The p15^INK4b/p16^INK4a/RB1 Pathway Is Frequently Deregulated in Human Pituitary Adenomas

Abstract

Pituitary adenomas are common benign intracranial neoplasms. However, their tumorigenesis is not yet clearly defined. Inactivation of genes involved in the negative cell-cycle regulatory p15(INK4b) - p16(INK4a) -cyclin D/CDK4-RB1-mediated pathway (RB1 pathway) is one of the most common and important mechanisms in the growth advantage of tumor cells. Recently, much attention has been focused on the importance of alternative mechanisms of gene inactivation, particularly promoter hypermethylation in the transcriptional silencing of such tumor-suppressor genes. Based on the rare occurrence of inactivation by gene mutations and deletions of the RB1 pathway in pituitary adenomas, we investigated the deregulation of the RB1 pathway in 42 sporadic human pituitary adenomas, especially focusing on the methylation status of this pathway as determined by a methylation-specific polymerase chain reaction assay. Homozygous deletion of the p15(INK4b) or p16(INK4a) gene was detected in one adenoma each. Amplification of the CDK4 gene was not apparent in any of the pituitary adenomas presently examined. Promoter hypermethylation of the p15(INK4b), p16(INK4a), and RB1 genes was detected in 15 (35.7%), 30 (71.4%), and 12 (28.6%) of the adenomas, respectively. Promoter hypermethylation of the p15(INK4b) gene coincided with p16(INK4a) alteration and/or RB1 methylation, whereas p16(INK4a) and RB1 methylations tended to be mutually exclusive (p = 0.019). Thus, the vast majority of the adenomas (38 of 42, 90.5%) displayed alterations of the RB1 pathway. None of the clinicopathologic features, including the proliferation cell index, was significantly correlated with any particular methylation status. Our results suggest that inactivation of the RB1 pathway may play a causal role in pituitary tumorigenesis, with hypermethylation of the p16(INK4a) gene being the most common deregulation, and further provide evidence that RB1 and p16(INK4a) methylations tend to be mutually exclusive but occasionally coincide with p15(INK4b) methylation.

Cell cycle control of pituitary development and disease

“…A detailed analysis suggested that CDKN2A methylation was confined to particular adenoma subtypes (Simpson et al 1999) and these findings were subsequently confirmed by several other groups concluding that hypermethylation of the CDKN2A is the most common epigenetic deregulation in these neoplasias (Morris et al 2005, Ogino et al 2005, Yoshino et al 2007 INK4a is able to inhibit cell proliferation in pituitary tumor cells in correlation with a shift in the phosphorylation status of pRB, suggesting the relevance of this CDK inhibitor in the activation of pRB and pituitary tumor suppression (Frost et al 1999).…”

“…Several studies based on immunodetection in tumor sections found abnormal expression of pRB in different pituitary adenomas. In some cases, decreased expression correlates with hypermethylation of the pRB promoter (Simpson et al 2000, Ogino et al 2005 or deletion within the protein-pocket binding domain (Simpson et al 2000).…”

Association between p16(CDKN2A) C540G polymorphism and tumor behavior in prolactinoma: A single-center study

“…In the latter group of tumors, the most common mechanism of CDKN2A inactivation involves methylation, while mutations or deletions are rare (13-16,26-28). Inactivation of the p16INK4a gene by methylation of the CpG islands in the promoter region has been reported to play a role in pituitary tumorigenesis, with a consequent decrease or abolishment of the p16 protein (7,27). Certain studies have shown an absence of the expression of p16INK4a at the level of mRNA and protein in all tumors examined.…”

“…Thus, the pRb pathway can be blocked through the loss of p16INK4a, pRb expression or CDK4 amplification. Such changes in the pRb pathway are expected to promote cell proliferation (6,7).…”

Role of the high mobility group A proteins in the regulation of pituitary cell cycle

“…INK4A by methylation is frequent in pituitary, mainly NFPA (Woloschak et al 1997, Simpson et al 1999a, Morris et al 2005, Ogino et al 2005, Yoshino et al 2007). Downregulation of p27 kip1 protein expression, likely due to a JAB1-mediated increased proteolysis (Korbonits et al 2002), is frequent in pituitary carcinomas and ACTH-secreting adenomas (Bamberger et al 1999, Lidhar et al 1999.…”