E200K Mutation Research Articles

Structurally targeted mutagenesis identifies key residues supporting -synuclein misfolding in multiple system atrophy.

Multiple system atrophy (MSA) and Parkinson's disease (PD) are caused by misfolded -synuclein spreading throughout the central nervous system. While familial PD is linked to several point mutations in -synuclein, there are no known mutations associated with MSA. Our previous work investigating differences in -synuclein misfolding between the two disorders showed that the familial PD mutation E46K inhibits replication of MSA prions both in vitro and in vivo, providing key evidence to support the hypothesis that -synuclein adopts unique strains in patients. Here, to further interrogate -synuclein misfolding, we engineered a panel of cell lines harboring both PD-linked and novel mutations designed to identify key residues that facilitate -synuclein misfolding in MSA. These data were paired with in silico analyses using Maestro software to predict the effect of each mutation on the ability of -synuclein to misfold into one of the reported MSA cryo-electron microscopy conformations. In many cases, our modeling accurately identified mutations that facilitated or inhibited MSA replication. However, Maestro was occasionally unable to predict the effect of a mutation on MSA propagation in vitro, demonstrating the challenge of using computational tools to investigate intrinsically disordered proteins. Finally, we used our cellular models to determine the mechanism underlying the E46K-driven inhibition of MSA replication, finding that the E46/K80 salt bridge is necessary to support -synuclein misfolding. Overall, our studies use a structure-based approach to investigate -synuclein misfolding, resulting in the creation of a powerful panel of cell lines that can be used to interrogate MSA strain biology.

Anti-prion drugs do not improve survival in novel knock-in models of inherited prion disease.

Prion diseases uniquely manifest in three distinct forms: inherited, sporadic, and infectious. Wild-type prions are responsible for the sporadic and infectious versions, while mutant prions cause inherited variants like fatal familial insomnia (FFI) and familial Creutzfeldt-Jakob disease (fCJD). Although some drugs can prolong prion incubation times up to four-fold in rodent models of infectious prion diseases, no effective treatments for FFI and fCJD have been found. In this study, we evaluated the efficacy of various anti-prion drugs on newly-developed knock-in mouse models for FFI and fCJD. These models express bank vole prion protein (PrP) with the pathogenic D178N and E200K mutations. We applied various drug regimens known to be highly effective against wild-type prions in vivo as well as a brain-penetrant compound that inhibits mutant PrPSc propagation in vitro. None of the regimens tested (Anle138b, IND24, Anle138b + IND24, cellulose ether, and PSCMA) significantly extended disease-free survival or prevented mutant PrPSc accumulation in either knock-in mouse model, despite their ability to induce strain adaptation of mutant prions. Our results show that anti-prion drugs originally developed to treat infectious prion diseases do not necessarily work for inherited prion diseases, and that the recombinant sPMCA is not a reliable platform for identifying compounds that target mutant prions. This work underscores the need to develop therapies and validate screening assays specifically for mutant prions, as well as anti-prion strategies that are not strain-dependent.

Creutzfeldt–Jakob Disease Associated with E200K Mutation and SARS-CoV-2 Infection: Pure Coincidence or Neurodegenerative Acceleration?

Several recent studies reported on some patients developing Creutzfeldt–Jakob disease (CJD) following coronavirus disease 2019, but, to the best of our knowledge, this case is the first reported in Italy on an onset of a CJD genetic form (gCJD) immediately after COVID-19 infection. We present a 51-year-old woman with a positive family history for CJD, who, two months after a mild SARS-CoV-2 infection, presented a rapidly progressing dementia diagnosed as CJD through clinical features, imaging, electroencephalography, and cerebrospinal fluid analysis. Genetic testing revealed the E200K mutation (p.Glu200Lys) c.598G>A, with homozygosity for methionine (MET) at codon 129, thus confirming the diagnosis of Creutzfeldt–Jakob disease. She passed away two months later. Interestingly, our case confirms that homozygous E200K gCJD patients are characterized by a relatively younger age of onset; moreover, it also sheds light on the neurodegeneration underlying both prion diseases and COVID-19 infection. In our opinion, the rising global prevalence of neurodegenerative complications following COVID-19 disease adds urgency to the study of this potential relationship, mostly in elderly patients who may experience worse long-lasting outcomes systemically and within the nervous system.

Immunocytokines with target cell-restricted IL-15 activity for treatment of B cell malignancies.

Despite the advances in cancer treatment achieved, for example, by the CD20 antibody rituximab, an urgent medical need remains to optimize the capacity of such antibodies to induce antibody-dependent cellular cytotoxicity (ADCC) that determines therapeutic efficacy. The cytokine IL-15 stimulates proliferation, activation, and cytolytic capacity of NK cells, but broad clinical use is prevented by short half-life, poor accumulation at the tumor site, and severe toxicity due to unspecific immune activation. We here report modified immunocytokines consisting of Fc-optimized CD19 and CD20 antibodies fused to an IL-15 moiety comprising an L45E-E46K double mutation (MIC+ format). The E46K mutation abrogated binding to IL-15Rα, thereby enabling substitution of physiological trans-presentation by target binding and thus conditional IL-15Rβγ stimulation, whereas the L45E mutation optimized IL-15Rβγ agonism and producibility. In vitro analysis of NK activation, anti-leukemia reactivity, and toxicity using autologous and allogeneic B cells confirmed target-dependent function of MIC+ constructs. Compared with Fc-optimized CD19 and CD20 antibodies, MIC+ constructs mediated superior target cell killing and NK cell proliferation. Mouse models using luciferase-expressing human NALM-6 lymphoma cells, patient acute lymphoblastic leukemia (ALL) cells, and murine EL-4 lymphoma cells transduced with human CD19/CD20 as targets and human and murine NK cells as effectors, respectively, confirmed superior and target-dependent anti-leukemic activity. In summary, MIC+ constructs combine the benefits of Fc-optimized antibodies and IL-15 cytokine activity and mediate superior NK cell immunity with potentially reduced side effects. They thus constitute a promising new immunotherapeutic approach shown here for B cell malignancies.

Early-Onset Parkinson Mutation Remodels Monomer-Fibril Interactions to Allosterically Amplify Synuclein's Amyloid Cascade.

Alpha synuclein (αS) aggregates are the main component of Lewy bodies (LBs) associated with Parkinson's disease (PD). A longstanding question about αS and PD pertains to the autosomal dominant E46K αS mutant, which leads to the early onset of PD and LB dementias. The E46K mutation not only promotes αS aggregation but also stabilizes αS monomers in "closed" conformers, which are compact and aggregation-incompetent. Hence, the mechanism of action of the E46K mutation is currently unclear. Here, we show that αS monomers harboring the E46K mutation exhibit more extensive interactions with fibrils compared to those of WT. Such monomer-fibril interactions are sufficient to allosterically drive transitions of αS monomers from closed to open conformations, enabling αS aggregation. We also show that E46K promotes head-to-tail monomer-monomer interactions in early self-association events. This multipronged mechanism provides a new framework to explain how the E46K mutation and possibly other αS variants trigger early-onset PD.

Elevated E200K Somatic Mutation of the Prion Protein Gene (PRNP) in the Brain Tissues of Patients with Sporadic Creutzfeldt-Jakob Disease (CJD).

Sporadic Creutzfeldt-Jakob disease (CJD) is a major human prion disease worldwide. CJD is a fatal neurodegenerative disease caused by an abnormal prion protein (PrPSc). To date, the exact etiology of sporadic CJD has not been fully elucidated. We investigated the E200K and V203I somatic mutations of the prion protein gene (PRNP) in sporadic CJD patients and matched healthy controls using pyrosequencing. In addition, we estimated the impact of somatic mutations on the human prion protein (PrP) using PolyPhen-2, PANTHER and PROVEAN. Furthermore, we evaluated the 3D structure and electrostatic potential of the human PrP according to somatic mutations using DeepView. The rates of PRNP K200 somatic mutation were significantly increased in the frontal cortex and hippocampus of sporadic CJD patients compared to the matched controls. In addition, the electrostatic potential of the human PrP was significantly changed by the K200 somatic mutation of the PRNP gene. To the best of our knowledge, this is the first report on an association of the PRNP K200 somatic mutation with sporadic CJD.

E46K Mutation of α-Synuclein Preorganizes the Intramolecular Interactions Crucial for Aggregation.

Aggregation of α-synuclein is central to the pathogenesis of Parkinson's disease. The most toxic familial mutation E46K accelerates the aggregation process by an unknown mechanism. Herein, we provide a clue by investigating the influence of E46K on monomeric α-synuclein and its relation to aggregation with molecular dynamics simulations. The E46K mutation suppresses β-sheet structures in the N-terminus while promoting those at the key fibrillization region named NACore. Even though WT and E46K monomers share conserved intramolecular interactions with fibrils, E46K abolishes intramolecular contacts within the N-terminus which are present in the WT monomer but absent in fibrils. Network analysis identifies residues 36-53 as the interaction core of the WT monomer. Upon mutation, residues 36-46 are expelled to water due to aggravated electrostatic repulsion in the 43KTKK46 segment. Instead, NACore (residues 68-78) becomes the interaction hub and connects preceding residues 47-56 and the C-terminus. Consequently, residues 47-95 which belong to the fibril core form more compact β-sheets. Overall, the interaction network of E46K is more like fibrils than WT, stabilizing the fibril-like conformations. Our work provides mechanistic insights into the faster aggregation of the E46K mutant. It implies a close link between monomeric conformations and fibrils, which would spur the development of therapeutic strategies.

Thalamic‐insomnia phenotype in genetic E200K Creutzfeldt‐Jakob disease: A PET/MRI study

AbstractBackgroundInsomnia and thalamic involvement are hallmarks of fatal familial insomnia; however, it was occasionally reported that genetic E200K Creutzfeldt‐Jakob disease (CJD) patients had also manifested thalamic‐insomnia phenotype. The study aimed to explore the clinical and neuroimage characteristics of genetic E200K CJD by comprehensive neuroimage analysis.MethodsSix genetic CJD patients carried Glu to Lys change at codon 200 (E200K) mutations on prion protein gene (PRNP), thirteen sporadic CJD (sCJD), and twenty‐two age‐ and sex‐matched normal controls were enrolled in the study. All participants completed hybrid positron emission tomography/ magnetic resonance imaging (PET/MRI) examinations. The high signal was visually inspected on diffusion weighted imaging (DWI) sequence. 18F‐fluorodeoxyglucose positron emission tomography (18F‐FDG‐PET) and structural images were quantitively analyzed by statistically parameter mapping (SPM). Clinical and image characteristics are compared between genetic E200K CJD patients and sporadic CJD patients.ResultsThere was no group difference in age (49.0±7.1 versus 55.0±4.9 versus 52.7±7.3, p = 0.193) and gender (3/3 versus 4/9 versus 11/11, p = 0.513) between genetic E200K CJD, sCJD, and normal controls. Insomnia was the main complaint in the genetic E200K CJD patients’ group (4/2 versus 1/12, p = 0.007). Hyperintensity, hypometabolism but not predominant atrophy of the thalamus was found by visual inspection in genetic E200K CJD patients. A decreased trend was found on gray matter volume (uncorrected p<0.0001) and metabolism (FWE corrected) of the thalamus in genetic E200K patients.ConclusionThe clinical and image characteristics of genetic E200K CJD mimic fatal familial insomnia, manifested as a thalamic‐insomnia phenotype, indicating some specific underlying pathological changes might exist in E200K mutations. PET measurement is a sensitive approach to help identify the functional changes of the thalamus in prion disease.

Bilateral tonic-clonic seizure and focal cortical hyperexcitability in familial Creutzfeldt-Jakob disease with E200K mutation of the prion protein.

Convulsive epileptic seizures are rare in Creutzfeldt-Jakob disease (CJD) and their clinical and EEG features have not been reported in detail. We describe a case of familial CJD with an E200K mutation of the prion protein who presented with bilateral tonic-clonic seizures (BTCS) during long-term video-EEG monitoring. Semiologically, BTCS showed focal clinical signs such as head turning and eye deviation to the left. The ictal EEG started with generalized polyspikes. Interictal EEG showed generalized periodic discharges with right fronto-temporal predominance (larger amplitude and earlier onset compared with other regions). MRI showed high-intensity signals persistently in the right temporo-parietal region on diffusion-weighted images (DWI). Interictal single-photon emission computed tomography (SPECT) showed hyperperfusion in the same region. Brain pathology revealed typical spongiform changes in CJD without other pathological findings of rapidly progressive dementia. Our case demonstrates that CJD can cause BTCS with generalized EEG changes and focal semiological/imaging abnormalities, suggesting that diffuse cortical and subcortical epileptic networks may develop in familial CJD.

Distinct Effects of Familial Parkinson's Disease-Associated Mutations on α-Synuclein Phase Separation and Amyloid Aggregation.

The Lewy bodies and Lewy neurites are key pathological hallmarks of Parkinson's disease (PD). Single-point mutations associated with familial PD cause α-synuclein (α-Syn) aggregation, leading to the formation of Lewy bodies and Lewy neurites. Recent studies suggest α-Syn nucleates through liquid-liquid phase separation (LLPS) to form amyloid aggregates in a condensate pathway. How PD-associated mutations affect α-Syn LLPS and its correlation with amyloid aggregation remains incompletely understood. Here, we examined the effects of five mutations identified in PD, A30P, E46K, H50Q, A53T, and A53E, on the phase separation of α-Syn. All other α-Syn mutants behave LLPS similarly to wild-type (WT) α-Syn, except that the E46K mutation substantially promotes the formation of α-Syn condensates. The mutant α-Syn droplets fuse to WT α-Syn droplets and recruit α-Syn monomers into their droplets. Our studies showed that α-Syn A30P, E46K, H50Q, and A53T mutations accelerated the formation of amyloid aggregates in the condensates. In contrast, the α-Syn A53E mutant retarded the aggregation during the liquid-to-solid phase transition. Finally, we observed that WT and mutant α-Syn formed condensates in the cells, whereas the E46K mutation apparently promoted the formation of condensates. These findings reveal that familial PD-associated mutations have divergent effects on α-Syn LLPS and amyloid aggregation in the phase-separated condensates, providing new insights into the pathogenesis of PD-associated α-Syn mutations.

The natural history study of preclinical genetic Creutzfeldt-Jakob Disease (CJD): a prospective longitudinal study protocol

BackgroundCreutzfeldt-Jakob Disease (CJD) is the most common prion disease in humans causing a rapidly progressive neurological decline and dementia and is invariably fatal. The familial forms (genetic CJD, gCJD) are caused by mutations in the PRNP gene encoding for the prion protein (PrP). In Israel, there is a large cluster of gCJD cases, carriers of an E200K mutation in the PRNP gene, and therefore the largest population of at-risk individuals in the world. The mutation is not necessarily sufficient for the formation and accumulation of the pathological prion protein (PrPsc), suggesting that other, genetic and non-genetic factors affect the age at symptoms onset. Here we present the protocol of a cross-sectional and longitudinal natural history study of gCJD patients and first-degree relatives of gCJD patients, aiming to identify biological markers of preclinical CJD and risk factors for phenoconversion.MethodsThe study has two groups: Patients diagnosed with gCJD, and first-degree healthy relatives (HR) (both carriers and non-carriers of the E200K mutation in the PRNP gene) of patients diagnosed with gCJD. At baseline, and at the end of every year, healthy participants are invited for an “in-depth” visit, which includes a clinical evaluation, blood and urine collection, gait assessment, brain MRI, lumbar puncture (LP), and Polysomnography (PSG). At 6 months from baseline, and then halfway through each year, participants are invited for a “brief” visit, which includes a clinical evaluation, short cognitive assessment, and blood and urine collection. gCJD patients will be invited for one “in-depth” visit, similar to the baseline visit of healthy relatives.DiscussionThis continuous follow-up of the participants and the frequent assessments will allow early identification and diagnosis in case of conversion into disease. The knowledge generated from this study is likely to advance the understanding of the underlying clinicopathological processes that occur at the very beginning of CJD, as well as potential genetic and environmental risk factors for the development of the disease, therefore advancing the development of safe and efficient interventions.Trial registrationThe study is an observational study. It has registered retrospectively in https://clinicaltrials.gov/ and has been assigned an identification number NCT05746715.

Interfacial properties of α-synuclein's Parkinsonian variants

Human alpha-synuclein (αS) is associated with the occurrence of Parkinson's disease. In the past decade, six autosomally dominant mutations have been identified in αS (SNCA) gene that translate into A30P, E46K, H50Q, G51D, A53E, and A53T mutations in the protein. These mutations alter the electrostatics and hydrophobicity of a cardinal region of the protein. A comprehensive comparison of interfacial properties of these Parkinsonian αS variants is crucial to understand their membrane dynamics. Here, we investigated the interfacial activity of these αS variants at air-aqueous interface. All the αS variants were found to possess comparable surface activity of ∼20–22 mN/m. Compression/expansion isotherms reveal a very distinct behaviour of the A30P variant compared to others. The Blodgett-deposited films were analysed using CD and LD spectroscopy as well as the atomic force microscopy. All the variants adopted predominantly α-helical conformation in these films. Atomic force microscopy of the Langmuir-Blodgett films revealed self-assembly at the interface. The lipid-penetration activity was also investigated using zwitterionic and negatively charged lipid monolayers.

Therapeutic Trial of anle138b in Mouse Models of Genetic Prion Disease.

Phenotypic screening has yielded small-molecule inhibitors of prion replication that are effective in vivo against certain prion strains but not others. Here, we sought to test the small molecule anle138b in multiple mouse models of prion disease. In mice inoculated with the RML strain of prions, anle138b doubled survival and durably suppressed astrogliosis measured by live-animal bioluminescence imaging. In knock-in mouse models of the D178N and E200K mutations that cause genetic prion disease, however, we were unable to identify a clear, quantifiable disease endpoint against which to measure therapeutic efficacy. Among untreated animals, the mutations did not impact overall survival, and bioluminescence remained low out to >20 months of age. Vacuolization and PrP deposition were observed in some brain regions in a subset of mutant animals but appeared to be unable to carry the weight of a primary endpoint in a therapeutic study. We conclude that not all animal models of prion disease are suited to well-powered therapeutic efficacy studies, and care should be taken in choosing the models that will support drug development programs. IMPORTANCE There is an urgent need to develop drugs for prion disease, a currently untreatable neurodegenerative disease. In this effort, there is a debate over which animal models can best support a drug development program. While the study of prion disease benefits from excellent animal models because prions naturally afflict many different mammals, different models have different capabilities and limitations. Here, we conducted a therapeutic efficacy study of the drug candidate anle138b in mouse models with two of the most common mutations that cause genetic prion disease. In a more typical model where prions are injected directly into the brain, we found anle138b to be effective. In the genetic models, however, the animals never reached a clear, measurable point of disease onset. We conclude that not all prion disease animal models are ideally suited to drug efficacy studies, and well-defined, quantitative disease metrics should be a priority.

Is There Horizontal Transmission of Creutzfeldt-Jakob Disease?

Background: Sporadic Creutzfeldt-Jakob disease (s-CJD) is a rare, fatal neurodegenerative disorder. Familial cases of Creutzfeldt-Jakob disease (f-CJD) due to mutations in the PRNP gene are even rarer around the world; however, in Israel there is a focus of f-CJD patients carrying the E200K mutation. As the number of CJD E200K carriers in Israel is high and increasing, transmission of CJD to normal people was suspected. If such transmission occurs, the incidence of s-CJD would be expected to increase as well, resulting in changes of the ratio of familial/sporadic cases. Methods: Using data from the National CJD Registry and official statistics on the Israeli population, we studied incidence rates of f-CJD and s-CJD for the period from 1985 to 2018 applying the Surveillance Epidemiology and End Results (SEER) statistical packet developed in the US National Cancer Institute. Results: In total, 621 CJD patients (405 f-CJD and 216 s-CJD) cases are included in the registry. In the cohort of f-CJD patients, the mean age-adjusted annual incidence rate over the abovementioned period was 1.88 ± 0.09 (95% CI: 1.7–2.08) per 1,000,000. In the cohort of s-CJD patients, the mean age-adjusted incidence rate over the same period was 0.93 ± 0.06 (95% CI: 0.81–1.06) per 1,000,000 people. No significant time trends were found over the observation period in either s-CJD or f-CJD. The ratio f-CJD/s-CJD decreases over the observation period from 2.2 to 1.80. Conclusion: Israel has a high predominance of f-CJD compared to s-CJD. The mean incidence rate of s-CJD in Israel is similar to most countries. Between 1985 and 2018, the annual age-adjusted incidence rates for both forms of CJD remained stable. Thus, there is no evidence that CJD is transmitted from affected individuals to others.

Prion Mutations in Republic of Republic of Korea, China, and Japan.

Prion gene (PRNP) mutations are associated with diverse disease phenotypes, including familiar Creutzfeldt-Jakob Disease (CJD), Gerstmann-Sträussler-Scheinker disease (GSS), and fatal familial insomnia (FFI). Interestingly, PRNP mutations have been reported in patients diagnosed with Alzheimer's disease, dementia with Lewy bodies, Parkinson's disease, and frontotemporal dementia. In this review, we describe prion mutations in Asian countries, including Republic of Republic of Korea, China, and Japan. Clinical phenotypes and imaging data related to these mutations have also been introduced in detail. Several prion mutations are specific to Asians and have rarely been reported in countries outside Asia. For example, PRNP V180I and M232R, which are rare in other countries, are frequently detected in Republic of Korea and Japan. PRNP T188K is common in China, and E200K is significantly more common among Libyan Jews in Israel. The A117V mutation has not been detected in any Asian population, although it is commonly reported among European GSS patients. In addition, V210I or octapeptide insertion is common among European CJD patients, but relatively rare among Asian patients. The reason for these differences may be geographical or ethical isolation. In terms of clinical phenotypes, V180I, P102L, and E200K present diverse clinical symptoms with disease duration, which could be due to other genetic and environmental influences. For example, rs189305274 in the ACO1 gene may be associated with neuroprotective effects in cases of V180I mutation, leading to longer disease survival. Additional neuroprotective variants may be possible in cases featuring the E200K mutation, such as KLKB1, KARS, NRXN2, LAMA3, or CYP4X1. E219K has been suggested to modify the disease course in cases featuring the P102L mutation, as it may result in the absence of prion protein-positive plaques in tissue stained with Congo red. However, these studies analyzed only a few patients and may be too preliminary. The findings need to be verified in studies with larger sample sizes or in other populations. It would be interesting to probe additional genetic factors that cause disease progression or act as neuroprotective factors. Further studies are needed on genetic modifiers working with prions and alterations from mutations.

Effects of the pathological E200K mutation on human prion protein: A computational screening and molecular dynamics approach.

The human prion protein gene (PRNP) is mapped to the short arm of chromosome 20 (20pter-12). Prion disease is associated with mutations in the prion protein-encoding gene sequence. Earlier studies found that the mutation G127V in the PRNP increases protein stability. In contrast, the mutation E200K, which has the highest mutation rate in the prion protein, causes Creutzfeldt-Jakob disease (CJD) in humans and induces protein aggregation.We aimed to identify the structural mechanisms of E200k and G127V mutations causing CJD.We used a variety of bioinformatic algorithms, including SIFT, PolyPhen, I-Mutant, PhD-SNP, and SNP& GO, to predict the association of the E200K mutation with prion disease. MD simulation is performed, and graphs for root mean square deviation, root mean square fluctuation, radius of gyration, DSSP, principal component analysis, porcupine, and free energy landscapeare generated to confirm and prove the stability of the wild-type and mutant protein structures.The protein is analyzed for aggregation, and the results indicate more fluctuations in the protein structure during the simulation owing to the E200K mutation; however, the G127V mutation makes the protein structure stable against aggregation during the simulation.

The E46K mutation modulates α-synuclein prion replication in transgenic mice.

In multiple system atrophy (MSA), the α-synuclein protein misfolds into a self-templating prion conformation that spreads throughout the brain, leading to progressive neurodegeneration. While the E46K mutation in α-synuclein causes familial Parkinson's disease (PD), we previously discovered that this mutation blocks in vitro propagation of MSA prions. Recent studies by others indicate that α-synuclein adopts a misfolded conformation in MSA in which a Greek key motif is stabilized by an intramolecular salt bridge between residues E46 and K80. Hypothesizing that the E46K mutation impedes salt bridge formation and, therefore, exerts a selective pressure that can modulate α-synuclein strain propagation, we asked whether three distinct α-synuclein prion strains could propagate in TgM47+/- mice, which express human α-synuclein with the E46K mutation. Following intracranial injection of these strains, TgM47+/- mice were resistant to MSA prion transmission, whereas recombinant E46K preformed fibrils (PFFs) transmitted neurological disease to mice and induced the formation of phosphorylated α-synuclein neuropathology. In contrast, heterotypic seeding following wild-type (WT) PFF-inoculation resulted in preclinical α-synuclein prion propagation. Moreover, when we inoculated TgM20+/- mice, which express WT human α-synuclein, with E46K PFFs, we observed delayed transmission kinetics with an incomplete attack rate. These findings suggest that the E46K mutation constrains the number of α-synuclein prion conformations that can propagate in TgM47+/- mice, expanding our understanding of the selective pressures that impact α-synuclein prion replication.

Stress and viral insults do not trigger E200K PrP conversion in human cerebral organoids.

Prion diseases are a group of rare, transmissible, and invariably fatal neurodegenerative diseases that affect both humans and animals. The cause of these diseases is misfolding of the prion protein into pathological isoforms called prions. Of all human prion diseases, 10–15% of cases are genetic and the E200K mutation, which causes familial Creutzfeldt-Jakob disease (CJD), is the most prevalent. For both sporadic and genetic disease, it remains uncertain as to how initial protein misfolding is triggered. Prior studies have linked protein misfolding with oxidative stress insults, deregulated interactions with cellular cofactors, and viral infections. Our previous work developed a cerebral organoid (CO) model using human induced pluripotent stem cells containing the E200K mutation. COs are three-dimensional human neural tissues that permit the study of host genetics and environmental factors that contribute to disease onset. Isogenically matched COs with and without the E200K mutation were used to investigate the propensity of E200K PrP to misfold following cellular insults associated with oxidative stress. Since viral infections have also been associated with oxidative stress and neurodegenerative diseases, we additionally investigated the influence of Herpes Simplex Type-1 virus (HSV1), a neurotropic virus that establishes life-long latent infection in its host, on E200K PrP misfolding. While COs proved to be highly infectable with HSV1, neither acute nor latent infection, or direct oxidative stress insult, resulted in evidence of E200K prion misfolding. We conclude that misfolding into seeding-active PrP species is not readily induced by oxidative stress or HSV1 in our organoid system.

Hereditary E200K mutation within the prion protein gene alters human iPSC derived cardiomyocyte function

Cardiomyopathy is a co-morbidity of some prion diseases including genetic disease caused by mutations within the PrP gene (PRNP). Although the cellular prion protein (PrP) has been shown to protect against cardiotoxicity caused by oxidative stress, it is unclear if the cardiomyopathy is directly linked to PrP dysfunction. We differentiated cardiomyocyte cultures from donor human induced pluripotent stem cells and found a direct influence of the PRNP E200K mutation on cellular function. The PRNP E200K cardiomyocytes showed abnormal function evident in the irregularity of the rapid repolarization; a phenotype comparable with the dysfunction reported in Down Syndrome cardiomyocytes. PRNP E200K cardiomyocyte cultures also showed increased mitochondrial superoxide accompanied by increased mitochondrial membrane potential and dysfunction. To confirm that the changes were due to the E200K mutation, CRISPR-Cas9 engineering was used to correct the E200K carrier cells and insert the E200K mutation into control cells. The isotype matched cardiomyocytes showed that the lysine expressing allele does directly influence electrophysiology and mitochondrial function but some differences in severity were apparent between donor lines. Our results demonstrate that cardiomyopathy in hereditary prion disease may be directly linked to PrP dysfunction.

Genetic Creutzfeldt-Jakob disease in Turkish Jews-demographic and clinical features.

The largest cluster of genetic Creutzfeldt- Jakob Disease (CJD) exists in Libyan Jews carrying the E200K mutation in the PRNP gene. However, there is another cluster of genetic CJD with E200K mutation in families of Turkish-Jewish origin. In this retrospective study, we aim to describe the demographic and clinical features of this population of patients. The Israeli National CJD database was searched for demographic, clinical, imaging, and laboratory data of genetic CJD patients of Libyan and Turkish ancestry with the E200K mutation. The data of Libyan and Turkish patients were compared with notice similar or different demographic or clinical courses. Four hundred and twenty-three patients with CJD of Libyan (L) ancestry and 27 patients with CJD of Turkish (T) ancestry were identified. There were no significant differences in demographic and clinical data between the two populations (age of onset: T=62 ± 8.8, L=60 ± 9.7; age of death: T=63 ± 8.6, L=61 ± 9.7; and disease duration: T=7.8 ± 8.4 months, L=9.6 ± 13.6 months). Rapidly progressive dementia was the most common presentation in both groups, followed by pure cerebellar onset. The levels of tau protein in CSF did not differ between groups (T=1290 ± 397.6 pg/ml, L=1276 ± 594.2 pg/ml). MRI and EEG showed classical CJD features in most patients in both groups. The E200K mutation is the most common mutation among gCJD patients and was reported in different ethnical populations, suggesting several independent haplotypes of the mutation. The Turkish-Jew cluster, first described in this study, shares similar demographic and clinical features with the bigger cluster of Libyan-Jews CJD patients. E200K gCJD patients of Turkish ancestry share similar demographic and clinical features to patients of Libyan descent, suggesting a common origin of both populations.