Molecular studies of mTOR and TAU pathways in focal cortical dysplasia

Author(s)

Marcella Gonçalves Mazutti

• Document type: Master's Dissertation
• Press: Campinas, SP.
• Institution: Universidade Estadual de Campinas (UNICAMP). Faculdade de Ciências Médicas
• Defense date: 2015-10-20

Examining board members

Íscia Teresinha Lopes Cendes; Fábio Rogério; Ana Lúcia Brunialti Godard

• Advisor: Íscia Teresinha Lopes Cendes; Fábio Rossi Torres

Abstract

Epilepsy is one of the most common neurological diseases, affecting 1.5 -2.0 % of the world population. Several types of malformations of cortical development (MCD), including Focal Cortical Dysplasia (FCD), cause epilepsy and are associated with the occurrence of refractory seizures. FCD is characterized by cortical architecture abnormalities observed in other MCDs, such as Tuberous Sclerosis (TS) and Hemimegalencephaly (HME). FCD, TS and HME also show aberrant expression of genes belonging to mTOR signaling pathway. Potential involvement of Tau pathway has also been reported in patients with FCD, due to the presence of tau aggregates, which are also identified in Alzheimer's disease. Therefore, the similarity in histological features and molecular pathways suggests that pathogenic mechanisms could be common to these three disorders. Furthermore, somatic mosaic mutations have been identified in patients with TS and HME. The identification of somatic mutations in a mosaic state might contribute to the understanding of complex diseases such as DCF; however, they are not usually detected by traditional methods such Sanger sequencing. Recently, Next-generation sequencing (NGS) technology has been reported to allow detection of somatic mutations in a mosaic state in the central nervous system. In addition to sequence variants , genomic structural variations called Copy Number Variations (CNV) have also been reported as a source of DNA variation which could be associated to disease. These variations are associated with many neurological disorders ranging from psychiatric disorders to malformations of the cerebral cortex, such as DCF. Some studies have identified structural variations in patients with DCF and HME.. Therefore, the main objective of this project is to investigate whether somatic mosaic mutations in genes belonging to the mTOR and Tau pathways, as well as, structural variations (CNVs) located in these and other genes could be involved in the molecular mechanisms leading to FCD. Deep Sequencing by NGS was performed in genomic DNA of both, FCD resected brain tissue and peripheral blood of patients in order to assess whether the somatic mutations are restricted to the central nervous system. Exon capturing of candidate genes were performed with Nextera® Expanded Kit (Illumina®) and NGS was performed on a Hiseq 2500 machine. CytoScan HD Kit (Affymetrix) was used for the identification of CNVs, following the manufacturer recommendations and Chromosome Analysis Suite (Affymetrix) was used for data analyzes. DNA sequencing was performed in four patients with FCD in both, samples resected brain tissue and peripheral blood. We identified a total of 758 variants in genes belonging to the mTOR pathway, 30 of these variants were found in a mosaic state - 13 were exclusively present in brain tissue and 17 were present only in blood DNA. In genes belonging to Tau pathway, we identified a total of 38 variants,seven of them were in a mosaic state - four were only present in brain tissue and three were present only in the blood DNA. In addition, we investigated the presence of CNVs in six patients in both, FCD resected brain tissue and peripheral blood. We identified a total of 131 CNVs, 61 of gain and 70 of loss, varying from of 100 - 1.237kbp. Of these, 12 CNVs have not yet been described in the Database of Genomic Variants (DGV). Therefore, we conclude that sequence variants in a mosaic state, as well as putative CNVs are indeed present in tissue resected from patients with FCD. Although additional studies are needed in order to unequivocally determine that these DNA variations are indeed causative of the molecular abnormalities which ultimately lead to FCD, the present work have contributed significantly by adding additional evidence supporting the involvement of mTOR and Tau pathways in the mechanisms leading to FCD (AU)

• FAPESP's process: 13/25782-1 - Molecular Studies of mTOR and Tau pathways in Focal Cortical Dysplasia
• Grantee: Marcella Gonçalves Mazutti
• Support Opportunities: Scholarships in Brazil - Master