Performance of a seed amplification assay for misfolded alpha-synuclein in cerebrospinal fluid and brain tissue in relation to Lewy body disease stage and pathology burden.

The development of in vitro seed amplification assays (SAA) detecting misfolded alpha-synuclein (αSyn) in cerebrospinal fluid (CSF) and other tissues has provided a pathology-specific biomarker for Lewy body disease (LBD). However, αSyn SAA diagnostic performance in early pathological stages or low Lewy body (LB) pathology load has only been assessed in small cohorts. Moreover, the relationship between SAA kinetic parameters, the number of αSyn brain seeds and the LB pathology burden assessed by immunohistochemistry has never been systematically investigated. We tested 269 antemortem CSF samples and 138 serially diluted brain homogenates from patients with and without neuropathological evidence of LBD in different stages by the αSyn Real-Time Quaking-Induced Conversion (RT-QuIC) SAA. Moreover, we looked for LB pathology by αSyn immunohistochemistry in a consecutive series of 604 Creutzfeldt-Jakob disease (CJD)-affected brains. αSyn CSF RT-QuIC showed 100% sensitivity in detecting LBD in limbic and neocortical stages. The assay sensitivity was significantly lower in patients in early stages (37.5% in Braak 1 and 2, 73.3% in Braak 3) or with focal pathology (50% in amygdala-predominant). The average number of CSF RT-QuIC positive replicates significantly correlated with LBD stage. Brain homogenate RT-QuIC showed higher sensitivity than immunohistochemistry for the detection of misfolded αSyn. In the latter, the kinetic parameter lag phase (time to reach the positive threshold) strongly correlated with the αSyn seed concentration in serial dilution experiments. Finally, incidental LBD prevalence was 8% in the CJD cohort. The present results indicate that (a) CSF RT-QuIC has high specificity and sufficient sensitivity to detect all patients with LB pathology at Braak stages > 3 and most of those at stage 3; (b) brain deposition of misfolded αSyn precedes the formation of LB and Lewy neurites; (c) αSyn SAA provides "quantitative" information regarding the LB pathology burden, with the lag phase and the number of positive replicates being the most promising variables to be used in the clinical setting.

Evaluation of the impact of CSF prion RT-QuIC and amended criteria on the clinical diagnosis of Creutzfeldt-Jakob disease: a 10-year study in Italy.

BACKGROUND: The introduction of the prion Real-Time Quaking-Induced Conversion assay (RT-QuIC) has led to a revision of the diagnostic criteria for sporadic Creutzfeldt-Jakob disease (sCJD).Validation studies are needed for the amended criteria, especially for their diagnostic value in the clinical setting. METHODS: We studied 1250 patients with suspected CJD referred for diagnosis to two Italian reference centres between 2010 and 2020. Focusing on the first diagnostic assessment, we compared the diagnostic value of the old and the amended criteria and that of different combinations of clinical variables and biomarker results. RESULTS: The studied cohort comprised 850 participants with CJD (297 definite sCJD, 151 genetic CJD, 402 probable sCJD) and 400 with non-CJD (61 with neuropathology). At first clinical evaluation, the sensitivity of the old criteria (76.8%) was significantly lower than that of the amended criteria (97.8%) in the definite CJD cohort with no difference between definite and probable sCJD cases. Specificity was ~94% for both criteria against the non-CJD cohort (82.0% against definite non-CJD group). Cerebrospinal fluid (CSF) RT-QuIC was highly sensitive (93.9%) and fully specific against definite non-CJD patients. Limiting the criteria to a positive RT-QuIC or/and the combination of a clinical course compatible with possible CJD with a positive MRI (Q-CM criteria) provided higher diagnostic accuracy than both the old and amended criteria, overcoming the suboptimal specificity of ancillary test results (ie, CSF protein 14-3-3). CONCLUSIONS: CSF RT-QuIC is highly sensitive and specific for diagnosing CJD in vitam. The Q-CM criteria provide a high diagnostic value for CJD.

Diagnostic accuracy of cerebrospinal fluid biomarkers in genetic prion diseases.

Genetic prion diseases are a rare and diverse group of fatal neurodegenerative disorders caused by pathogenic sequence variations in the prion protein gene, PRNP. Data on CSF biomarkers in patients with genetic prion diseases are limited and conflicting results have been reported for unclear reasons. Here, we aimed to analyse the diagnostic accuracy of CSF biomarkers currently used in prion clinical diagnosis in 302 symptomatic genetic prion disease cases from 11 prion diagnostic centres, encompassing a total of 36 different pathogenic sequence variations within the open reading frame of PRNP. CSF samples were assessed for the surrogate markers of neurodegeneration, 14-3-3 protein (14-3-3), total-tau protein (t-tau) and α-synuclein and for prion seeding activity through the real-time quaking-induced conversion assay. Biomarker results were compared with those obtained in healthy and neurological controls. For the most prevalent PRNP pathogenic sequence variations, biomarker accuracy and associations between biomarkers, demographic and genetic determinants were assessed. Additionally, the prognostic value of biomarkers for predicting total disease duration from symptom onset to death was investigated. High sensitivity of the four biomarkers was detected for genetic Creutzfeldt-Jakob disease associated with the E200K and V210I mutations, but low sensitivity was observed for mutations associated with Gerstmann-Sträussler-Scheinker syndrome and fatal familial insomnia. All biomarkers showed good to excellent specificity using the standard cut-offs often used for sporadic Creutzfeldt-Jakob disease. In genetic prion diseases related to octapeptide repeat insertions, the biomarker sensitivity correlated with the number of repeats. New genetic prion disease-specific cut-offs for 14-3-3, t-tau and α-synuclein were calculated. Disease duration in genetic Creutzfeldt-Jakob disease-E200K, Gerstmann-Sträussler-Scheinker-P102L and fatal familial insomnia was highly dependent on PRNP codon 129 MV polymorphism and was significantly associated with biomarker levels. In a large cohort of genetic prion diseases, the simultaneous analysis of CSF prion disease biomarkers allowed the determination of new mutation-specific cut-offs improving the discrimination of genetic prion disease cases and unveiled genetic prion disease-specific associations with disease duration.

Concordance of cerebrospinal fluid real-time quaking-induced conversion across the European Creutzfeldt-Jakob Disease Surveillance Network.

BACKGROUND AND PURPOSE: Cerebrospinal fluid (CSF) real-time quaking-induced conversion (RT-QuIC) has a high degree of sensitivity and specificity for the diagnosis of sporadic Creutzfeldt-Jakob disease (sCJD) and this has led to its being included in revised European CJD Surveillance Network diagnostic criteria for sCJD. As CSF RT-QuIC becomes more widely established, it is crucial that the analytical performance of individual laboratories is consistent. The aim of this ring-trial was to ascertain the degree of concordance between European countries undertaking CSF RT-QuIC. METHODS: Ten identical CSF samples, seven from probable or neuropathologically confirmed sCJD and three from non-CJD cases, were sent to 13 laboratories from 11 countries for RT-QuIC analysis. A range of instrumentation and different recombinant prion protein substrates were used. Each laboratory analysed the CSF samples blinded to the diagnosis and reported the results as positive or negative. RESULTS: All 13 laboratories correctly identified five of the seven sCJD cases and the remaining two sCJD cases were identified by 92% of laboratories. Of the two sCJD cases that were not identified by all laboratories, one had a disease duration >26 months with a negative 14-3-3, whilst the remaining case had a 4-month disease duration and a positive 14-3-3. A single false positive CSF RT-QuIC result was observed in this study. CONCLUSIONS: This study shows that CSF RT-QuIC demonstrates an excellent concordance between centres, even when using a variety of instrumentation, recombinant prion protein substrates and CSF volumes. The adoption of CSF RT-QuIC by all CJD surveillance centres is recommended.

Biochemical and Neuropathological Findings in a Creutzfeldt-Jakob Disease Patient with the Rare Val180Ile-129Val Haplotype in the Prion Protein Gene.

Genetic Creutzfeldt-Jakob disease (gCJD) associated with the V180I mutation in the prion protein (PrP) gene (PRNP) in phase with residue 129M is the most frequent cause of gCJD in East Asia, whereas it is quite uncommon in Caucasians. We report on a gCJD patient with the rare V180I-129V haplotype, showing an unusually long duration of the disease and a characteristic pathological PrP (PrPSc) glycotype. Family members carrying the mutation were fully asymptomatic, as commonly observed with this mutation. Neuropathological examination showed a lesion pattern corresponding to that commonly reported in Japanese V180I cases with vacuolization and gliosis of the cerebral cortexes, olfactory areas, hippocampus and amygdala. PrP was deposited with a punctate, synaptic-like pattern in the cerebral cortex, amygdala and olfactory tract. Western blot analyses of proteinase-K-resistant PrP showed the characteristic two-banding pattern of V180I gCJD, composed of mono- and un-glycosylated isoforms. In line with reports on other V180I cases in the literature, Real-Time Quaking Induced Conversion (RT-QuIC) analyses did not demonstrate the presence of seeding activity in the cerebrospinal fluid and olfactory mucosa, suggesting that this haplotype also may result in a reduced seeding efficiency of the pathological PrP. Further studies are required to understand the origin, penetrance, disease phenotype and transmissibility of 180I-129V haplotype in Caucasians.

Genetic Creutzfeldt-Jakob disease in Sardinia: a case series linked to the PRNP R208H mutation due to a single founder effect.

In genetic prion diseases (gPrD), five genetic variants (E200K, V210I, V180I, P102L, and D178N) are responsible for about 85% of cases. The R208H is one of the several additional rare mutations and to date, only 16 cases carrying this mutation have been reported worldwide. To describe the phenotypic features of 5 affected patients belonging to apparently unrelated Sardinian (Italian) families with R208H gPrD, and provide evidence for a possible founder effect are the aims of this study. The R208H PRNP mutation has a much higher relative frequency in Sardinia than elsewhere in Italy (72% vs. 4.4% of gCJD cases). Our cohort shared similar phenotypic features to the previously described patients with R208H-129M haplotype with most patients showing the classical Creutzfeldt-Jakob disease (CJD) phenotype. The analysis of 10 controls and 5 patients by NGS sequencing identified 4 haplotypes, 3 associated with the wild type variant, and one (H1) shared by all patients carrying the 208His variant. This is the first report of a regional cluster for R208H mutation in gPrD and the first report of the presence of a common ancestor for this Sardinian R208H cluster, confirming the probable consequences of genetic isolation process even for rare diseases.

Comparison between plasma and cerebrospinal fluid biomarkers for the early diagnosis and association with survival in prion disease.

OBJECTIVE: To compare the diagnostic accuracy and the prognostic value of blood and cerebrospinal fluid (CSF) tests across prion disease subtypes. METHODS: We used a single-molecule immunoassay to measure tau and neurofilament light chain (NfL) protein levels in the plasma and assessed CSF total(t)-tau, NfL and protein 14-3-3 levels in patients with prion disease (n=336), non-prion rapidly progressive dementias (n=106) and non-neurodegenerative controls (n=37). We then evaluated each plasma and CSF marker for diagnosis and their association with survival, taking into account the disease subtype, which is a strong independent prognostic factor in prion disease. RESULTS: Plasma tau and NfL concentrations were higher in patients with prion disease than in non-neurodegenerative controls and non-prion rapidly progressive dementias. Plasma tau showed higher diagnostic value than plasma NfL, but a lower accuracy than the CSF proteins t-tau and 14-3-3. In the whole prion cohort, both plasma (tau and NfL) and CSF (t-tau, 14-3-3 and NfL) markers were significantly associated with survival and showed similar prognostic values. However, the intrasubtype analysis revealed that only CSF t-tau in sporadic Creutzfeldt-Jakob disease (sCJD) MM(V)1, plasma tau and CSF t-tau in sCJD VV2, and plasma NfL in slowly progressive prion diseases were significantly associated with survival after accounting for covariates. CONCLUSIONS: Plasma markers have lower diagnostic accuracy than CSF biomarkers. Plasma tau and NfL and CSF t-tau are significantly associated with survival in prion disease in a subtype-specific manner and can be used to improve clinical trial stratification and clinical care.

Prion Strain Characterization of a Novel Subtype of Creutzfeldt-Jakob Disease.

In 2007, we reported a patient with an atypical form of Creutzfeldt-Jakob disease (CJD) heterozygous for methionine-valine (MV) at codon 129 who showed a novel pathological prion protein (PrPTSE) conformation with an atypical glycoform (AG) profile and intraneuronal PrP deposition. In the present study, we further characterize the conformational properties of this pathological prion protein (PrPTSE MVAG), showing that PrPTSE MVAG is composed of multiple conformers with biochemical properties distinct from those of PrPTSE type 1 and type 2 of MV sporadic CJD (sCJD). Experimental transmission of CJD-MVAG to bank voles and gene-targeted transgenic mice carrying the human prion protein gene (TgHu mice) showed unique transmission rates, survival times, neuropathological changes, PrPTSE deposition patterns, and PrPTSE glycotypes that are distinct from those of sCJD-MV1 and sCJD-MV2. These biochemical and experimental data suggest the presence of a novel prion strain in CJD-MVAGIMPORTANCE Sporadic Creutzfeldt-Jakob disease is caused by the misfolding of the cellular prion protein, which assumes two different major conformations (type 1 and type 2) and, together with the methionine/valine polymorphic codon 129 of the prion protein gene, contribute to the occurrence of distinct clinical-pathological phenotypes. Inoculation in laboratory rodents of brain tissues from the six possible combinations of pathological prion protein types with codon 129 genotypes results in the identification of 3 or 4 strains of prions. We report on the identification of a novel strain of Creutzfeldt-Jakob disease isolated from a patient who carried an abnormally glycosylated pathological prion protein. This novel strain has unique biochemical characteristics, does not transmit to humanized transgenic mice, and shows exclusive transmission properties in bank voles. The identification of a novel human prion strain improves our understanding of the pathogenesis of the disease and of possible mechanisms of prion transmission.

Age at onset of genetic (E200K) and sporadic Creutzfeldt-Jakob diseases is modulated by the CYP4X1 gene.

OBJECTIVES: The Glu to Lys change at codon 200 (E200K) of the PRNP gene is the most frequent mutation associated to genetic Creutzfeldt-Jakob disease (CJD) and the only one responsible for geographical clusters. Patients carrying this mutation develop disease at different ages and show variable clinical phenotypes that are not affected by the methione/valine polymorphism at codon 129 of the PRNP gene suggesting the influence of other factors. The objective of this study is to look for genes other than PRNP that might be responsible of this variability. METHODS: We searched for other genes by performing genome-wide analyses (GWA) on 19 patients with genetic CJD and 18 healthy subjects carrying the E200K mutation of PRNP and belonging to the Calabrian cluster in Italy. We then validate this result in 32 patients with E200K CJD from non-cluster areas and 259 patients with sporadic CJD referred to the Italian CJD national registry. RESULTS AND CONCLUSIONS: We identified two single nucleotide polymorphisms on the CYP4X1 gene locus as candidate disease modifiers in patients with E200K CJD of the cluster area and confirmed this finding in 32 patients with E200K CJD from non-cluster areas and 259 patients with sporadic CJD. Our results indicate that the CYP4X1 gene modulates the onset of disease in patients with E200K genetic and sporadic CJD. This finding improves our understanding on the pathogenesis of CJD, suggests new targets for developing novel therapeutic strategies and might be useful for the stratification of patients in future preventive treatment trials.

Cerebrospinal fluid real-time quaking-induced conversion is a robust and reliable test for sporadic creutzfeldt-jakob disease: An international study.

Real-time quaking-induced conversion (RT-QuIC) has been proposed as a sensitive diagnostic test for sporadic Creutzfeldt-Jakob disease; however, before this assay can be introduced into clinical practice, its reliability and reproducibility need to be demonstrated. Two international ring trials were undertaken in which a set of 25 cerebrospinal fluid samples were analyzed by a total of 11 different centers using a range of recombinant prion protein substrates and instrumentation. The results show almost complete concordance between the centers and demonstrate that RT-QuIC is a suitably reliable and robust technique for clinical practice. Ann Neurol 2016;80:160-165.

Heidenhain variant in two patients with inherited V210I Creutzfeldt-Jakob disease.

OBJECTIVE: To report two members of the same family carrying the valine to isoleucine point mutation of the prion protein gene (PRNP) and presenting with visual symptoms as initial manifestation as in the "Heidenhain variant" of sporadic Creutzfeldt-Jakob disease (CJD). METHODS: Patients underwent neurological examination, electroencephalogram (EEG), brain magnetic resonance images (MRI) and cerebrospinal fluid (CSF) analysis including the Real Time Quaking Induced Conversion (RT-QuIC) test. Disease-specific mutations and polymorphism at codon 129 of the PRNP gene were also studied. RESULTS: Isolated visual symptoms characterized disease onset of both patients followed by progressive neurological signs, dementia and death in 3 (proband) and 9 (his aunt) months. RT-QuIC analysis of CSF samples of both patients revealed the presence of the pathological prion protein and DNA analysis the V210I point mutation of the PRNP and methionine homozygosity at the polymorphic codon 129. CONCLUSIONS: This report suggests to consider the diagnosis of V210I genetic CJD in patients presenting with the Heidenhain form of CJD and highlights the importance of genetic testing in all patients with isolated visual manifestations at onset followed by progressive neurological signs and dementia.

Mutant PrPCJD prevails over wild-type PrPCJD in the brain of V210I and R208H genetic Creutzfeldt-Jakob disease patients.

Creutzfeldt-Jakob disease (CJD) is a neurodegenerative disorder characterized by the deposition of the pathological conformer (PrP(CJD)) of the host encoded cellular prion protein (PrP(C)). In genetic CJD associated with V210I or R208H PrP substitutions, the pathogenic role of mutant residues is still poorly understood. To understand how V210I or R208H PrP mutations facilitate the development of the disease, we determined by mass spectrometry the quantitative ratio of mutant/wild-type PrP(CJD) allotypes in brains from affected subjects. We found that the mutant PrP(CJD) allotypes moderately exceeds of 2- or 3-fold the amount of the wild-type counterpart suggesting that these mutations mainly exert their pathogenic effect on the onset of the pathogenic cascade. Different mechanisms can be hypothesized to explain the pathogenic role of mutant residues: V210I and R208H substitutions can increase the concentration of PrP(C) and the probability to form insoluble aggregates, or they may facilitate the formation of pathological intermediates, or, alternatively, they may increase the affinity for ligands that are involved in the initial phases of PrP(CJD) formation and aggregation. Whatever the mechanism, the enrichment found for the mutated PrP(CJD) species indicates that these altered structures are more prone, with respect to the non-mutated ones, to be captured in the polymerization process either at the onset or during the development of the disease.

Progressive supranuclear palsy phenotype as an atypical clinical presentation of Creutzfeldt-Jakob disease: A case report and review of the literature.

Creutzfeldt-Jakob disease (CJD) is a rare, rapidly progressive neurodegenerative disorder, characterized by the accumulation of abnormal prion proteins in the brain. While CJD has some typical clinical features, its presentation can be quite heterogeneous, particularly in the early stages of the disease, posing challenges in diagnosis. Atypical manifestations of CJD can mimic various neurodegenerative disorders, including atypical parkinsonisms. In this case report, we present an 81-year-old man who exhibited an atypical clinical presentation of sporadic CJD, initially resembling progressive supranuclear palsy (PSP). The patient presented with symmetric parkinsonism, postural instability, and ocular motor dysfunction, accompanied by rapid clinical deterioration. Alongside the case report, we also provide a review of the literature on atypical presentations of CJD as PSP, highlighting the importance of recognizing these manifestations in clinical practice.

The Use of Real-Time Quaking-Induced Conversion for the Diagnosis of Human Prion Diseases.

Prion diseases are rapidly progressive, invariably fatal, transmissible neurodegenerative disorders associated with the accumulation of the amyloidogenic form of the prion protein in the central nervous system (CNS). In humans, prion diseases are highly heterogeneous both clinically and neuropathologically. Prion diseases are challenging to diagnose as many other neurologic disorders share the same symptoms, especially at clinical onset. Definitive diagnosis requires brain autopsy to identify the accumulation of the pathological prion protein, which is the only specific disease biomarker. Although brain post-mortem investigation remains the gold standard for diagnosis, antemortem clinical, instrumental, and laboratory tests showing variable sensitivities and specificity, being surrogate disease biomarkers, have been progressively introduced in clinical practice to reach a diagnosis. More recently, the ultrasensitive Real-Time Quaking-Induced Conversion (RT-QuIC) assay, exploiting, for the first time, the detection of misfolded prion protein through an amplification strategy, has highly improved the "in-vitam" diagnostic process, reaching in cerebrospinal fluid (CSF) and olfactory mucosa (OM) around 96% sensitivity and close to 100% specificity. RT-QuIC also improved the detection of the pathologic prion protein in several peripheral tissues, possibly even before the clinical onset of the disease. The latter aspect is of great interest for the early and even preclinical diagnosis in subjects at genetic risk of developing the disease, who will likely be the main target population in future clinical trials. This review presents an overview of the current knowledge and future perspectives on using RT-QuIC to diagnose human prion diseases.

Penetrance of the V203I variant of the PRNP gene: report of a patient with stroke-like onset of Creutzfeld-Jacob Disease and review of published cases.

Creutzfeldt-Jakob disease (CJD) is usually sporadic, but 10-15% of cases are caused by autosomal-dominant pathogenic variants in the prion protein gene (PRNP). A few PRNP variants show low penetrance. We report the case of a 64-year-old man, admitted to the ward with acute onset of aphasia; death occurred 6 weeks later. Brain MRI, EEG pattern and brain pathology were consistent with CJD diagnosis. Genetic analysis revealed a heterozygous V203I variant. We summarized the key clinical findings in patients carrying the V203I variant who were described to date. We also discuss the hypothesis as to whether V203I is a risk factor for CJD rather than a Mendelian disease-associated variant, as well as the possible implications of such hypothesis in the clinical scenario.

Diagnostic and prognostic performance of CSF α-synuclein in prion disease in the context of rapidly progressive dementia.

INTRODUCTION: Surrogate cerebrospinal fluid (CSF) biomarkers of neurodegeneration still have a central role in the first-line screening of patients with suspected Creutzfeldt-Jakob disease (CJD). Recently, CSF α-synuclein, a marker of synaptic damage, showed a close to optimal performance in distinguishing between CJD and other neurodegenerative dementias. METHODS: We evaluated the diagnostic value of CSF α-synuclein in patients with prion disease, non-prion rapidly progressive dementias, and non-neurodegenerative controls. Additionally, we studied its distribution across the different prion disease subtypes and evaluated its association with survival. RESULTS: CSF α-synuclein levels were significantly higher in patients with prion disease than in the other groups but showed a lower diagnostic value than CSF total tau or 14-3-3. Moreover, CSF α-synuclein was significantly associated with survival in the whole prion cohort and the most frequent clinicopathological subtypes. DISCUSSION: In the clinical setting, CSF α-synuclein does not exceed the diagnostic performance of currently used surrogate markers, but it might constitute a robust prognostic indicator.

Selecting antidepressants according to a drug-by-environment interaction: A comparison of fluoxetine and minocycline effects in mice living either in enriched or stressful conditions.

Selective serotonin reuptake inhibitors (SSRIs) are the first-line treatment for major depressive disorder. It has been recently proposed that these drugs, by enhancing neural plasticity, amplify the influences of the living conditions on mood. Consequently, SSRI outcome depends on the quality of the environment, improving symptomatology mainly in individuals living in favorable conditions. In adverse conditions, drugs with a different mechanism of action might have higher efficacy. The antibiotic minocycline, with neuroprotective and anti-inflammatory properties, has been recently proposed as a novel potential antidepressant treatment. To explore the drug-by-environment interaction, we compared the effects on depressive-like behavior and neural plasticity of the SSRI fluoxetine and minocycline in enriched and stressful conditions. We first exposed C57BL/6 adult female mice to 14 days of chronic unpredictable mild stress to induce a depressive-like profile. Afterward, mice received vehicle, fluoxetine, or minocycline for 21 days, while exposed to either enriched or stressful conditions. During the first five days, fluoxetine led to an improvement in enrichment but not in stress. By contrast, minocycline led to an improvement in both conditions. After 21 days, all groups showed a significant improvement in enrichment while fluoxetine worsened the depressive like behavior in stress. The effects of the drugs on neural plasticity, measured as long-term potentiation, were also environment-dependent. Overall, we show that the environment affects fluoxetine but not minocycline outcome, indicating that the latter represents a potential alternative to SSRIs to treat depressed patients living in adverse conditions. From a translation perspective, our finding call for considering the drug-by-environment interaction to select the most effective pharmacological treatment.

Cathepsin D (C224T) polymorphism in sporadic and genetic Creutzfeldt-Jakob disease.

Accumulation of cathepsin D immunoreactive lysosomes correlates with tissue pathology in sporadic Creutzfeldt-Jakob disease (CJD) brains. The C-to-T transition within exon 2 of the cathepsin D (CTSD) gene is associated with altered enzymatic activity. Possession of the TT genotype is a risk factor for variant CJD. To verify the association between the CTSD position 224T allele and the risk for and survival in sporadic and genetic CJD, we genotyped 540 sporadic, 101 genetic CJD, and 723 control individuals. Genotype data and duration of illness were compared using multiple logistic regression and Kruskal-Wallis test. Multivariate survival analysis was performed using Cox's regression model. The distribution of CTSD position 224 alleles was approximately the same in all groups. We observed a trend for shorter survival in sporadic CJD patients harboring the T allele at position 224 of the CTSD gene in particular in sporadic CJD patients with the prion protein gene position 129 MM genotype. We conclude that the CTSD position 224 polymorphism alone is not a significant risk or disease-modifying factor in sporadic or genetic CJD.

Ring trial of 2nd generation RT-QuIC diagnostic tests for sporadic CJD.

OBJECTIVE: Real-time quaking-induced conversion (RT-QuIC) assays detect prion-seeding activity in a variety of human biospecimens, including cerebrospinal fluid and olfactory mucosa swabs. The assay has shown high diagnostic accuracy in patients with prion disorders. Recently, advances in these tests have led to markedly improved diagnostic sensitivity and reduced assay times. Accordingly, an algorithm has been proposed that entails the use of RT-QuIC analysis of both sample types to diagnose sporadic Creutzfeldt-Jakob disease with nearly 100% accuracy. Here we present a multi-center evaluation (ring trial) of the reproducibility of these improved "second generation" RT-QuIC assays as applied to these diagnostic specimens. METHODS: Cerebrospinal fluid samples were analyzed from subjects with sporadic Creutzfeldt-Jakob (n = 55) or other neurological diseases (n = 45) at multiple clinical centers. Olfactory mucosa brushings collected by multiple otolaryngologists were obtained from nine sporadic Creutzfeldt-Jakob disease cases and 19 controls. These sample sets were initially tested blindly by RT-QuIC by a coordinating laboratory, recoded, and then sent to five additional testing laboratories for blinded ring trial testing. RESULTS: Unblinding of the results by a third party indicated 98-100% concordance between the results obtained by the testing of these cerebrospinal fluid and nasal brushings at the six laboratories. INTERPRETATION: This second-generation RT-QuIC assay is highly transferrable, reproducible, and therefore robust for the diagnosis of sporadic Creutzfeldt-Jakob disease in clinical practice.

Diagnostic Accuracy of Prion Disease Biomarkers in Iatrogenic Creutzfeldt-Jakob Disease.

Human prion diseases are classified into sporadic, genetic, and acquired forms. Within this last group, iatrogenic Creutzfeldt-Jakob disease (iCJD) is caused by human-to-human transmission through surgical and medical procedures. After reaching an incidence peak in the 1990s, it is believed that the iCJD historical period is probably coming to an end, thanks to lessons learnt from past infection sources that promoted new prion prevention and decontamination protocols. At this point, we sought to characterise the biomarker profile of iCJD and compare it to that of sporadic CJD (sCJD) for determining the value of available diagnostic tools in promptly recognising iCJD cases. To that end, we collected 23 iCJD samples from seven national CJD surveillance centres and analysed the electroencephalogram and neuroimaging data together with a panel of seven CSF biomarkers: 14-3-3, total tau, phosphorylated/total tau ratio, alpha-synuclein, neurofilament light, YKL-40, and real-time quaking induced conversion of prion protein. Using the cut-off values established for sCJD, we found the sensitivities of these biomarkers for iCJD to be similar to those described for sCJD. Given the limited relevant information on this issue to date, the present study validates the use of current sCJD biomarkers for the diagnosis of future iCJD cases.