Investigating the correlation between genotype and phenotype in Prader-Willi syndrome: a study of 45 cases from Brazil.

BACKGROUND: Prader-Willi syndrome (PWS) is a genetic disorder characterized by abnormalities in the 15q11-q13 region. Understanding the correlation between genotype and phenotype in PWS is crucial for improved genetic counseling and prognosis. In this study, we aimed to investigate the correlation between genotype and phenotype in 45 PWS patients who previously underwent methylation-sensitive high-resolution melting (MS-HRM) for diagnosis. RESULTS: We employed methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) and Sanger sequencing, along with collecting phenotypic data from the patients for comparison. Among the 45 patients, 29 (64%) exhibited a deletion of 15q11-q13, while the remaining 16 (36%) had uniparental disomy. No statistically significant differences were found in the main signs and symptoms of PWS. However, three clinical features showed significant differences between the groups. Deletion patients had a higher prevalence of myopia than those with uniparental disomy, as well as obstructive sleep apnea and an unusual skill with puzzles. CONCLUSIONS: The diagnostic tests (MS-HRM, MS-MLPA, and Sanger sequencing) yielded positive results, supporting their applicability in PWS diagnosis. The study's findings indicate a general similarity in the genotype-phenotype correlation across genetic subtypes of PWS.

Establishing a Standardized DNA Extraction Method Using NaCl from Oral Mucosa Cells for Its Application in Imprinting Diseases Such as Prader-Willi and Angelman Syndromes: A Preliminary Investigation.

BACKGROUND: Diagnosing imprinting defects in neonates and young children presents challenges, often necessitating molecular analysis for a conclusive diagnosis. The isolation of genetic material from oral swabs becomes crucial, especially in settings where blood sample collection is impractical or for vulnerable populations like newborns, who possess limited blood volumes and are often too fragile for invasive procedures. Oral swab samples emerge as an excellent source of DNA, effectively overcoming obstacles associated with rare diseases. METHODS: In our study, we specifically addressed the determination of the quality and quantity of DNA extracted from oral swab samples using NaCl procedures. RESULTS: We compared these results with extractions performed using a commercial kit. Subsequently, the obtained material underwent MS-HRM analysis for loci associated with imprinting diseases such as Prader-Willi and Angelman syndromes. CONCLUSIONS: Our study emphasizes the significance of oral swab samples as a reliable source for obtaining DNA for MS-HRM analysis. NaCl extraction stands out as a practical and cost-effective method for genetic studies, contributing to a molecular diagnosis that proves particularly beneficial for patients facing delays in characterization, ultimately influencing their treatment.

Correlação entre o genótipo e o fenótipo dos pacientes com a Síndrome de Prader-Willi / Correlation between the genotype and phenotype of patients with Prader-Willi Syndrome

A síndrome de Prader-Willi (SPW) é uma doença multissistêmica, cujas manifestações principais incluem hipotonia, obesidade, leve atraso mental, hipogonadismo e insuficiência do hormônio de crescimento. A SPW foi a primeira desordem genética descrita envolvida com o imprinting genômico. O imprinting genômico é uma modificação epigenética do DNA responsável por metilar as ilhas CpG, presentes em regiões promotoras dos genes, inativando a expressão deste gene. Na SPW, o indivíduo possui o alelo materno quimicamente inativado através do imprinting, além disso, o indivíduo perde a função dos mesmos genes no alelo paterno devido a 3 possíveis mecanismos genéticos: Deleção, dissomia uniparental materna (DUM), e microdeleções ou defeitos no centro de controle do imprinting. Ainda há muito a se entender sobre as bases genéticas da SPW e sua correlação com os fenótipos clínicos vistos nestes pacientes, e esta comparação entre o perfil molecular e os sintomas clínicos vistos em pacientes com a SPW vem sendo um tema muito discutido dentro da literatura. Muitos achados suportam uma possível correlação entre o genótipo e o fenótipo destes pacientes. Estabelecendo uma possível correlação entre genótipo e fenótipo irá trazer uma maior compreensão da SPW, provendo um melhor aconselhamento genético e consequentemente melhorando o prognóstico para estes indivíduos e suas famílias. Este estudo tem como objetivo identificar a associação dos diferentes mecanismos genéticos da SPW com os diversos sintomas clínicos, contribuindo para um melhor prognóstico da doença. Um estudo descritivo de pesquisa básica e quantitativa a partir de amostras de sangue periférico de 45 pacientes com padrão de metilação compatível com a SPW acompanhados no Centro de Genética Médica do IFF/FIOCRUZ e pelo Instituto Estadual de Diabetes e Endocrinologia do Estado do rio de Janeiro (IEDE/RJ). O estudo vemsendo desenvolvido no Laboratório de Alta Complexidade do IFF (LACIFF). A abordagem metodológica consistiu no rastreamento destes 45 pacientes utilizando a técnica de MS HRM; MS-MLPA visando identificar as deleções nos pacientes; e o sequenciamento de Sanger visando identificar as dissomias uniparentais maternas e os defeitos no centro de controle do imprinting. Posteriormente foi realizado a coleta de dados fenotípicos dos pacientes que apresentaram alterações compatíveis com a SPW. O trabalho em questão tem como resultados esperados encontrar uma correlação genótipo ­ fenótipo, visando um melhor entendimento sobre as bases genéticas da síndrome e um melhor prognóstico para estes pacientes e suas famílias.

Prader-Willi syndrome (PWS) is a multisystemic disease, the main manifestations of which include hypotonia, obesity, mild mental retardation, hypogonadism, and insufficient growth hormone. SPW was the first described genetic disorder involved with genomic imprinting. Genomic imprinting is an epigenetic modification of the DNA responsible for the methylation of the CpG islands, present in promoter regions of the genes, inactivating the expression of this gene. In PWS, the individual has the maternally allele chemically inactivated through imprinting, in addition, the individual loses the function of the same genes in the paternal allele due to 3 possible genetic mechanisms: Deletion, maternal uniparental disomy (matUPD), and micro deletions or defects in the imprinting control center. There is still a lot to understand about the genetic bases of PWS and its correlation with the clinical phenotypes seen in these patients, and this parallel between the molecular profile and the clinical symptoms seen in patients with PWS has been a very discussed topic in the literature. Many findings support a possible correlation between the genotype and phenotype of these patients. Establishing a possible correlation between genotype and phenotype, will bring a greater understanding of PWS, providing better genetic counseling and consequently improving the prognosis for these individuals and their families. This study aims to identify the association of the different genetic mechanisms of PWS with the different clinical symptoms, contributing to a better prognosis of the disease. A descriptive study of basic and quantitative research based on peripheral blood samples from 45 patients with methylation status compatible with PWS followed at the Medical Genetics Center of IFF / FIOCRUZ and by the State Institute of Diabetes and Endocrinology of the State of Rio de Janeiro (IEDE / RJ). The study has been developed at the IFF High Complexity Laboratory (LACIFF). The methodological approach consisted of tracking these 45 patients using the MS-HRM technique; MS MLPA to identify deletions in patients; and the Sanger sequencing aiming to identify maternal uniparental dissomies and defects in the imprinting control center. Subsequently, phenotypic data were collected from patients who presented changes compatible with PWS. The work in question has as expected results to find this genotype - phenotype correlation, aiming at a better understanding about the genetic bases of the syndrome and a better prognosis for these patients and their families.

Genotype-Phenotype Relationships and Endocrine Findings in Prader-Willi Syndrome.

Prader-Willi syndrome (PWS) is a complex imprinting disorder related to genomic errors that inactivate paternally-inherited genes on chromosome 15q11-q13 with severe implications on endocrine, cognitive and neurologic systems, metabolism, and behavior. The absence of expression of one or more genes at the PWS critical region contributes to different phenotypes. There are three molecular mechanisms of occurrence: paternal deletion of the 15q11-q13 region; maternal uniparental disomy 15; or imprinting defects. Although there is a clinical diagnostic consensus criteria, DNA methylation status must be confirmed through genetic testing. The endocrine system can be the most affected in PWS, and growth hormone replacement therapy provides improvement in growth, body composition, and behavioral and physical attributes. A key feature of the syndrome is the hypothalamic dysfunction that may be the basis of several endocrine symptoms. Clinical and molecular complexity in PWS enhances the importance of genetic diagnosis in therapeutic definition and genetic counseling. So far, no single gene mutation has been described to contribute to this genetic disorder or related to any exclusive symptoms. Here we proposed to review individually disrupted genes within the PWS critical region and their reported clinical phenotypes related to the syndrome. While genes such as MKRN3, MAGEL2, NDN, or SNORD115 do not address the full spectrum of PWS symptoms and are less likely to have causal implications in PWS major clinical signs, SNORD116 has emerged as a critical, and possibly, a determinant candidate in PWS, in the recent years. Besides that, the understanding of the biology of the PWS SNORD genes is fairly low at the present. These non-coding RNAs exhibit all the hallmarks of RNA methylation guides and can be incorporated into ribonucleoprotein complexes with possible hypothalamic and endocrine functions. Also, DNA conservation between SNORD sequences across placental mammals strongly suggests that they have a functional role as RNA entities on an evolutionary basis. The broad clinical spectrum observed in PWS and the absence of a clear genotype-phenotype specific correlation imply that the numerous genes involved in the syndrome have an additive deleterious effect on different phenotypes when deficiently expressed.