Clinical and molecular characterization of myotonia congenita using whole-exome sequencing in Egyptian patients.

BACKGROUND: Myotonia Congenita (MC) is a rare disease classified into two major forms; Thomsen and Becker disease caused by mutations in the CLCN1 gene, which affects muscle excitability and encodes voltage-gated chloride channels (CLC-1). While, there are no data regarding the clinical and molecular characterization of myotonia in Egyptian patients. METHODS: Herein, we report seven Egyptian MC patients from six unrelated families. Following the clinical diagnosis, whole-exome sequencing (WES) was performed for genetic diagnosis. Various in silico prediction tools were utilized to interpret variant pathogenicity. The candidate variants were then validated using Sanger sequencing technique. RESULTS: In total, seven cases were recruited. The ages at the examination were ranged from eight months to nineteen years. Clinical manifestations included warm-up phenomenon, hand grip, and percussion myotonia. Electromyography was performed in all patients and revealed myotonic discharges. Molecular genetic analysis revealed five different variants. Of them, we identified two novel variants in the CLCN1 gene ( c.1583G > C; p.Gly528Ala and c.2203_2216del;p.Thr735ValfsTer57) and three known variants in the CLCN1 and SCN4A gene. According to in silico tools, the identified novel variants were predicted to have deleterious effects. CONCLUSIONS: As the first study to apply WES among Egyptian MC patients, our findings reported two novel heterozygous variants that expand the CLCN1 mutational spectrum for MC diagnosis. These results further confirm that genetic testing is essential for early diagnosis of MC, which affects follow-up treatment and prognostic assessment in clinical practice.

Next generation sequencing panel as an effective approach to genetic testing in patients with a highly variable phenotype of neuromuscular disorders.

Neuromuscular disorders (NMDs) include a wide range of diseases affecting the peripheral nervous system. The genetic diagnoses are increasingly obtained with using the next generation sequencing (NGS). We applied the custom-design targeted NGS panel including 89 genes, together with genotyping and multiplex ligation-dependent probe amplification (MLPA) to identify a genetic spectrum of NMDs in 52 Polish patients. As a result, the genetic diagnosis was determined by NGS panel in 29 patients so its diagnostic utility is estimated at 55.8%. The most pathogenic variants were found in CLCN1, followed by CAPN3, SCN4A, and SGCA genes. Genotyping of myotonic dystrophy type 1 and 2 (DM1 and DM2) as a secondary approach has been performed. The co-occurrence of CAPN3 and CNBP mutations in one patient as well as DYSF and CNBP mutations in another suggests possibly more complex inheritance as well as expression of a phenotype. In 7 individuals with single nucleotide variant found in NGS testing, the MLPA of the CAPN3 gene was performed detecting the deletion encompassing exons 2-8 in the CAPN3 gene in one patient, confirming recessive limb-girdle muscular dystrophy type 1 (LGMDR1). Thirty patients obtained a genetic diagnosis (57.7%) after using NGS testing, genotyping and MLPA analysis. The study allowed for the identification of 27 known and 4 novel pathogenic/likely pathogenic variants and variants of uncertain significance (VUS) associated with NMDs.In conclusion, the diagnostic approach with diverse molecular techniques enables to broaden the mutational spectrum and maximizes the diagnostic yield. Furthermore, the co-occurrence of DM2 and LGMD has been detected in 2 individuals.

Toxic to the touch: The makings of lethal mantles in pitohui birds and poison dart frogs.

How do chemically defended animals resist their own toxins? This intriguing question on the concept of autotoxicity is at the heart of how species interactions evolve. In this issue of Molecular Ecology (Molecular Ecology, 2024, 33), Bodawatta and colleagues report on how Papua New Guinean birds coopted deadly neurotoxins to create lethal mantles that protect against predators and parasites. Combining chemical screening of the plumage of a diverse collection of passerine birds with genome sequencing, the researchers unlocked a deeper understanding of how some birds sequester deadly batrachotoxin (BTX) from their food without poisoning themselves. They identified that birds impervious to BTX bear amino acid substitutions in the toxin-binding site of the voltage-gated sodium channel Nav1.4, whose function is essential for proper contraction and relaxation of vertebrate muscles. Comparative genetic and molecular docking analyses show that several of the substitutions associated with insensitivity to BTX may have become prevalent among toxic birds through positive selection. Intriguingly, poison dart frogs that also co-opted BTX in their lethal mantles were found to harbour similar toxin insensitivity substitutions in their Nav1.4 channels. Taken together, this sets up a powerful model system for studying the mechanisms behind convergent molecular evolution and how it may drive biological diversity.

Genetic analysis of 37 cases with primary periodic paralysis in Chinese patients.

BACKGROUND: Primary periodic paralysis (PPP) is an inherited disorders of ion channel dysfunction characterized by recurrent episodes of flaccid muscle weakness, which can classified as hypokalemic (HypoPP), normokalemic (NormoPP), or hyperkalemic (HyperPP) according to the potassium level during the paralytic attacks. However, PPP is charactered by remarkable clinical and genetic heterogeneity, and the diagnosis of suspected patients is based on the characteristic clinical presentation then confirmed by genetic testing. At present, there are only limited cohort studies on PPP in the Chinese population. RESULTS: We included 37 patients with a clinical diagnosis of PPP. Eleven (29.7%) patients were tested using a specific gene panel and 26 (70.3%) by the whole-exome sequencing (WES). Twenty-two cases had a genetic variant identified, representing a diagnostic rate of 59.5% (22/37). All the identified mutations were either in the SCN4A or the CACNA1S gene. The overall detection rate was comparable between the panel (54.5%: 6/11) and WES (61.5%: 16/26). The remaining patients unresolved through panel sequencing were further analyzed by WES, without the detection of any mutation. The novel atypical splicing variant c.2020-5G > A affects the normal splicing of the SCN4A mRNA, which was confirmed by minigene splicing assay. Among 21 patients with HypoPP, 15 patients were classified as HypoPP-2 with SCN4A variants, and 6 HypoPP-1 patients had CACNA1S variants. CONCLUSIONS: Our results suggest that SCN4A alleles are the main cause in our cohort, with the remainder caused by CACNA1S alleles, which are the predominant cause in Europe and the United States. Additionally, this study identified 3 novel SCN4A and 2 novel CACNA1S variants, broadening the mutation spectrum of genes associated with PPP.

[Prenatal diagnosis of a case with Congenital myasthenic syndrome due to compound heterozygous variants of SCN4A gene].

OBJECTIVE: To explore the clinical and genetic characteristics of a fetus diagnosed with Congenital myasthenic syndrome type 16 (CMS16). METHODS: A couple who had visited Tianjin Medical University General Hospital in February 2018 due to "adverse outcome of two pregnancies" was selected as the study subject. Clinical data was gathered. Peripheral blood and amniotic fluid samples were collected and subjected to whole exome sequencing (WES). Candidate variant was verified by Sanger sequencing. Low-depth whole-genome sequencing was carried out to detect copy number variation (CNV) in the fetus. RESULTS: The couple's first pregnancy had resulted in a miscarriage at 27+5 weeks, when ultrasound had revealed pleural effusion and polyhydramnios in the fetus. Their second pregnancy was terminated at 30+5 weeks due to fetal hand malformations, polyhydramnios and pleural fluid. Both couple had denied family history of genetic conditions. For their third pregnancy, no CNV abnormality was detected, whilst a compound heterozygous variants, including a maternally derived c.3172C>T (p.R1058W) and paternal c.1431delG (p.K477fs*89) in the SCN4A gene were detected. Based on the guidelines from the American College of Medical Genetics and Genomics, the c.3172C>T (p.R1058W) was predicted as a likely pathogenic variant (PM1+PM2_supporting+PP3+PP4), whilst the c.1431delG (p.K477fs*89) was predicted as a pathogenic variant (PVS1+PM2_supporting+PP4). CONCLUSION: The c.3172C>T (p.R1058W) and c.1431delG (p.K477fs*89) compound heterozygous variants of the SCN4A gene probably underlay the CMS16 in the third fetus.

Idiopathic generalized epilepsy in a family with SCN4A-related myotonia.

OBJECTIVES: Myotonia is a clinical sign typical of a group of skeletal muscle channelopathies, the non-dystrophic myotonias. These disorders are electrophysiologically characterized by altered membrane excitability, due to specific genetic variants in known causative genes (CLCN1 and SCN4A). Juvenile Myoclonic Epilepsy (JME) is an epileptic syndrome identified as idiopathic generalized epilepsy, its genetics is complex and still unclarified. The co-occurrence of these two phenotypes is rare and the causes likely have a genetic background. In this study, we have genetically investigated an Italian family in which co-segregates myotonia, JME, or abnormal EEG without seizures was observed. METHODS: All six individuals of the family, 4 affected and 2 unaffected, were clinically evaluated; EMG and EEG examinations were performed. For genetic testing, Exome Sequencing was performed for the six family members and Sanger sequencing was used to confirm the candidate variant. RESULTS: Four family members, the mother and three siblings, were affected by myotonia. Moreover, EEG recordings revealed interictal generalized sharp-wave discharges in all affected individuals, and two siblings were affected by JME. All four affected members share the same identified variant, c.644 T > C, p.Ile215Thr, in SCN4A gene. Variants that could account for the epileptic phenotype alone, separately from the myotonic one, were not identified. SIGNIFICANCE: These results provide supporting evidence that both myotonic and epileptic phenotypes could share a common genetic background, due to variants in SCN4A gene. SCN4A pathogenic variants, already known to be causative of myotonia, likely increase the susceptibility to epilepsy in our family. PLAIN LANGUAGE SUMMARY: This study analyzed all members of an Italian family, in which the mother and three siblings had myotonia and epilepsy. Genetic analysis allowed to identify a variant in the SCN4A gene, which appears to be the cause of both clinical signs in this family.

A c.1775C > T Point Mutation of Sodium Channel Alfa Subunit Gene (SCN4A) in a Three-Generation Sardinian Family with Sodium Channel Myotonia.

Background: The nondystrophic myotonias are rare muscle hyperexcitability disorders caused by gain-of-function mutations in the SCN4A gene or loss-of-function mutations in the CLCN1 gene. Clinically, they are characterized by myotonia, defined as delayed muscle relaxation after voluntary contraction, which leads to symptoms of muscle stiffness, pain, fatigue, and weakness. Diagnosis is based on history and examination findings, the presence of electrical myotonia on electromyography, and genetic confirmation. Methods: Next-generation sequencing including the CLCN1 and SCN4A genes was performed in patients with clinical neuromuscular disorders. Electromyography, Short Exercise Test, in vivo and in vitro electrophysiology, site-directed mutagenesis and heterologous expression were collected. Results: A heterozygous point mutation (c.1775C > T, p.Thr592Ile) of muscle voltage-gated sodium channel α subunit gene (SCN4A) has been identified in five female patients over three generations, in a family with non-dystrophic myotonia. The muscle stiffness and myotonia involve mainly the face and hands, but also affect walking and running, appearing early after birth and presenting a clear cold sensitivity. Very hot temperatures, menstruation and pregnancy also exacerbate the symptoms; muscle pain and a warm-up phenomenon are variable features. Neither paralytic attacks nor post-exercise weakness has been reported. Muscle hypertrophy with cramp-like pain and increased stiffness developed during pregnancy. The symptoms were controlled with both mexiletine and acetazolamide. The Short Exercise Test after muscle cooling revealed two different patterns, with moderate absolute changes of compound muscle action potential amplitude. Conclusions: The p.Thr592Ile mutation in the SCN4A gene identified in this Sardinian family was responsible of clinical phenotype of myotonia.

Sodium Channel Gene Variants in Fetuses with Abnormal Sonographic Findings: Expanding the Prenatal Phenotypic Spectrum of Sodium Channelopathies.

Voltage-gated sodium channels (VGSCs) are responsible for the initiation and propagation of action potentials in the brain and muscle. Pathogenic variants in genes encoding VGSCs have been associated with severe disorders including epileptic encephalopathies and congenital myopathies. In this study, we identified pathogenic variants in genes encoding the α subunit of VGSCs in the fetuses of two unrelated families with the use of trio-based whole exome sequencing, as part of a larger cohort study. Sanger sequencing was performed for variant confirmation as well as parental phasing. The fetus of the first family carried a known de novo heterozygous missense variant in the SCN2A gene (NM_001040143.2:c.751G>A p.(Val251Ile)) and presented intrauterine growth retardation, hand clenching and ventriculomegaly. Neonatally, the proband also exhibited refractory epilepsy, spasms and MRI abnormalities. The fetus of the second family was a compound heterozygote for two parentally inherited novel missense variants in the SCN4A gene (NM_000334.4:c.4340T>C, p.(Phe1447Ser), NM_000334.4:c.3798G>C, p.(Glu1266Asp)) and presented a severe prenatal phenotype including talipes, fetal hypokinesia, hypoplastic lungs, polyhydramnios, ear abnormalities and others. Both probands died soon after birth. In a subsequent pregnancy of the latter family, the fetus was also a compound heterozygote for the same parentally inherited variants. This pregnancy was terminated due to multiple ultrasound abnormalities similar to the first pregnancy. Our results suggest a potentially crucial role of the VGSC gene family in fetal development and early lethality.

Muscle MRI in periodic paralysis shows myopathy is common and correlates with intramuscular fat accumulation.

INTRODUCTION/AIMS: The periodic paralyses are muscle channelopathies: hypokalemic periodic paralysis (CACNA1S and SCN4A variants), hyperkalemic periodic paralysis (SCN4A variants), and Andersen-Tawil syndrome (KCNJ2). Both episodic weakness and disabling fixed weakness can occur. Little literature exists on magnetic resonance imaging (MRI) in muscle channelopathies. We undertake muscle MRI across all subsets of periodic paralysis and correlate with clinical features. METHODS: A total of 45 participants and eight healthy controls were enrolled and underwent T1-weighted and short-tau-inversion-recovery (STIR) MRI imaging of leg muscles. Muscles were scored using the modified Mercuri Scale. RESULTS: A total of 17 patients had CACNA1S variants, 16 SCN4A, and 12 KCNJ2. Thirty-one (69%) had weakness, and 9 (20%) required a gait-aid/wheelchair. A total of 78% of patients had intramuscular fat accumulation on MRI. Patients with SCN4A variants were most severely affected. In SCN4A, the anterior thigh and posterior calf were more affected, in contrast to the posterior thigh and posterior calf in KCNJ2. We identified a pattern of peri-tendinous STIR hyperintensity in nine patients. There were moderate correlations between Mercuri, STIR scores, and age. Intramuscular fat accumulation was seen in seven patients with no fixed weakness. DISCUSSION: We demonstrate a significant burden of disease in patients with periodic paralyses. MRI intramuscular fat accumulation may be helpful in detecting early muscle involvement, particularly in those without fixed weakness. Longitudinal studies are needed to assess the role of muscle MRI in quantifying disease progression over time and as a potential biomarker in clinical trials.

Optical measurement of gating pore currents in hypokalemic periodic paralysis model cells.

Hypokalemic periodic paralysis (HypoPP) is a rare genetic disease associated with mutations in CACNA1S or SCN4A encoding the voltage-gated Ca2+ channel Cav1.1 or the voltage-gated Na+ channel Nav1.4, respectively. Most HypoPP-associated missense changes occur at the arginine residues within the voltage-sensing domain (VSD) of these channels. It is established that such mutations destroy the hydrophobic seal that separates external fluid and the internal cytosolic crevices, resulting in the generation of aberrant leak currents called gating pore currents. Presently, the gating pore currents are thought to underlie HypoPP. Here, based on HEK293T cells and by using the Sleeping Beauty transposon system, we generated HypoPP-model cell lines that co-express the mouse inward-rectifier K+ channel (mKir2.1) and HypoPP2-associated Nav1.4 channel. Whole-cell patch-clamp measurements confirmed that mKir2.1 successfully hyperpolarizes the membrane potential to levels comparable to those of myofibers, and that some Nav1.4 variants induce notable proton-based gating pore currents. Importantly, we succeeded in fluorometrically measuring the gating pore currents in these variants by using a ratiometric pH indicator. Our optical method provides a potential in vitro platform for high-throughput drug screening, not only for HypoPP but also for other channelopathies caused by VSD mutations.

Sinus arrest in familial hypokalemic periodic paralysis caused by SCN4A mutation: a case report.

BACKGROUND: Primary hypokalemic periodic paralysis (HypoPP), a rare skeletal muscle channelopathy resulting in episodic muscle weakness or paralysis under hypokalemic conditions, is caused by autosomal-dominant genetic mutations. HypoPP limits physical activity, and cardiac arrhythmias during paralytic attacks have been reported. We describe a rare familial HypoPP case complicated by sinus arrest and syncope requiring urgent temporary pacemaker implantation. CASE REPORT: A 27-year-old Vietnamese man with a family history of periodic paralysis presented with his third attack of muscle weakness triggered by intense football training the previous day. Clinical and laboratory features justified a HypoPP diagnosis. During intravenous potassium replacement, the patient experienced syncopal sinus arrest requiring urgent temporary pacemaker implantation. The patient gradually improved, responding favorably to oral potassium supplements. Genetic testing revealed an Arg1132Gln mutation in the sodium ion channel (SCN4A, chromosome 17: 63947091). At discharge, the patient received expert consultation regarding nonpharmacological preventive strategies, including avoidance of vigorous exercise and carbohydrate-rich diet. CONCLUSIONS: No evidence has established a relationship between hypokalemia and sinus arrest, and no specific treatment exists for familial HypoPP due to SCN4A mutation. Clinician awareness of this rare condition will promote appropriate diagnostic approaches and management strategies for acute paralytic attacks. Treatment should be tailored according to HypoPP phenotypes and genotypes.

Clinical and Pathologic Features of Congenital Myasthenic Syndromes Caused by 35 Genes-A Comprehensive Review.

Congenital myasthenic syndromes (CMS) are a heterogeneous group of disorders characterized by impaired neuromuscular signal transmission due to germline pathogenic variants in genes expressed at the neuromuscular junction (NMJ). A total of 35 genes have been reported in CMS (AGRN, ALG14, ALG2, CHAT, CHD8, CHRNA1, CHRNB1, CHRND, CHRNE, CHRNG, COL13A1, COLQ, DOK7, DPAGT1, GFPT1, GMPPB, LAMA5, LAMB2, LRP4, MUSK, MYO9A, PLEC, PREPL, PURA, RAPSN, RPH3A, SCN4A, SLC18A3, SLC25A1, SLC5A7, SNAP25, SYT2, TOR1AIP1, UNC13A, VAMP1). The 35 genes can be classified into 14 groups according to the pathomechanical, clinical, and therapeutic features of CMS patients. Measurement of compound muscle action potentials elicited by repetitive nerve stimulation is required to diagnose CMS. Clinical and electrophysiological features are not sufficient to identify a defective molecule, and genetic studies are always required for accurate diagnosis. From a pharmacological point of view, cholinesterase inhibitors are effective in most groups of CMS, but are contraindicated in some groups of CMS. Similarly, ephedrine, salbutamol (albuterol), amifampridine are effective in most but not all groups of CMS. This review extensively covers pathomechanical and clinical features of CMS by citing 442 relevant articles.

Muscle fat replacement and contractility in patients with skeletal muscle sodium channel disorders.

Skeletal muscle sodium channel disorders give rise to episodic symptoms such as myotonia and/or periodic paralysis. Chronic symptoms with permanent weakness are not considered characteristic of the phenotypes. Muscle fat replacement represents irreversible damage that inevitably will impact on muscle strength. This study investigates muscle fat replacement and contractility in patients with pathogenic SCN4A variants compared to healthy controls. T1-weighted and 2-point Dixon MRI of the legs were conducted to assess fat replacement. Stationary dynamometry was used to assess muscle strength. Contractility was determined by maximal muscle contraction divided by cross-sectional muscle area. The average cross-sectional intramuscular fat fraction was greater in patients compared with controls by 2.5% in the calves (95% CI 0.74-4.29%, p = 0.007) and by 2.0% in the thighs (95% CI 0.75-3.2%, p = 0.003). Muscle contractility was less in patients vs. controls by 14-27% (p < 0.05). Despite greater fat fraction and less contractility, absolute strength was not significantly less. This study quantitatively documents greater fat fraction and additionally describes difference in muscle contractility in a large cohort of patients with skeletal muscle sodium channel disorders. The clinical impact of these abnormal findings is likely limited as muscle hypertrophy in the patients served to preserve absolute muscle strength. Subgroup analysis indicated significant difference in phenotype by genotype, however these findings lack statistical significance and serve as inspiration for future researchers to probe into the geno- phenotype relationship in these disorders.Trial registration: The study was registered at http://clinicaltrials.gov (identifier: NCT04808388).

Prevalence of genetically confirmed skeletal muscle channelopathies in the era of next generation sequencing.

We provide an up-to-date and accurate minimum point prevalence of genetically defined skeletal muscle channelopathies which is important for understanding the population impact, planning for treatment needs and future clinical trials. Skeletal muscle channelopathies include myotonia congenita (MC), sodium channel myotonia (SCM), paramyotonia congenita (PMC), hyperkalemic periodic paralysis (hyperPP), hypokalemic periodic paralysis (hypoPP) and Andersen- Tawil Syndrome (ATS). Patients referred to the UK national referral centre for skeletal muscle channelopathies and living in UK were included to calculate the minimum point prevalence using the latest data from the Office for National Statistics population estimate. We calculated a minimum point prevalence of all skeletal muscle channelopathies of 1.99/100 000 (95% CI 1.981-1.999). The minimum point prevalence of MC due to CLCN1 variants is 1.13/100 000 (95% CI 1.123-1.137), SCN4A variants which encode for PMC and SCM is 0.35/100 000 (95% CI 0.346 - 0.354) and for periodic paralysis (HyperPP and HypoPP) 0.41/100 000 (95% CI 0.406-0.414). The minimum point prevalence for ATS is 0.1/100 000 (95% CI 0.098-0.102). There has been an overall increase in point prevalence in skeletal muscle channelopathies compared to previous reports, with the biggest increase found to be in MC. This can be attributed to next generation sequencing and advances in clinical, electrophysiological and genetic characterisation of skeletal muscle channelopathies.

Blockers of Skeletal Muscle Nav1.4 Channels: From Therapy of Myotonic Syndrome to Molecular Determinants of Pharmacological Action and Back.

The voltage-gated sodium channels represent an important target for drug discovery since a large number of physiological processes are regulated by these channels. In several excitability disorders, including epilepsy, cardiac arrhythmias, chronic pain, and non-dystrophic myotonia, blockers of voltage-gated sodium channels are clinically used. Myotonia is a skeletal muscle condition characterized by the over-excitability of the sarcolemma, resulting in delayed relaxation after contraction and muscle stiffness. The therapeutic management of this disorder relies on mexiletine and other sodium channel blockers, which are not selective for the Nav1.4 skeletal muscle sodium channel isoform. Hence, the importance of deepening the knowledge of molecular requirements for developing more potent and use-dependent drugs acting on Nav1.4. Here, we review the available treatment options for non-dystrophic myotonia and the structure-activity relationship studies performed in our laboratory with a focus on new compounds with potential antimyotonic activity.

Hyperkalemic periodic paralysis associated with a novel missense variant located in the inner pore of Nav1.4.

BACKGROUND: Hyperkalemic periodic paralysis (HyperPP) is an autosomal dominantly inherited disease characterized by episodic paralytic attacks with hyperkalemia, and is caused by mutations of the SCN4A gene encoding the skeletal muscle type voltage-gated sodium channel Nav1.4. The pathological mechanism of HyperPP was suggested to be associated with gain-of-function changes for Nav1.4 gating, some of which are defects of slow inactivation. CASE PRESENTATION & METHODS: We identified a HyperPP family consisting of the proband and his mother, who showed a novel heterozygous SCN4A variant, p.V792G, in an inner pore lesion of segment 6 in Domain II of Nav1.4. Clinical and neurophysiological evaluations were conducted for the proband and his mother. We explored the pathogenesis of the variant by whole-cell patch clamp technique using HEK293T cells expressing the mutant Nav1.4 channel. RESULTS: Functional analysis of Nav1.4 with the V792G mutation revealed a hyperpolarized shift of voltage-dependent activation and fast inactivation. Moreover, steady-state slow inactivation in V792G was impaired with larger residual currents in comparison with wild-type Nav1.4. CONCLUSION: V792G in SCN4A is a pathogenic variant associated with the HyperPP phenotype and the inner pore lesion of Nav1.4 plays a crucial role in slow inactivation.

Coexistence of SCN4A and CLCN1 mutations in a family with atypical myotonic features: A clinical and functional study.

Non-dystrophic myotonias include several entities with possible clinical overlap, i.e. myotonia congenita caused by CLCN1 gene mutations, as well as paramyotonia congenita and sodium channel myotonia caused by SCN4A gene mutations. Herein, we describe the clinical features of five relatives affected by clinical and neurophysiological myotonia, with an aspecific and mixed phenotype. Next-generation sequencing identified the novel p.K1302R variant in SCN4A and the p.H838P variant in CLCN1. Segregation of the two mutations with the disease was confirmed by genotyping affected and non-affected family members. Patch-clamp experiments showed that sodium currents generated by p.K1302R and WT hNav1.4 were very similar. Mutant channel showed a small negative shift (5 mV) in the voltage-dependence of activation, which increased the likelihood of the channel to open at more negative voltages. The p.H838P mutation caused a reduction in chloride current density and a small voltage-dependence shift towards less negative potentials, in agreement with its position into the CBS2 domain of the C-terminus. Our results demonstrated that the mild functional alterations induced by p.K1302R and p.H838P in combination may be responsible for the mixed myotonic phenotypes. The K1302R mutant was sensitive to mexiletine and lamotrigine, suggesting that both drugs might be useful for the K1302R carriers.

Acute myotonic reaction during succinylcholine anaesthesia.

A 21-year-old woman developed an acute myotonic reaction while undergoing anaesthesia using succinylcholine. Examination later showed she had shoulder, neck and calf hypertrophy, bilateral symmetrical ptosis and eyelid, handgrip and percussion myotonia. Peripheral neurophysiology studies identified significant, continuous myotonic discharges in both upper and lower limbs. Genetic analysis identified a c.3917G>A (p.Gly1306Glu) mutation in the SCN4A gene, confirming a diagnosis of sodium channel myotonia. Succinylcholine and other depolarising agents can precipitate life-threatening acute myotonic reactions when given to patients with myotonia. Patients with neuromuscular disorders are at an increased risk of perioperative anaesthetic complications. We report a woman who developed an acute myotonic reaction whilst undergoing anaesthesia, in the context of an unrecognised myotonic disorder. We then discuss an approach to the diagnosis of myotonic disorders.

An ion-channel-gene-based prediction model for head and neck squamous cell carcinoma: Prognostic assessment and treatment guidance.

Purpose: Head and neck squamous cell carcinoma (HNSCC) is a very diverse malignancy with a poor prognosis. The purpose of this study was to develop a new signature based on 12 ion channel genes to predict the outcome and immune status of HNSCC patients. Methods: Clinicopathological information and gene sequencing data of HNSCC patients were generated from the Cancer Genome Atlas and Gene Expression Omnibus databases. A set of 323 ion channel genes was obtained from the HUGO Gene Nomenclature Committee database and literature review. Using univariate Cox regression analysis, the ion channel genes related to HNSCC prognosis were identified. A prognostic signature and nomogram were then created using machine learning methods. Kaplan-Meier analysis was used to explore the relevance of the risk scores and overall survival (OS). We also investigated the association between risk scores, tumor immune infiltration, and gene mutational status. Finally, we detected the expression levels of the signature genes by quantitative real-time polymerase chain reaction, western blotting, and immunohistochemistry. Results: We separated the patients into high- and low-risk groups according to the risk scores computed based on these 12 ion channel genes, and the OS of the low-risk group was significantly longer (p<0.001). The area under the curve for predicting 3-year survival was 0.729. Univariate and multivariate analyses showed that the 12-ion-channel-gene risk model was an independent prognostic factor. We also developed a nomogram model based on risk scores and clinicopathological variables to forecast outcomes. Furthermore, immune cell infiltration, gene mutation status, immunotherapy response, and chemotherapeutic treatment sensitivity were all linked to risk scores. Moreover, high expression levels of ANO1, AQP9, and BEST2 were detected in HNSCC tissues, whereas AQP5, SCNN1G, and SCN4A expression was low in HNSCC tissues, as determined by experiments. Conclusion: The 12-ion-channel-gene prognostic signatures have been demonstrated to be highly efficient in predicting the prognosis, immune microenvironment, gene mutation status, immunotherapy response, and chemotherapeutic sensitivity of HNSCC patients.