Expanding the clinical and molecular spectrum of FOXG1- and ZBTB18-associated neurodevelopmental disorders.

Introduction The zinc finger BTB domain-containing protein ZBTB18 binds to FOXG1 to form a transcriptional repressive complex involved in neuronal differentiation. Disruption of the components of this complex results in chromosome 1q43-q44 deletion syndrome/intellectual developmental disorder 22 or in FOXG1 syndrome. Case presentation This study reports on five patients with cognitive and behavioral impairment, seizures, microcephaly, and/or congenital brain abnormalities. Whole exome sequencing identified deleterious ZBTB18 variants in three patients and deleterious FOXG1 variants in the remaining patients. We have detected a missense variant within the BTB domain of ZBTB18 in two affected monozygotic twins. In addition, we observed agenesis of the septum pellucidum in a missense FOXG1 carrier with a severe FOXG1 syndrome. Conclusion Although the ZBTB18 zinc finger domains harbor the majority of known deleterious variants, we report a novel de novo rare missense variant within the BTB domain. The agenesis of the septum pellucidum observed in a missense FOXG1 carrier could be considered as a novel clinical feature associated with FOXG1 syndrome. The severe FOXG1 syndrome in this patient contrasts with the milder phenotype expected for a missense. Genetic or environmental factors may explain this phenotypic variability in FOXG1 syndrome.

Trio-based exome sequencing reveals a high rate of the de novo variants in intellectual disability.

Intellectual disability (ID), a neurodevelopmental disorder affecting 1-3% of the general population, is characterized by limitations in both intellectual function and adaptive skills. The high number of conditions associated with ID underlines its heterogeneous origin and reveals the difficulty of obtaining a rapid and accurate genetic diagnosis. However, the Next Generation Sequencing, and the whole exome sequencing (WES) in particular, has boosted the diagnosis rate associated with ID. In this study, WES performed on 244 trios of patients clinically diagnosed with isolated or syndromic ID and their respective unaffected parents has allowed the identification of the underlying genetic basis of ID in 64 patients, yielding a diagnosis rate of 25.2%. Our results suggest that trio-based WES facilitates ID's genetic diagnosis, particularly in patients who have been extensively waiting for a definitive molecular diagnosis. Moreover, genotypic information from parents provided by trio-based WES enabled the detection of a high percentage (61.5%) of de novo variants inside our cohort. Establishing a quick genetic diagnosis of ID would allow early intervention and better clinical management, thus improving the quality of life of these patients and their families.

From neural stem cells to glioblastoma: A natural history of GBM recapitulated in vitro.

Due to its aggressive and invasive nature glioblastoma (GBM), the most common and aggressive primary brain tumour in adults, remains almost invariably lethal. Significant advances in the last several years have elucidated much of the molecular and genetic complexities of GBM. However, GBM exhibits a vast genetic variation and a wide diversity of phenotypes that have complicated the development of effective therapeutic strategies. This complex pathogenesis makes necessary the development of experimental models that could be used to further understand the disease, and also to provide a more realistic testing ground for potential therapies. In this report, we describe the process of transformation of primary mouse embryo astrocytes into immortalized cultures with neural stem cell characteristics, that are able to generate GBM when injected into the brain of C57BL/6 mice, or heterotopic tumours when injected IV. Overall, our results show that oncogenic transformation is the fate of NSC if cultured for long periods in vitro. In addition, as no additional hit is necessary to induce the oncogenic transformation, our model may be used to investigate the pathogenesis of gliomagenesis and to test the effectiveness of different drugs throughout the natural history of GBM.

Immortalization of a cell line with neural stem cell characteristics derived from mouse embryo brain.

BACKGROUND: Neural stem cells (NSC) have been extensively used as a tool to investigate the mechanisms responsible for neural repair, and they have been also considered as the source for a series of promising replacement therapies in various neurodegenerative diseases. However, their use is limited by their relative rarity and anatomical localization, and also because, the methods for isolation and characterization are usually time consuming and have some technical limitations. RESULTS: In this study, we describe a resource and method for obtaining immortalized cells with NSC characteristics obtained from mouse brain embryo. CONCLUSIONS: Because these cells can be maintained indefinitely in culture, they may constitute a permanent source of NSC that can be used for research studies on neural development and regeneration.

Expanding the clinical and molecular spectrum of the CWC27-related spliceosomopathy.

Cyclophilins are a type of peptidyl-prolyl cis-trans isomerases. CWC27, one of the known human cyclophilins, is recruited by the spliceosome for the pre-mRNA splicing process. Biallelic deleterious variants in CWC27 lead to a spectrum of overlapping phenotypes including retinal degeneration, skeletal anomalies, short stature, and neurological defects. The present work reports a woman showing these clinical features, in addition to hypergonadotropic hypogonadism, hypoplastic/agenesic teeth, and cataracts, not previously associated with such phenotypic spectrum. Whole exome sequencing on this patient identified a novel CWC27 homozygous variant predicted to originate a severely truncated protein and the consequent loss of functionality. The clinical and genetic characterization of such patient could provide further insight into the underlying causes of the spliceosomopathies.

PLGA:poloxamer and PLGA:poloxamine blend nanoparticles: new carriers for gene delivery.

The main objective of the present work was the development of new nanoparticulate carrier systems for the delivery of plasmid DNA. These new carriers consist of a blend matrix formed by a poly(lactic-co-glycolic acid) (PLGA) copolymer and polyoxyethylene derivatives. More specifically, we have prepared nanostructures with different PLGA:poloxamer and PLGA:poloxamine compositions by an optimized emulsification-solvent diffusion technique and studied the potential of these carriers for the encapsulation and controlled release of plasmid DNA. Depending on the particle composition, the encapsulation efficiency of the model plasmid pEGFP-C1 varied between 30% and 45%. All formulations provided continuous and controlled release of the plasmid with minimal burst effect. In addition, the release rate and duration was dependent on the composition of the particle matrix. Moreover, gel electrophoresis and cell culture (MCF-7 cell line) assays allowed us to confirm that the biologically active form of the plasmid was preserved during the particle preparation process and also during its release. Cell culture experiments also indicated that the new nanoparticles do not exhibit toxic effects on these cells at concentrations up to 5 mg/mL. Altogether, these results indicate that these composite nanostructures present a promising approach for the delivery of plasmid DNA.

Inhibition of Cdk4 activity enhances translation of p27kip1 in quiescent Rb-negative cells.

We show in this work that the inhibition of Cdk4 (6) in Rb(-/-) 3T3 cells enhances the accumulation of the p27(kip1) cyclin-dependent kinase inhibitor when these cells are induced into quiescence. Two different forms of inhibition of Cdk4 (6), namely overexpression of the Cdk4 (6) inhibitor p16 and overexpression of a dominant negative mutant of Cdk4 (Cdk4(N158)), result in this effect. This suggests that the relevant activity of Cdk4 (6) that has to be inactivated in this setting is its kinase activity. The accumulation of p27(kip1) is due to enhanced translation of the protein, mediated by the 3'-untranslated region of the p27(kip1) mRNA. Moreover, the cells that overexpress p16(ink4a) or Cdk4(N158) show a delay in G(1) when made quiescent and restimulated to proliferate. This delay is overcome by transfection of a plasmid expressing antisense p27(kip1), which shows that the accumulation of p27(kip1) in these cells is related to their G(1) delay. In summary, we report a new functional link between two important cell cycle regulators, Cdk4 and p27(kip1), and provide a mechanistic explanation to the previously reported epistatic relations between these two proteins.