Banca de QUALIFICAÇÃO: HOZANA BEATRIZ DANTAS BARBALHO

Uma banca de QUALIFICAÇÃO de MESTRADO foi cadastrada pelo programa.
STUDENT : HOZANA BEATRIZ DANTAS BARBALHO
DATE: 03/06/2026
TIME: 15:00
LOCAL: Sala Carl Peter von Dietrich
TITLE:

Development of a molecular test for the diagnosis of Silver-Russell Syndrome

KEY WORDS:

Silver-Russell Syndrome; Imprinting; Hypomethylation; Molecular diagnosis; MS-MLPA; Sodium bisulfite; qPCR; MS-HRM.

PAGES: 63
BIG AREA: Ciências Biológicas
AREA: Bioquímica
SUMMARY:

Silver-Russell Syndrome (SRS) is a rare condition of pre- and postnatal growth restriction associated with genomic imprinting alterations. Ap- proximately 50% of cases are caused by hypomethylation at the 11p15.5 locus, within imprinting control region 1 (ICR1), which regulates the expression of the H19 and IGF2 genes. Currently, molecular diagnosis is performed primarily through Methylation-Sensitive Multiplex Ligationdependent Probe Amplification (MS-MLPA). Despite its widespread use, this technique has relevant limitations, including high cost and limited coverage, as it depends on HhaI enzyme binding sites targeting only seven specific cytosines within ICR1. Therefore, this study aimed to identify differentially methylated cytosines in the 11p15.5 region, develop a more accessible PCR-based molecular tool for hypomethylation detection, and conduct scientific outreach on Instagram. A differential methylation analysis was performed using three public methylation cohorts and controls from MethBank to identify and validate differentially methylated cytosines (DMCs) in the H19, IGF2, and CTCF regions. Differential methylation was assessed using simple linear regression models (limma package, R), applying stringent cutoffs of effect size ≥0.1 and p≤4.3×10−5. Based on this analysis, primer pairs were designed following the criteria of Wojdacz et al. (2008). Genomic DNA from participants with clinical suspicion of SRS (n=4) and controls (n=6) was subjected to sodium bisulfite treatment and amplified by qPCR with SYBR Green, followed by melting curve analysis. Fluorescence data were normalized to a 0–100% scale and compared between groups using the Wilcoxon rank-sum test, with the area under the curve (AUC) as the primary metric, and represented graphically. Methylome analysis identified potentially relevant differentially methylated cytosines for detecting hypomethylation at 11p15.5, particularly in the IGF2 gene. The developed assay covered a greater number of target cytosines compared to MS-MLPA (26 vs. 7). Results demonstrated satisfactory amplification for H19 and IGF2. For H19, no significant difference was observed between SRS and control groups (W=8; p=0.713). For IGF2, a statistically significant difference was found (W=1; p=0.037), indicating greater capacity to distinguish between SRS and control samples. Main limitations include the use of SYBR Green instead of HRM-specific saturating dyes, the small sample size, and the absence of molecular confirmation for suspected SRS cases. In parallel, a scientific outreach project about the syndrome was developed through Instagram, demonstrating potential in facilitating access to information about SRS for different audiences (general public and healthcare professionals). These findings indicate that the developed approach has potential for qualitative detection of hypomethylation at the 11p15.5 locus, representing a possible more accessible alternative for the molecular diagnosis of SRS, with additional studies involving larger sample sizes needed for validation.

COMMITTEE MEMBERS:
Presidente - 1880243 - DANIEL CARLOS FERREIRA LANZA
Externo ao Programa - 2566638 - CARLOS EDUARDO MAIA GOMES - UFRNExterna ao Programa - 1714243 - DANIELLA REGINA ARANTES MARTINS SALHA - UFRN