Kleefstra syndrome is a rare genetic disorder attributed to loss of function of EHMT1, either due to a point mutation or a microdeletion in the chromosome region 9q34.3. This gene encodes an enzyme that modifies histone function and is essential for normal development. Individuals with Kleefstra syndrome typically present intellectual disability (from moderate to severe), language delay, autism spectrum disorders, generalized hypotonia, and distinctive facial dysmorphic features. Additional manifestations in children may include cardiac defects, renal and urological malformations, genital anomalies, respiratory infections, epilepsy (including febrile seizures), and psychiatric disorders. Panayiotopoulos syndrome is a specific type of epilepsy, usually presenting in early to mid-childhood with benign focal seizures. These seizures are characterized by primarily autonomic symptoms, abnormal EEG findings showing shifts or multiple seizure foci (often located in occipital lobe), and other autonomic manifestations such as pallor, redness or cyanosis, mydriasis or miosis, heart and breathing problems, thermoregulatory changes, urinary and/or fecal incontinence, hypersalivation, and altered gut motility. We present the case of a child with Kleefstra syndrome and Panayiotopoulos epilepsy. The patient is a 12-year-old male born from a full-term pregnancy to non-consanguineous healthy parents with a family history of neurodevelopmental disorders. At birth, he presented dysmorphic facial features including receding forehead, low-set ears and lingual protrusion. From 6 months of age, he manifested predominantly axial and lower limb hypotonia, associated with a delay in acquiring psychomotor developmental milestones. Genetic counseling was requested, and array-CGH was then performed. Molecular analysis detected a 9q34.3 microdeletion which included the EHMT1 gene, leading to Kleefstra syndrome diagnosis. From the age of 6 years, he began experiencing seizures with features typical of Panayiotopoulos epilepsy and started treatment with valproic acid. We highlight the association between Panayiotopoulos epilepsy and Kleefstra syndrome, which has not been previously reported in the literature. Although this kind of epilepsy is quite frequent in pediatric age and the possibility of a casual co-occurrence should be considered, however in Kleefstra syndrome patients carrying 9q34.3 microdeletion a potential additional role of genetic (besides EHMT1) and epigenetic factors in developing seizures cannot be excluded. The present data expand the genomic and phenotypical features of the syndrome, providing new insights about research, which are useful to achieve genotype/phenotype correlations and better management of affected subjects.

Ventricular septal defects (VSDs) are recognized as one of the commonest congenital heart diseases (CHD), accounting for up to 40% of all cardiac malformations, and occur as isolated CHDs as well as together with other cardiac and extracardiac congenital malformations in individual patients and families. The genetic etiology of VSD is complex and extraordinarily heterogeneous. Chromosomal abnormalities such as aneuploidy and structural variations as well as rare point mutations in various genes have been reported to be associated with this cardiac defect. This includes both well-defined syndromes with known genetic cause (e.g., DiGeorge syndrome and Holt-Oram syndrome) and so far undefined syndromic forms characterized by unspecific symptoms. Mutations in genes encoding cardiac transcription factors (e.g., NKX2-5 and GATA4) and signaling molecules (e.g., CFC1) have been most frequently found in VSD cases. Moreover, new high-resolution methods such as comparative genomic hybridization enabled the discovery of a high number of different copy number variations, leading to gain or loss of chromosomal regions often containing multiple genes, in patients with VSD. In this chapter, we will describe the broad genetic heterogeneity observed in VSD patients considering recent advances in this field.

Novel approaches to uncover the molecular etiology of neurodevelopmental disorders (NDD) are highly needed. Even using a powerful tool such as whole exome sequencing (WES), the diagnostic process may still prove long and arduous due to the high clinical and genetic heterogeneity of these conditions. The main strategies to improve the diagnostic rate are based on family segregation, re-evaluation of the clinical features by reverse-phenotyping, re-analysis of unsolved NGS-based cases and epigenetic functional studies. In this article, we described three selected cases from a cohort of patients with NDD in which trio WES was applied, in order to underline the typical challenges encountered during the diagnostic process: (1) an ultra-rare condition caused by a missense variant in MEIS2, identified through the updated Solve-RD re-analysis; (2) a patient with Noonan-like features in which the NGS analysis revealed a novel variant in NIPBL causing Cornelia de Lange syndrome; and (3) a case with de novo variants in genes involved in the chromatin-remodeling complex, for which the study of the epigenetic signature excluded a pathogenic role. In this perspective, we aimed to (i) provide an example of the relevance of the genetic re-analysis of all unsolved cases through network projects on rare diseases; (ii) point out the role and the uncertainties of the reverse phenotyping in the interpretation of the genetic results; and (iii) describe the use of methylation signatures in neurodevelopmental syndromes for the validation of the variants of uncertain significance.

Kleefstra syndrome (KS; OMIM #610253), formerly known as the 9q subtelomeric deletion syndrome, is an autosomal dominant cause of intellectual disability (ID) characterized by hypotonia and facial dysmorphisms.(1,2) The cause of KS is attributed to haploinsufficiency of the euchromatin histone methyltransferase 1 (EHMT1) gene (OMIM *607001) located at chromosome 9q34.3 (i.e., distal long arm of chromosome 9), either by microdeletion or point mutation. EHMT1 encodes a histone H3 methyltransferase at position Lys-9 (H3K9).(1-3).