Study of TETs genes expression and hydroxymethylation levels in medulloblastoma

Author(s)

Karina Bezerra Salomão

• Document type: Doctoral Thesis
• Press: Ribeirão Preto.
• Institution: Universidade de São Paulo (USP). Faculdade de Medicina de Ribeirão Preto (PCARP/BC)
• Defense date: 2017-09-15

Examining board members

Luiz Gonzaga Tone; Vitor Marcel Faça; Andréia Machado Leopoldino; Rodrigo Alexandre Panepucci; Elvis Terci Valera

• Advisor: Luiz Gonzaga Tone

Abstract

Medulloblastoma (MB) is an embryonic tumor that originates from genetic alterations in pathways that are important to the neurogenesis of cerebellum, such as Sonic hedgehog (Shh) e Wingless (Wnt). These alterations allow us to classify MB based in expression and mutational profile in four subgroups: SHH, WNT, group 3 and group 4. The activation of these pathways could be related to hypermethylation of negative regulators. Hydroxymethylation dynamics was described during cerebellum development, but there are not reports in the literature about hydroxymethylation levels in MB samples. The main aims of this study were: to evaluate TET and IDH genes expression in MB using qPCR; to investigate hydroxymethylation levels using immunohistochemistry and dot-blot; to evaluate mutations in exon 4 of IDH1 and IDH2 genes through sequencing analysis; to analyze methylation and hydroxymethylation levels in genes that regulate SHH, WNT, NOTCH and BMP pathways; to modulate TET genes activity through ascorbic acid and verify its functional and epigenetic influence in MB cell lines. We observed a decrease in TET and IDH genes expression in MB samples compared to fetal cerebellum, but not according to non-fetal cerebellum. MB cell lines presented a decrease when compared to both control groups. The classification of MB samples allowed us to verify a subgroup-specific gene expression. TET3 expression was associated with disease status; and higher levels of IDH2 gene expression were associated with metastasis. We did not find mutations in exon 4 of IDH1 and exon 4 of IDH2 genes in MB samples. Hydroxymethylation levels were decreased in MB samples and cell lines when compared to non-neoplastic cerebellum; however, they were not associated with clinical characteristics of the patients. We did not detect hydroxymethylation levels in the studied genes. Ascorbic acid effects are cell linespecific: we did not observe increase in hydroxymethylation levels, but alterations in TET3 gene expression. In conclusion, hydroxymethylation and expression levels of TET and IDH genes are important for MB. Though, functional assays that target these genes are required to elucidate their function in MB. (AU)

• FAPESP's process: 13/15125-3 - Study of TET gene expression and hydroxymethylation levels in medulloblastoma.
• Grantee: Karina Bezerra Salomão Xavier
• Support Opportunities: Scholarships in Brazil - Doctorate