Objective: To validate the International Parkinson Disease and Movement Disorders society (MDS) diagnostic criteria against a gold-standard expert clinical diagnosis and to compare concordance/accuracy of the MDS criteria to 1988 United Kingdom brain bank criteria. Background: In 2015, the MDS published the new clinical diagnostic criteria for Parkinson’s disease (PD). These criteria aimed to codify/reproduce the expert clinical diagnostic process, to help standardize diagnosis in research and clinical settings. Their accuracy compared to expert clinical diagnosis has not been tested. Design/Methods: From 8 centers, we recruited 626 patients with parkinsonism (434 with PD and 192 with other causes, diagnosed by an expert treating physician). A second, less experienced neurologist evaluated the presence/absence of each individual item from the diagnostic criteria. The overall accuracy/concordance rate, sensitivity, and specificity of diagnostic criteria were calculated. Results: Of 434 patients diagnosed with PD, 94.5% met MDS criteria for probable PD (i.e. a 5.5% false-negative rate). Of 192 non-PD patients, 88.5% were identified as non-PD by the criteria (i.e. a 11.5% false-positive rate). The overall accuracy for probable PD was 92.6%. For the clinically-established PD category, 59.3% of PD patients and only 1.6% of non-PD patients met MDS criteria. Compared to MDS probable PD criteria, the UK brain bank criteria had significantly lower sensitivity (89.2%, p=0.008), specificity (79.2%, p=0.018), and overall accuracy (86.4%, p Conclusions: The MDS criteria demonstrated high sensitivity and specificity compared to gold-standard expert diagnosis. To meet the unmet gap of specific diagnostic criteria for early PD, we suggest additional ‘Clinically-Established Early PD’ criteria to be used for clinical trials of early PD. Study Supported by: Michael J. Fox Foundation Disclosure: Dr. Postuma has received personal compensation for consulting, serving on a scientific advisory board, speaking, or other activities with Biotie, Roche/Prothena, Teva Neurosciences, Jazz Pharmaceuticals, Novartis Canada, Theranexus, GE HealthCare, . Dr. Poewe has nothing to disclose. Dr. Litvan has received personal compensation for serving onthe scientific steering committee of the Biotie/Parkinson Study Group clinical trial Dr. Lewis has nothing to disclose. Dr. Lang has received personal compensation for consulting, serving on a scientific advisory board, speaking, or other activities with Abbvie, Acorda, Avanir Pharmaceuticals, Biogen, Bristol Myers Squibb, Sun Pharma, Cipla, Intekrin, Merck, Medichem, Medtronic, Teva, UCB, and Sunovion, . Dr. Halliday has nothing to disclose. Dr. Goetz has nothing to disclose. Dr. Chan has nothing to disclose. Dr. Slow has nothing to disclose. Dr. Seppi has nothing to disclose. Dr. Schaeffer has nothing to disclose. Dr. Berg has received personal compensation for consulting, serving on a scientific advisory board, speaking, or other activities with UCB Pharma GmbH, Lundbeck, Prexton Therapeutics, GE-Healthcare. Dr. Rios Romenets has received personal compensation for consulting, serving on a scientific advisory board, speaking, or other activities with NIA, Genentech/Roche. Dr. Rios Romenets has received research support from NIA, Genentech/Roche. Dr. Mi has nothing to disclose. Dr. Maetzler has nothing to disclose. Dr. Li has nothing to disclose. Dr. Heim has nothing to disclose. Dr. Bledsoe has nothing to disclose.

Validation of the MDS Clinical Diagnostic Criteria for Parkinson’s Disease (S3.001)

Lysosomal proteases play a crucial role in maintaining cell homeostasis. Human cathepsin D manages protein turnover degrading misfolded and aggregated proteins and favors apoptosis in the case of proteostasis disruption. However, when cathepsin D regulation is affected, it can contribute to numerous disorders. The down-regulation of human cathepsin D is associated with neurodegenerative disorders, such as neuronal ceroid lipofuscinosis. On the other hand, its excessive levels outside lysosomes and the cell membrane lead to tumor growth, migration, invasion and angiogenesis. Therefore, targeting cathepsin D could provide significant diagnostic benefits and new avenues of therapy. Herein, we provide a brief overview of cathepsin D structure, regulation, function, and its role in the progression of many diseases and the therapeutic potentialities of natural and synthetic inhibitors and activators of this protease.

https://www.mdpi.com/1999-4923/13/6/837/pdf

Cathepsin D—Managing the Delicate Balance

ABSTRACT Parkinson disease (PD) is a neurodegenerative disorder characterized by the abnormal intracellular accumulation of SNCA/α-synuclein. While the exact mechanisms underlying SNCA pathology are not fully understood, increasing evidence suggests the involvement of autophagy as well as lysosomal deficiencies. Because CTSD (cathepsin D) has been proposed to be the major lysosomal protease involved in SNCA degradation, its deficiency has been linked to the presence of insoluble SNCA conformers in the brain of mice and humans as well as to the transcellular transmission of SNCA aggregates. We here postulate that SNCA degradation can be enhanced by the application of the recombinant human proform of CTSD (rHsCTSD). Our results reveal that rHsCTSD is efficiently endocytosed by neuronal cells, correctly targeted to lysosomes and matured to an enzymatically active protease. In dopaminergic neurons derived from induced pluripotent stem cells (iPSC) of PD patients harboring the A53T mutation within the SNCA gene, we confirm the reduction of insoluble SNCA after treatment with rHsCTSD. Moreover, we demonstrate a decrease of pathological SNCA conformers in the brain and within primary neurons of a ctsd-deficient mouse model after dosing with rHsCTSD. Boosting lysosomal CTSD activity not only enhanced SNCA clearance in human and murine neurons as well as tissue, but also restored endo-lysosome and autophagy function. Our findings indicate that CTSD is critical for SNCA clearance and function. Thus, enzyme replacement strategies utilizing CTSD may also be of therapeutic interest for the treatment of PD and other synucleinopathies aiming to decrease the SNCA burden. Abbreviations: aa: amino acid; SNCA/α-synuclein: synuclein alpha; APP: amyloid beta precursor protein; BBB: blood brain barrier; BF: basal forebrain; CBB: Coomassie Brilliant Blue; CLN: neuronal ceroid lipofuscinosis; CNL10: neuronal ceroid lipofuscinosis type 10; Corr.: corrected; CTSD: cathepsin D; CTSB: cathepsin B; DA: dopaminergic; DA-iPSn: induced pluripotent stem cell-derived dopaminergic neurons; dox: doxycycline; ERT: enzyme replacement therapy; Fx: fornix, GBA/β-glucocerebrosidase: glucosylceramidase beta; h: hour; HC: hippocampus; HT: hypothalamus; i.c.: intracranially; IF: immunofluorescence; iPSC: induced pluripotent stem cell; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LSDs: lysosomal storage disorders; MAPT: microtubule associated protein tau; M6P: mannose-6-phosphate; M6PR: mannose-6-phosphate receptor; MB: midbrain; mCTSD: mature form of CTSD; neurofil.: neurofilament; PD: Parkinson disease; proCTSD: proform of CTSD; PRNP: prion protein; RFU: relative fluorescence units; rHsCTSD: recombinant human proCTSD; SAPC: Saposin C; SIM: structured illumination microscopy; T-insol: Triton-insoluble; T-sol: Triton-soluble; TEM: transmission electron microscopy, TH: tyrosine hydroxylase; Thal: thalamus

https://www.tandfonline.com/doi/pdf/10.1080/15548627.2022.2045534?needAccess=true

Recombinant pro-CTSD (cathepsin D) enhances SNCA/α-Synuclein degradation in α-Synucleinopathy models

Proteostasis in Parkinson's disease: Recent development and possible implication in diagnosis and therapeutics

In Parkinson's disease, aggregates of α-synuclein within Lewy bodies and Lewy neurites represent neuropathological hallmarks. However, the cellular and molecular mechanisms triggering oligomeric and fibrillary α-synuclein aggregation are not fully understood. Recent evidence indicates that oxidative stress induced by metal ions and post-translational modifications such as phosphorylation, ubiquitination, nitration, glycation, and SUMOylation affect α-synuclein conformation along with its aggregation propensity and neurotoxic profiles. In addition, proteolytic cleavage of α-synuclein by specific proteases results in the formation of a broad spectrum of fragments with consecutively altered and not fully understood physiological and/or pathological properties. In the present review, we summarize the current knowledge on proteolytical α-synuclein cleavage by neurosin, calpain-1, cathepsin D, and matrix metalloproteinase-3 in health and disease. We also shed light on the contribution of the same enzymes to proteolytical processing of pathogenic proteins in Alzheimer's disease and report potential cross-disease mechanisms of pathogenic protein aggregation.

Proteolytic α-Synuclein Cleavage in Health and Disease.

To date, no reliable clinically applicable biomarker has been established for Parkinson's disease. Our results indicate that a long anticipated blood test for Parkinson's disease may be realized. Following the isolation of neuron-derived extracellular vesicles of Parkinson's disease patients and non-Parkinson's disease individuals, immunoblot analyses were performed to detect extracellular vesicle-derived α-synuclein. Pathological α-synuclein forms derived from neuronal extracellular vesicles could be detected under native conditions and were significantly increased in all individuals with Parkinson's disease and clearly distinguished disease from the non-disease state. By performing an α-synuclein seeding assay these soluble conformers could be amplified and seeding of pathological protein folding was demonstrated. Amplified α-synuclein conformers exhibited β-sheet-rich structures and a fibrillary appearance. Our study demonstrates that the detection of pathological α-synuclein conformers from neuron-derived extracellular vesicles from blood plasma samples has the potential to evolve into a blood-biomarker of Parkinson's disease that is still lacking so far. Moreover, the distribution of seeding-competent α-synuclein within blood exosomes sheds a new light of pathological disease mechanisms in neurodegenerative disorders.

Detection of neuron-derived pathological α-synuclein in blood.

Lysosomes are ubiquitous organelles with a fundamental role in maintaining cellular homeostasis by mediating degradation and recycling processes. Cathepsins are the most abundant lysosomal hydrolyses and are responsible for the bulk degradation of various substrates. A correct autophagic function is essential for neuronal survival, as most neurons are post-mitotic and thus susceptible to accumulate cellular components. Increasing evidence suggests a crucial role of the lysosome in neurodegeneration as a key regulator of aggregation-prone and disease-associated proteins, such as α-synuclein, β-amyloid and huntingtin. Particularly, alterations in lysosomal cathepsins CTSD, CTSB and CTSL can contribute to the pathogenesis of neurodegenerative diseases as seen for neuronal ceroid lipofuscinosis, synucleinopathies (Parkinson's disease, Dementia with Lewy Body and Multiple System Atrophy) as well as Alzheimer's and Huntington's disease. In this review, we provide an overview of recent evidence implicating CTSD, CTSB and CTSL in neurodegeneration, with a special focus on the role of these enzymes in α-synuclein metabolism. In addition, we summarize the potential role of lysosomal cathepsins as clinical biomarkers in neurodegenerative diseases and discuss potential therapeutic approaches by targeting lysosomal function. 

The role of lysosomal cathepsins in neurodegeneration: Mechanistic insights, diagnostic potential and therapeutic approaches.

Cathepsin D (CTSD) is a lysosomal protease important for the degradation of various substrates, including disease-associated proteins like α-synuclein (a-syn), amyloid precursor protein (APP) and tau, all of which tend to aggregate if not efficiently degraded. Hence, it is not surprising that genetic variants within the CTSD gene have been linked to neurodegenerative diseases, like Parkinson's and Alzheimer's disease (PD, AD), as well as the lysosomal storage disorder neuronal ceroid lipofuscinosis type-10 (NCL10). Although recent studies have shown the molecular dependence of substrate degradation via CTSD within autophagic pathways, only little is known about the precise role of lysosomal CTSD function in disease development. We here performed biochemical, cellular and structural analyses of eleven disease-causing CTSD point mutations found in genomic sequencing data of patients to understand their role in neurodegeneration. These CTSD variants were analyzed for cellular localization, maturation and enzymatic activity in overexpression analyses. Moreover, for PD-associated mutants, intracellular degradation of a-syn was monitored. In summary, our results suggest that NCL10-associated CTSD variants are significantly impaired in lysosomal maturation and enzymatic activity, whereas the AD- and PD-associated variants seemed rather unaffected, indicating normal maturation, and lysosomal presence. Interestingly, a PD-associated CTSD variant (A239V) exhibited increased enzymatic activity accompanied by enhanced a-syn degradation. By structural analyses of this mutant utilizing molecular dynamics simulation (MDS), we identified a structural change within a loop adjacent to the catalytic center leading to a higher flexibility and potentially accelerated substrate exchange rates. Our data sheds light onto the role of CTSD in disease development and helps to understand the structural regulation of enzymatic function, which could be utilized for targeted CTSD activation. Because of the degradative function of CTSD, this enzyme is especially interesting for therapeutic strategies tackling protein aggregates in neurodegenerative disorders.

/pdf/cathepsin-d-variants-associated-with-neurodegenerative-2g31o8qo61.pdf

Cathepsin D Variants Associated With Neurodegenerative Diseases Show Dysregulated Functionality and Modified α-Synuclein Degradation Properties


 To date, no reliable clinically applicable biomarker has been established for Parkinson’s disease (PD). Our results indicate that a long hoped blood test for Parkinson’s disease may be realized. We here assess the potential of pathological α-synuclein originating from neuron-derived exosomes from blood plasma as a possible biomarker. Following the isolation of neuron-derived exosomes from plasma of PD patients and non-PD individuals immunoblot analyses were performed to detect exosomal α-synuclein. Under native conditions significantly increased signals of disease-associated α-synuclein forms in neuron-derived exosomes were measured in all individuals with PD and clearly distinguished PD samples from controls. By performing a protein misfolding cyclic amplification assay these aggregates could be amplified and seeding could be demonstrated. Moreover, the aggregates exhibited β-sheet-rich structures and showed a fibrillary appearance. Our study demonstrates that the detection of pathological α-synuclein conformers from neuron-derived exosomes from plasma samples has the potential of a promising blood-biomarker of PD.

/pdf/a-blood-marker-for-parkinson-s-disease-neuronal-exosome-eumdc2fhfa.pdf

Josina Bunk

Papers

Detection of neuron-derived pathological α-synuclein in blood.

The role of lysosomal cathepsins in neurodegeneration: Mechanistic insights, diagnostic potential and therapeutic approaches.

Cathepsin D Variants Associated With Neurodegenerative Diseases Show Dysregulated Functionality and Modified α-Synuclein Degradation Properties

A blood marker for Parkinson’s Disease: Neuronal exosome-derived α-synuclein