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Background: Lewy Body Dementia (LBD) is the second most common neurodegenerative dementia. To date, no validated biochemical marker is available to support clinical diagnosis. The development of the Real‐Time Quaking‐Induced Conversion (RT‐QuIC) assay for detecting alpha‐synuclein (aSyn) seeds in biological samples can be a sensitive biomarker specific for the diagnosis easily applicable in a clinical setting. We aimed to describe the diagnostic performance of RT‐QuIC aSyn assay in cerebrospinal fluid (CSF) to diagnose LBD in a clinical cohort with cognitive impairment. Method: A cohort of subjects with cognitive impairment (neurodegenerative and non‐neurodegenerative) with available CSF sample at the moment of first evaluation was selected by convenience sample in our database. Subjects had clinical follow‐ups ranging from 6 months to 12 years and current clinical diagnosis according to established consensus criteria. The diagnostic performance of RT‐QuIC aSyn for the diagnosis of LBD were evaluated. Result: The test was evaluated in 155 subjects (age 65 years (SD 11), MMSE 24 (SD 4.5), 56% male) with the following clinical diagnoses: LBD (n = 40), Alzheimer's disease (AD) (n = 73)(all with compatible CSF profile), frontotemporal dementia (FTD) (n = 7), non‐neurodegenerative mild cognitive impairment (MCI) (n = 21), other neurodegenerative diagnoses (n = 12) and healthy controls (n = 2). aSyn was detected in 33/40 (83%) LBD patients, 7/73 (10%) AD, 0/7 FTD patients, 0/21 MCI patients, 0/12 patients with other neurodegenerative diseases and 0/2 healthy controls. Only 3/7 (42%) LBD subjects with a negative RT‐QuIC asyn fulfilled criteria for LBD (established or prodromal) at the moment of first evaluation as compared with 23/33 (69%) of LBD with positive RTQuIC suggesting a more initial disease in the negative group at the moment of CSF sampling. The sensitivity and specificity of the assay were 83% and 94% respectively for the diagnosis of LBD, with a PPV of 83% and a NPV of 94%. Conclusion: Detection of αSyn seeds by RT‐QuIC has a good performance in identifying patients with LBD in a clinical cohort of cognitively impaired subjects. The test also identifies aSyn co‐pathology in a subgroup of subjects diagnosed with AD. [ABSTRACT FROM AUTHOR]

This study quantified the occurrence of an underlying synucleinopathy in 50 patients with idiopathic normal pressure hydrocephalus by means of real‐time quaking‐induced conversion, a highly sensitive and specific technique capable of detecting and amplifying misfolded aggregated forms of α‐synuclein in the cerebrospinal fluid. Seven patients were positive and they did not differ from negative cases, except for a more frequent L‐dopa responsiveness and gait characterized by a wider base. The two groups did not differ in terms of response rate to tap test or shunt surgery, although step length and gait velocity improved by a lesser extent in positive cases. ANN NEUROL 2022;92:985–991 [ABSTRACT FROM AUTHOR]

Background and purpose: Real‐time quaking‐induced conversion (RT‐QuIC) assays offer a sensitive and specific means for detection of prions, although false negative results are recognized in clinical practice. We profile the clinical, laboratory, and pathologic features associated with false negative RT‐QuIC assays and extend these to frame the diagnostic approach to patients with suspected prion disease. Methods: A total of 113 patients with probable or definite prion disease were assessed at Mayo Clinic (Rochester, MN; Jacksonville, FL; Scottsdale, AZ) or Washington University School of Medicine (Saint Louis, MO) from 2013 to 2021. RT‐QuIC testing for prions was performed in cerebrospinal fluid (CSF) at the National Prion Disease Pathology Surveillance Center (Cleveland, OH). Results: Initial RT‐QuIC testing was negative in 13 of 113 patients (sensitivity = 88.5%). RT‐QuIC negative patients were younger (median = 52.0 years vs. 66.1 years, p < 0.001). Other demographic and presenting features, and CSF cell count, protein, and glucose levels were similar in RT‐QuIC negative and positive patients. Frequency of 14‐3‐3 positivity (4/13 vs. 77/94, p < 0.001) and median CSF total tau levels were lower in RT‐QuIC negative patients (2517 vs. 4001 pg/mL, p = 0.020), and time from symptom onset to first presentation (153 vs. 47 days, p = 0.001) and symptomatic duration (710 vs. 148 days, p = 0.001) were longer. Conclusions: RT‐QuIC is a sensitive yet imperfect measure necessitating incorporation of other test results when evaluating patients with suspected prion disease. Patients with negative RT‐QuIC had lower markers of neuronal damage (CSF total tau and protein 14‐3‐3) and longer symptomatic duration of disease, suggesting that false negative RT‐QuIC testing associates with a more indolent course. [ABSTRACT FROM AUTHOR]

Background: An unmet clinical need in Parkinson's disease (PD) is to identify biomarkers for diagnosis, preferably in peripherally accessible tissues such as skin. Immunohistochemical studies have detected pathological α‐synuclein (αSyn) in skin biopsies from PD patients albeit sensitivity needs to be improved. Objective: Our study provides the ultrasensitive detection of pathological αSyn present in the skin of PD patients, and thus, pathological αSyn in skin could be a potential biomarker for PD. Methods: The real‐time quaking‐induced conversion assay was used to detect pathological αSyn present in human skin tissues. Further, we optimized this ultra‐sensitive and specific assay for both frozen and formalin‐fixed paraffin‐embedded sections of skin tissues. We determined the seeding kinetics of the αSyn present in the skin from autopsied subjects consisting of frozen skin tissues from 25 PD and 25 controls and formalin‐fixed paraffin‐embedded skin sections from 12 PD and 12 controls. Results: In a blinded study of skin tissues from autopsied subjects, we correctly identified 24/25 PD and 24/25 controls using frozen skin tissues (96% sensitivity and 96% specificity) compared to 9/12 PD and 10/12 controls using formalin‐fixed paraffin‐embedded skin sections (75% sensitivity and 83% specificity). Conclusions: Our blinded study results clearly demonstrate the feasibility of using skin tissues for clinical diagnosis of PD by detecting pathological αSyn. Moreover, this peripheral biomarker discovery study may have broader translational value in detecting misfolded proteins in skin samples as a longitudinal progression marker. © 2020 International Parkinson and Movement Disorder Society [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Background: alpha‐synuclein co‐pathology may influence Alzheimer's disease (AD) phenotype but its presence has never been tested in large series of patients. Real‐time quaking‐induced conversion (RT‐QuIC) recently provided evidence of alpha‐synuclein seeding activity in CSF and olfactory mucosa of patients with alpha‐synucleinopathies Method: a preliminary set of 82 AD patients (mean age 70,3 + 7.3 years) underwent CSF analyses for Tau, P‐tau and A‐beta amyloid and an extensive cognitive, behavioral and motor assessment. RT‐QuiC for alpha‐synuclein was performed on CSF samples. Result: alpha synuclein RT‐QuIC resulted positive in thirty‐nine out of 82 AD patients (47.6%). AD patients with positive alpha‐synuclein were comparable for age and disease severity those negative to RT‐QuIC assay including Tau/P‐Tau and A‐beta CSF markers in cross‐sectional analyses but exhibited higher prevalence of parkinsonism (p=0.02). Conclusion: these preliminary findings indicate that alpha‐synuclein co‐pathology might be detected in more than one third of Alzheimer's disease patients. These findings might potentially explain the clinical variability of AD including age at onset, progression and response to treatment observed in clinical series. Larger longitudinal studies are warranted to confirm and extend these findings. [ABSTRACT FROM AUTHOR]

Background: Evidence suggests that skin represents a suitable matrix for demonstrating α‐synuclein oligomers as a diagnostic biomarker for Lewy body disease. Objective: The objective of this study was to evaluate the diagnostic performance of skin α‐syn real‐time quaking‐induced conversion assay in patients with Lewy body disease. Methods: We analyzed skin punches taken in vitam (n = 69) or postmortem (n = 49) from patients with PD, dementia with Lew bodies (DLB), incidental Lewy body pathology, and neurological controls. Seventy‐nine patients underwent both CSF and skin α‐synuclein real‐time quaking‐induced conversion assay. Results: Overall, the skin α‐synuclein real‐time quaking‐induced conversion assay distinguished Lewy body disease patients with 94.1% accuracy (sensitivity, 89.2%; specificity, 96.3%). Assay sensitivity reached 94.1% in the 17 Lewy body disease patients analyzed in the cervical region. In patients with both CSF and skin samples, the 2 real‐time quaking‐induced conversion assay protocols yielded similar diagnostic accuracy (skin, 97.5%; CSF, 98.7%). Conclusion: Skin punch biopsies might represent a valid and convenient alternative to CSF analysis to demonstrate Lew body‐related α‐synuclein deposition in patients with Lewy body disease. © 2021 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society [ABSTRACT FROM AUTHOR]

We applied RT‐QuIC assay to detect α‐synuclein aggregates in cerebrospinal fluid (CSF) of patients with suspected Creutzfeldt–Jakob disease who had a neuropathological diagnosis of dementia with Lewy bodies (DLB) (n = 7), other neurodegenerative diseases with α‐synuclein mixed pathology (n = 20), or without Lewy‐related pathology (n = 49). The test had a sensitivity of 92.9% and specificity of 95.9% in distinguishing α‐synucleinopathies from non‐α‐synucleinopathies. When performed in the CSF of patients with DLB (n = 36), RT‐QuIC was positive in 17/20 with probable DLB, 0/6 with possible DLB, and 0/10 with Alzheimer disease. These results indicate that RT‐QuIC for α‐synuclein is an accurate test for DLB diagnosis. [ABSTRACT FROM AUTHOR]

Background: Leucine‐rich kinase 2 (LRRK2)‐linked Parkinson's disease (PD) is clinically indistinguishable from idiopathic PD (IPD). A pleiotropic neuropathology has been recognized but the majority of studies in LRRK2 p.G2019S patients reveal Lewy‐type synucleinopathy as its principal histological substrate. To date no in vivo biomarkers of synucleinopathy have been found in LRRK2 mutation carriers. Objectives: We used real‐time quaking‐induced conversion (RT‐QuIC) technique to assess the presence of alpha‐synuclein (a‐syn) aggregates in cerebrospinal fluid (CSF) of LRRK2 p.G2019S carriers. Methods: CSF samples of 51 subjects were analyzed: 15 LRRK2 p.G2019S PD, 10 IPD, 16 LRRK2 p.G2019S nonmanifesting carriers (NMC) and 10 healthy controls. The presence of parkinsonism and prodromal symptoms was assessed in all study subjects. Results: Forty percent (n = 6) LRRK2‐PD, and 18.8% (n = 3) LRRK2‐NMC had a positive a‐syn RT‐QuIC response. RT‐QuIC detected IPD with 90% sensitivity and 80% specificity. No clinical differences were detected between LRRK2‐PD patients with positive and negative RT‐QuIC. A positive RT‐QuIC result in LRRK2‐NMC occurred in a higher proportion of subjects meeting the Movement Disorder Society research criteria for prodromal PD. Interpretation: RT‐QuIC detects a‐syn aggregation in CSF in a significant number of patients with LRRK2‐PD, but less frequently than in IPD. A small percentage of LRRK2‐NMC tested also positive. If appropriately validated in long‐term studies with large number of mutation carriers, and hopefully, postmortem or in vivo confirmation of histopathology, RT‐QuIC could contribute to the selection of candidates to receive disease modifying drugs, in particular treatments targeting a‐syn deposition. [ABSTRACT FROM AUTHOR]

Objective: The detection of prion seeding activity in CSF and olfactory mucosal brushings using real‐time quaking‐induced conversion assays allows highly accurate clinical diagnosis of sporadic Creutzfeldt–Jakob disease. To gauge transmission risks associated with these biospecimens and their testing, we have bioassayed prion infectivity levels in patients' brain tissue, nasal brushings, and CSF, and assessed the pathogenicity of amplified products of real‐time quaking‐induced conversion assays seeded with Creutzfeldt–Jakob disease prions. Methods: We obtained olfactory mucosal brushings and CSF from patients with a final diagnosis of sporadic Creutzfeldt–Jakob disease subtype MM1 (n = 3). Samples were inoculated intracerebrally into Tg66 transgenic mice that overexpress the homologous human 129M prion protein. The mice were evaluated for clinical, neuropathological, and biochemical evidence of prion infection. Results: Patients' brain tissue at 102 to 105 fold dilutions affected 47/48 Tg66 mice. In contrast, maximum acutely tolerable doses of insoluble pellets from their olfactory mucosa brushings caused evidence of prion disease in only 4/28 inoculated mice, and no effects were seen with 10‐fold dilutions. No clinical prion disease was observed in mice inoculated with antemortem CSF samples or prion‐seeded real‐time quaking‐induced conversion assay products. Interpretation: Pellets from patients' olfactory mucosa brushings had ≥10,000‐fold lower infectivity per unit volume than brain tissue, while CSF lacked detectable infectivity. Nonetheless, the results suggest that appropriate precautions may be warranted in surgical interventions involving the olfactory areas. The lack of pathogenic infectivity in the real‐time quaking‐induced conversion assay products provides evidence that the assay does not replicate biohazardous prions in vitro. [ABSTRACT FROM AUTHOR]

Background: Skin biopsy is a potential tool for the premortem confirmation of an α‐synucleinopathy. Objective: The aim was to assess the aggregation assay real‐time quaking‐induced conversion (RT‐QuIC) of skin biopsy lysates to confirm isolated rapid eye movement sleep behavior disorder (iRBD) as an α‐synucleinopathy. Methods: Skin biopsies of patients with iRBD, Parkinson's disease (PD), and controls were analyzed using RT‐QuIC and immunohistochemical detection of phospho‐α‐synuclein. Results: α‐Synuclein aggregation was detected in 97.4% of iRBD patients (78.4% of iRBD biopsies), 87.2% of PD patients (70% of PD biopsies), and 13% of controls (7.9% of control biopsies), with a higher seeding activity in iRBD compared to PD. RT‐QuIC was more sensitive but less specific than immunohistochemistry. Conclusions: Dermal RT‐QuIC is a sensitive method to detect α‐synuclein aggregation in iRBD, and high seeding activity may indicate a strong involvement of dermal nerve fibers in these patients. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Background: Misfolded α‐synuclein in Parkinson's disease (PD) and dementia with Lewy bodies (DLB) can be detected using the real‐time quaking‐induced conversion (RT‐QuIC) technique in cerebrospinal fluid (CSF). Objectives: The objectives are (1) to examine misfolded CSF α‐synuclein incidence, and (2) to compare clinical presentation, sports history, brain volumes, and RT‐QuIC α‐synuclein positivity in former athletes. Methods: Thirty former athletes with magnetic resonance imaging, neuropsychological testing, and CSF analyzed for phosphorylated tau 181 (p‐tau), total tau (t‐tau), amyloid‐β 42 (Aβ42), and neurofilament light chain (NfL). CSF α‐synuclein was detected using RT‐QuIC. Results: Six (20%) former athletes were α‐synuclein positive. α‐Synuclein positive athletes were similar to α‐synuclein negative athletes on demographics, sports history, clinical features, CSF p‐tau, t‐tau, Aβ42, and NfL; however, had lower grey matter volumes in the right inferior orbitofrontal, right anterior insula and right olfactory cortices. Conclusions: α‐Synuclein RT‐QuIC analysis of CSF may be useful as a prodromal biofluid marker of PD and DLB. © 2023 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Background: Aggregation of α-synuclein (oligomeric α-syn) has been considered as the pathological hallmark of Parkinson's disease (PD) and multiple system atrophy (MSA). Studies showed oligomeric α-syn/total α-syn ratio was increased in the saliva of patients with PD, suggesting that seeding activity of salivary oligomeric α-syn may be a novel biomarker for the diagnosis of PD and MSA.Objective: This study aimed to evaluate the diagnostic value of salivary α-syn seeding activity in patients with PD and MSA.Methods: A total of 75 patients with PD, 18 patients with MSA, and 36 nonneurodegenerative healthy control subjects underwent salivary α-syn real-time quaking-induced conversion (RT-QuIC) assay.Results: Salivary α-syn RT-QuIC assay distinguished patients with PD with 76.0% sensitivity (95% confidence interval [CI], 66.1-85.9) and 94.4% specificity (95% CI, 86.6-100.0). RT-QuIC assay sensitivity reached 61.1% (95% CI, 36.2-86.1) in patients with MSA. No significant differences were observed in the diameter of salivary α-syn fibrils examined by electron microscopy and in thioflavin T fluorescence intensity of salivary α-syn fibrils detected by RT-QuIC assay between patients with PD and MSA. Notably, the lag phase of RT-QuIC assay from patients with PD was significantly shorter than that of patients with MSA, which might be clinically applicable to the discrimination between PD and MSA.Conclusions: Salivary α-syn seeding activity may serve as a novel biomarker for the clinical diagnosis of PD and MSA.© 2022 International Parkinson and Movement Disorder Society © 2022 International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Background: The neuropathology of Parkinson's disease (PD) associated with leucine‐rich repeat kinase 2 (LRRK2) mutations (LRRK2‐PD) is heterogeneous and varies with the type of mutation. There are only a few studies evaluating seeding aggregation assays to detect α‐synuclein (α‐syn) in patients with LRRK2‐PD. Objective: We aimed to investigate whether α‐syn real‐time quaking induced conversion (RT‐QuIC) is a sensitive biomarker of synucleinopathy in LRRK2‐PD. Methods: We studied α‐syn RT‐QuIC in brain tissue and postmortem ventricular cerebrospinal fluid (CSF) of LRRK2‐PD cases with and without Lewy‐type pathology. Results: The accuracy of α‐syn RT‐QuIC in substantia nigra and CSF samples of patients with LRRK2‐PD was 100%. The test also obtained 100% sensitivity to detect misfolded α‐syn in substantia nigra of cases with idiopathic PD and was negative in the substantia nigra of all the control brains without Lewy‐type pathology. Conclusions: Substantia nigra and ventricular CSF RT‐QuIC discriminates with high sensitivity and specificity LRRK2 cases with Lewy‐type pathology from those without it. RT‐QuIC assay could be of particular interest in the selection of cases for clinical trials in this genetic form of PD. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

Background: Real‐time quaking‐induced conversion (RT‐QuIC) and protein misfolding cyclic amplification (PMCA) have been developed to detect minute amounts of amyloidogenic proteins via amplification techniques and have been used to detect misfolded α‐synuclein (αSyn) aggregates in the cerebrospinal fluid (CSF) and other source materials of patients with Parkinson's Disease and other synucleinopathies. Objectives: The aim of this systematic review and meta‐analysis was to evaluate the diagnostic accuracy of αSyn seed amplification assays (αSyn‐SAAs), including RT‐QuIC and PMCA, using CSF as source material to differentiate synucleinopathies from controls. Methods: The electronic MEDLINE database PubMed was searched for relevant articles published until June 30, 2022. Study quality assessment was performed using the QUADAS‐2 toolbox. A random effects bivariate model was exploited for data synthesis. Results: Our systematic review identified 27 eligible studies according to the predefined inclusion criteria, of which 22 were included in the final analysis. Overall, 1855 patients with synucleinopathies and 1378 non‐synucleinopathies as control subjects were included in the meta‐analysis. The pooled sensitivity and specificity to differentiate synucleinopathies from controls with αSyn‐SAA were 0.88 (95% CI, 0.82–0.93) and 0.95 (95% CI, 0.92–0.97), respectively. Evaluating the diagnostic performance of RT‐QuIC in a subgroup analysis for the detection of patients with multiple system atrophy the pooled sensitivity decreased to 0.30 (95% CI, 0.11–0.59). Conclusions: While our study clearly demonstrated a high diagnostic performance of RT‐QuIC and PMCA for differentiating synucleinopathies with Lewy bodies from controls, results for the diagnosis of multiple system atrophy were less robust. [ABSTRACT FROM AUTHOR]

Background: The recent development of IQ‐CSF, the second generation of Real Time Quaking‐Induced Conversion (RT‐QuIC) using cerebrospinal fluid (CSF), for the diagnosis of Creutzfeldt Jakob Disease (CJD) represents a major diagnostic advance in the field. High accuracy results have been reported with encouraging reproducibility among different centers. The aim of this research is to report the pilot use of IQ‐CSF in a small cohort of Brazilian patients with possible or probable CJD, implementing a reference center in the country. Method: We stored CSF samples from patients with possible, probable or genetic CJD (one case) during the timeframe of December 2016 through June 2018. All CSF samples were processed according to standardized protocols without access to the clinical data. Eight patients presented to our team with a rapidly progressive dementia and typical neurological signs of CJD. We used CSF samples from 8 patients with other neurological conditions as controls. Result: 7 out of 9 suspected cases had positive tests; among controls, there was 1 false‐positive (a CSF sample from a 4‐year‐old child with leukemia under treatment). The occurrence of a false positive in one of the negative control samples raises the possibility of the presence of interfering components in the CSF sample from patients with non‐neurodegenerative pathologies. Conclusion: Our pilot results illustrate the feasibility of having CJD CSF samples tested in Brazilian centers and highlight the importance of inter‐institutional collaboration to pursue a higher diagnostic accuracy in CJD in Brazil and Latin American. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Presenting symptoms of sporadic Creutzfeldt–Jakob disease (sCJD) are variable, and as imaging and EEG may be normal in the early to middle stages of the disease process, serial testing is vital when there is clinical suspicion for sCJD. We present a case of probable Heidenhain variant of sCJD (HvCJD) with notable rapid progression. A 72‐year‐old woman presented with neurological decline following new–onset visual changes. Over the course of 3 weeks, she developed ataxia followed by paranoia, memory impairment, and visual hallucinations. An extensive workup from 1 week prior at an outside hospital was unrevealing and included two magnetic resonance imaging (MRI) studies read as normal and an EEG without periodic sharp wave complexes. Repeat of imaging at our hospital showed cortical restricted diffusion in the right occipital lobe. In combination with new periodic sharp wave complexes visualized on prolonged EEG, concern was raised for sCJD. Palliative care was consulted early in the hospitalization, and the patient was transitioned to comfort care and discharged 3 days after admission. She declined quickly and passed away at home within a week, one day before her send out CSF sample resulted with a positive real‐time quaking‐induced conversion (RT‐QuiC) and markedly elevated T‐tau protein and 14‐3‐3 gamma. As there is no treatment for this fatal disease, palliative engagement and discussion of goals of care in cases of CJD is critical in providing compassionate care for the patient and their family. High clinical suspicion warrants discussion of comfort care measures even prior to confirmation with RT‐QuiC. [ABSTRACT FROM AUTHOR]

The lack of a dedicated surveillance program for prion disease, particularly in low‐ and middle‐income countries (LMICs), has hindered the global effort to address this public health threat. Although cerebrospinal fluid (CSF) Real‐time quaking‐induced conversion (RT‐QuIC) is considered the most reliable test for sporadic Creutzfeldt‐Jakob disease (sCJD), its availability in LMICs is limited because of its cost and technical difficulty in generating the recombinant prion protein substrate (recPrP). This study aimed to evaluate the performance of RT‐QuIC with recPrP produced in‐house through a small‐scale method—that is, the application of reusable prepacked chromatography columns and subsequent dialysis. Here, CSF specimens from patients suspected of having prion disease were consecutively collected and stored between October 2015 and January 2023. Electronic medical record data were reviewed to clinically classify participants as probable sCJD or non‐sCJD. CSF RT‐QuIC was performed using in‐house recPrP. Its specificity and sensitivity for diagnosing probable sCJD were reported, along with details of other clinical data and investigations. We found that among 39 eligible participants, with a median (interquartile range) age of 64 (56–70) years and 16 (41%) female, 13 had probable sCJD and the remaining 26 unequivocally suffered from nonprion disorders. Magnetic resonance imaging and electroencephalogram were suggestive of sCJD in 100% (13/13) and 46.2% (6/13) of sCJD participants, respectively. RT‐QuIC was positive in 12/13 sCJD participants (sensitivity 0.92, 95% confidence interval [CI] 0.67–0.99) and negative in all non‐sCJD participants (specificity 1.00, 95% CI 0.87–1.00). CSF tau/p‐tau ratio showed sensitivity and specificity of 0.62–1.0 and 0.85–1.0, respectively. In summary, RT‐QuIC using recPrP generated through a small‐scale workflow demonstrated great performance in detecting sCJD. Given its performance results along with its low cost, this technique could feasibly be implemented in LMICs and potentially be the first step toward establishing local prion disease surveillance programs. [ABSTRACT FROM AUTHOR]

Objectives: Currently, the exact reasons why different α-synucleinopathies exhibit variable pathologies and phenotypes are still unknown. A potential explanation may be the existence of distinctive α-synuclein conformers or strains. Here, we intend to analyze the seeding activity of dementia with Lewy bodies (DLB) and Parkinson's disease (PD) brain-derived α-synuclein seeds by real-time quaking-induced conversion (RT-QuIC) and to investigate the structure and morphology of the α-synuclein aggregates generated by RT-QuIC.Methods: A misfolded α-synuclein-enriched brain fraction from frontal cortex and substantia nigra pars compacta tissue, isolated by several filtration and centrifugation steps, was subjected to α-synuclein/RT-QuIC analysis. Our study included neuropathologically well-characterized cases with DLB, PD, and controls (Ctrl). Biochemical and morphological analyses of RT-QuIC products were conducted by western blot, dot blot analysis, Raman spectroscopy, atomic force microscopy, and transmission electron microscopy.Results: Independently from the brain region, we observed different seeding kinetics of α-synuclein in the RT-QuIC in patients with DLB compared to PD and Ctrl. Biochemical characterization of the RT-QuIC product indicated the generation of a proteinase K-resistant and fibrillary α-synuclein species in DLB-seeded reactions, whereas PD and control seeds failed in the conversion of wild-type α-synuclein substrate.Interpretation: Structural variances of α-synuclein seeding kinetics and products in DLB and PD indicated, for the first time, the existence of different α-synuclein strains in these groups. Therefore, our study contributes to a better understanding of the clinical heterogeneity among α-synucleinopathies, offers an opportunity for a specific diagnosis, and opens new avenues for the future development of strain-specific therapies. Ann Neurol 2019;85:691-703. [ABSTRACT FROM AUTHOR]

Background: Identification of a peripheral biomarker is a major roadblock in the diagnosis of PD. Immunohistological identification of p-serine 129 α-synuclein in the submandibular gland tissues of PD patients has been recently reported.Objective: We report on a proof-of-principle study for using an ultra-sensitive and specific, real-time quaking-induced conversion assay to detect pathological α-synuclein in the submandibular gland tissues of PD patients.Methods: The α-synuclein real-time quaking-induced conversion assay was used to detect and quantify pathological α-synuclein levels in PD, incidental Lewy body disease, and control submandibular gland tissues as well as in formalin-fixed paraffin-embedded sections.Results: We determined the quantitative seeding kinetics of pathological α-synuclein present in submandibular gland tissues from autopsied subjects using the α-synuclein real-time quaking-induced conversion assay. A total of 32 cases comprising 13 PD, 3 incidental Lewy body disease, and 16 controls showed 100% sensitivity and 94% specificity. Interestingly, both PD and incidental Lewy body disease tissues showed 100% concordance for elevated levels of pathological α-synuclein seeding activity compared to control tissues. End-point dilution kinetic analyses revealed that the submandibular gland had a wide dynamic range of pathological α-synuclein seeding activity.Conclusions: Our results are the first to demonstrate the utility of using the real-time quaking-induced conversion assay on peripherally accessible submandibular gland tissues and formalin-fixed paraffin-embedded tissue sections to detect PD-related pathological changes with high sensitivity and specificity. Additionally, the detection of seeding activity from incidental Lewy body disease cases containing immunohistochemically undetected pathological α-synuclein demonstrates the α-synuclein real-time quaking-induced conversion assay's potential utility for identifying prodromal PD in submandibular gland tissues. © 2019 International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]

A reliable biomarker is needed for accurate and early differentiation between Parkinson disease and the various forms of atypical parkinsonism. We used a novel real-time quaking-induced conversion (RT-QuIC) assay to detect α-synuclein (α-syn) aggregates in cerebrospinal fluid (CSF) of 118 patients with parkinsonism of uncertain clinical etiology and 52 controls. Diagnostic accuracy to distinguish α-synucleinopathies from non-α-synucleinopathies and controls was 84% (sensitivity = 75%, specificity = 94%, area under the curve = 0.84, 95% confidence interval = 0.78-0.91, p < 0.0001, positive predictive value = 93%). CSF α-syn RT-QuIC could be a useful diagnostic tool to help clinicians differentiate α-synucleinopathies from other forms of parkinsonism when the clinical picture is uncertain. Ann Neurol 2019;85:777-781. [ABSTRACT FROM AUTHOR]

Background: There are no systematic reviews of cerebrospinal fluid and blood biomarkers for sporadic Creutzfeldt–Jakob disease (sCJD) in specialized care settings that compare diagnostic accuracies in a network meta‐analysis (NMA). Methods: We searched Medline, Embase, and Cochrane Library for diagnostic studies of sCJD biomarkers. Studies had to use established diagnostic criteria for sCJD and for diseases in the non‐CJD groups, which had to represent a consecutive population of patients suspected as a CJD case, as reference standard. Risk of bias was assessed with QUADAS‐2. We conducted individual biomarker meta‐analyses with generalized bivariate models. To investigate heterogeneity, we performed subgroup analyses based on QUADAS‐2 quality and clinical criteria. For the NMA, we applied a Bayesian beta‐binomial ANOVA model. The study protocol was registered at PROSPERO (CRD42019118830). Results: Of 2976 publications screened, we included 16 studies, which investigated 14–3‐3β (n = 13), 14–3‐3γ (n = 3), neurofilament light chain (NfL, n = 1), neuron‐specific enolase (n = 1), p‐tau181/t‐tau ratio (n = 2), RT‐QuIC (n = 7), S100B (n = 3), t‐tau (n = 12), and t‐tau/Aβ42 ratio (n = 1). Excluded diagnostic studies had strong limitations in study design. In the NMA, RT‐QuIC (0.91; 95% CI [0.83, 0.95]) and NfL (0.93 [0.78, 0.99]) were the most sensitive biomarkers for the diagnosis of definite, probable, and possible sCJD cases. RT‐QuIC was the most specific biomarker (0.97 [0.89, 1.00]). Heterogeneity in accuracy estimates was high between studies. Conclusions: We identified RT‐QuIC as the most accurate biomarker, partially confirming currently applied diagnostic criteria. The shortcomings identified in many diagnostic studies for sCJD biomarkers need to be addressed in future studies. [ABSTRACT FROM AUTHOR]

Objective: To assess the proportion of clinically diagnosed MM2‐type sporadic Creutzfeldt–Jakob disease (sCJD) in a Chinese cohort, describe the clinical features of MM2‐cortical (MM2C) and MM2‐thalamic (MM2T) type sCJD to improve the early detection of MM2‐type sCJD. Methods: A total of 209 patients with sCJD admitted to the Xuanwu Hospital between February 2012 and August 2022 were reviewed. The patients were classified into probable MM2C, MM2T‐type sCJD, and other types of sCJD according to current clinical diagnostic criteria. Clinical and ancillary data were compared between the groups. Results: Fifty‐one (24.4%) patients were clinically diagnosed with MM2‐type sCJD, of which 44 were diagnosed with MM2C‐type sCJD and 7 with MM2T‐type sCJD. In the absence of RT‐QuIC, 27 (61.3%) patients of MM2C‐type sCJD did not meet the US CDC sCJD criteria for possible sCJD on admission, even though the mean period from onset to admission was 6.0 months. However, all of these patients had cortical hyperintensity on DWI. Compared to the other types of sCJD, MM2C‐type sCJD was associated with slower disease progression and the absence of the typical clinical features of sCJD; the MM2T‐type sCJD group had a higher proportion of males, earlier age of onset, longer duration of disease, and a higher incidence of bilateral thalamic hypometabolism/hypoperfusion. Interpretation: In the absence of multiple typical sCJD symptoms within 6 months, the presence of cortical hyperintensity on DWI should raise concerns for MM2C‐type sCJD after excluding other etiologies. Bilateral thalamic hypometabolism/hypoperfusion may be more helpful in the clinical diagnosis of MM2T‐type sCJD. [ABSTRACT FROM AUTHOR]

Objective: Current cerebrospinal fluid (CSF) tests for sporadic Creutzfeldt-Jakob disease (sCJD) are based on the detection of surrogate markers of neuronal damage such as CSF 14-3-3, which are not specific for sCJD. A number of prion protein conversion assays have been developed, including real time quaking-induced conversion (RT-QuIC). The objective of this study is to investigate whether CSF RT-QuIC analysis could be used as a diagnostic test in sCJD. Methods: An exploratory study was undertaken that analyzed 108 CSF samples from patients with neuropathologically confirmed sCJD or from control patients. Of the 108 CSF samples, 56 were from sCJD patients (30 female, 26 male; aged 31-84 years; mean age, 62.3 ± 13.5 years), and 52 were from control patients (26 female, 26 male; aged 43-84 years; mean age, 67.8 ± 10.4 years). A confirmatory group of 118 patients was subsequently examined that consisted of 67 cases of neuropathologically confirmed sCJD (33 female, 34 male; aged 39-82 years; mean age, 67.5 ± 9.0 years) and 51 control cases (26 female, 25 male; aged 36-87 years; mean age, 63.5 ± 11.6 years). Results: The exploratory study showed that RT-QuIC analysis had a sensitivity of 91% and a specificity of 98% for the diagnosis of sCJD. These results were confirmed in the confirmatory study, which showed that CSF RT-QuIC analysis had a sensitivity and specificity of 87% and 100%, respectively. Interpretation: This study shows that CSF RT-QuIC analysis has the potential to be a more specific diagnostic test for sCJD than current CSF tests. ANN NEUROL 2012;72:278-285. [ABSTRACT FROM AUTHOR]

The typical clinical manifestations of sporadic Creutzfeldt‐Jakob disease (sCJD) are rapid‐progressive dementia and myoclonus. However, the diagnosis of atypical sCJD can be challenging due to its wide phenotypic variations. We report an autopsy case of variably protease‐sensitive prionopathy (VPSPr) with Met/Met homogeneity at codon 129. An 81‐year‐old woman presented with memory loss without motor symptoms. Seventeen months after the onset, her spontaneous language production almost disappeared. Diffusion‐weighted images (DWI) showed hyperintensity in the cerebral cortex while electroencephalogram (EEG) showed nonspecific change. 14‐3‐3 protein and real‐time qualing‐induced conversion (RT‐QuIC) of cerebrospinal fluid were negative. She died at age 85, 3.5 years after the onset. Pathological investigation revealed spongiform change, severe neuronal loss, and gliosis in the cerebral cortex. Mild to moderate neuronal loss and gliosis were observed in the basal ganglia. PrP immunostaining revealed plaque‐like, dotlike, and synaptic structures in the cerebral cortex and small plaque‐like structures in the molecular layer of the cerebellum. Analysis of PRNP showed no pathogenic mutations, and Western blot examination revealed the lack of a diglycosylated band consistent with VPSPr. The present case, which is the first report on a VPSPr case in Japan, supports previously published evidence that VPSPr cases can present variable and nonspecific clinical presentations. Because a small number of VPSPr cases can show typical magnetic resonance imaging (MRI) change in sCJD. We should investigate the possibility of VPSPr in a differential diagnosis with atypical dementia that presented DWIs of high intensity in the cortex, even though 14‐3‐3 proteins and RT‐QuIC are both negative. In addition, VPSPr cases can take a longer clinical course compared to that of sCJD, and long‐term follow‐up is important. [ABSTRACT FROM AUTHOR]

Objective: Several prion amplification systems have been proposed for detection of prions in cerebrospinal fluid (CSF), most recently, the measurements of prion seeding activity with second-generation real-time quaking-induced conversion (RT-QuIC). The objective of this study was to investigate the diagnostic performance of the RT-QuIC prion test in the broad phenotypic spectrum of prion diseases.Methods: We performed CSF RT-QuIC testing in 2,141 patients who had rapidly progressive neurological disorders, determined diagnostic sensitivity and specificity in 272 cases that were autopsied, and evaluated the impact of mutations and polymorphisms in the PRNP gene, and type 1 or type 2 human prions on diagnostic performance.Results: The 98.5% diagnostic specificity and 92% sensitivity of CSF RT-QuIC in a blinded retrospective analysis matched the 100% specificity and 95% sensitivity of a blind prospective study. The CSF RT-QuIC differentiated 94% of cases of sporadic Creutzfeldt-Jakob disease (sCJD) MM1 from the sCJD MM2 phenotype, and 80% of sCJD VV2 from sCJD VV1. The mixed prion type 1-2 and cases heterozygous for codon 129 generated intermediate CSF RT-QuIC patterns, whereas genetic prion diseases revealed distinct profiles for each PRNP gene mutation.Interpretation: The diagnostic performance of the improved CSF RT-QuIC is superior to surrogate marker tests for prion diseases such as 14-3-3 and tau proteins, and together with PRNP gene sequencing the test allows the major prion subtypes to be differentiated in vivo. This differentiation facilitates prediction of the clinicopathological phenotype and duration of the disease-two important considerations for envisioned therapeutic interventions. ANN NEUROL 2017;81:79-92. [ABSTRACT FROM AUTHOR]

Background: Intravascular large B‐cell lymphoma (IVLBCL) is a rare extranodal lymphoma that is characterized by the selective growth of neoplastic cells in blood vessels, representing a potentially treatable cause of rapidly progressive dementia (RPD). Given its diverse clinical and instrumental presentation, it is often misdiagnosed with more common RPD causes, for example, Creutzfeldt‐Jakob disease (CJD) or vascular dementia. Methods: This study presents the clinical and histopathological characteristics of four IVLBCL cases that we diagnosed post‐mortem over 20 years among over 600 brain samples received as suspected CJD cases at our prion disease reference center. Results: Our patients exhibited various presenting symptoms, including behavioral disturbances, disorientation, and alertness fluctuations. The diagnostic tests performed at the time, including blood work, cerebrospinal fluid (CSF) analyses, electroencephalography, and neuroimaging, yielded nonspecific and occasionally misleading results. Consequently, the patients were repeatedly diagnosed as variably having CJD, epilepsy, vascular dementia, and encephalitis. The stored CSF samples of two patients tested negative at prion real‐time quaking‐induced conversion (RT‐QuIC), which we performed afterwards for research purposes. Neuropathological analysis revealed a differential involvement of various brain areas, with frontotemporal neocortices being the most affected. Conclusions: Our results confirm the significant clinical and instrumental heterogeneity of IVLBCL. Neuropathological evidence of the preferential involvement of frontotemporal neocortices, potentially conditioning the clinical phenotype, could be relevant to reach an early diagnosis. Finally, given the therapeutic implications of its misdiagnosis with CJD, we emphasize the utility of prion RT‐QuIC as a test for ruling out CJD in these patients. [ABSTRACT FROM AUTHOR]

Key Clinical Message: Creutzfeldt–Jakob disease is a neurodegenerative disorder caused by brain accumulation of a misfolded form of the cellular prion protein, whose diagnosis is challenging, particularly in early stages, due to the variability and nonspecificity of the clinical and radiological features. 18F‐fluorodeoxyglucose positron‐emitted tomography has the potential to be considered a crucial investigation in these patients, revealing metabolic abnormalities earlier than the conventional neuroimaging analysis. A 59‐year‐old man, the military officer, was referred to our Units for the onset of neurological symptoms rapidly evolving within a month, characterized by akinetic mutism, constructional apraxia, and disorders of spatial orientation. Brain 18F‐fluorodeoxyglucose (18F‐FDG) positron‐emitted tomography (PET)/CT depicted an asymmetric hypometabolism in the left fronto‐temporo‐parietal cortex, as well as in the left thalamus and the right cerebellar hemisphere, while the glucose metabolism appears to be preserved in the somatosensory cortex and the basal ganglia. Laboratory routine analyses, cerebrospinal fluid routine, infective tests, electroencephalography (EEG), and brain magnetic resonance (MR) were all unremarkable. A positive RT‐QuIC result on cerebro‐spinal fluid (CSF) was subsequently shown, without any pathogenic gene mutations and, therefore, the result was consistent with a diagnosis of sporadic Creutzfeld–Jacob disease. The clinical evolution was quickly unfavorable, and the patient died about 4 months after hospital admission. FDG PET/computed tomography (CT) has the potential to be considered a crucial investigation in these patients, documenting metabolic changes long time before other diagnostic investigations such as CSF, EEG, brain CT, and brain MR, thus suggesting a greater sensitivity of glucose metabolic evaluation in the early stage of the disease in question. [ABSTRACT FROM AUTHOR]

Prion real‐time quaking‐induced conversion (RT‐QuIC) is an ultrasensitive assay detecting pathological aggregates of misfolded prion protein in biospecimens. We studied 71 punch biopsy skin samples of 35 patients with Creutzfeldt–Jakob disease (CJD), including five assessed in vitam. The results confirmed the high value of skin prion RT‐QuIC for CJD diagnosis (89% sensitivity and 100% specificity) and support its use in clinical practice. Preliminary data based on a limited number of cases suggest that prion‐seeding activity in the skin varies according to the prion strain, being higher in sporadic CJD subtypes linked to the V2 strain (VV2 and MV2K) than in typical CJDMM1. [ABSTRACT FROM AUTHOR]

INTRODUCTION: Amyloid beta and tau pathology are the hallmarks of sporadic Alzheimer's disease (AD) and autosomal dominant AD (ADAD). However, Lewy body pathology (LBP) is found in ≈ 50% of AD and ADAD brains. METHODS: Using an α‐synuclein seed amplification assay (SAA) in cerebrospinal fluid (CSF) from asymptomatic (n = 26) and symptomatic (n = 27) ADAD mutation carriers, including 12 with known neuropathology, we investigated the timing of occurrence and prevalence of SAA positive reactivity in ADAD in vivo. RESULTS: No asymptomatic participant and only 11% (3/27) of the symptomatic patients tested SAA positive. Neuropathology revealed LBP in 10/12 cases, primarily affecting the amygdala or the olfactory areas. In the latter group, only the individual with diffuse LBP reaching the neocortex showed α‐synuclein seeding activity in CSF in vivo. DISCUSSION: Results suggest that in ADAD LBP occurs later than AD pathology and often as amygdala‐ or olfactory‐predominant LBP, for which CSF α‐synuclein SAA has low sensitivity. Highlights: Cerebrospinal fluid (CSF) real‐time quaking‐induced conversion (RT‐QuIC) detects misfolded α‐synuclein in ≈ 10% of symptomatic autosomal dominant Alzheimer's disease (ADAD) patients.CSF RT‐QuIC does not detect α‐synuclein seeding activity in asymptomatic mutation carriers.Lewy body pathology (LBP) in ADAD mainly occurs as olfactory only or amygdala‐predominant variants.LBP develops late in the disease course in ADAD.CSF α‐synuclein RT‐QuIC has low sensitivity for focal, low‐burden LBP. [ABSTRACT FROM AUTHOR]

Introduction: Multiple system atrophy (MSA) is a rapidly progressive neurodegenerative disorder characterized by the presence of glial cytoplasmic inclusions (GCIs) containing aggregated α‐synuclein (α‐Syn). Accurate diagnosis and monitoring of MSA present significant challenges, which can lead to potential misdiagnosis and inappropriate treatment. Biomarkers play a crucial role in improving the accuracy of MSA diagnosis, and phosphorylated α‐synuclein (p‐syn) has emerged as a promising biomarker for aiding in diagnosis and disease monitoring. Methods: A literature search was conducted on PubMed, Scopus, and Google Scholar using specific keywords and MeSH terms without imposing a time limit. Inclusion criteria comprised various study designs including experimental studies, case‐control studies, and cohort studies published only in English, while conference abstracts and unpublished sources were excluded. Results: Increased levels of p‐syn have been observed in various samples from MSA patients, such as red blood cells, cerebrospinal fluid, oral mucosal cells, skin, and colon biopsies, highlighting their diagnostic potential. The α‐Syn RT‐QuIC assay has shown sensitivity in diagnosing MSA and tracking its progression. Meta‐analyses and multicenter investigations have confirmed the diagnostic value of p‐syn in cerebrospinal fluid, demonstrating high specificity and sensitivity in distinguishing MSA from other neurodegenerative diseases. Moreover, combining p‐syn with other biomarkers has further improved the diagnostic accuracy of MSA. Conclusion: The p‐syn stands out as a promising biomarker for MSA. It is found in oligodendrocytes and shows a correlation with disease severity and progression. However, further research and validation studies are necessary to establish p‐syn as a reliable biomarker for MSA. If proven, p‐syn could significantly contribute to early diagnosis, disease monitoring, and assessing treatment response. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

We report the general autopsy findings of abnormal prion protein (PrP) deposits with their seeding activities, as assessed by the real‐time quaking‐induced conversion (RT‐QuIC) method, in a 72‐year‐old female patient with sporadic Creutzfeldt–Jakob disease (sCJD). At 68 years of age, she presented with gait disturbance and visual disorders. Electroencephalography showed periodic synchronous discharge. Myoclonus was also observed. A genetic test revealed that PRNP codon 129 was methionine/methionine (MM). She died of pneumonia three years and four months after disease onset, and a general autopsy was performed. The brain weighed 650 g and appeared markedly atrophic. Immunohistochemistry for PrP revealed synaptic PrP deposits and coarse PrP deposits in the cerebral cortices, basal ganglia, cerebellum, and brainstem. Western blot analysis identified type 1 proteinase‐K‐resistant PrP in frontal cortex samples. PrP deposits were also observed in systemic organs, including the femoral nerve, psoas major muscle, abdominal skin, adrenal medulla, zona reticularis of the adrenal gland, islet cells of the pancreas, and thyroid gland. The RT‐QuIC method revealed positive seeding activities in all examined organs, including the frontal cortex, femoral nerve, psoas major muscle, scalp, abdominal skin, adrenal gland, pancreas, and thyroid gland. The following 50% seeding dose (SD50) values were 9.5 (frontal cortex); 8 ± 0.53 (femoral nerve); 7 ± 0.53 (psoas major muscle); and 7.88 ± 0.17 (scalp). The SD50 values for the adrenal gland, dermis, pancreas, and thyroid gland were 6.12 ± 0.53, 5.25, 4.75, and 4.5, respectively. PrP deposits in general organs may be associated with long‐term disease duration. This case indicated the necessity for general autopsies in sCJD cases to establish strict infection control procedures for surgical treatment and to examine certain organs. [ABSTRACT FROM AUTHOR]

Human prion diseases, including Creutzfeldt‐Jakob disease (CJD), occur in sporadic, genetic, and acquired forms. Variant Creutzfeldt‐Jakob disease (vCJD) first reported in 1996 in the United Kingdom (UK), resulted from contamination of food with bovine spongiform encephalopathy. There is a concern that UK national surveillance mechanisms might miss some CJD cases (including vCJD), particularly in the older population where other neurodegenerative disorders are more prevalent. We developed a highly sensitive protocol for analysing autopsy brain tissue for the misfolded prion protein (PrPSc) associated with prion disease, which could be used to screen for prion disease in the elderly. Brain tissue samples from 331 donors to the Edinburgh Brain and Tissue Bank (EBTB), from 2005 to 2022, were analysed, using immunohistochemical analysis on fixed tissue, and five biochemical tests on frozen specimens from six brain regions, based on different principles for detecting PrPSc. An algorithm was established for classifying the biochemical results. To test the effectiveness of the protocol, several neuropathologically confirmed prion disease controls, including vCJD, were included and blinded in the study cohort. On unblinding, all the positive control cases had been correctly identified. No other cases tested positive; our analysis uncovered no overlooked prion disease cases. Our algorithm for classifying cases was effective for handling anomalous biochemical results. An overall analysis suggested that a reduced biochemical protocol employing only three of the five tests on only two brain tissue regions gave sufficient sensitivity and specificity. We conclude that this protocol may be useful as a UK‐wide screening programme for human prion disease in selected brains from autopsies in the elderly. Further improvements to the protocol were suggested by enhancements of the in vitro conversion assays made during the course of this study. [ABSTRACT FROM AUTHOR]

α‐Synuclein aggregation is a hallmark of Parkinson's disease and a promising biomarker for early detection and assessment of disease progression. The prospect of a molecular test for Parkinson's disease is materializing with the recent developments of detection methods based on amplification of synuclein seeds (e.g. RT‐QuIC or PMCA). Here we adapted single‐molecule counting methods for the detection of α‐synuclein aggregates in cerebrospinal fluid (CSF), using a simple 3D printed microscope. Single‐molecule methods enable to probe the early events in the amplification process used in RT‐QuIC and a precise counting of ThT‐positive aggregates. Importantly, the use of single‐molecule counting also allows a refined characterization of the samples and fingerprinting of the protein aggregates present in CSF of patients. The fingerprinting of size and reactivity of individual aggregate shows a unique signature for each PD patients compared to controls and may provide new insights on synucleinopathies in the future. [ABSTRACT FROM AUTHOR]

Background: The lack of a dedicated surveillance program for prion disease, particularly in low‐ and middle‐income countries (LMICs), has hindered the global effort to address this public health threat. Although CSF Real‐time quaking‐induced conversion (RT‐QuIC) is considered the most reliable test for sporadic Creutzfeldt‐Jakob disease (sCJD), its availability in LMICs is limited due to its cost and technical difficulty in generating the recombinant prion protein substrate (recPrP). This study aimed to evaluate the performance of RT‐QuIC with recPrP produced in‐house through a small‐scale method – i.e. the application of reusable pre‐packed chromatography columns and subsequent dialysis. Method: CSF specimens from patients suspected of having prion disease were consecutively collected and stored between October 2015 and November 2022. Those with available CSF for RT‐QuIC analysis were included in this study. Electronic medical record data were reviewed to clinically classified participants as probable sCJD or non‐sCJD. CSF RT‐QuIC was performed using in‐house recPrP and 2nd generation (improved‐QuIC) protocol (Orrú et al., 2015). Its specificity and sensitivity for diagnosing probable sCJD were reported, along with details of other clinical data and investigations. Result: Among 38 eligible participants, with a median (interquartile range) age of 64 (56.5‐70) years and 15 (39.5%) female, 12 had probable sCJD and the remaining 26 unequivocally suffered from non‐prion disorders. MRI and EEG were suggestive of sCJD in 100% (12/12) and 50% (6/12) of sCJD participants, respectively. PRNP gene sequencing was performed in all sCJD and 7 non‐sCJD participants, all of whom had MM genotype without mutation. RT‐QuIC was positive in 11/12 sCJD participants (sensitivity 0.92, 95% confidence interval (CI) 0.65‐0.99) and negative in all non‐sCJD participants (specificity 1.00, 95%CI 0.87‐1.00). CSF tau/p‐tau ratio showed sensitivity and specificity of 0.67‐1.0 and 0.85‐1.0, respectively. Conclusion: RT‐QuIC using recPrP generated through a small‐scale workflow demonstrated great performance in detecting sCJD. Given its performance results along with its low cost, this technique could feasibly be implemented in LMICs and potentially be the first step towards establishing local prion disease surveillance programs that will strengthen the global efforts. [ABSTRACT FROM AUTHOR]

BACKGROUND: The cis‐conformer of tau phosphorylated at threonine‐231 (cis‐pT231 tau) is hypothesized to contribute to tauopathies. PNT001 is a humanized, monoclonal antibody that recognizes cis‐pT231 tau. PNT001 was characterized to assess clinical development readiness. METHODS: Affinity and selectivity were assessed by surface plasmon resonance and enzyme‐linked immunosorbent assay. Immunohistochemistry (IHC) was performed with brain sections from human tauopathy patients and controls. Real‐time quaking‐induced conversion (RT‐QuIC) was used to assess whether PNT001 reduced tau seeds from Tg4510 transgenic mouse brain. Murine PNT001 was evaluated in vivo in the Tg4510 mouse. RESULTS: The affinity of PNT001 for a cis‐pT231 peptide was 0.3 to 3 nM. IHC revealed neurofibrillary tangle‐like structures in tauopathy patients with no detectable staining in controls. Incubation of Tg4510 brain homogenates with PNT001 lowered seeding in RT‐QuIC. Multiple endpoints were improved in the Tg4510 mouse. No adverse findings attributable to PNT001 were detected in Good Laboratory Practice safety studies. DISCUSSION: The data support clinical development of PNT001 in human tauopathies. [ABSTRACT FROM AUTHOR]

We have developed a novel real-time quaking-induced conversion RT-Qu IC-based assay to detect alpha-synuclein aggregation in brain and cerebrospinal fluid from dementia with Lewy bodies and Parkinson's disease patients. This assay can detect alpha-synuclein aggregation in Dementia with Lewy bodies and Parkinson's disease cerebrospinal fluid with sensitivities of 92% and 95%, respectively, and with an overall specificity of 100% when compared to Alzheimer and control cerebrospinal fluid. Patients with neuropathologically confirmed tauopathies (progressive supranuclear palsy; corticobasal degeneration) gave negative results. These results suggest that RT-QuiC analysis of cerebrospinal fluid is potentially useful for the early clinical assessment of patients with alpha-synucleinopathies. [ABSTRACT FROM AUTHOR]

We report a subtype of immune‐mediated encephalitis associated with COVID‐19, which closely mimics acute‐onset sporadic Creutzfeldt–Jakob disease. A 64‐year‐old man presented with confusion, aphasia, myoclonus, and a silent interstitial pneumonia. He tested positive for SARS‐CoV‐2. Cognition and myoclonus rapidly deteriorated, EEG evolved to generalized periodic discharges and brain MRI showed multiple cortical DWI hyperintensities. CSF analysis was normal, except for a positive 14‐3‐3 protein. RT‐QuIC analysis was negative. High levels of pro‐inflammatory cytokines were present in the CSF and serum. Treatment with steroids and intravenous immunoglobulins produced EEG and clinical improvement, with a good neurological outcome at a 6‐month follow‐up. [ABSTRACT FROM AUTHOR]

Objective: Comprehensively describe the phenotypic spectrum of sporadic fatal insomnia (sFI) to facilitate diagnosis and management of this rare and peculiar prion disorder.Methods: A survey among major prion disease reference centers in Europe identified 13 patients diagnosed with sFI in the past 20 years. We undertook a detailed analysis of clinical and histopathological features and the results of diagnostic investigations.Results: Mean age at onset was 43 years, and mean disease duration 30 months. Early clinical findings included psychiatric, sleep, and oculomotor disturbances, followed by cognitive decline and postural instability. In all tested patients, video-polysomnography demonstrated a severe reduction of total sleep time and/or a disorganized sleep. Cerebrospinal fluid (CSF) levels of proteins 14-3-3 and t-tau were unrevealing, the concentration of neurofilament light protein (NfL) was more consistently increased, and the real-time quaking-induced conversion assay (RT-QuIC) revealed a positive prion seeding activity in 60% of cases. Electroencephalography and magnetic resonance imaging showed nonspecific findings, whereas fluorodeoxyglucose positron emission tomography (FDG-PET) demonstrated a profound bilateral thalamic hypometabolism in 71% of cases. Molecular analyses revealed PrPSc type 2 and methionine homozygosity at PRNP codon 129 in all cases.Interpretation: sFI is a disease of young or middle-aged adults, which is difficult to reconcile with the hypothesis of a spontaneous etiology related to stochastic, age-related PrP misfolding. The combination of psychiatric and/or sleep-related symptoms with oculomotor abnormalities represents an early peculiar clinical feature of sFI to be valued in the differential diagnosis. Video-polysomnography, FDG-PET, and especially CSF prion RT-QuIC and NfL constitute the most promising supportive diagnostic tests in vivo. Ann Neurol 2018;84:347-360. [ABSTRACT FROM AUTHOR]

Parkinson's disease (PD) is a progressive neurodegenerative disease which is histologically characterized by loss of dopaminergic neurons in the substantia nigra and deposition of aggregated alpha‐synuclein (aSyn) in the brain. The detection of aSyn in well accessible fluids has been one of the central approaches in the development of biomarkers for PD. Recently, real‐time quaking‐induced conversion (RT‐QuIC) has been successfully adapted for use with aSyn seeds. Here, we systematically analysed parameters potentially impacting the reliability of this assay by using quantitative real‐time quaking‐induced conversion (qRT‐QuIC) with in vitro‐formed aSyn seeds. Seeds diluted in cerebrospinal fluid (CSF) accelerated the seeding reaction and slightly increased the sensitivity without affecting specificity. Repeated freeze–thaw cycles decreased the apparent lag times of seeds diluted in ddH2O but did not alter the seeding activity of seeds diluted in CSF. High levels of artificial contamination with blood resulted in prolonged apparent lag times, while sensitivity and specificity were unaffected. Altogether, qRT‐QuIC with aSyn seems to be robust concerning sensitivity and specificity in our model system, but quantitative interpretation might be limited under certain conditions. [ABSTRACT FROM AUTHOR]

Real-Time Quaking-Induced Conversion ( RT-Qu IC) testing of human cerebrospinal fluid ( CSF) is highly sensitive and specific in discriminating sporadic CJD patients from those without prion disease. Here, using CSF samples from 113 CJD and 64 non-prion disease patients, we provide the first direct and concurrent comparison of our improved RT-Qu IC assay to our previous assay, which is similar to those commonly used internationally for CJD diagnosis. This extended comparison demonstrated a ~21% increase in diagnostic sensitivity, a 2-day reduction in average detection time, and 100% specificity. [ABSTRACT FROM AUTHOR]

Background: PD diagnosis is based primarily on clinical criteria and can be inaccurate. Biological markers, such as α-synuclein aggregation, that reflect ongoing pathogenic processes may increase diagnosis accuracy and allow disease progression monitoring. Though α-synuclein aggregation assays have been published, reproducibility, standardization, and validation are key challenges for their development as clinical biomarkers.Objective: To cross-validate two α-synuclein seeding aggregation assays developed to detect pathogenic oligomeric α-synuclein species in CSF using samples from the same PD patients and healthy controls from the BioFIND cohort.Methods: CSF samples were tested by two independent laboratories in a blinded fashion. BioFIND features standardized biospecimen collection of clinically typical moderate PD patients and nondisease controls. α-synuclein aggregation was measured by protein misfolding cyclic amplification (Soto lab) and real-time quaking-induced conversion (Green lab). Results were analyzed by an independent statistician.Results: Measuring 105 PD and 79 healthy control CSF samples, these assays showed 92% concordance. The areas under the curve from receiver operating characteristic curve analysis for the diagnosis of PD versus healthy controls were 0.93 for protein misfolding cyclic amplification, 0.89 for real-time quaking-induced conversion, and 0.95 when considering only concordant assay results. Clinical characteristics of false-positive and -negative subjects were not different from true-negative and -positive subjects, respectively.Conclusions: These α-synuclein seeding aggregation assays are reliable and reproducible for PD diagnosis. Assay parameters did not correlate with clinical parameters, including disease severity or duration. This assay is highly accurate for PD diagnosis and may impact clinical practice and clinical trials. © 2019 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society. [ABSTRACT FROM AUTHOR]