Rett Syndrome (RTT) is a severe neurodevelopmental disorder predominately diagnosed in females and primarily caused by pathogenic variants in the X-linked gene Methyl-CpG Binding Protein 2 (MECP2). Most often, the disease causing the MECP2 allele resides on the paternal X chromosome while a healthy copy is maintained on the maternal X chromosome with inactivation (XCI), resulting in mosaic expression of one allele in each cell. Preferential inactivation of the paternal X chromosome is theorized to result in reduced disease severity; however, establishing such a correlation is complicated by known MECP2 genotype effects and an age-dependent increase in severity. To mitigate these confounding factors, we developed an age- and genotype-normalized measure of RTT severity by modeling longitudinal data collected in the US Rett Syndrome Natural History Study. This model accurately reflected individual increase in severity with age and preserved group-level genotype specific differences in severity, allowing for the creation of a normalized clinical severity score. Applying this normalized score to a RTT XCI dataset revealed that XCI influence on disease severity depends on MECP2 genotype with a correlation between XCI and severity observed only in individuals with MECP2 variants associated with increased clinical severity. This normalized measure of RTT severity provides the opportunity for future discovery of additional factors contributing to disease severity that may be masked by age and genotype effects.

Rett syndrome (RTT) is a progressive neurodevelopmental disorder, and pathogenic Methyl-CpG-binding Protein 2 (MECP2) variants are identified in >95% of individuals with typical RTT. Most of RTT-causing variants in MECP2 are de novo and usually on the paternally inherited X chromosome. While paternal age has been reported to be associated with increased risk of genetic disorders, it is unknown whether parental age contributes to the risk of the development of RTT. Clinical data including parental age, RTT diagnostic status, and clinical severity are collected from 1226 participants with RTT and confirmed MECP2 variants. Statistical analyses are performed using Student t-test, single factor analysis of variance (ANOVA), and multi-factor regression. No significant difference is observed in parental ages of RTT probands compared to that of the general population. A small increase in parental ages is observed in participants with missense variants compared to those with nonsense variants. When we evaluate the association between clinical severity and parental ages by multiple regression analysis, there is no clear association between clinical severity and parental ages. Advanced parental ages do not appear to be a risk factor for RTT, and do not contribute to the clinical severity in individuals with RTT.

Chromosome 2p (chr2p) duplication, also known as trisomy 2p, is a rare chromosome abnormality associated with developmental delay, intellectual disability, behavioral problems, and distinctive facial features. Most of the reported cases involving trisomy 2p include additional copy number variants (CNVs) in other regions of the genome and are usually small in size. Little is known about the clinical outcomes of large duplications of chr2p as the sole cytogenetic abnormality. In this study, 193 samples at the Greenwood Genetic Center (GGC) with CNVs involving chr2p were evaluated, out of which 86 had chr2p duplications. Among them, 8 patients were identified with large chr2p duplications ranging in size from 9.3 Mb to 89 Mb, and no deletions or duplications involving other chromosomes were identified in those patients. These duplications were associated with inverted duplication, tandem duplication, and duplication as the result of translocation, with no additional CNVs identified by microarray analysis. Confirmation by conventional cytogenetics was performed in 7 of the 8 patients, and the translocations were confirmed by fluorescence in situ hybridization. Interestingly, 1 patient was found to have mosaic complete trisomy 2p as the result of an unbalanced de novo (X;2) chromosomal translocation. X-inactivation was skewed toward the derivative X chromosome, yet it did not appear to extend into the chromosome 2 material. Various shared clinical manifestations were observed in the individuals in this study, including developmental delay, hemifacial hypoplasia, cleft palate, and short stature, and they also have distinct features such as hypotonia, cerebellar hypogenesis, and corpus callosum agenesis, which might result from a gene dosage effect of the duplication. In conclusion, single-event large chr2p duplications can result from different mechanisms, including inverted or tandem duplications within chromosome 2, or translocations involving chromosome 2 and other chromosomes. Partial or complete trisomy 2p is commonly associated with developmental delay, and additional clinical features may be related to gene dosage effects.

Ankyrins are a family of proteins that link integral membrane proteins to the underlying spectrin-actin cytoskeleton and play a key role in activities such as cell motility, activation, proliferation, cell–cell contact, and the maintenance of specialized membrane domains.Ankyrin 3 (ANK3)is one of the three major subtypes of the ankyrin protein family. Ankryin genes are ubiquitously expressed, but their expression is highest in the brain. In the central nervous system, ankyrins have critical roles at the axonal initial segment, the nodes of Ranvier, and at synapses. To date, pathogenic variants inANK3have been reported in individuals with neuropsychiatric, cognitive, and neurodevelopmental disorders. The clinical severity is variable in these individuals with both autosomal recessive and autosomal dominant patterns of inheritance observed. These findings have suggested genotype–phenotype correlations and even isoform-specific implications for individuals withANK3pathogenic variants. Here, we report a patient with speech delay, autism spectrum disorder, and a language disorder in which a de novo nonsenseANK3alteration was discovered by exome sequencing. Interestingly, the next-generation sequencing data suggested the change was mosaic in the affected child, and it was confirmed by digital polymerase chain reaction (dPCR) at 22% allelic fraction. To our knowledge, this is the first case of an individual with a pathogenic mosaicANK3variant. This finding expands upon the existing genotype–phenotype information available for theANK3gene while also highlighting potential gene expression correlations with phenotype.

AbstractBackgroundRett syndrome (RTT) is a rare neurodevelopmental disorder associated with pathogenic MECP2 variants. Because the MECP2 gene is subject to X‐chromosome inactivation (XCI), factors including MECP2 genotypic variation, tissue differences in XCI, and skewing of XCI all likely contribute to the clinical severity of individuals with RTT.MethodsWe analyzed the XCI patterns from blood samples of 320 individuals and their mothers. It includes individuals with RTT (n = 287) and other syndromes sharing overlapping phenotypes with RTT (such as CDKL5 Deficiency Disorder [CDD, n = 16]). XCI status in each proband/mother duo and the parental origin of the preferentially inactivated X chromosome were analyzed.ResultsThe average XCI ratio in probands was slightly increased compared to their unaffected mothers (73% vs. 69%, p = .0006). Among the duos with informative XCI data, the majority of individuals with classic RTT had their paternal allele preferentially inactivated (n = 180/220, 82%). In sharp contrast, individuals with CDD had their maternal allele preferentially inactivated (n = 10/12, 83%). Our data indicate a weak positive correlation between XCI skewing ratio and clinical severity scale (CSS) scores in classic RTT patients with maternal allele preferentially inactivated XCI (rs = 0.35, n = 40), but not in those with paternal allele preferentially inactivated XCI (rs = −0.06, n = 180). The most frequent MECP2 pathogenic variants were enriched in individuals with highly/moderately skewed XCI patterns, suggesting an association with higher levels of XCI skewing.ConclusionThese results extend our understanding of the pathogenesis of RTT and other syndromes with overlapping clinical features by providing insight into the both XCI and the preferential XCI of parental alleles.

Consensus donor splice site alterations have been studied for their impact on RNA splicing and transcriptional products, and +1 and +2 position variants have been well known to affect the mRNA splicing. However, less was known about the +5 position variants and their role in mediating genetic disorders.

Beckwith-Wiedemann syndrome (BWS) and Russell-Silver syndrome (RSS) are congenital imprinting disorders commonly caused by methylation defects in the same region of chromosome 11p15. BWS is an overgrowth syndrome that includes clinical features such as macroglossia, organomegaly, umbilical hernia, hypoglycemia, and an increased risk of childhood tumors. RSS is characterized by intrauterine and postnatal growth retardation, relative macrocephaly, fifth finger clinodactyly, triangular facies, and body asymmetry.

Recurrent microdeletions and microduplications on chromosome 16p11.2 (MIM: 611913 and 614671) confer susceptibility to autism spectrum disorder (ASD) in ∼1% of individuals with ASD, in addition to other clinical features. To further analyze copy number variants (CNVs) in this region, we studied 41 individuals with 16p11.2 duplications and 53 individuals with 16p11.2 deletions at Greenwood Genetic Center. They share common clinical features including: developmental delay, intellectual disability, autism spectrum disorder, seizures, and some features that are unique to either deletions or duplications. We summarize unique CNVs in individuals with either deletion (n=41) or duplication (n=35) of these regions. By overlapping the del/dup regions, we identify six subtypes of CNVs and classify them by potential relevant breakpoints based on size and location. Types 1, 2 and 3 are more common with both deletions and duplications found in these regions, and they overlap with BP1-BP3, BP1-BP2 and BP4-BP5 regions, respectively, in the literature. Types 4, 5 and 6 are less frequent, locating at 31.5-32.5 Mb, 29.6-47.5 Mb and 28.48-28.49 Mb regions on chromosome 16. Type 5 is found only in individuals with duplications and type 6 only in individuals with deletions. Type 4 variants are more frequently observed in individuals of African American decent, yet no clear association could be determined due to small sample size in this study. Genotype-phenotype correlation analysis was also performed to narrow down the potentially causative genes for specific clinical features. It is also worth noting that de novo CNVs are observed with much higher frequency in individuals with 16p11.2 deletions (68.2%, n=22) compared to those with duplications (28.6%, n=14). In conclusion, our analysis explores 16p11.2 CNVs in more detail for rarely observed regions, and could broaden the understanding of the clinical significance of genes and non-coding regions in this chromosomal region.

Hypospadias and urethral stricture are common urological diseases which seriously affect voiding function and life quality of the patients, yet current clinical treatments often result in unsatisfactory clinical outcome with frequent complications. In vitro experiments confirmed that ICG-001 (a well-established Wnt signaling inhibitor) could effectively suppress fibroblast proliferation and fibrotic protein expression. In this study, we applied a novel drug-delivering nanoyarn scaffold in urethroplasty in dog model, which continuously delivers ICG-001 during tissue reconstruction, and could effectively promote urethral recovery and resume fully functional urethra within 12 weeks. Such attempts are essential to the development of regenerative medicine for urological disorders and for broader clinical applications in human patients.

Hypospadias and urethral stricture are common urological diseases which seriously affect voiding function and life quality of the patients. Current clinical treatments using autologous tissues or tissue engineered grafts often result in unsatisfactory clinical outcome with frequent complications at tissue-harvesting organ(s). In the present study, we performed RNA profiling (mRNA and miRNA sequencing) using urethral stricture animal model. Functional enrichment analysis of differential expressed genes showed that CTNNB1-centered Wnt signaling were significantly regulated during the scar formation and fibrosis, suggesting Wnt pathway as a promising therapeutic target for clinical treatment of urethral defects. In vitro experiments confirmed that ICG-001 (a well-established Wnt signaling inhibitor) could effectively prohibit fibroblast proliferation and the expression of fibrosis associated proteins. To testify whether inhibition of Wnt signaling could benefit tissue reconstruction from urethral defects in vivo, we applied a novel drug-delivering nanoyarn scaffold in urethroplasty in dog model, which could continuously deliver ICG-001 during the tissue reconstruction. The ICG-001 delivering nanoyarn could effectively promote urethral recovery and resume fully functional urethra within 12 weeks. Such attempts could be essential in the Next Generation Sequencing (NGS)-guided regenerative medicine to cure urethral defects and might be helpful for broader clinical applications in precision medicine.