Aberrant p27^Kip1 promoter methylation in malignant melanoma

Abstract

p27^Kip1 is a regulator of the mammalian cell cycle and a putative tumor suppressor. Distinct altered patterns of p27^Kip1 protein expression are found in a variety of human carcinomas, and p27^Kip1 expression levels usually correlate directly with disease-free survival. The mechanism(s) by which p27^Kip1 expression is reduced or lost during tumorigenesis remains unclear. Specific alterations of the p27^Kip1 gene, including mutations and homozygous deletions, are exceedingly rare in human cancers. We have used methylation-specific PCR and bisulfite genomic sequencing to examine the methylation status of p27^Kip1 in 61 primary and metastatic tumors and 35 cell lines from patients with malignant melanoma. Dense methylation of a CpG island in the promoter region of p27^Kip1 was detected in four of 45 metastatic tumors (9%) and three of the cell lines (9%), including two cell lines established from two different metastases from the same patient. Examination of a naturally occurring, allele-specific sequence variant demonstrated that p27^Kip1 methylation is associated with transcriptional silencing in situ. Cell lines with p27^Kip1 methylation showed retention of the wild-type allele and detectable p27^Kip1 protein whose abundance was reduced compared with normal melanocytes. Collectively, our data suggest that DNA methylation may be a cause of monoallelic p27^Kip1 silencing in malignant melanoma, which would support a role for p27^Kip1 haploinsufficiency in human cancer.