Communication of individuals with CDKL5 deficiency disorder as observed by caregivers: A descriptive qualitative study.

CDKL5 deficiency disorder (CDD) is a genetically caused developmental epileptic encephalopathy that causes severe communication impairments. Communication of individuals with CDD is not well understood in the literature and currently available measures are not well validated in this population. Accurate and sensitive measurement of the communication of individuals with CDD is important for understanding this condition, clinical practice, and upcoming interventional trials. The aim of this descriptive qualitative study was to understand how individuals with CDD communicate, as observed by caregivers. Participants were identified through the International CDKL5 Disorder Database and invited to take part if their child had a pathogenic variant of the CDKL5 gene and they had previously completed the Communication and Symbolic Behavior Checklist (CSBS-DP ITC). The sample comprised caregivers of 23 individuals with CDD, whose ages ranged from 2 to 30 years (median 13 years), 15 were female, and most did not use words. Semistructured interviews were conducted via videoconference and analyzed using a conventional content analysis. Three overarching categories were identified: mode, purpose and meaning, and reciprocal exchanges. These categories described the purposes and mechanism of how some individuals with CDD communicate, including underpinning influential factors. Novel categories included expressing a range of emotions, and reciprocal exchanges (two-way interactions that varied in complexity). Caregivers observed many communication modes for multiple purposes. Understanding how individuals with CDD communicate improves understanding of the condition and will guide research to develop accurate measurement for clinical practice and upcoming medication trials.

Long-term treatment with ganaxolone for seizures associated with cyclin-dependent kinase-like 5 deficiency disorder: Two-year open-label extension follow-up.

OBJECTIVE: In the placebo-controlled, double-blind phase of the Marigold study (NCT03572933), ganaxolone significantly reduced major motor seizure frequency (MMSF) in patients with cyclin-dependent kinase-like 5 deficiency disorder (CDD). We report 2-year safety and clinical outcomes data from the open-label extension (OLE) phase of Marigold. METHODS: Patients with CDD who completed the double-blind phase were eligible to continue in the OLE. Efficacy assessments included MMSF reduction from prerandomization baseline, responder rates, and Clinical Global Impression-Improvement scores, including assessment of seizure intensity and duration (CGI-CSID). Safety assessments included treatment-emergent adverse events (TEAEs) and TEAEs leading to discontinuation. RESULTS: Of 101 patients who enrolled in Marigold, 88 (87.1%) entered the OLE (median age = 5 years, 79.5% female). Median 28-day MMSF at baseline was 50.6. At 2 years in the OLE (months 22-24), MMSF was reduced by a median of 48.2% (n = 50); when missing data were imputed, median reduction in MMSF was 43.8% using a mixed effects model and 27.4% using a last observation carried forward model. During months 22-24, 23 of 50 (46.0%) patients experienced reductions in MMSF of ≥50%; 12 of 50 (24.0%) patients experienced MMSF reductions of ≥75%. During months 22-24, 40 of 49 (81.6%) patients were rated by caregivers as having improvement in seizure-related outcomes based on CGI-CSID scores. Thirty-seven patients discontinued ganaxolone due to lack of efficacy (n = 13), withdrawal by caregiver (n = 12), adverse event (n = 10), physician decision (n = 1), or death (n = 1; unrelated to study drug). The most common treatment-related TEAEs were somnolence (17.0%), seizure (11.4%), and decreased appetite (5.7%). Patients reported serious TEAEs (n = 28, 31.8%); those reported in ≥3% of patients were seizure (n = 6), pneumonia (n = 5), acute respiratory failure (n = 3), aspiration pneumonia (n = 3), and dehydration (n = 3). SIGNIFICANCE: Sustained reductions in MMSF at 2 years in the OLE support the efficacy of ganaxolone in seizures associated with CDD. Safety findings in the OLE were consistent with the double-blind phase.

CDKL5 deficiency disorder and other infantile-onset genetic epilepsies.

AIM: To differentiate phenotypic features of individuals with CDKL5 deficiency disorder (CDD) from those of individuals with other infantile-onset epilepsies. METHOD: We performed a retrospective cohort study and ascertained individuals with CDD and comparison individuals with infantile-onset epilepsy who had epilepsy gene panel testing. We reviewed records, updated variant classifications, and compared phenotypic features. Wilcoxon rank-sum tests and χ2 or Fisher's exact tests were performed for between-cohort comparisons. RESULTS: We identified 137 individuals with CDD (110 females, 80.3%; median age at last follow-up 3 year 11 months) and 313 individuals with infantile-onset epilepsies (156 females, 49.8%; median age at last follow-up 5 years 2 months; 35% with genetic diagnosis). Features reported significantly more frequently in the CDD group than in the comparison cohort included developmental and epileptic encephalopathy (81% vs 66%), treatment-resistant epilepsy (95% vs 71%), sequential seizures (46% vs 6%), epileptic spasms (66% vs 42%, with hypsarrhythmia in 30% vs 48%), regression (52% vs 29%), evolution to Lennox-Gastaut syndrome (23% vs 5%), diffuse hypotonia (72% vs 36%), stereotypies (69% vs 11%), paroxysmal movement disorders (29% vs 17%), cerebral visual impairment (94% vs 28%), and failure to thrive (38% vs 22%). INTERPRETATION: CDD, compared with other suspected or confirmed genetic epilepsies presenting in the first year of life, is more often characterized by a combination of treatment-resistant epilepsy, developmental and epileptic encephalopathy, sequential seizures, spasms without hypsarrhythmia, diffuse hypotonia, paroxysmal movement disorders, cerebral visual impairment, and failure to thrive. Defining core phenotypic characteristics will improve precision diagnosis and treatment.

Analysis of electrocardiograms in individuals with CDKL5 deficiency disorder.

CDKL5 deficiency disorder (CDD) is an epileptic encephalopathy associated with medically refractory epilepsy. We sought to determine whether prolonged corrected QT interval (QTc) or other cardiac conduction abnormalities were seen in CDD in a clinical cohort. A cohort of individuals with CDD was evaluated in the Children's Hospital Colorado's International Foundation for CDKL5 Research designated Center of Excellence clinic with routine electrocardiograms obtained as part of routine clinical care. Retrospective review of electrocardiograms was completed. ECGs from 44 individuals (7 male, 37 female, age range 0-34.5 years) with pathogenic mutations and findings consistent with CDD were evaluated. Multiple ECGs were available from the 44 individuals obtained from 1996 to 2020. Prolonged QTc was found in two individuals (4.5%) and either resolved or was not confirmed on Holter monitor; no additional interventions were performed. A total of 11 individuals had echocardiograms for a variety of indications including unexplained tachycardia and ECG abnormalities; all were normal. Two individuals in the cohort died during the study with no abnormal findings on ECG. The incidence of prolonged QTc or other significant actionable cardiac abnormalities was rare in a cohort of individuals with CDD though was higher than the prevalence seen within the general population. Further studies in a larger, confirmatory cohort over a longer period are needed.

Epileptic spasms in CDKL5 deficiency disorder: Delayed treatment and poor response to first-line therapies.

OBJECTIVE: We aimed to assess the treatment response of infantile-onset epileptic spasms (ES) in CDKL5 deficiency disorder (CDD) vs other etiologies. METHODS: We evaluated patients with ES from the CDKL5 Centers of Excellence and the National Infantile Spasms Consortium (NISC), with onset from 2 months to 2 years, treated with adrenocorticotropic hormone (ACTH), oral corticosteroids, vigabatrin, and/or the ketogenic diet. We excluded children with tuberous sclerosis complex, trisomy 21, or unknown etiology with normal development because of known differential treatment responses. We compared the two cohorts for time to treatment and ES remission at 14 days and 3 months. RESULTS: We evaluated 59 individuals with CDD (79% female, median ES onset 6 months) and 232 individuals from the NISC database (46% female, median onset 7 months). In the CDD cohort, seizures prior to ES were common (88%), and hypsarrhythmia and its variants were present at ES onset in 34%. Initial treatment with ACTH, oral corticosteroids, or vigabatrin started within 1 month of ES onset in 27 of 59 (46%) of the CDD cohort and 182 of 232 (78%) of the NISC cohort (p < .0001). Fourteen-day clinical remission of ES was lower for the CDD group (26%, 7/27) than for the NISC cohort (58%, 106/182, p = .0002). Sustained ES remission at 3 months occurred in 1 of 27 (4%) of CDD patients vs 96 of 182 (53%) of the NISC cohort (p < .0001). Comparable results were observed with longer lead time (≥1 month) or prior treatment. Ketogenic diet, used within 3 months of ES onset, resulted in ES remission at 1 month, sustained at 3 months, in at least 2 of 13 (15%) individuals with CDD. SIGNIFICANCE: Compared to the broad group of infants with ES, children with ES in the setting of CDD often experience longer lead time to treatment and respond poorly to standard treatments. Development of alternative treatments for ES in CDD is needed.

Assessing the landscape of STXBP1-related disorders in 534 individuals.

Disease-causing variants in STXBP1 are among the most common genetic causes of neurodevelopmental disorders. However, the phenotypic spectrum in STXBP1-related disorders is wide and clear correlations between variant type and clinical features have not been observed so far. Here, we harmonized clinical data across 534 individuals with STXBP1-related disorders and analysed 19 973 derived phenotypic terms, including phenotypes of 253 individuals previously unreported in the scientific literature. The overall phenotypic landscape in STXBP1-related disorders is characterized by neurodevelopmental abnormalities in 95% and seizures in 89% of individuals, including focal-onset seizures as the most common seizure type (47%). More than 88% of individuals with STXBP1-related disorders have seizure onset in the first year of life, including neonatal seizure onset in 47%. Individuals with protein-truncating variants and deletions in STXBP1 (n = 261) were almost twice as likely to present with West syndrome and were more phenotypically similar than expected by chance. Five genetic hotspots with recurrent variants were identified in more than 10 individuals, including p.Arg406Cys/His (n = 40), p.Arg292Cys/His/Leu/Pro (n = 30), p.Arg551Cys/Gly/His/Leu (n = 24), p.Pro139Leu (n = 12), and p.Arg190Trp (n = 11). None of the recurrent variants were significantly associated with distinct electroclinical syndromes, single phenotypic features, or showed overall clinical similarity, indicating that the baseline variability in STXBP1-related disorders is too high for discrete phenotypic subgroups to emerge. We then reconstructed the seizure history in 62 individuals with STXBP1-related disorders in detail, retrospectively assigning seizure type and seizure frequency monthly across 4433 time intervals, and retrieved 251 anti-seizure medication prescriptions from the electronic medical records. We demonstrate a dynamic pattern of seizure control and complex interplay with response to specific medications particularly in the first year of life when seizures in STXBP1-related disorders are the most prominent. Adrenocorticotropic hormone and phenobarbital were more likely to initially reduce seizure frequency in infantile spasms and focal seizures compared to other treatment options, while the ketogenic diet was most effective in maintaining seizure freedom. In summary, we demonstrate how the multidimensional spectrum of phenotypic features in STXBP1-related disorders can be assessed using a computational phenotype framework to facilitate the development of future precision-medicine approaches.

De novo DHDDS variants cause a neurodevelopmental and neurodegenerative disorder with myoclonus.

Subcellular membrane systems are highly enriched in dolichol, whose role in organelle homeostasis and endosomal-lysosomal pathway remains largely unclear besides being involved in protein glycosylation. DHDDS encodes for the catalytic subunit (DHDDS) of the enzyme cis-prenyltransferase (cis-PTase), involved in dolichol biosynthesis and dolichol-dependent protein glycosylation in the endoplasmic reticulum. An autosomal recessive form of retinitis pigmentosa (retinitis pigmentosa 59) has been associated with a recurrent DHDDS variant. Moreover, two recurring de novo substitutions were detected in a few cases presenting with neurodevelopmental disorder, epilepsy and movement disorder. We evaluated a large cohort of patients (n = 25) with de novo pathogenic variants in DHDDS and provided the first systematic description of the clinical features and long-term outcome of this new neurodevelopmental and neurodegenerative disorder. The functional impact of the identified variants was explored by yeast complementation system and enzymatic assay. Patients presented during infancy or childhood with a variable association of neurodevelopmental disorder, generalized epilepsy, action myoclonus/cortical tremor and ataxia. Later in the disease course, they experienced a slow neurological decline with the emergence of hyperkinetic and/or hypokinetic movement disorder, cognitive deterioration and psychiatric disturbances. Storage of lipidic material and altered lysosomes were detected in myelinated fibres and fibroblasts, suggesting a dysfunction of the lysosomal enzymatic scavenger machinery. Serum glycoprotein hypoglycosylation was not detected and, in contrast to retinitis pigmentosa and other congenital disorders of glycosylation involving dolichol metabolism, the urinary dolichol D18/D19 ratio was normal. Mapping the disease-causing variants into the protein structure revealed that most of them clustered around the active site of the DHDDS subunit. Functional studies using yeast complementation assay and in vitro activity measurements confirmed that these changes affected the catalytic activity of the cis-PTase and showed growth defect in yeast complementation system as compared with the wild-type enzyme and retinitis pigmentosa-associated protein. In conclusion, we characterized a distinctive neurodegenerative disorder due to de novo DHDDS variants, which clinically belongs to the spectrum of genetic progressive encephalopathies with myoclonus. Clinical and biochemical data from this cohort depicted a condition at the intersection of congenital disorders of glycosylation and inherited storage diseases with several features akin to of progressive myoclonus epilepsy such as neuronal ceroid lipofuscinosis and other lysosomal disorders.

Psychometric properties of QI-Disability in CDKL5 Deficiency Disorder: Establishing readiness for clinical trials.

CDKL5 Deficiency Disorder (CDD) is a rare genetic disorder with symptoms of epilepsy, developmental impairments, and other comorbidities. Currently, there are no outcome measures for CDD with comprehensive evidence of validation. This study aimed to evaluate the psychometric properties of the Quality of Life Inventory-Disability (QI-Disability) in CDD. Quality of Life Inventory-Disability was administered to 152 parent caregivers registered with the International CDKL5 Disorder Database (ICDD). Confirmatory factor analysis was conducted and the goodness of fit of the factor structure was assessed. Fixed-effects linear regression models examined the responsiveness of QI-Disability to reported changes in child health. A subset of parent caregivers (n = 56) completed QI-Disability, as well as additional health-related questions, on two occasions separated by four weeks to evaluate test-retest reliability. Test-retest reliability was assessed using intra-class correlations (ICCs) calculated from QI-Disability scores. Based upon adjustments for changes in child health, ICCs were recalculated to estimate responsiveness to change. Confirmatory factor analysis, internal consistency, and divergent validity were mostly satisfactory, except divergent validity was not satisfactory for the Social Interactions and Independence domains. The Physical Health, Social Interactions, Leisure, and Total scores responded to changes in the child's Physical health, and the Negative Emotions and Leisure domains responded to changes in the child's behavior. Unadjusted and adjusted ICC values were above 0.8 for the Positive Emotions, Negative Emotions, Social Interactions, Leisure, Independence domains and Total score, and above 0.6 for the Physical Health domain. Findings suggest that QI-Disability is suitable to assess the quality of life of children and adults with CDD and could be of value for upcoming clinical trials.

Efficacy, safety, and tolerability of soticlestat as adjunctive therapy for the treatment of seizures in patients with Dup15q syndrome or CDKL5 deficiency disorder in an open-label signal-finding phase II study (ARCADE).

OBJECTIVE: Chromosome 15q duplication (Dup15q) syndrome and cyclin­dependent kinase-like 5 deficiency disorder (CDD) are rare neurodevelopmental disorders associated with epileptic encephalopathies, with a lack of specifically approved treatment options. ARCADE assessed the efficacy and safety of adjunctive soticlestat (TAK-935) for the treatment of seizures in patients with Dup15q syndrome or CDD (NCT03694275). METHODS: ARCADE was a phase II, open-label, pilot study of soticlestat (≤300 mg/day twice daily, weight-adjusted) in pediatric and adult patients 2-55 years of age with Dup15q syndrome or CDD who experienced ≥3 motor seizures per month in the 3 months before screening and at baseline. The 20-week treatment period consisted of a dose-optimization period and a 12-week maintenance period. Efficacy endpoints included the change from baseline in motor seizure frequency during the maintenance period and the proportion of treatment responders. Safety endpoints included the incidence of treatment-emergent adverse effects (TEAEs). RESULTS: The modified-intent-to-treat population included 20 participants who received ≥1 dose of soticlestat and had ≥1 efficacy assessment (Dup15q syndrome, n = 8; CDD, n = 12). Soticlestat administration during the maintenance period was associated with a median change from baseline in motor seizure frequency of +11.7% in the Dup15q syndrome group and -23.6% in the CDD group. Reductions in all seizure frequency of -23.4% and -30.5% were also observed during the maintenance period in the Dup15q syndrome group and the CDD group, respectively. Most TEAEs were of mild or moderate severity. Serious TEAEs were reported by three patients (15.0%); none were considered drug related. The most common TEAEs were constipation, rash, and seizure. No deaths were reported. CONCLUSIONS: Adjunctive soticlestat treatment was associated with a decrease in motor seizure frequency from baseline in patients with CDD and a decrease in all seizure frequency in both patient groups. Soticlestat treatment was associated with an increase in motor seizure frequency in patients with Dup15q syndrome.

Precision medicine for genetic epilepsy on the horizon: Recent advances, present challenges, and suggestions for continued progress.

The genetic basis of many epilepsies is increasingly understood, giving rise to the possibility of precision treatments tailored to specific genetic etiologies. Despite this, current medical therapy for most epilepsies remains imprecise, aimed primarily at empirical seizure reduction rather than targeting specific disease processes. Intellectual and technological leaps in diagnosis over the past 10 years have not yet translated to routine changes in clinical practice. However, the epilepsy community is poised to make impressive gains in precision therapy, with continued innovation in gene discovery, diagnostic ability, and bioinformatics; increased access to genetic testing and counseling; fuller understanding of natural histories; agility and rigor in preclinical research, including strategic use of emerging model systems; and engagement of an evolving group of stakeholders (including patient advocates, governmental resources, and clinicians and scientists in academia and industry). In each of these areas, we highlight notable examples of recent progress, new or persistent challenges, and future directions. The future of precision medicine for genetic epilepsy looks bright if key opportunities on the horizon can be pursued with strategic and coordinated effort.

Negative impact of insomnia and daytime sleepiness on quality of life in individuals with the cyclin-dependent kinase-like 5 deficiency disorder.

Cyclin-dependent kinase-like 5 (CDKL5) gene pathogenic variants result in CDKL5 deficiency disorder (CDD). Early onset intractable epilepsy and severe developmental delays are prominent symptoms of CDD. Comorbid sleep disturbances are a major concerning symptom for families. We aimed to explore the relationship between insomnia, daytime sleepiness, sleep medications and quality of life in children with CDD. Caregivers of 129 children with CDD in the International CDKL5 Disorder Database completed the Quality-of-Life Inventory-Disability (QI-Disability) questionnaire and "Disorders of Maintaining Sleep" (DIMS) and the "Disorders of Excessive Somnolence" (DOES) items of the Sleep Disturbance Scale for Children. Adjusting for covariates, a unit increase in DOES score was associated with reduced quality of life total (coefficient -3.06, 95% confidence interval [CI] 1.35-7.80), physical health (coefficient -7.20, 95% CI -10.64, -3.76) and negative emotions (coefficient -3.90, 95% CI -7.38, -0.42) scores. Adjusting for covariates, a unit increase in DIMS score was associated with reduced negative emotions (coefficient -6.02, 95% CI -10.18, -2.86). Use of sleep medications had small influences on the effect sizes. This study highlights the importance of sleep problems as a determinant of quality of life in children with CDD, consistent with effects observed for other groups of children with intellectual disability. Excessive daytime sleepiness was particularly associated with detrimental effects on quality of life. Further research in optimal behavioural and pharmaceutical management of sleep problems for this population is required.

Whole-exome sequencing and adrenocorticotropic hormone therapy in individuals with infantile spasms.

AIM: To identify additional genes associated with infantile spasms using a cohort with defined infantile spasms. METHOD: Whole-exome sequencing (WES) was performed on 21 consented individuals with infantile spasms and their unaffected parents (a trio-based study). Clinical history and imaging were reviewed. Potentially deleterious exonic variants were identified and segregated. To refine potential candidates, variants were further prioritized on the basis of evidence for relevance to disease phenotype or known associations with infantile spasms, epilepsy, or neurological disease. RESULTS: Likely pathogenic de novo variants were identified in NR2F1, GNB1, NEUROD2, GABRA2, and NDUFAF5. Suggestive dominant and recessive candidate variants were identified in PEMT, DYNC1I1, ASXL1, RALGAPB, and STRADA; further confirmation is required to support their relevance to disease etiology. INTERPRETATION: This study supports the utility of WES in uncovering the genetic etiology in undiagnosed individuals with infantile spasms with an overall yield of five out of 21. High-priority candidates were identified in an additional five individuals. WES provides additional support for previously described disease-associated genes and expands their already broad mutational and phenotypic spectrum.

Exploring genotype-phenotype relationships in the CDKL5 deficiency disorder using an international dataset.

Characterized by early-onset seizures, global developmental delay and severe motor deficits, CDKL5 deficiency disorder is caused by pathogenic variants in the cyclin-dependent kinase-like 5 gene. Previous efforts to investigate genotype-phenotype relationships have been limited due to small numbers of recurrent mutations and small cohort sizes. Using data from the International CDKL5 Disorder Database we examined genotype-phenotype relationships for 13 recurrent CDKL5 variants and the previously analyzed historic variant groupings. We have applied the CDKL5 Developmental Score (CDS) and an adapted version of the CDKL5 Clinical Severity Assessment (CCSA), to grade the severity of phenotype and developmental outcomes for 285 individuals with CDKL5 variants. Comparisons of adapted CCSA and CDS between recurrent variants and variant groups were performed using multiple linear regression adjusting for age and sex. Individuals with the missense variant, p.Arg178Trp, had the highest mean adapted CCSA and lowest mean developmental scores. Other variants producing severe phenotypes included p.Arg559* and p.Arg178Gln. Variants producing milder phenotypes included p.Arg134*, p.Arg550*, and p.Glu55Argfs*20. There are observed differences in phenotype severity and developmental outcomes for individuals with different CDKL5 variants. However, the historic variant groupings did not seem to reflect differences in phenotype severity or developmental outcomes as clearly as analyzed by individual variants.

Randomized Clinical Trial of First-Line Genome Sequencing in Pediatric White Matter Disorders.

OBJECTIVE: Genome sequencing (GS) is promising for unsolved leukodystrophies, but its efficacy has not been prospectively studied. METHODS: A prospective time-delayed crossover design trial of GS to assess the efficacy of GS as a first-line diagnostic tool for genetic white matter disorders took place between December 1, 2015 and September 27, 2017. Patients were randomized to receive GS immediately with concurrent standard of care (SoC) testing, or to receive SoC testing for 4 months followed by GS. RESULTS: Thirty-four individuals were assessed at interim review. The genetic origin of 2 patient's leukoencephalopathy was resolved before randomization. Nine patients were stratified to the immediate intervention group and 23 patients to the delayed-GS arm. The efficacy of GS was significant relative to SoC in the immediate (5/9 [56%] vs 0/9 [0%]; Wild-Seber, p < 0.005) and delayed (control) arms (14/23 [61%] vs 5/23 [22%]; Wild-Seber, p < 0.005). The time to diagnosis was significantly shorter in the immediate-GS group (log-rank test, p = 0.04). The overall diagnostic efficacy of combined GS and SoC approaches was 26 of 34 (76.5%, 95% confidence interval = 58.8-89.3%) in <4 months, greater than historical norms of <50% over 5 years. Owing to loss of clinical equipoise, the trial design was altered to a single-arm observational study. INTERPRETATION: In this study, first-line GS provided earlier and greater diagnostic efficacy in white matter disorders. We provide an evidence-based diagnostic testing algorithm to enable appropriate clinical GS utilization in this population. ANN NEUROL 2020;88:264-273.

PLPHP deficiency: clinical, genetic, biochemical, and mechanistic insights.

Biallelic pathogenic variants in PLPBP (formerly called PROSC) have recently been shown to cause a novel form of vitamin B6-dependent epilepsy, the pathophysiological basis of which is poorly understood. When left untreated, the disease can progress to status epilepticus and death in infancy. Here we present 12 previously undescribed patients and six novel pathogenic variants in PLPBP. Suspected clinical diagnoses prior to identification of PLPBP variants included mitochondrial encephalopathy (two patients), folinic acid-responsive epilepsy (one patient) and a movement disorder compatible with AADC deficiency (one patient). The encoded protein, PLPHP is believed to be crucial for B6 homeostasis. We modelled the pathogenicity of the variants and developed a clinical severity scoring system. The most severe phenotypes were associated with variants leading to loss of function of PLPBP or significantly affecting protein stability/PLP-binding. To explore the pathophysiology of this disease further, we developed the first zebrafish model of PLPHP deficiency using CRISPR/Cas9. Our model recapitulates the disease, with plpbp-/- larvae showing behavioural, biochemical, and electrophysiological signs of seizure activity by 10 days post-fertilization and early death by 16 days post-fertilization. Treatment with pyridoxine significantly improved the epileptic phenotype and extended lifespan in plpbp-/- animals. Larvae had disruptions in amino acid metabolism as well as GABA and catecholamine biosynthesis, indicating impairment of PLP-dependent enzymatic activities. Using mass spectrometry, we observed significant B6 vitamer level changes in plpbp-/- zebrafish, patient fibroblasts and PLPHP-deficient HEK293 cells. Additional studies in human cells and yeast provide the first empirical evidence that PLPHP is localized in mitochondria and may play a role in mitochondrial metabolism. These models provide new insights into disease mechanisms and can serve as a platform for drug discovery.

Factors in the disease severity of ATP1A3 mutations: Impairment, misfolding, and allele competition.

Dominant mutations of ATP1A3, a neuronal Na,K-ATPase α subunit isoform, cause neurological disorders with an exceptionally wide range of severity. Several new mutations and their phenotypes are reported here (p.Asp366His, p.Asp742Tyr, p.Asp743His, p.Leu924Pro, and a VUS, p.Arg463Cys). Mutations associated with mild or severe phenotypes [rapid-onset dystonia-parkinsonism (RDP), alternating hemiplegia of childhood (AHC), or early infantile epileptic encephalopathy (EIEE)] were expressed in HEK-293 cells. Paradoxically, the severity of human symptoms did not correlate with whether there was enough residual activity to support cell survival. We hypothesized that distinct cellular consequences may result not only from pump inactivation but also from protein misfolding. Biosynthesis was investigated in four tetracycline-inducible isogenic cell lines representing different human phenotypes. Two cell biological complications were found. First, there was impaired trafficking of αß complex to Golgi apparatus and plasma membrane, as well as changes in cell morphology, for two mutations that produced microcephaly or regions of brain atrophy in patients. Second, there was competition between exogenous mutant ATP1A3 (α3) and endogenous ATP1A1 (α1) so that their sum was constant. This predicts that in patients, the ratio of normal to mutant ATP1A3 proteins will vary when misfolding occurs. At the two extremes, the results suggest that a heterozygous mutation that only impairs Na,K-ATPase activity will produce relatively mild disease, while one that activates the unfolded protein response could produce severe disease and may result in death of neurons independently of ion pump inactivation.

CDKL5 deficiency disorder: Relationship between genotype, epilepsy, cortical visual impairment, and development.

OBJECTIVE: The cyclin-dependent kinase like 5 (CDKL5) gene is a known cause of early onset developmental and epileptic encephalopathy, also known as CDKL5 deficiency disorder (CDD). We sought to (1) provide a description of seizure types in patients with CDD, (2) provide an assessment of the frequency of seizure-free periods and cortical visual impairment (CVI), (3) correlate these features with genotype and gender, and (4) correlate these features with developmental milestones. METHODS: This is a cohort study of patients with CDD. Phenotypic features were explored and correlated with gene variant grouping and gender. A developmental score was created based on achieving seven primary milestones. Phenotypic variables were correlated with the developmental score to explore markers of better developmental outcomes. Multivariate linear regression was used to account for age at last visit. RESULTS: Ninety-two patients with CDD were seen during the enrollment period. Eighteen were male (19%); median age at last visit was 5 years (interquartile range = 2.0-11.0). Eighty-one percent of patients developed epileptic spasms, but only 47% of those also had hypsarrhythmia. Previously described hypermotor-tonic-spasms sequence was seen in only 24% of patients, but 56% of patients had seizures with multiple phases (often tonic and spasms). Forty-three percent of patients experienced a seizure-free period ranging from 1 to >12 months, but only 6% were still seizure-free at the last visit. CVI was present in 75% of all CDD patients. None of these features was associated with genotype group or gender. CVI was correlated with reduced milestone achievement after adjusting for age at last visit and a history of hypsarrhythmia. SIGNIFICANCE: The most common seizure types in CDD are epileptic spasms (often without hypsarrhythmia) and tonic seizures that may cluster together. CVI is a common feature in CDD and is correlated with achieving fewer milestones.

The impact of hypsarrhythmia on infantile spasms treatment response: Observational cohort study from the National Infantile Spasms Consortium.

OBJECTIVE: The multicenter National Infantile Spasms Consortium prospective cohort was used to compare outcomes and phenotypic features of patients with infantile spasms with and without hypsarrhythmia. METHODS: Patients aged 2 months to 2 years were enrolled prospectively with new-onset infantile spasms. Treatment choice and categorization of hypsarrhythmia were determined clinically at each site. Response to therapy was defined as resolution of clinical spasms (and hypsarrhythmia if present) without relapse 3 months after initiation. RESULTS: Eighty-two percent of patients had hypsarrhythmia, but this was not associated with gender, mean age, preexisting developmental delay or epilepsy, etiology, or response to first-line therapy. Infants with hypsarrhythmia were more likely to receive standard treatment (adrenocorticotropic hormone, prednisolone, or vigabatrin [odds ratio (OR) 2.6, 95% confidence interval (CI) 1.4-4.7] and preexisting epilepsy reduced the likelihood of standard treatment (OR 3.2, 95% CI 1.9-5.4). Hypsarrhythmia was not a determinant of response to treatment. A logistic regression model demonstrated that later age of onset (OR 1.09 per month, 95% CI 1.03-1.15) and absence of preexisting epilepsy (OR 1.7, 95% CI 1.06-2.81) had a small impact on the likelihood of responding to the first-line treatment. However, receiving standard first-line treatment increased the likelihood of responding dramatically: vigabatrin (OR 5.2 ,95% CI 2-13.7), prednisolone (OR 8, 95% CI 3.1-20.6), and adrenocorticotropic hormone (ACTH; OR 10.2, 95% CI 4.1-25.8) . SIGNIFICANCE: First-line treatment with standard therapy was by far the most important variable in determining likelihood of response to treatment of infantile spasms with or without hypsarrhythmia.

Factors influencing the attainment of major motor milestones in CDKL5 deficiency disorder.

This study investigated the influence of factors at birth and in infancy on the likelihood of achieving major motor milestones in CDKL5 Deficiency Disorder (CDD). Data on 350 individuals with a pathogenic CDKL5 variant was sourced from the International CDKL5 Disorder Database. A first model included factors available at birth (e.g., sex, variant group and mosaicism) and the second additionally included factors available during infancy (e.g., age at seizure onset, number of anti-seizure medications used, experience of a honeymoon period and formal therapy). Cox regression was used to model the time to achieve the milestones. The probability of attaining the outcomes at specific ages was estimated by evaluating the time-to-event function at specific covariate values. Independent sitting and walking were achieved by 177/350 and 57/325 children respectively. By seven years of age, 67.1% of females but only 37.3% of males could sit independently. About a quarter each of females and males achieved independent walking by eight and six years, respectively. When observed from birth, female gender, a late truncating variant and mosaicism impacted most positively on the likelihood of independent sitting. When observed from one year, later seizure onset and experiencing a honeymoon period also improved the likelihood of independent sitting. Factors that favoured sitting (except gender) also improved walking. Having a truncating variant between aa178 and aa781 reduced the likelihood of achieving independent sitting and walking. It is possible to utilise factors occurring early in life to inform the likelihood of future motor development in CDD.

Validating the Communication and Symbolic Behavior Scales-Developmental Profile Infant-Toddler Checklist (CSBS-DP ITC) Beyond Infancy in the CDKL5 Deficiency Disorder.

CDKL5 deficiency disorder (CDD) results in early-onset epilepsy and lifelong cognitive and motor impairments. With no validated measure for communication in CDD, this study evaluated the psychometric properties of the Communication and Symbolic Behavior Scales-Developmental Profile Infant Toddler Checklist (CSBS-DP ITC). Caregivers (n = 150; affected individuals aged 1-29 years) completed the CSBS-DP ITC. Distribution of scores indicated a floor effect. There was poor divergent validity for the three-factor model but goodness of fit and convergent validity data were satisfactory for the one-factor model. Individuals with poorer overall functional abilities scored lower on the CSBS-DP ITC. Test-retest reliability was excellent. The floor effect could explain the very high reliability, suggesting problems as a sensitive outcome measure in clinical trials for CDD.