Skeletal muscle vulnerability in a child with Pitt-Hopkins syndrome.

BACKGROUND: TCF4 acts as a transcription factor that binds to the immunoglobulin enhancer Mu-E5/KE5 motif. Dominant variants in TCF4 are associated with the manifestation of Pitt-Hopkins syndrome, a rare disease characterized by severe mental retardation, certain features of facial dysmorphism and, in many cases, with abnormalities in respiratory rhythm (episodes of paroxysmal tachypnea and hyperventilation, followed by apnea and cyanosis). Frequently, patients also develop epilepsy, microcephaly, and postnatal short stature. Although TCF4 is expressed in skeletal muscle and TCF4 seems to play a role in myogenesis as demonstrated in mice, potential myopathological findings taking place upon the presence of dominant TCF4 variants are thus far not described in human skeletal muscle. METHOD: To address the pathological effect of a novel deletion affecting exons 15 and 16 of TCF4 on skeletal muscle, histological and immunofluorescence studies were carried out on a quadriceps biopsy in addition to targeted transcript studies and global proteomic profiling. RESULTS: We report on muscle biopsy findings from a Pitt-Hopkins patient with a novel heterozygous deletion spanning exon 15 and 16 presenting with neuromuscular symptoms. Microscopic characterization of the muscle biopsy revealed moderate fiber type I predominance, imbalance in the proportion of fibroblasts co-expressing Vimentin and CD90, and indicate activation of the complement cascade in TCF4-mutant muscle. Protein dysregulations were unraveled by proteomic profiling. Transcript studies confirmed a mitochondrial vulnerability in muscle and confirmed reduced TCF4 expression. CONCLUSION: Our combined findings, for the first time, unveil myopathological changes as phenotypical association of Pitt-Hopkins syndrome and thus expand the current clinical knowledge of the disease as well as support data obtained on skeletal muscle of a mouse model.

DNA methylation episignature and comparative epigenomic profiling for Pitt-Hopkins syndrome caused by TCF4 variants.

Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by pathogenic variants in TCF4, leading to intellectual disability, specific morphological features, and autonomic nervous system dysfunction. Epigenetic dysregulation has been implicated in PTHS, prompting the investigation of a DNA methylation (DNAm) "episignature" specific to PTHS for diagnostic purposes and variant reclassification and functional insights into the molecular pathophysiology of this disorder. A cohort of 67 individuals with genetically confirmed PTHS and three individuals with intellectual disability and a variant of uncertain significance (VUS) in TCF4 were studied. The DNAm episignature was developed with an Infinium Methylation EPIC BeadChip array analysis using peripheral blood cells. Support vector machine (SVM) modeling and clustering methods were employed to generate a DNAm classifier for PTHS. Validation was extended to an additional cohort of 11 individuals with PTHS. The episignature was assessed in relation to other neurodevelopmental disorders and its specificity was examined. A specific DNAm episignature for PTHS was established. The classifier exhibited high sensitivity for TCF4 haploinsufficiency and missense variants in the basic-helix-loop-helix domain. Notably, seven individuals with TCF4 variants exhibited negative episignatures, suggesting complexities related to mosaicism, genetic factors, and environmental influences. The episignature displayed degrees of overlap with other related disorders and biological pathways. This study defines a DNAm episignature for TCF4-related PTHS, enabling improved diagnostic accuracy and VUS reclassification. The finding that some cases scored negatively underscores the potential for multiple or nested episignatures and emphasizes the need for continued investigation to enhance specificity and coverage across PTHS-related variants.

Tcf4 dysfunction alters dorsal and ventral cortical neurogenesis in Pitt-Hopkins syndrome mouse model showing sexual dimorphism.

Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by haploinsufficiency of transcription factor 4 (TCF4). In this work, we focused on the cerebral cortex and investigated in detail the progenitor cell dynamics and the outcome of neurogenesis in a PTHS mouse model. Labeling and quantification of progenitors and newly generated neurons at various time points during embryonic development revealed alterations affecting the dynamic of cortical progenitors since the earliest stages of cortex formation in PTHS mice. Consequently, establishment of neuronal populations and layering of the cortex were found to be altered in heterozygotes subjects at birth. Interestingly, defective layering process of pyramidal neurons was partially rescued by reintroducing TCF4 expression using focal in utero electroporation in the cerebral cortex. Coincidentally with a defective dorsal neurogenesis, we found that ventral generation of interneurons was also defective in this model, which may lead to an excitation/inhibition imbalance in PTHS. Overall, sex-dependent differences were detected with more marked effects evidenced in males compared with females. All of this contributes to expand our understanding of PTHS, paralleling the advances of research in autism spectrum disorder and further validating the PTHS mouse model as an important tool to advance preclinical studies.

[Genetic analysis of a fetus with Pitt-Hopkins syndrome due to a 18q21.2q21.31 microdeletion].

OBJECTIVE: To carry out invasive prenatal diagnosis for a fetus with ultrasound-indicated agenesis of corpus callosum and explore its genetic etiology. METHODS: A pregnant woman presented at the Affiliated Hospital of Putian College on December 16, 2022 was selected as the study subject. Amniotic fluid and peripheral blood samples from the fetus and the couple were collected. Conventional G-banded chromosomal karyotyping was carried out, and whole-genome copy number variation analysis was performed using single nucleotide polymorphism microarray (SNP-array). RESULTS: The karyotypes of the fetus and the couple were normal by the G-banding analysis. SNP-array analysis of the amniotic fluid sample revealed a 4.5 Mb microdeletion in the 18q21.2q21.31 region of the fetus. SNP-array analysis of peripheral blood samples from the couple did not find any abnormality. CONCLUSION: Through G-banded chromosomal karyotyping and SNP-array analysis, a fetus with 18q21.2q21.31 microdeletion was identified, which has conformed to the diagnosis of Pitt-Hopkins syndrome. Above finding has provided a basis for genetic counseling for the couple.

Clinical and genetic characterization of 47 Chinese pediatric patients with Pitt-Hopkins syndrome: a retrospective study.

BACKGROUND: Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder that remains underdiagnosed and its clinical presentations and mutation profiles in a diverse population are yet to be evaluated. This retrospective study aims to investigate the clinical and genetic characteristics of Chinese patients with PTHS. METHODS: The clinical, biochemical, genetic, therapeutic, and follow-up data of 47 pediatric patients diagnosed with PTHS between 2018 and 2021 were retrospectively analyzed. RESULTS: The Chinese PTHS patients presented with specific facial features and exhibited global developmental delay of wide severity range. The locus heterogeneity of the TCF4 gene in the patients was highlighted, emphasizing the significance of genetic studies for accurate diagnosis, albeit no significant correlations between genotype and phenotype were observed in this cohort. The study also reports the outcomes of patients who underwent therapeutic interventions, such as ketogenic diets and biomedical interventions. CONCLUSIONS: The findings of this retrospective analysis expand the phenotypic and molecular spectra of PTHS patients. The study underscores the need for a long-term prospective follow-up study to assess potential therapeutic interventions.

Complex Craniosynostosis in Pitt-Hopkins Syndrome: Case Report in Twins.

INTRODUCTION: Pitt-Hopkins syndrome (PTHS) is a rare genetic syndrome associated with neurodevelopmental disorders and craniofacial dysmorphisms caused by variations in the TCF4 transition factor. The aim of this article was to report the case of two twin infants diagnosed with PTHS, confirmed by the identification of a heterozygous pathogenic variant in the TCF4 gene through DNA extracted from a buccal swab. CASE PRESENTATION: Both infants presented with craniofacial asymmetry with a metopic crest and cranial deformity. During the diagnostic investigation, computed tomography with three-dimensional reconstruction of the skull showed premature fusion of the left coronal and metopic sutures in both twins. They underwent craniofacial reconstruction at the 9th month of age using a combination of techniques. The postoperative outcomes were satisfactory in both cases. CONCLUSION: To the best of our knowledge, this is the first case report to describe the occurrence of complex craniosynostosis (CCS) in children with PTHS. Further studies are needed to determine whether the co-occurrence of PTHS and CCS described here indicates an association or is explained by chance.

A patient with Pitt-Hopkins syndrome with concomitant common variable immunodeficiency.

In patients with 18q deletion syndrome (18q-), immunodeficiency, autoimmunity, and allergies have been described in a subset. Pitt-Hopkins syndrome represents a specific subset of patients with 18q- who have a proximal deletion involving the TCF4 gene or a TCF4 variant. Immunodeficiency has been reported in the overall 18q- population; however, immunodeficiency with Pitt-Hopkins syndrome has not been highlighted. This case report details the immunologic evaluations and the associated infections seen in a young adult with Pitt-Hopkins syndrome to underscore the challenges of managing adults with a complex phenotype who develop frequent infections. This patient with Pitt-Hopkins syndrome ultimately fulfilled the diagnostic criteria for common variable immunodeficiency. Immunoglobulin replacement has led to a somewhat improved infection pattern, although she continues to have aspiration events leading to pneumonia. This case highlights the clinical evolution of Pitt-Hopkins syndrome and serves as a reminder that immunodeficiency can occur in this syndrome.

Pathogenic variants in SOX11 mimicking Pitt-Hopkins syndrome phenotype.

Pitt-Hopkins syndrome (PTHS) is a rare neurodevelopmental disorder characterised by severe intellectual disability (ID), distinctive facial features and autonomic nervous system dysfunction, caused by TCF4 haploinsufficiency. We clinically diagnosed with PTHS a 14 6/12 -year-old female, who had a normal status of TCF4. The pathogenic c.667del (p.Asp223MetfsTer45) variant in SOX11 was identified through whole exome sequencing (WES). SOX11 variants were initially reported to cause Coffin-Siris syndrome (CSS), characterised by growth restriction, moderate ID, coarse face, hypertrichosis and hypoplastic nails. However, recent studies have provided evidence that they give rise to a distinct neurodevelopmental disorder. To date, SOX11 variants are associated with a variable phenotype, which has been described to resemble CSS in some cases, but never PTHS. By reviewing both clinically and genetically 32 out of 82 subjects reported in the literature with SOX11 variants, for whom detailed information are provided, we found that 7/32 (22%) had a clinical presentation overlapping PTHS. Furthermore, we made a confirmation that overall SOX11 abnormalities feature a distinctive disorder characterised by severe ID, high incidence of microcephaly and low frequency of congenital malformations. Purpose of the present report is to enhance the role of clinical genetics in assessing the individual diagnosis after WES results.

Fatal gastrointestinal complications in Pitt-Hopkins syndrome.

Pitt-Hopkins syndrome (PTHS) is a rare neurodevelopmental disorder caused by mutations of the transcription factor 4 (Tcf4) gene. Individuals with PTHS often suffer from severe abdominal bloating and constipation. In this short communication, we discuss two individuals with PTHS who died unexpectedly due to gastrointestinal complications. We aim to increase awareness among healthcare professionals who care for individuals with PTHS, to ensure adequate screening and management of gastrointestinal symptoms in this population. Moreover, we discuss how fatal gastrointestinal complications may be related to PTHS and provide an overview of the literature.

A typical variant in TCF4 exon 18 is not associated with Pitt-Hopkins syndrome but with a familial case of mild and nonspecific neurodevelopmental disorder.

TCF4 gene encodes a class I helix-loop-helix transcription factor critical for the developing brain. Common polymorphisms in TCF4 and disruptive variants in the proximal region of the gene have been linked to relatively mild neuropsychiatric or neurodevelopmental disorders. In contrast, variants impacting distal exons are associated with Pitt-Hopkins syndrome (PTHS), a severe autosomal dominant condition characterized by profound intellectual disability, developmental delay, limited or absent speech, distinctive facies, and disordered breathing. Although phenotypic variability has been observed in PTHS, intellectual impairment and significant speech and motor delays are invariably present. In contrast to the typical de novo variants causing TCF4-related disorder and PTHS, we report a familial form of TCF4-related disorder where the missense variant arose de novo in the father and was inherited by two of his children. Although this family's variant's position in exon 18 predicted a typical PTHS phenotype, none of the affected individuals met the clinical diagnostic criteria for PTHS suggested by Zollino et al. in the first international consensus statement (as in the study by Zollino et al. in 2019). Rather, the three affected family members exhibited remarkably variable and milder phenotypes than would have been predicted from the position of their TCF4 variant. Thus, the clinical spectrum of PTHS-associated TCF4 variants may be broader than previously reported.

Evaluation of Nav1.8 as a therapeutic target for Pitt Hopkins Syndrome.

Pitt Hopkins Syndrome (PTHS) is a rare syndromic form of autism spectrum disorder (ASD) caused by autosomal dominant mutations in the Transcription Factor 4 (TCF4) gene. TCF4 is a basic helix-loop-helix transcription factor that is critical for neurodevelopment and brain function through its binding to cis-regulatory elements of target genes. One potential therapeutic strategy for PTHS is to identify dysregulated target genes and normalize their dysfunction. Here, we propose that SCN10A is an important target gene of TCF4 that is an applicable therapeutic approach for PTHS. Scn10a encodes the voltage-gated sodium channel Nav1.8 and is consistently shown to be upregulated in PTHS mouse models. In this perspective, we review prior literature and present novel data that suggests inhibiting Nav1.8 in PTHS mouse models is effective at normalizing neuron function, brain circuit activity and behavioral abnormalities and posit this therapeutic approach as a treatment for PTHS.

Disruption and deletion of the proximal part of TCF4 are associated with mild intellectual disability: About three new patients.

TCF4 gene (18q21.1) encodes for a transcription factor with multiple isoforms playing a critical role during neurodevelopment. Molecular alterations of this gene are associated with Pitt-Hopkins syndrome, a severe condition characterized by intellectual disability, specific facial features and autonomic nervous system dysfunction. We report here three patients presenting with structural variations of the proximal part of TCF4 associated with a mild phenotype. The first patient is a six-years-old girl carrier of a pericentric inversion of chromosome 18, 46,XX,inv(18)(p11.2q21.1). Whole genome sequencing (WGS) characterized the breakpoint at the base-pair level at chr18:1262334_1262336 and chr18:53254747_53254751 (hg19). This latter breakpoint disrupted the proximal promotor region of TCF4 in the first intron of the gene. The second and third patients are a son and his mother, carrier of a 46 kb deletion characterized by high-resolution chromosomal micro-array and WGS (chr:18:53243454_53287927, hg19) encompassing the first three exon of TCF4 gene and including the proximal promotor region. Expression studies on blood lymphocytes in these patients showed a marked decrease of mRNA level for long isoforms of TCF4 and an increased level for shorter isoforms. The patients described here, together with previously reported patients with proximal structural alterations of TCF4, help to delineate a phenotype of mild ID with non-specific facial dysmorphism without characteristic features of PTHS. It also suggests a gradient of phenotypic severity inversely correlated with the number of intact TCF4 promotor regions, with expression of short isoforms compensating in part the loss of longer isoforms.

Two types of early epileptic encephalopathy in a Pitt-Hopkins syndrome patient with a novel TCF4 mutation.

INTRODUCTION: Pitt-Hopkins syndrome (PTHS) is a neurodevelopmental disorder caused by mutations in TCF4. Seizures have been found to vary among patients with PTHS. We report the case of a PTHS patient with a novel missense mutation in the gene TCF4, presenting with two types of early epileptic encephalopathy. CASE REPORT: The patient was a Japanese boy. His first seizure was reported at 17 days of age, with twitching of the left eyelid and tonic-clonic seizures on either side of his body. An ictal electroencephalogram (EEG) showed epileptic discharges arising independently from both hemispheres, occasionally resembling migrating partial seizures of infancy (MPSI) that migrated from one side to the other. Brain magnetic resonance imaging revealed agenesis of the corpus callosum. His facial characteristics included a distinctive upper lip and thickened helices. His seizures were refractory, and psychomotor development was severely delayed. At the age of 10 months, he developed West syndrome with spasms and hypsarrhythmia. After being prescribed topiramate (TPM), his seizures and EEG abnormalities dramatically improved. Also, psychomotor development progressed. Whole-exome sequencing revealed a novel de novo missense mutation in exon 18 (NM_001083962.2:c.1718A > T, p.(Asn573Ile)), corresponding to the basic region of the basic helix-loop-helix domain, which may be a causative gene for epileptic encephalopathy. CONCLUSIONS: To our knowledge, this is the first report of a patient with PTHS treated with TPM, who presented with both MPSI as well as West syndrome. This may help provide new insights regarding the phenotypes caused by mutations in TCF4.

Functional consequences of TCF4 missense substitutions associated with Pitt-Hopkins syndrome, mild intellectual disability, and schizophrenia.

Transcription factor 4 (TCF4) is a basic helix-loop-helix transcription factor essential for neurocognitive development. The aberrations in TCF4 are associated with neurodevelopmental disorders including schizophrenia, intellectual disability, and Pitt-Hopkins syndrome, an autism-spectrum disorder characterized by developmental delay. Several disease-associated missense mutations in TCF4 have been shown to interfere with TCF4 function, but for many mutations, the impact remains undefined. Here, we tested the effects of 12 functionally uncharacterized disease-associated missense mutations and variations in TCF4 using transient expression in mammalian cells, confocal imaging, in vitro DNA-binding assays, and reporter assays. We show that Pitt-Hopkins syndrome-associated missense mutations within the basic helix-loop-helix domain of TCF4 and a Rett-like syndrome-associated mutation in a transcription activation domain result in altered DNA-binding and transcriptional activity of the protein. Some of the missense variations found in schizophrenia patients slightly increase TCF4 transcriptional activity, whereas no effects were detected for missense mutations linked to mild intellectual disability. We in addition find that the outcomes of several disease-related mutations are affected by cell type, TCF4 isoform, and dimerization partner, suggesting that the effects of TCF4 mutations are context-dependent. Together with previous work, this study provides a basis for the interpretation of the functional consequences of TCF4 missense variants.

Disordered breathing in a Pitt-Hopkins syndrome model involves Phox2b-expressing parafacial neurons and aberrant Nav1.8 expression.

Pitt-Hopkins syndrome (PTHS) is a rare autism spectrum-like disorder characterized by intellectual disability, developmental delays, and breathing problems involving episodes of hyperventilation followed by apnea. PTHS is caused by functional haploinsufficiency of the gene encoding transcription factor 4 (Tcf4). Despite the severity of this disease, mechanisms contributing to PTHS behavioral abnormalities are not well understood. Here, we show that a Tcf4 truncation (Tcf4tr/+) mouse model of PTHS exhibits breathing problems similar to PTHS patients. This behavioral deficit is associated with selective loss of putative expiratory parafacial neurons and compromised function of neurons in the retrotrapezoid nucleus that regulate breathing in response to tissue CO2/H+. We also show that central Nav1.8 channels can be targeted pharmacologically to improve respiratory function at the cellular and behavioral levels in Tcf4tr/+ mice, thus establishing Nav1.8 as a high priority target with therapeutic potential in PTHS.

Molecular and Cellular Function of Transcription Factor 4 in Pitt-Hopkins Syndrome.

Transcription factor 4 (TCF4, also known as ITF2 or E2-2) is a type I basic helix-loop-helix transcription factor. Autosomal dominant mutations in TCF4 cause Pitt-Hopkins syndrome (PTHS), a rare syndromic form of autism spectrum disorder. In this review, we provide an update on the progress regarding our understanding of TCF4 function at the molecular, cellular, physiological, and behavioral levels with a focus on phenotypes and therapeutic interventions. We examine upstream and downstream regulatory networks associated with TCF4 and discuss a range of in vitro and in vivo data with the aim of understanding emerging TCF4-specific mechanisms relevant for disease pathophysiology. In conclusion, we provide comments about exciting future avenues of research that may provide insights into potential new therapeutic targets for PTHS.

Pitt-Hopkins syndrome: phenotypic and genotypic description of four unrelated patients and structural analysis of corresponding missense mutations.

Pitt-Hopkins syndrome is an underdiagnosed neurodevelopmental disorder which is characterized by specific facial features, early-onset developmental delay, and moderate to severe intellectual disability. The genetic cause, a deficiency of the TCF4 gene, has been established; however, the underlying pathological mechanisms of this disease are still unclear. Herein, we report four unrelated children with different de novo mutations (T606A, K607E, R578C, and V617I) located at highly conserved sites and with clinical phenotypes which present variable degrees of developmental delay and intellectual disability. Three of these four missense mutations have not yet been reported. The patient with V617I mutation exhibits mild intellectual disability and has attained more advanced motor and verbal skills, which is significantly different from other cases reported to date. Molecular dynamics simulations are used to explore the atomic level mechanism of how missense mutations impair the functions of TCF4. Mutations T606A, K607E, and R578C are found to affect DNA binding directly or indirectly, while V617I only induces subtle conformational changes, which is consistent with the milder clinical phenotype of the corresponding patient. The study expands the mutation spectrum and phenotypic characteristics of Pitt-Hopkins syndrome, and reinforces the genotype-phenotype correlation and strengthens the understanding of phenotype variability, which is helpful for further investigation of pathogenetic mechanisms and improved genetic counseling.

Transcription factor 4 controls positioning of cortical projection neurons through regulation of cell adhesion.

The establishment of neural circuits depends on precise neuronal positioning in the cortex, which occurs via a tightly coordinated process of neuronal differentiation, migration, and terminal localization. Deficits in this process have been implicated in several psychiatric disorders. Here, we show that the transcription factor Tcf4 controls neuronal positioning during brain development. Tcf4-deficient neurons become mispositioned in clusters when their migration to the cortical plate is complete. We reveal that Tcf4 regulates the expression of cell adhesion molecules to control neuronal positioning. Furthermore, through in vivo extracellular electrophysiology, we show that neuronal functions are disrupted after the loss of Tcf4. TCF4 mutations are strongly associated with schizophrenia and cause Pitt-Hopkins syndrome, which is characterized by severe intellectual disability. Thus, our results not only reveal the importance of neuronal positioning in brain development but also provide new insights into the potential mechanisms underlying neurological defects linked to TCF4 mutations.

CDKL5 mutations may mimic Pitt-Hopkins syndrome phenotype.

Genetic conditions comprise a wide spectrum of different phenotypes, rapidly expanding due to new diagnostic methodologies. Patients' facial features and clinical history represent the key elements leading clinicians to the right diagnosis. CDKL5-early onset epilepsy and Pitt-Hopkins syndrome are two well-known genetic conditions, with a defined phenotype sharing some common characteristics like early-onset epilepsy and hyperventilation episodes. Whilst facial features represent a diagnostic handle in patients with Pitt-Hopkins syndrome, clinical history is crucial in patients carrying a mutation in CDKL5. Here we present the clinical case of a girl evaluated for the first time when she was 24-years old, with a clinical phenotype mimicking Pitt-Hopkins syndrome. Her facial features have become coarser while she was growing up, leading geneticists to raise different clinical hypotheses and to perform several molecular tests before getting the diagnosis of CDKL5-early-epileptic encephalopathy. This finding highlights that although typical facial gestalt has not so far extensively been described in CDKL5 mutated adult patients, peculiar facial features could be present later in life and may let CDKL5-related disorder mimic Pitt Hopkins. Thus, considering atypical Rett syndrome in the differential diagnosis of Pitt Hopkins syndrome could be important to solve complex clinical cases.