Evaluation of rs1362756 SALL1, rs1192415 CDC7/TGFBR3, rs9607469 CARD10 and rs1900004 ATOH7 variants in the determination of optic disc parameters and circumpapillary retinal nerve fiber layer thickness in normal individuals in a sample of the Brazilian population

Orientador: Jose Paulo Cabral de VascocellosDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências MédicasResumo: Introdução: A avaliação da cabeça do nervo óptico é parte importante do exame clínico oftalmológico, pois ela pode ser acometida por diversas doenças, particularmente o glaucoma. Há grande variabilidade na anatomia e tamanho do disco óptico (DO) mesmo entre indivíduos normais, o que dificulta o diagnóstico de afecções oculares cujo acometimento inclua esta estrutura. O glaucoma é uma delas caracterizando-se clinicamente pelo aumento da relação escavação disco secundário a perda de rima neural e pela redução da espessura da camada de fibras nervosas da retina. Discute-se na literatura a associação de características anatômicas do DO como fator de risco para o glaucoma primário de ângulo aberto e estudos epidemiológicos tem sugerido que estas variações anatômicas tem base genética na sua determinação. Diversos estudos de associação de amplitude genômica (GWAS) já demonstraram associações genéticas que podem contribuir para melhor caracterização deste fenótipo. Entre elas encontram-se variantes dos genes SALL1 (spalt like transcription factor1) [602218], ATOH7 (atonal homolog 7) [609875], CDC7/TGFBR3 (cell division cycle 7/ transforming growth factor beta receptor 3) [603311] e CARD10 (caspase recruitment domain family 10) [607209]. No entanto, como estes estudos utilizaram diferentes metodologias para as medidas do DO, esta falta de padronização na avaliação de seus parâmetros pode ter afetado a precisão das associações genéticas já demonstradas. Recentemente, foram propostos novos parâmetros para medidas estruturais do disco óptico obtidas pela tomografia de coerência óptica de domínio espectral (TCODE). Entre os parâmetros avaliados incluem-se a área da abertura da Membrana de Bruch (AMB) ou Bruch¿s Membrane Opening (BMO) e a largura mínima da rima neural (LMRN) global e setorizada ou minimum rim width (MRW). Estas variantes nestes genes supracitados ainda não foram replicadas em uma população heterogênea como a brasileira, além disso, elas ainda não foram avaliadas em nenhuma população com os novos parâmetros anatômicos do DO obtidos a partir TCODE. Objetivos: O objetivo do presente estudo é avaliar a associação de variantes dos genes SALL1, CDC7/TGFBR3, CARD10 e ATOH7 com: a área da AMB ajustada pela medida do comprimento axial; a LMRN global e setorizada (temporal superior e inferior) e a espessura da camada de fibras nervosas da retina (CFNR) peripapilar global e setorizada (temporal superior e inferior); ambas ajustadas para idade e para a área de AMB em uma coorte de 189 indivíduos da população brasileira sem doenças oculares por meio de TCODE. Materiais e Métodos: O DNA genômico dos indivíduos incluídos no estudo foi extraído de sangue periférico e as regiões de interesse foram amplificadas por reação em cadeia da polimerase (polymerase chain reaction - PCR) e sequenciadas pelo método de Sanger. As imagens e medidas dos parâmetros do DO avaliados foram obtidas por TCODE Spectralis (Heidelberg Engineering, GmbH, Heidelberg, Germany) incluindo a área de AMB, LMRN global e setorizada (temporal superior e inferior) e a espessura da CFNR peripapilar global e setorizada (temporal superior e inferior). Resultados: A Análise de Regressão Multivariada mostrou associação estatisticamente significativa da variante rs1362756 do gene SALL1 com a área AMB. O alelo C desta variante e o genótipo CC correspondente estão associados a áreas AMB maiores. Não houve associações significativas desta variante com os outros parâmetros analisados (espessura da CFNR e LMRN). Também se observou uma associação estatisticamente significativa entre a variante rs1900004 do gene ATOH7 e a LMRN global. As outras variantes analisadas no presente estudo (rs1192415 do gene CDC7/TGFBR3 e rs9607469 do gene CARD10) não apresentaram associação significativa com nenhum dos parâmetros avaliados. Conclusão: a variante rs1362756 do gene SALL1 apresentou associação significativa com a área de AMB e seu alelo C e genótipo CC correspondentes estão associados a áreas maiores em uma coorte de 189 indivíduos sem doenças oculares da população brasileira. A variante rs1900004 de ATOH7 está associada a LMRN global na mesma coorteAbstract: Background: Optic disc (OD) assessment is an important part of a routine ophthalmological examination, as it might be affected by a number of diseases, including glaucoma. The great variability in OD size and anatomy even amongst healthy individuals makes the diagnosis of OD affections especially challenging, in particular glaucoma. This condition manifests itself clinically by both an increase in vertical cup-to-disc ratio and thinning of the neuroretinal rim, which are consequences of the loss of retinal ganglion cells, as well as disc hemorrhage, notching, or an asymmetry of the vertical cup-to-disc ratio greater than 0.2µm. The association between optic disc anatomic parameters and increased risk of primary open angle glaucoma has been extensively discussed in previously published data and epidemiological studies have suggested that these anatomic variations might have a genetic background in their determination. Many genome wide association studies (GWAS) have already demonstrated genetic associations that may contribute to a better characterization of these phenotypes, including genetic variants in SALL1 gene (spalt like transcription factor1) [602218], ATOH7 gene (atonal homolog 7) [609875], CDC7/TGFBR3 gene (cell division cycle 7/ transforming growth factor beta receptor 3) [603311] and CARD10 gene (caspase recruitment domain family 10) [607209]. However, since most of the previously published studies have utilized different technologies and techniques to measure optic disc area (ODA), this non-standardized evaluation of OD parameters may have led to less precise genetic associations. Recently, new OD parameters obtained by spectral domain optical coherence tomography (SDOCT) have been proposed, including Bruch¿s Membrane Opening (BMO), minimum rim width (MRW), among others. The genetic variants cited above have not yet been replicated in an admixed population like the Brazilian population. Besides, these genetic variants have never been evaluated in any population with the new anatomic parameters of the optic disc obtained with SDOCT. Objectives: To evaluate the association of SALL1, CDC7/TGFBR3, CARD10 and ATOH7 variants with BMO area (adjusted by axial length), global minimum rim width (GMRW), superotemporal minimum rim width (STMRW), inferotemporal minimum rim width (ITMRW), global retinal nerve fiber layer thickness (GRNFLT), superotemporal retinal nerve fiber layer thickness (STRNFLT), inferotemporal retinal nerve fiber layer thickness (ITRNFLT) (adjusted by age and BMO area), in a cohort of 189 normal individuals in a Brazilian admixed population. Methods: Genomic DNA was amplified by polymerase chain reaction (PCR) and the genetic variants were analyzed by Sanger sequencing. OD parameters and RNFLT measurements were acquired by SDOCT Spectralis (Heidelberg Engineering, GmbH, Heidelberg, Germany), including BMO area, GMRW, STMRW, ITMRW, GRNFLT, STRNFLT, and ITRNFLT. We performed a multivariate regression analysis to evaluate the association between the OD parameters and the four polymorphisms analyzed. Results: The regression analysis showed that there is a statistically significant association between the SALL1 rs1362756 variant and BMO area and that the C allele of this variant and the corresponding CC genotype are associated with greater BMO areas. A statistically significant association between the ATOH7 rs 1900004 and GMRW has also been observed. The other parameters were not statistically associated with SALL1 variant. The other variants evaluated in our study (ATOH7 rs 1900004, CDC7/TGFBR3 rs1192415 and CARD10 rs9607469) did not show a significant association with any of the parameters evaluated. Conclusions: SALL1 rs1362756 is significantly associated with BMO area and the corresponding C allele and CC genotype is associated with greater BMO area in a cohort of normal individuals. ATOH7 rs 1900004 is statistically associated with GMRW in the same cohortMestradoOftalmologiaMestra em Ciência

SATB2-Associated Syndrome Due to a c.715C>T:p(Arg239*) Variant in Adulthood: Natural History and Literature Review

SATB2-associated syndrome (SAS) is a rare condition, and it is characterized by severe developmental delay/intellectual disability, especially severe speech delay/or absence, craniofacial abnormalities, and behavioral problems. Most of the published reports are limited to children, with little information about the natural history of the disease and the possible novel signs and symptoms or behavioral changes in adulthood. We describe the management and follow-up of a 25-year-old male with SAS due to a de novo heterozygous nonsense variant SATB2:c.715C>T:p.(Arg239*) identified by whole-exome sequencing and review the literature. The case herein described contributes to a better characterization of the natural history of this genetic condition and in addition to the genotype–phenotype correlation of the SATB2:c.715C>T:p.(Arg239*) variant in SAS, highlights some particularities of its management

Molecular investigation in individuals with orofacial clefts and microphthalmia-anophthalmia-coloboma spectrum

This study describes genomic findings among individuals with both orofacial clefts (OC) and microphthalmia/anophthalmia/coloboma (MAC) recorded in the Brazilian Database on Craniofacial Anomalies (BDCA). Chromosomal microarray analysis (CMA) and Whole Exome Sequencing (WES) were performed in 17 individuals with OC-MAC. Clinical interpretation of molecular findings was based on data available at the BDCA and on re-examination. No copy number variants (CNVs) classified as likely pathogenic or pathogenic were detected by CMA. WES allowed a conclusive diagnosis in six individuals (35.29%), two of them with variants in the CHD7 gene, and the others with variants in the TFAP2A, POMT1, PTPN11, and TP63 genes with the following syndromes: CHARGE, CHD7-spectrum, Branchiooculofacial, POMT1-spectrum, LEOPARD, and ADULT. Variants of uncertain significance (VUS) possibly associated to the phenotypes were found in six other individuals. Among the individuals with VUSes, three individuals presented variants in genes associated to defects of cilia structure and/or function, including DYNC2H1, KIAA0586, WDR34, INTU, RPGRIP1L, KIF7, and LMNA. These results show that WES was the most effective molecular approach for OC-MAC in this cohort. This study also reinforces the genetic heterogeneity of OC-MAC, and the importance of genes related to ciliopathies in this phenotype