Antenatal diagnosis and maternal sirolimus treatment of polyhydramnios, megalencephaly, and symptomatic epilepsy (PMSE) syndrome

Pregnancy · 2026-04-29

Abstract Background Polyhydramnios, megalencephaly, and symptomatic epilepsy (PMSE) syndrome is a rare autosomal recessive mTORopathy caused by biallelic STE20‐related kinase adaptor alpha ( STRADA ) loss‐of‐function variants. Animal models demonstrate that in utero mechanistic target of rapamycin (mTOR) inhibition can prevent cortical dyslamination, suggesting a prenatal therapeutic window. Methods A gravida 3 para 2 patient presented at 32+2 weeks with medication‐refractory preterm labor and severe polyhydramnios. Fetal magnetic resonance imaging (MRI) demonstrated macrocephaly and an enlarged cavum septum pellucidum (CSP). Amniocentesis with microarray revealed homozygosity across the STRADA locus; sequencing confirmed the founder STRADA exons 9–13 deletion. After counseling, maternal sirolimus therapy was initiated at 35+6 weeks using a 20 mg loading regimen followed by 6–8 mg/day maintenance dosing. Serial ultrasound monitored fetal biometry and CSP size. Delivery occurred at 39+1 weeks. Pediatric follow‐up was extended nearly 5 years. Results Maternal sirolimus was well tolerated. Amniotic fluid index (AFI) declined from 36.1 to 26.2 cm, head circumference normalized from the 89th to 35th percentile, and CSP diameter decreased from 14.0 to 12.3 mm. A term vaginal delivery produced a 3554 g infant with normocephaly. Cord blood sirolimus level (3.9 mcg/L) approximated maternal level (3.4 mcg/L). Postnatally, sirolimus was continued from day of life 7. The first seizure occurred at 4.5 months, coincident with subtherapeutic trough levels; seizure control was achieved thereafter. At nearly 5 years, the child shows adequate growth but severe global developmental delay typical of PMSE. This case demonstrates the feasibility and pharmacokinetic effectiveness of maternal sirolimus during late gestation, with no identified maternal or fetal adverse consequences in this first case of third‐trimester prenatal mTOR inhibition in confirmed PMSE.

Mechanistic Target of Rapamycin and Megalencephaly: Novel Research Strategies for Therapeutic Discovery

Epiliepsy currents/Epilepsy currents · 2026-02-26

Megalencephaly (ME) is a malformation of cortical development defined by an enlarged brain. Individuals with ME often suffer from drug resistant epilepsy, intellectual disability, and autism spectrum disorder. Several clinical ME subtypes result from pathogenic variants in mTOR pathway genes (MPG) which cause diffuse brain overgrowth likely as a consequence of hyperactive mechanistic target of rapamycin (mTOR) signaling during brain development. Unfortunately, resected surgical or post-mortem ME brain tissue specimens are not widely available, and thus, there is only limited understanding of the histopathology of MPG associated ME. Thus, research strategies including new mouse models and human cerebral organoids have been developed to study the developmental pathogenesis of ME linked to MPG variants. These model systems provide a platform to study the mechanisms leading to brain overgrowth in ME as well as the establishment of the epileptic network. Perhaps most compelling, pre-clinical research approaches in ME models may pave the way for therapeutic development that could be deployed in utero to prevent ME formation.

The Clinical Spectrum and Neurodevelopmental Pathogenesis of <scp> <i>KPTN</i> </scp> ‐Related Disorder in a Mouse Model

Annals of Neurology · 2026-02-16

OBJECTIVE: Pathogenic variants in Kaptin (KPTN) cause KPTN-related disorder (KRD). KPTN modulates mTOR signaling activation within the KICSTOR complex in response to cellular amino acid levels. We define the clinical spectrum and investigate the developmental pathogenesis of KRD. METHODS: We report the genotype and clinical phenotype of 71 KRD individuals (28 female subjects, ages 1 to 55 years) including 48 newly identified KRD individuals. The effects of Kptn knockout on brain development were assayed in vitro and in vivo. RESULTS: We defined 15 novel KPTN variants. Intellectual disability (ID) was identified in all KRD individuals. Macrocephaly and epilepsy were observed in 46% and 47%, respectively. Neuroimaging revealed megalencephaly but no overt structural abnormalities. Ketotic hypoglycemia and endocrinopathies were identified in KRD. Increased head size was detected in unaffected parents heterozygous for KPTN variants. Two KRD individuals with drug-resistant epilepsy were treated with the mTOR inhibitor sirolimus but did not exhibit improved seizure control. CRISPR/Cas9 Kptn knockout in vitro induced mTOR activation and an mTOR-dependent increase in cell size. Kptn-/- mice exhibited increased cortical mTOR signaling that was reduced by rapamycin. Heterotopic neurons were identified in the subcortical white matter in the Kptn -/- mouse. Focal CRISPR/Cas9 Kptn knockout in cortex via in utero electroporation resulted in white matter heterotopic neurons. Electroencephalogram (EEG) did not detect ictal or inter-ictal abnormalities. INTERPRETATION: KRD is a multisystem neurodevelopmental disorder associated with ID, macrocephaly, epilepsy, mTOR signaling hyperactivation, and in a mouse model, subtle structural alterations in cerebral cortical cytoarchitecture. ANN NEUROL 2026;99:1287-1302.

Tuberous sclerosis complex

2025-04-01

Abstract Tuberous sclerosis complex (TSC) is an autosomal dominant or sporadic, multisystem disorder affecting predominantly the brain, lung, heart, skin, and kidney. TSC results from loss-of-function mutations in the TSC1 or TSC2 genes which encode proteins that are direct modulators of the mechanistic target of rapamycin (mTOR) pathway. The neurological and neuropsychiatric features of TSC include epilepsy in 80% of affected individuals, autism spectrum disorder in 50%, and intellectual disability in 50%. These clinical phenotypes are closely related to cortical dysplasias found in TSC cerebral cortex (“tubers”) that arise during embryonic brain development. Subependymal giant cell astrocytomas are intraventricular low-grade tumors that appear in approximately 10% of TSC patients and can enlarge to cause hydrocephalus, increased intracranial pressure, focal neurological deficits, and death. Until the approval of pharmacological mTOR inhibitors such as sirolimus and everolimus for the treatment of subependymal giant cell astrocytomas, resection was often required to ameliorate progressive neurological decline.

Clinical Center for Adults With Neurodevelopmental Disorders

Neurology · 2025-09-30 · 1 citations

Adults with neurodevelopmental disorders (ANDDs), including intellectual disability and autism spectrum disorder, are largely underserved by dedicated specialty centers in the United States. Many ANDDs also have seizures and sensorimotor deficits. To fill the gaps in care for ANDDs, we developed the Clinical Center for Adults with Neurodevelopmental Disorders (CCAND) to provide a multidisciplinary care program. Funding for CCAND staff salaries was solicited from and allocated by the Maryland General Assembly and Governor after our fair market salary analysis determined that physician professional fees would not be adequate to pay staff salaries. We retrospectively (2019-2024) present our clinical cohort at CCAND to demonstrate one strategy to improve quality of care for ANDDs and to illustrate a fiscal model and blueprint for similar centers around the United States. CCAND operates within an academic medical center. ANDDs aged 18 and older were referred to CCAND from regional pediatric providers through care transition, from area neurologists or psychiatrists, or by self-referral. CCAND staff includes a neurologist/epileptologist and psychiatrist, 2 advanced practice providers (certified registered nurse practitioner), a social worker, a certified genetic counselor, and an administrative assistant. Over 5 years, 305 ANDDs were evaluated in CCAND with >90% follow-up rate. During the coronavirus disease 2019 pandemic, all care was delivered successfully through telemedicine. Patients were evaluated for behavioral health needs and medication management. Social work services provided counseling, connection with state-offered services, and assistance with transition to semi-independent living. A total of 131 individuals had a history of seizures requiring ongoing treatment. In total, 150 individuals had not previously undergone standard-of-care genetic ascertainment, and in 17 of 86 individuals who were genetically ascertained by chromosomal microarray or whole-exome sequencing, a new genetic diagnosis was made including copy number and single-nucleotide variants. Care for ANDDs requires a multidisciplinary team approach. Because provider professional billing does not fully cover the salaries needed for support staff, state or other sources of support are necessary. We submit that advocacy to state governments to support other CCANDs could greatly influence access to quality care for ANDDs in the United States.

<i>SLC35A2</i> loss-of-function variants affect glycomic signatures, neuronal fate and network dynamics

Brain · 2025-05-25 · 7 citations

SLC35A2 encodes a uridine diphosphate (UDP)-galactose transporter essential for glycosylation of proteins and galactosylation of lipids and glycosaminoglycans. Germline genetic SLC35A2 variants have been identified in congenital disorders of glycosylation and somatic SLC35A2 variants have been linked to intractable epilepsy associated with malformations of cortical development. However, the functional consequences of these pathogenic variants on brain development and network integrity remain unknown. In this study, we used an isogenic human-induced pluripotent stem cell-derived neuron model to comprehensively interrogate the functional impact of loss-of-function variants in SLC35A2 through the integration of cellular and molecular biology, protein glycosylation analysis, neural network dynamics and single-cell electrophysiology. We show that loss-of-function variants in SLC35A2 result in disrupted glycomic signatures and precocious neurodevelopment, yielding hypoactive, asynchronous neural networks. This aberrant network activity is attributed to an inhibitory/excitatory imbalance as characterization of neural composition revealed preferential differentiation of SLC35A2 loss-of-function variants towards a GABAergic fate. Furthermore, electrophysiological recordings of synaptic activity and gene expression differences suggest network phenotypes are driven by changes occurring at the synapse. Our study is the first to provide mechanistic insight regarding the early development and functional connectivity of SLC35A2 loss-of-function variant harbouring human neurons, providing important groundwork for future exploration of potential therapeutic interventions.

mTOR pathway gene knockout results in mTOR-dependent cellular aggregation

bioRxiv (Cold Spring Harbor Laboratory) · 2025-11-04

Abstract Malformations of cortical development (MCD) caused by variants in mTOR pathway genes (MPGs) are a leading cause of drug-resistant epilepsy. Characteristic histopathological features of MPG-associated MCD include cytomegaly and cortical dyslamination often with neurons in abnormally close apposition (aggregates). We hypothesized that cellular aggregation is an mTOR-dependent phenotype. Tsc2, Nprl3, Stradα , or Kptn were knocked out (KO) using CRISPR/Cas9 in N2a cells in vitro . Levels of phosphorylated ribosomal S6 protein (PS6; Ser240/244), a marker for mTOR activation, were defined via Western blotting in vitro . Timelapse live-cell imaging was used to observe aggregate formation, with or without mTORC1 inhibition (rapamycin). EdU-base cell proliferation assay and cell death assays were performed to determine whether aggregation was the result of changes in cell cycle or increased cell death. Liquid chromatography-mass spectrometry (LC-MS/MS) was used to define changes in the cell aggregate proteome. Human MCD brain tissue specimens were stained with PS6 to assay mTOR signaling in neuronal clusters. All knockout lines formed multi-cell aggregates compared to control lines within 24-48 hours of plating in vitro . Aggregation was abolished with mTOR inhibitor treatment, establishing the mTOR-dependency of aggregate formation. Aggregation was not driven by cell proliferation, apoptosis/necrosis, or the presence of extracellular DNA in culture media. LC-MS/MS analysis revealed altered expression of protein across KO lines including adhesion molecules (e.g., contactin-3), cytoskeletal proteins (e.g., stathmin-2), and protein processing/transport (e.g., Uevld). Our findings establish aberrant cellular aggregation as an mTOR-dependent phenotype across multiple MPG associated with MCD. Changes in expression of adhesion molecules may contribute to abnormal cell aggregation and cortical lamination in MCD and results in abnormal network formation that leads to seizures. Highlights In human MCD specimens, neurons are frequently observed in clustered groups. In vitro models of mTORopathies show mTOR-dependent changes in cellular aggregation. Proteomic analysis revealed changes in protein levels in adhesion molecules and other molecules relevant to cellular dynamics and protein transport.

Loss of Slc35a2 alters development of the mouse cerebral cortex

Neuroscience Letters · 2024-06-22 · 18 citations

Brain somatic variants in SLC35A2, an intracellular UDP-galactose transporter, are commonly identified mutations associated with drug-resistant neocortical epilepsy and developmental brain malformations, including focal cortical dysplasia type I and mild malformation of cortical development with oligodendroglial hyperplasia in epilepsy (MOGHE). However, the causal effects of altered SLC35A2 function on cortical development remain untested. We hypothesized that focal Slc35a2 knockout (KO) or knockdown (KD) in the developing mouse cortex would disrupt cortical development and change network excitability. Through two independent studies, we used in utero electroporation (IUE) to introduce CRISPR/Cas9/targeted guide RNAs or short-hairpin RNAs into the embryonic mouse brain at day 14.5-15.5 to achieve Slc35a2 KO or KD, respectively, from neural precursor cells. Slc35a2 KO or KD caused disrupted radial migration of electroporated neurons evidenced by heterotopic cells located in lower cortical layers and in the sub-cortical white matter. Slc35a2 KO in neurons did not induce changes in oligodendrocyte number, importantly suggesting that the oligodendroglial hyperplasia observed in MOGHE originates from distinct cell autonomous effects of Slc35a2 mutations. Adult KO mice were implanted with EEG electrodes for 72-hour continuous recording. Spontaneous seizures were not observed in focal Slc35a2 KO mice, but there was reduced seizure threshold following pentylenetetrazol injection. Here we demonstrate that focal Slc35a2 KO or KD in vivo disrupts corticogenesis through altered neuronal migration and that KO leads to reduced seizure threshold. Together these results demonstrate a direct causal role for SLC35A2 in cortical development.

SLC35A2 loss of function variants affect glycomic signatures, neuronal fate, and network dynamics

bioRxiv (Cold Spring Harbor Laboratory) · 2024-12-27

Abstract SLC35A2 encodes a UDP-galactose transporter essential for glycosylation of proteins and galactosylation of lipids and glycosaminoglycans. Germline genetic SLC35A2 variants have been identified in congenital disorders of glycosylation and somatic SLC35A2 variants have been linked to intractable epilepsy associated with malformations of cortical development. However, the functional consequences of these pathogenic variants on brain development and network integrity remain elusive. In this study, we use an isogenic human induced pluripotent stem cell-derived neuron model to comprehensively interrogate the functional impact of loss of function variants in SLC35A2 through the integration of cellular and molecular biology, protein glycosylation analysis, neural network dynamics, and single cell electrophysiology. We show that loss of function variants in SLC35A2 result in disrupted glycomic signatures and precocious neurodevelopment, yielding hypoactive, asynchronous neural networks. This aberrant network activity is attributed to an inhibitory/excitatory imbalance as characterization of neural composition revealed preferential differentiation of SLC35A2 loss of function variants towards the GABAergic fate. Additionally, electrophysiological recordings of synaptic activity reveal a shift in excitatory/inhibitory balance towards increased inhibitory drive, indicating changes occurring specifically at the pre-synaptic terminal. Our study is the first to provide mechanistic insight regarding the early development and functional connectivity of SLC35A2 loss of function variant harboring human neurons, providing important groundwork for future exploration of potential therapeutic interventions.

Somatic variants in diverse genes leads to a spectrum of focal cortical malformations

UNC Libraries · 2024-07-17

Post-zygotically acquired genetic variants, or somatic variants, that arise during cortical development have emerged as important causes of focal epilepsies, particularly those due to malformations of cortical development. Pathogenic somatic variants have been identified in many genes within the PI3K-AKT-mTOR-signalling pathway in individuals with hemimegalencephaly and focal cortical dysplasia (type II), and more recently in SLC35A2 in individuals with focal cortical dysplasia (type I) or non-dysplastic epileptic cortex. Given the expanding role of somatic variants across different brain malformations, we sought to delineate the landscape of somatic variants in a large cohort of patients who underwent epilepsy surgery with hemimegalencephaly or focal cortical dysplasia. We evaluated samples from 123 children with hemimegalencephaly (n=16), focal cortical dysplasia type I and related phenotypes (n=48), focal cortical dysplasia type II (n=44), or focal cortical dysplasia type III (n=15). We performed high-depth exome sequencing in brain tissue-derived DNA from each case and identified somatic single nucleotide, indel and large copy number variants. In 75% of individuals with hemimegalencephaly and 29% with focal cortical dysplasia type II, we identified pathogenic variants in PI3K-AKT-mTOR pathway genes. Four of 48 cases with focal cortical dysplasia type I (8%) had a likely pathogenic variant in SLC35A2. While no other gene had multiple disease-causing somatic variants across the focal cortical dysplasia type I cohort, four individuals in this group had a single pathogenic or likely pathogenic somatic variant in CASK, KRAS, NF1 and NIPBL, genes previously associated with neurodevelopmental disorders. No rare pathogenic or likely pathogenic somatic variants in any neurological disease genes like those identified in the focal cortical dysplasia type I cohort were found in 63 neurologically normal controls (P=0.017), suggesting a role for these novel variants. We also identified a somatic loss-of-function variant in the known epilepsy gene, PCDH19, present in a small number of alleles in the dysplastic tissue from a female patient with focal cortical dysplasia IIIa with hippocampal sclerosis. In contrast to focal cortical dysplasia type II, neither focal cortical dysplasia type I nor III had somatic variants in genes that converge on a unifying biological pathway, suggesting greater genetic heterogeneity compared to type II. Importantly, we demonstrate that focal cortical dysplasia types I, II and III are associated with somatic gene variants across a broad range of genes, many associated with epilepsy in clinical syndromes caused by germline variants, as well as including some not previously associated with radiographically evident cortical brain malformations.