lymphoid and non-lymphoid malignancies, using 74 primary tumors and 11 tumor cell lines. Cloning of feline p16 and subsequent sequence analysis revealed 11 germline sequence polymorphisms in control cats. Bisulfite sequencing analysis of the p16 promoter region in a feline lymphoma cell line revealed that promoter methylation was associated with decreased mRNA expression. Treatment with a demethylating agent restored mRNA expression of the silenced p16. PCR amplification and sequencing analysis detected homozygous loss (five tumors, 6.7%) and a missense mutation (one tumor, 1.4%) in the 74 primary tumors analyzed. Methylation-specific PCR analysis revealed promoter methylation in 10 primary tumors (14%). Promoter methylation was frequent in B cell lymphoid tumors (7/21 tumors, 33%). These genetic and epigenetic alterations were also observed in lymphoma and mammary gland carcinoma cell lines, but not detected in non-neoplastic control specimens. These data indicate that molecular alterations of the p16 locus may be involved in the development of specific types of feline cancer, and warrant further studies to evaluate the clinical value of this evolutionarily-conserved molecular alteration in feline cancers.