Objective: This study aimed to evaluate the diagnostic yield of whole genome sequencing (WGS) in patients with developmental epileptic encephalopathy manifesting with epileptic spasms (DEE-ES) and infantile epileptic spasms syndrome (IESS) with unknown etiology, and to assess treatment outcomes of patients with and without a genetic diagnosis.Methods: We included patients clinically diagnosed with ES, dividing them into the DEE-ES and IESS groups with no identifiable causes from prior neuroimaging, genetic, or metabolic testing. WGS was performed for all patients, and their parents underwent targeted sequencing.Results: WGS identified pathogenic or likely pathogenic variants in 15 of 56 (26.7%) patients. Specifically, the yield for the DEE-ES and IESS groups was 6 of 13 (46.7%) and 9 of 43 (20.9%), respectively. Of those with a genetic diagnosis, 12 had single nucleotide variants, 3 had copy number variants, and 1 had both. KCNQ2 was the most frequently affected gene in the DEE-ES group. Clinical management changed for 4 (26.7%) of these patients. Cessation of ES after the first treatment was similar regardless of genetic diagnosis. A clinical response to the first treatment was associated with long-term favorable seizure outcomes, particularly in the IESS group, although patients with a genetic diagnosis had a higher rate of developmental delay at the last follow-up.Conclusions: WGS identified genetic causes in 46.7% of DEE-ES patients and 20.9% IESS patients, with limited impact on clinical management. Patients in the IESS group with a genetic diagnosis exhibited poorer long-term developmental outcomes.

BackgroundThis study aims to assess the effectiveness of non-invasive preimplantation genetic testing (niPGT) for detecting aneuploidy at two different time points, 48- and 54-hour following the cleavage stage of embryo development.MethodsA cohort of 15 infertile women was enrolled, involving a total of 58 embryos. All embryos underwent sequential culture media procedures and received assisted hatching during the cleavage stage on Day 3. After biopsy examination, conducted either 48 or 54 hours post cleavage, spent culture media (SCM) were gathered and processed for the amplification and quantification of cell-free DNA. This was followed by low-pass whole genome sequencing.ResultsThe cell-free DNA content within SCM remained consistent across both time points post cleavage. The accuracy of niPGT in ploidy detection, in comparison to trophectoderm biopsy, was 53.71%. No significant distinction in ploidy detection accuracy was observed between SCM collected from embryos at 48 hours versus those at 54 hours post cleavage. The overall accuracy for sex determination reached 79.63%.ConclusionsThe concentration of cell-free DNA within SCM was found to be consistent at both 48- and 54-hours after embryo cleavage. However, the accuracy of ploidy determination, when contrasted with the conventional trophectoderm biopsy, did not yield satisfactory outcomes.

Congenital Adrenal Hyperplasia (CAH) due to 21-hydroxylase deficiency (21-OHD CAH) is an autosomal recessive disorder resulting from pathogenic variants in the CYP21A2 gene. The disorder exhibits variable clinical severity, with the classical form manifesting as salt-wasting crisis in neonates, while inducing ambiguous genitalia in females and precocious puberty in males through simple virilization. Identifying at-risk couples during the preconception stage holds significance for optimizing reproductive choices. Methods: This study included 204 unrelated preconception individuals undergoing carrier screening. A robust molecular approach was devised for rapid detection of nine prevalent CYP21A2 pathogenic variants, utilizing Amplification-Refractory Mutation System (ARMS) PCR and mass spectrometry (MS) genotyping. Complementary quantitative real-time PCR (qPCR) and PCR-based Restriction Fragment Length Polymorphism (PCR-based RFLP) assays were employed for comprehensive gene deletion analysis. The concordance of pathogenic variant detection between ARMS-PCR and MS, as well as the consistency observed in molecular insights from qPCR and PCR-based RFLP, fortified the accuracy of our methodologies. Results: Our combined method could detect common pathogenic variants and large gene deletions with high concordance between ARMS-PCR, MS genotyping, qPCR, and PCR-based RFLP assays. Remarkably, two carriers exhibited significant large-scale deletions, while another manifested a carrier state due to minor-scale gene conversion. The estimated carrier frequency in our cohort using these methods was approximately 1 in 65 individuals. Conclusions: The methods used for 21-OHD CAH carrier screening offer a reliable, swift, and cost-effective approach for detecting common pathogenic variants and large deletions. Despite some limitations, such as the inability to detect all rare mutations, the techniques provide a practical solution for carrier screening, with an estimated carrier frequency of 1 in 65 in our study population. These findings support the potential adoption of these methods in national carrier screening programs, offering a practical balance between efficiency and affordability.

The MUi040-A human induced pluripotent stem cell (hiPSC) line was established from CD34 + hematopoietic stem cells of a male patient with senile amyloidosis. Patient carries a unique chromosome inversion at bands 9p12 and 9q13, which is one of the most common variants of the normal karyotype of chromosome 9 (inv[9]). This hiPSC line maintains an identical karyotype to that of the parental cells. MUi040-A expressed pluripotency markers and demonstrated the capacity to differentiate into all three germ layers. This cell line offers a valuable in vitro for studying the pathophysiology of senile systemic amyloidosis and for evaluating potential therapeutic approaches.

Schwartz-Jampel syndrome (SJS) is a rare autosomal recessive disorder characterized by typical facial dysmorphism, generalized muscle stiffness, joint contracture, and skeletal abnormalities. This condition is caused by mutations in the heparan sulfate proteoglycan 2 (HSPG2) gene, which encodes perlecan, a component of the basement membrane. The management of patients with SJS primarily aims to alleviate symptoms related to muscle stiffness. In this report, we describe a male patient with SJS type 1A. Trio whole-exome sequencing identified a pathogenic mutation (NM_001291860.1: c.10897C>T; p.Arg3633Ter) and variants of unknown significance (NM_001291860.2: c.413+10G>T). The patient experienced difficulty in opening his eyes and mouth, which significantly limited his daily activities. Botulinum toxin A injection was administered and demonstrated significant clinical improvement after the treatment.

Fragile X syndrome (FXS) is a neurodevelopmental disorder, caused by an CGG repeat expansion (FM, > 200 CGG) in the fragile X messenger ribonucleoprotein 1 (FMR1) gene. Female carriers of a premutation (PM; 55–200 CGG) can transmit the PM allele, which, depending on the CGG allele size, can expand to an allele in the FM range in the offspring. Carrier screening for FMR1 PM is not available in Thailand. This study aimed to investigate the prevalence of PM carriers among Thai reproductive women at the tertiary hospital. A total of 1250 females participated in this study; ages ranged from 20 to 45 years, mean of 30 years (S.D. = 6.27). Two carriers of a premutation allele, with 32,62 and 32,69 CGG repeats respectively, were identified. This corresponds to 1 in 600 women or 0.17

BACKGROUND:Epileptic spasms are a devastating form of early infantile epileptic encephalopathy (EIEE) with various etiologies. Early diagnosis and a shorter lead time to treatment are crucial to stop the seizures and optimize the neurodevelopmental outcome. Genetic testing has become an integral part of epilepsy care that directly guides management and family planning and discovers new targeted treatments. Neuronal differentiation Factor 2 (NEUROD2) variants have recently been a cause of neurodevelopmental disorders (NDDs) and EIEEs with distinctive features. However, there is limited information about the clinical and electroencephalographic response of epileptic spasm treatment in NEUROD2-related NDD syndrome. CASE PRESENTATION:We report a female patient of Southeast Asian ethnicity with global developmental delay and epileptic spasms commencing in the first few months of life. A novel de novo heterozygous pathogenic NEUROD2 variant, p. E130Q, was subsequently identified by whole-exome sequencing. Electroencephalogram before treatment showed multifocal independent spikes predominantly in both posterior head regions and demonstrated marked improvement following combined vigabatrin and high-dose prednisolone treatment. However, multiple courses of relapse occurred after weaning off the antiseizure medication. CONCLUSIONS:We propose that epileptic spasms related to de novo NEUROD2 pathogenic variant respond well to combined vigabatrin and high-dose prednisolone therapy. These findings may imply the benefit of using combination therapy to treat epileptic spasms in NEUROD2-related NDD syndrome.

Hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM) are the most common referrals in the Inherited Cardiovascular Condition (ICC) Genetics Service. Several issues must be discussed with patients and their families during the genetic consultation session, including the options for genetic testing and cardiovascular surveillance in family members. We developed an ICC registry and performed next-generation-based DNA sequencing for all patients affected by non-syndromic HCM and idiopathic DCM in our joint specialist genetics service. The target gene sequencing panel relied on the Human Phenotype Ontology with 237 genes for HCM (HP:0001639) and 142 genes for DCM (HP:0001644). All subjects were asked to contact their asymptomatic first-degree relatives for genetic counseling regarding their risks and to initiate cardiovascular surveillance and cascade genetic testing. The study was performed from January 1, 2014, to December 31, 2020, and a total of 62 subjects (31-HCM and 31-DCM) were enrolled. The molecular detection frequency was 48.39% (32.26% pathogenic/likely pathogenic, 16.13% variant of uncertain significance or VUS for HCM, and 25.81% (16.13% pathogenic/likely pathogenic, 9.68% VUS) for DCM. The most prevalent gene associated with HCM was MYBPC3 . The others identified in this study included ACTN2 , MYL2 , MYH7 , TNNI3 , TPM1 , and VCL . Among the DCM subjects, variants were detected in two cases with the TTN nonsense variants, while the others were missense and identified in MYH7 , DRSP3 , MYBPC3 , and SCN5A . Following the echocardiogram surveillance and cascade genetic testing in the asymptomatic first-degree relatives, the detection rate of new cases was 8.82% and 6.25% in relatives of HCM and DCM subjects, respectively. Additionally, a new pre-symptomatic relative belonging to an HCM family was identified, although the genomic finding in the affected case was absent. Thus, ICC service is promising for the national healthcare system, aiming to prevent morbidity and mortality in asymptomatic family members.

AIM:Osteogenesis imperfecta (OI) is a hereditary connective tissue disorder primarily caused by mutations in COL1A1 or COL1A2, which encode type I collagen. These mutations affect the quantity and/or quality of collagen composition in bones, leading to bone fragility. Currently, there is still a lack of treatment that addresses disease-causing factors due to an insufficient understanding of the pathological mechanisms involved.MAIN METHODS:Induced pluripotent stem cells (iPSCs) were generated from OI patients with glycine substitution mutations in COL1A1 and COL1A2 and developed into mesenchymal stem cells (iPS-MSCs). OI-derived iPS-MSCs underwent in vitro osteogenic induction to study cell growth, osteogenic differentiation capacity, mRNA expression of osteogenic and unfolded protein response (UPR) markers and apoptosis. The effects of 4-phenylbutyric acid (4-PBA) were examined after treatment of OI iPS-MSCs during osteogenesis.KEY FINDINGS:OI-derived iPS-MSCs exhibited decreased cell growth and impaired osteogenic differentiation and collagen expression. Expression of UPR genes was increased, which led to an increase in apoptotic cell death. 4-PBA treatment decreased apoptotic cells and reduced expression of UPR genes, including HSPA5, XBP1, ATF4, DDIT3, and ATF6. Osteogenic phenotypes, including RUNX2, SPP1, BGLAP, and IBPS expression, as well as calcium mineralization, were also improved.SIGNIFICANCE:MSCs differentiated from disease-specific iPSCs have utility as a disease model for identifying disease-specific treatments. In addition, the ER stress-associated UPR could be a pathogenic mechanism associated with OI. Treatment with 4-PBA alleviated OI pathogenesis by attenuating UPR markers and apoptotic cell death.

Pantothenate kinase-associated neurodegeneration (PKAN) is linked to brain iron accumulation caused by PANK2 gene mutation. Despite the importance of genetic testing to confirm PKAN and identify at risk parents, genetic screening is financially burdensome for developing countries like Thailand. Because genetic screeners are expensive and not reimbursed by the universal health care coverage system, they are not typically performed. To investigate clinical symptoms, radiological findings and mutation analysis for patients based in Thailand with unknown genetic status but suspected PKAN based on clinical symptoms. Genetic testing was performed for cases suspected for PKAN and their biological parents by direct genomic sequencing of PANK2 at Maharat Nakhon Ratchasima Hospital during 2017-2018. Clinical evaluation and documentation were performed by pediatric neurologists. Five children had classical onset form of PKAN. Most presented with gait dystonia. Three patients diagnosed after 4 years showed the eye-of-the-tiger sign in their brain MRI, whereas two younger patients revealed only isolated hyperintensity bilateral globus pallidus. However, PANK2 mutations were identified in all cases: the most common mutation was c.982-1G>C. This mutation was detected in four unrelated individuals but not reported in other studies. Genetic testing is recommended to confirm diagnoses in cases with supporting clinical features of PKAN with or without the classical 'eye-of-the-tiger-sign'. A novel PANK2 mutation (c.982-1G>C) was identified in South East Asian populations based in Thailand, suggesting that this genetic variant is a founder genotype in this population. Moreover, genetic diagnosis is helpful to provide appropriate genetic counseling to families. (C) 2019 Elsevier Ltd. All rights reserved.

Mucopolysaccharidosis Type VII (MPS7, also called β‐glucuronidase deficiency or Sly syndrome; MIM 253220) is an extremely rare autosomal recessive lysosomal storage disease, caused by mutations in the GUSB gene. β‐glucuronidase (GUSB) is a lysosomal hydrolase involved in the stepwise degradation of glucuronic acid‐containing glycosaminoglycans (GAGs). Patients affected with MPS VII are not able to completely degrade glucuronic acid‐containing GAGs, including chondroitin 4‐sulfate, chondroitin 6‐sulfate, dermatan sulfate, and heparan sulfate. The accumulation of these GAGs in lysosomes of various tissues leads to cellular and organ dysfunctions. Characteristic features of MPS VII include short stature, macrocephaly, hirsutism, coarse facies, hearing loss, cloudy cornea, short neck, valvular cardiac defects, hepatosplenomegaly, and dysostosis multiplex. Oral manifestations in patients affected with MPS VII have never been reported. Oral manifestations observed in three patients consist of wide root canal spaces, taurodontism, hyperplastic dental follicles, malposition of unerupted permanent molars, and failure of tooth eruption with malformed roots. The unusual skeletal features of the patients include maxillary hypoplasia, hypoplastic midface, long mandibular length, mandibular prognathism, hypoplastic and aplastic mandibular condyles, absence of the dens of the second cervical vertebra, and erosion of the cortex of the lower border of mandibles. Dogs affected with MPS VII had anterior and posterior open bite, maxillary hypoplasia, premolar crowding, and mandibular prognathism. Unlike patients with MPS VII, the dogs had unremarkable mandibular condyles. This is the first report of oral manifestations in patients affected with MPS VII.

Thalassemia and hemoglobinopathy is a group of hereditary blood disorder with diverse clinical manifestation inherited by autosomal recessive manner. The Beta thalassemia/Hemoglobin E disease (HbE/βthal) causes a variable degree of hemolysis and the most severe form of HbE/βthal disease develop a lifelong transfusion-dependent anemia. Preimplantation genetic testing (PGT) is an established procedure of embryo genetic analysis to avoid the risk of passing on this particular condition from the carrier parents to their offspring. Preimplantation genetic testing for chromosomal aneuploidy (PGT-A) also facilitates the selection of embryos without chromosomal aberration resulting in the successful embryo implantation rate. Herein, we study the clinical outcome of using combined PGT-M and PGT-A in couples at risk of passing on HbE/βthal disease. The study was performed from January 2016 to December 2017. PGT-M was developed using short tandem repeat linkage analysis around the beta globin gene cluster and direct mutation testing using primer extension-based mini-sequencing. Thereafter, we recruited 15 couples at risk of passing on HbE/βthal disease who underwent a combined total of 22 IVF cycles. PGT was performed in 106 embryos with a 3.89% allele drop-out rate. Using combined PGT-M and PGT-A methods, 80% of women obtained satisfactory genetic testing results and were able to undergo embryo transfer within the first two cycles. The successful implantation rate was 64.29%. PGT accuracy was evaluated by prenatal and postnatal genetic confirmation and 100% had a genetic status consistent with PGT results. The overall clinical outcome of successful live birth for couples at risk of producing offspring with HbE/βthal was 53.33%. Conclusively, combined PGT-M and PGT-A is a useful technology to prevent HbE/βthal disease in the offspring of recessive carriers.

Thalassemia and hemoglobinopathy is a group of hereditary blood disorder with diverse clinical manifestation inherited by autosomal recessive manner. The Beta thalassemia/Hemoglobin E disease (HbE/beta thal) causes a variable degree of hemolysis and the most severe form of HbE/beta thal disease develop a lifelong transfusion-dependent anemia. Preimplantation genetic testing (PGT) is an established procedure of embryo genetic analysis to avoid the risk of passing on this particular condition from the carrier parents to their offspring. Preimplantation genetic testing for chromosomal aneuploidy (PGT-A) also facilitates the selection of embryos without chromosomal aberration resulting in the successful embryo implantation rate. Herein, we study the clinical outcome of using combined PGT-M and PGT-A in couples at risk of passing on HbE/beta thal disease. The study was performed from January 2016 to December 2017. PGT-M was developed using short tandem repeat linkage analysis around the beta globin gene cluster and direct mutation testing using primer extension-based mini-sequencing. Thereafter, we recruited 15 couples at risk of passing on HbE/beta thal disease who underwent a combined total of 22 IVF cycles. PGT was performed in 106 embryos with a 3.89% allele drop-out rate. Using combined PGT-M and PGT-A methods, 80% of women obtained satisfactory genetic testing results and were able to undergo embryo transfer within the first two cycles. The successful implantation rate was 64.29%. PGT accuracy was evaluated by prenatal and postnatal genetic confirmation and 100% had a genetic status consistent with PGT results. The overall clinical outcome of successful live birth for couples at risk of producing offspring with HbE/beta thal was 53.33%. Conclusively, combined PGT-M and PGT-A is a useful technology to prevent HbE/beta thal disease in the offspring of recessive carriers.