EZH2 as a regulator of cell proliferation in pituitary tumors

Abstract

Abstract: 

Pituitary adenomas are the most frequent intracranial neoplasms mainly benign in nature. However, they can occasionally lead to malignant endocrinopathies. Epigenetic modifications are described among the various factors proposed as guidelines for pituitary tumorigenesis.

EZH2, the enzyme responsible for introducing the epigenetic mark H3K27m3, is overexpressed in various tumor phenotypes and significantly associated with increased proliferation and poor prognosis. In order to analyze the regulatory mechanisms involved in the change of the epigenetic profile and its impact on the proliferative behavior of pituitary adenomas, the variations of EZH2 and H3K27me3, cell cycle regulators, metabolic and proliferative activity of tumor pituitary cells were analyzed.

The experimental model of estrogen-induced prolactinoma in adult male Fisher rats and the GH3 somatolactotropic pituitary tumor line were used. Through western blot, the protein levels of EZH2 and H3K27me3 were analyzed in control and adenomatous glands. On these samples, the mRNA levels of the following cell cycle regulators were also determined: cdkn1a, cdk4, ccdn1 and tp53. The GH3 cell line was treated with GSK343, a specific EZH2 inhibitor, in order to analyze the impact that EZH2 has on cell metabolism and proliferation through the MTT and BrDU incorporation assays, respectively. H3K27me3 levels were examined by Western Blot.

It was determined that the levels of EZH2 and H3K27me3 were increased in the adenomatous glands, while the levels of cdkn1a decreased compared to what was observed in control pituitary glands. The tests carried out with GSK343 ​​demonstrated a reduction in H3K27me3 along with a decrease in cell metabolism and proliferation. The methodologies were performed in triplicate and analyzed by t test (Western Blot and qPCR) or ANOVA-Post test Tukey (MTT and BrDU) with significance of p <0.05.

The results obtained suggest EZH2 as a regulator of cell proliferation in pituitary tumors, where the presence of the repressive mark H3K27me3 on the cdkn1a promoter could be responsible for mediating this regulation, postulating EZH2 as a possible therapeutic target.