Monogenic Causes in Familial Stroke Across Intracerebral Hemorrhage and Ischemic Stroke Subtypes Identified by Whole-Exome Sequencing.

Whole exome sequencing (WES) has been used to detect rare causative variants in neurological diseases. However, the efficacy of WES in genetic diagnosis of clinically heterogeneous familial stroke remains inconclusive. We prospectively searched for disease-causing variants in unrelated probands with defined familial stroke by candidate gene/hotspot screening and/or WES, depending on stroke subtypes and neuroimaging features at a referral center. The clinical significance of each variant was determined according to the American College of Medical Genetics guidelines. Among 161 probands (mean age at onset 53.2 ± 13.7 years; male 63.4%), 33 participants (20.5%) had been identified with 19 pathogenic/likely pathogenic variants (PVs; WES applied 152/161 = 94.4%). Across subtypes, the highest hit rate (HR) was intracerebral hemorrhage (ICH, 7/18 = 38.9%), particularly with the etiological subtype of structural vasculopathy (4/4 = 100%, PVs in ENG, KRIT1, PKD1, RNF213); followed by ischemic small vessel disease (SVD, 15/48 = 31.3%; PVs in NOTCH3, HTRA1, HBB). In contrast, large artery atherosclerosis (LAA, 4/44 = 9.1%) and cardioembolism (0/11 = 0%) had the lowest HR. NOTCH3 was the most common causative gene (16/161 = 9.9%), presenting with multiple subtypes of SVD (n = 13), ICH (n = 2), or LAA (n = 1). Importantly, we disclosed two previously unreported PVs, KRIT1 p.E379* in a familial cerebral cavernous malformation, and F2 p.F382L in a familial cerebral venous sinus thrombosis. The contribution of monogenic etiologies was particularly high in familial ICH and SVD subtypes in our Taiwanese cohort. Utilizing subtype-guided hotspot screening and/or subsequent WES, we unraveled monogenic causes in 20.5% familial stroke probands, including 1.2% novel PVs. Genetic diagnosis may enable early diagnosis, management and lifestyle modification. Among 161 familial stroke probands, 33 (20.5%) had been identified pathogenic or likely pathogenic monogenic variants related to stroke. The positive hit rate among all subtypes was high in intracerebral hemorrhage (ICH) and ischemic small vessel disease (SVD). Notably, two previously unreported variants, KRIT1 p.E379* in a familial cerebral cavernous malformation and F2 p.F382L in familial cerebral venous sinus thrombosis, were disclosed. CVT cerebral venous thrombosis; HTN Hypertensive subtype; LAA large artery atherosclerosis; SV structural vasculopathy; U Undetermined.

Plasma Matrix Metalloproeteinase-9 Is Associated with Seizure and Angioarchitecture Changes in Brain Arteriovenous Malformations.

Both angiogenesis and inflammation contribute to activation of matrix metalloproeteinase-9 (MMP-9), which dissolves the extracellular matrix, disrupts the blood-brain barrier, and plays an important role in the pathogenesis of brain arteriovenous malformations (BAVMs). The key common cytokine in both angiogenesis and inflammation is interleukin 6 (IL-6). Previous studies have shown elevated systemic MMP-9 and decreased systemic vascular endothelial growth factor (VEGF) in BAVM patients. However, the clinical utility of plasma cytokines is unclear. The purpose of this study is to explore the relationship between plasma cytokines and the clinical presentations of BAVMs. Prospectively, we recruited naive BAVM patients without hemorrhage as the experimental group and unruptured intracranial aneurysm (UIA) patients as the control group. All patients received digital subtraction angiography, and plasma cytokines were collected from the lesional common carotid artery. Plasma cytokine levels were determined using a commercially available, monoclonal antibody-based enzyme-linked immunosorbent assay. Subgroup analysis based on hemorrhagic presentation and angiograchitecture was done for the BAVM group. Pearson correlations were calculated for the covariates. Means and differences for continuous and categorical variables were compared using Student's t and χ2 tests respectively. Plasma MMP-9 levels were significantly higher in the BAVM group (42,945 ± 29,991 pg/mL) than in the UIA group (28,270 ± 17,119 pg/mL) (p < 0.001). Plasma MMP-9 levels in epileptic BAVMs (57,065 ± 35,732; n = 9) were higher than in non-epileptic BAVMs (35,032 ± 28,301; n = 19) (p = 0.049). Lower plasma MMP-9 levels were found in cases of BAVM with angiogenesis and with peudophlebitis. Plasma MMP-9 is a good biomarker reflecting ongoing vascular remodeling in BAVMs. Angiogenesis and pseudophlebitis are two angioarchitectural signs that reflect MMP-9 activities and can potentially serve as imaging biomarkers for epileptic BAVMs.

Clinical characteristics and long-term outcome of cerebral cavernous malformations-related epilepsy.

OBJECTIVE: Cerebral cavernous malformations (CCMs) present variably, and epileptic seizures are the most common symptom. The factors contributing to cavernoma-related epilepsy (CRE) and drug resistance remain inconclusive. The outcomes of CRE after different treatment modalities have not yet been fully addressed. This study aimed to characterize the clinical features of patients with CRE and the long-term seizure outcomes of medical and surgical treatment strategies. METHODS: This was a retrospective cohort of 135 patients with CCM who were diagnosed in 2007-2011 and followed up for 93.6 months on average. The patients were divided into drug-resistant epilepsy (DRE; n = 29), non-DRE (n = 45), and no epilepsy (NE; n = 61). RESULTS: Temporal CCM was the factor most strongly associated with the development of both CRE and DRE. The majority of patients with single temporal CCMs had CRE (86.8%, n = 33), and 50% had DRE, whereas only 14.7% (n = 5) with a nontemporal supratentorial CCM had DRE (p < .05). The most common lesion site in the DRE group was the mesiotemporal lobe (50%). Multiple CCMs were more frequently observed in the CRE (29.2%) than the NE (11.5%) group (p < .05). In patients with CRE, multiple lesions were associated with a higher rebleeding rate (odds ratio = 11.1), particularly in those with DRE (odds ratio = 15.4). The majority of patients who underwent resective surgery for DRE (76.5%, n = 13) achieved International League Against Epilepsy Class I and II seizure outcomes even after a long disease course. SIGNIFICANCE: Temporal CCM not only predisposes to CRE but also is a major risk factor for drug resistance. The mesiotemporal lobe is the most epileptogenic zone. Multiple CCMs are another risk factor for CRE and increase the rebleeding risk in these patients. Surgical resection could provide beneficial long-term seizure outcomes in patients with DRE.

Acute withdrawal of new-generation antiepileptic drugs in epilepsy monitoring units: Safety and efficacy.

INTRODUCTION: Acute withdrawal of antiepileptic drugs (AEDs) is a safe and effective approach to provoking seizures in order to complete video-electroencephalogram (V-EEG) studies in a timely manner. Previous studies have focused only on withdrawal from conventional AEDs, and the effects of withdrawal from new-generation AEDs have not been extensively studied. MATERIALS AND METHODS: This study examined adult patients with drug-resistant epilepsy admitted to an epilepsy monitoring unit between 2015 and 2018. Patients were classified according to whether they received conventional AEDs (Con; n = 13) or new-generation AEDs (N-Gen; n = 26). We then compared the effects of withdrawing these two types of AEDs over a period of one week in terms of efficacy (time to complete V-EEG monitoring) and safety, including the incidence of cluster seizures (CS), focal to bilateral tonic-clonic seizures (FBTCS) and status epilepticus (SE). RESULTS: In both groups, approximately one week was required to complete V-EEG analysis: N-Gen group (5.6 days) and Con group (6.3 days). No differences were observed between the two groups in terms of the median number of seizures, the onset of the 1st seizure, the distribution of CS, FBTCS, or SE. Following acute withdrawal of medication, a high percentage of patients with a history of CS or FBTCS, respectively, presented CS or FBTCS. CONCLUSIONS: We did not observe significant differences between patients taking new-generation AEDs and those taking conventional AEDs following withdrawal during V-EEG recording. In the current study, we employed a standard protocol for the rapid withdrawal of AEDs (daily dose reduction of 50%), which was sufficient for 80% of patients to complete V-EEG monitoring within one week.

Cellular secretion and cytotoxicity of transthyretin mutant proteins underlie late-onset amyloidosis and neurodegeneration.

Transthyretin amyloidosis (ATTR) is a progressive life-threatening disease characterized by the deposition of transthyretin (TTR) amyloid fibrils. Several pathogenic variants have been shown to destabilize TTR tetramers, leading to aggregation of misfolded TTR fibrils. However, factors that underlie the differential age of disease onset amongst amyloidogenic TTR variants remain elusive. Here, we examined the biological properties of various TTR mutations and found that the cellular secretory pattern of the wild-type (WT) TTR was similar to those of the late-onset mutant (Ala97Ser, p. Ala117Ser), stable mutant (Thr119Met, p. Thr139Met), early-onset mutant (Val30Met, p. Val50Met), but not in the unstable mutant (Asp18Gly, p. Asp38Gly). Cytotoxicity assays revealed their toxicities in the order of Val30Met > Ala97Ser > WT > Thr119Met in neuroblastoma cells. Surprisingly, while early-onset amyloidogenic TTR monomers (M-TTRs) are retained by the endoplasmic reticulum quality control (ERQC), late-onset amyloidogenic M-TTRs can be secreted extracellularly. Treatment of thapsigargin (Tg) to activate the unfolded protein response (UPR) alleviates Ala97Ser M-TTR secretion. Interestingly, Ala97Ser TTR overexpression in Drosophila causes late-onset fast neurodegeneration and a relatively short lifespan, recapitulating human disease progression. Our study demonstrates that the escape of TTR monomers from the ERQC may underlie late-onset amyloidogenesis in patients and suggests that targeting ERQC could mitigate late-onset ATTR.

Gamma Knife radiosurgery for cerebral cavernous malformation.

This is a retrospective study examining the efficacy and safety of Gamma Knife radiosurgery (GKS) in treating patients with cerebral cavernous malformations (CCMs). Between 1993 and 2018, 261 patients with 331 symptomatic CCMs were treated by GKS. The median age was 39.9 years and females were predominant (54%). The median volume of CCMs was 3.1 mL. The median margin dose was 11.9 Gy treat to a median isodose level of 59%. Median clinical and imaging follow-up times were 69 and 61 months, respectively. After the initial hemorrhage that led to CCM diagnosis, 136 hemorrhages occurred in the period prior to GKS (annual incidence = 23.6%). After GKS, 15 symptomatic hemorrhages occurred within the first 2 years of follow-up (annual incidence = 3.22%), and 37 symptomatic hemorrhages occurred after the first 2 years of follow-up (annual incidence = 3.16%). Symptomatic radiation-induced complication was encountered in 8 patients (3.1%). Mortality related to GKS occurred in 1 patient (0.4%). In conclusion, GKS decreased the risk of hemorrhage in CCM patients presenting with symptomatic hemorrhage. GKS is a viable alternative treatment option for patients with surgically-inaccessible CCMs or significant medical comorbidities.

Biophysical characterization and modulation of Transthyretin Ala97Ser.

OBJECTIVE: Ala97Ser (A97S) is the major transthyretin (TTR) mutation in Taiwanese patients of familial amyloid polyneuropathy (FAP), characterized by a late-onset but rapidly deteriorated neuropathy. Tafamidis can restore the stability of some mutant TTR tetramers and slow down the progression of TTR-FAP. However, there is little understanding of the biophysical features of A97S-TTR mutant and the pharmacological modulation effect of tafamidis on it. This study aims to delineate the biophysical characteristics of A97S-TTR and the pharmacological modulation effect of tafamidis on this mutant. METHOD: The stability of TTR tetramers was assessed by urea denaturation and differential scanning calorimetry. Isothermal titration calorimetry (ITC) was used to measure the binding constant of tafamidis to TTR. Nuclear magnetic resonance spectroscopy (NMR) titration experiment was used to map out the tafamidis binding site. RESULTS: Chemical and thermal denaturation confirmed the destabilization effect of A97S. Consistent with other the amyloidogenic mutant, A97S-TTR has slightly lower conformational stability. NMR revealed the binding site of A97S-TTR with tafamidis is at the thyroxine binding pocket. The ITC experiments documented the high affinity of the binding which can effectively stabilize the A97S-TTR tetramer. INTERPRETATION: This study confirmed the structural modulation effect of tafamidis on A97S-TTR and implied the potential therapeutic benefit of tafamidis for A97S TTR-FAP. This approach can be applied to investigate the modulation effect of tafamidis on other rare TTR variants and help to make individualized choices of available treatments for FAP patients.

PDXK mutations cause polyneuropathy responsive to pyridoxal 5'-phosphate supplementation.

OBJECTIVE: To identify disease-causing variants in autosomal recessive axonal polyneuropathy with optic atrophy and provide targeted replacement therapy. METHODS: We performed genome-wide sequencing, homozygosity mapping, and segregation analysis for novel disease-causing gene discovery. We used circular dichroism to show secondary structure changes and isothermal titration calorimetry to investigate the impact of variants on adenosine triphosphate (ATP) binding. Pathogenicity was further supported by enzymatic assays and mass spectroscopy on recombinant protein, patient-derived fibroblasts, plasma, and erythrocytes. Response to supplementation was measured with clinical validated rating scales, electrophysiology, and biochemical quantification. RESULTS: We identified biallelic mutations in PDXK in 5 individuals from 2 unrelated families with primary axonal polyneuropathy and optic atrophy. The natural history of this disorder suggests that untreated, affected individuals become wheelchair-bound and blind. We identified conformational rearrangement in the mutant enzyme around the ATP-binding pocket. Low PDXK ATP binding resulted in decreased erythrocyte PDXK activity and low pyridoxal 5'-phosphate (PLP) concentrations. We rescued the clinical and biochemical profile with PLP supplementation in 1 family, improvement in power, pain, and fatigue contributing to patients regaining their ability to walk independently during the first year of PLP normalization. INTERPRETATION: We show that mutations in PDXK cause autosomal recessive axonal peripheral polyneuropathy leading to disease via reduced PDXK enzymatic activity and low PLP. We show that the biochemical profile can be rescued with PLP supplementation associated with clinical improvement. As B6 is a cofactor in diverse essential biological pathways, our findings may have direct implications for neuropathies of unknown etiology characterized by reduced PLP levels. ANN NEUROL 2019;86:225-240.

Clinical and genetic profiles of hereditary transthyretin amyloidosis in Taiwan.

OBJECTIVE: The clinical and genetic profiles of hereditary transthyretin amyloidosis (ATTR) in Chinese populations remain elusive. We aim to characterize the features of ATTR in a Taiwanese cohort of Han Chinese descent. METHODS: Seventy-nine patients with molecularly confirmed ATTR from 57 Taiwanese families were identified by sequencing the transthyretin gene (TTR). The clinical and electrophysiological data were scrutinized. Cardiac involvement of ATTR was evaluated by echocardiography and cardiac scintigraphy. Four microsatellite and seven single-nucleotide polymorphism markers flanking TTR were genotyped to investigate the founder effect of the TTR Ala97Ser mutation. RESULTS: Most of the patients had a peripheral neuropathy with variable autonomic symptoms. The average age at disease onset (AO) was 58.2 ± 7.2 years, and the male patients had an earlier AO than female patients (56.6 ± 5.7 years vs. 61.8 ± 8.9 years, P = 0.013). Electrophysiological studies revealed a generalized axonal sensorimotor polyneuropathy and isolated median neuropathy in 84.5% and 15.5% of the patients, respectively. Up to 80% of the patients with ATTR had symptomatic or subclinical cardiac involvement. Six TTR mutations were identified in the participants including one novel mutation Glu89Asp. Among them, Ala97Ser was the most common mutation, accounting for 91.2% of the ATTR pedigrees. Detailed haplotype analyses demonstrated a shared haplotype in the 47 patients with the Ala97Ser mutation, suggesting a founder effect. INTERPRETATION: The present study delineates the distinct features of ATTR in Taiwan and provides useful information for the diagnosis and management of ATTR, especially in patients of Chinese descent.

Contributions of Animal Models to the Mechanisms and Therapies of Transthyretin Amyloidosis.

Transthyretin amyloidosis (ATTR amyloidosis) is a fatal systemic disease caused by amyloid deposits of misfolded transthyretin, leading to familial amyloid polyneuropathy and/or cardiomyopathy, or a rare oculoleptomeningeal amyloidosis. A good model system that mimic the disease phenotype is crucial for the development of drugs and treatments for this devastating degenerative disorder. The present models using fruit flies, worms, rodents, non-human primates and induced pluripotent stem cells have helped researchers understand important disease-related mechanisms and test potential therapeutic options. However, the challenge of creating an ideal model still looms, for these models did not recapitulates all symptoms, particularly neurological presentation, of ATTR amyloidosis. Recently, knock-in techniques was used to generate two humanized ATTR mouse models, leading to amyloid deposition in the nerves and neuropathic manifestation in these models. This review gives a recent update on the milestone, progress, and challenges in developing different models for ATTR amyloidosis research.

Genetic and clinical characteristics of NEFL-related Charcot-Marie-Tooth disease.

OBJECTIVES: To analyse and describe the clinical and genetic spectrum of Charcot-Marie-Tooth disease (CMT) caused by mutations in the neurofilament light polypeptide gene (NEFL). METHODS: Combined analysis of newly identified patients with NEFL-related CMT and all previously reported cases from the literature. RESULTS: Five new unrelated patients with CMT carrying the NEFL mutations P8R and N98S and the novel variant L311P were identified. Combined data from these cases and 62 kindreds from the literature revealed four common mutations (P8R, P22S, N98S and E396K) and three mutational hotspots accounting for 37 (55%) and 50 (75%) kindreds, respectively. Eight patients had de novo mutations. Loss of large-myelinated fibres was a uniform feature in a total of 21 sural nerve biopsies and 'onion bulb' formations and/or thin myelin sheaths were observed in 14 (67%) of them. The neurophysiological phenotype was broad but most patients with E90K and N98S had upper limb motor conduction velocities <38 m/s. Age of onset was ≤3 years in 25 cases. Pyramidal tract signs were described in 13 patients and 7 patients were initially diagnosed with or tested for inherited ataxia. Patients with E90K and N98S frequently presented before age 3 years and developed hearing loss or other neurological features including ataxia and/or cerebellar atrophy on brain MRI. CONCLUSIONS: NEFL-related CMT is clinically and genetically heterogeneous. Based on this study, however, we propose mutational hotspots and relevant clinical-genetic associations that may be helpful in the evaluation of NEFL sequence variants and the differential diagnosis with other forms of CMT.

Acute simultaneous multiple lacunar infarcts as the initial presentation of cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is an adult-onset, dominantly inherited small-vessel disease of the brain caused by NOTCH3 mutations and characterized by recurrent subcortical infarctions, dementia, migraine with aura, and mood disturbance. We report a patient with unusual presentation of CADASIL with acute simultaneous multiple subcortical lacunar infarcts as the first manifestation. A 69-year-old man developed confusion, drowsiness, right hemiparesis, and slurred speech following orthopedic surgeries. Brain magnetic resonance imaging revealed diffuse leukoencephalopathy and multiple acute subcortical lacunar infarcts. Brain magnetic resonance angiography, echocardiography and 24-hour electrocardiography were unremarkable. The symptoms improved quickly after treatment with fluid hydration and antiplatelet agent, and his consciousness and mentality totally recovered within 3 days. The NOTCH3 genetic testing showed a heterozygous missense mutation, c.1630C>T (p. Arg544Cys). The experience in this case suggests that brain imaging is important in managing postoperative confusion, and any patient with diffuse leukoencephalopathy of unknown etiology may need to be tested for NOTCH3 mutations. Surgery is an important factor of encephalopathy and acute infarction in individuals with NOTCH3 mutations. Comprehensive presurgical evaluations and proactive perioperative precautions to avoid dehydration and anemia are necessary for patients with CADASIL who are about to receive anesthesia and surgery.

Transthyretin Ala97Ser in Chinese-Taiwanese patients with familial amyloid polyneuropathy: genetic studies and phenotype expression.

BACKGROUND: Transthyretin (TTR) variants of familial amyloid neuropathies (FAP) form a heterogenous group of autosomal dominantly inherited diseases. TTR gene analysis in several nationalities (Japanese, Portuguese, French, and British) has shown many distinguishing characteristics in the genotype-phenotype correlation. In Chinese, there are only a few reports of private TTR gene mutations belonging to single kindred. MATERIALS AND METHODS: We collected five patients with autosomal dominant inheritant sensorimotor polyneuropathy and tissue-proved amyloid deposition. The diagnosis of FAP was established on the mutation of the TTR gene detected by direct sequencing. Haplotype analysis was conducted in four of these patients. RESULTS AND CONCLUSIONS: These five FAP patients shared an identical missense mutation, Ala97Ser, in the TTR gene. This mutation presented with a constellation of late-onset polyneuropathy, preceding carpal tunnel syndrome, and outstanding autonomic dysfunction. Heart was the most frequently involved vital organ. Haplotype analysis hinted independent origins although the numbers were limited. Our study is the first case series gathering from the Chinese-Taiwanese population. We proposed a possible hot-spot mutation of the TTR gene, Ala97Ser, in this ethnic.