[Diagnosis of acute coronary syndrome]. / Diagnose des akuten Koronarsyndroms.

Acute coronary syndrome is one of the most important differential diagnostic considerations in emergency medicine. It describes the constellation of newly occurring clinical symptoms, often accompanied by typical 12-lead ECG changes and the release of cardiac troponins. The spectrum includes unstable angina pectoris, non-ST-segment elevation myocardial infarction (NSTEMI), and ST-segment elevation myocardial infarction (STEMI). It is important to consistently carry out the diagnostic steps that are crucial for further therapeutic procedures to avoid delaying life-saving invasive coronary diagnostics, without losing sight of the diverse, sometimes time-critical differential diagnoses. Anamnesis and clinical examination form the basis of the further procedure. Further developments of biomarker assays with personalized limit values, new imaging modalities with ever higher resolution and faster imaging methods as well as advances in automated ECG analysis with integration of all findings through artificial intelligence will continue to offer many optimization options in the future diagnosis of acute coronary syndrome.

Analysis of cathepsin B and cathepsin L treatment to clear toxic lysosomal protein aggregates in neuronal ceroid lipofuscinosis.

Proteolysis mediated by lysosomal cathepsin proteases maintains a physiological flow in autophagy, phagocytosis and endocytosis. Neuronal Ceroid Lipofuscinosis (NCL) is a childhood neurodegenerative disorder characterized by disturbed autophagic flow and pathological accumulation of proteins. We demonstrated a therapeutic clearance of protein aggregates after dosing NCL10 mice with recombinant human pro-cathepsin-D. Prompted by these results and speculating that cathepsins may act in a redundant and in an hierarchical manner we envisaged that a treatment with human recombinant cysteine proteases pro-cathepsin-L (proCTSL) and pro-cathepsin-B (proCTSB) could similarly be used to induce protein degradation. Both enzymes were taken up by mannose 6-phosphate receptor- and LRP-receptor-mediated endocytosis and processed to the lysosomal mature cathepsins. In murine NCL10 astrocytes an abnormal increase in LAMP1 and saposin expression was revealed. Although proCTSB application did not improve this phenotype, proCTSL treatment led to reduced saposin-C levels in this model as well as in an acute brain slice model. Intracerebral dosing in a NCL10 mouse model revealed cellular and lysosomal uptake of both enzymes. Only proCTSL mildly reduced saposin-C levels and attenuated reactive astrogliosis. Application of both proteases did not improve weight loss and mortality of mutant mice. Our data reveal that although recombinant lysosomal proteases can be efficiently delivered to neuronal lysosomes cysteine proteases are less efficient in protein aggregates clearance as compared to the cathepsin-D treatment. Our data including in vitro degradation assays support the idea that bulk proteolysis requires a hierarchical process in which both aspartyl and cysteine hydrolases play a role.

Enzyme replacement therapy with recombinant pro-CTSD (cathepsin D) corrects defective proteolysis and autophagy in neuronal ceroid lipofuscinosis.

CTSD (cathepsin D) is one of the major lysosomal proteases indispensable for the maintenance of cellular proteostasis by turning over substrates of endocytosis, phagocytosis and autophagy. Consequently, CTSD deficiency leads to a strong impairment of the lysosomal-autophagy machinery. In mice and humans CTSD dysfunction underlies the congenital variant (CLN10) of neuronal ceroid lipofuscinosis (NCL). NCLs are distinct lysosomal storage disorders (LSDs) sharing various hallmarks, namely accumulation of protein aggregates and ceroid lipofuscin leading to neurodegeneration and blindness. The most established and clinically approved approach to treat LSDs is enzyme replacement therapy (ERT) aiming to replace the defective hydrolase with an exogenously applied recombinant protein. Here we reveal that recombinant human pro-CTSD produced in a mammalian expression system can be efficiently taken up by a variety of cell models, is correctly targeted to lysosomes and processed to the active mature form of the protease. In proof-of-principle experiments we provide evidence that recombinant human CTSD (rhCTSD) can improve the biochemical phenotype of CTSD-deficient hippocampal slice cultures in vitro and retinal cells in vivo. Furthermore, we demonstrate that dosing of rhCTSD in the murine CLN10 model leads to a correction of lysosomal hypertrophy, storage accumulation and impaired autophagic flux in the viscera and central nervous system (CNS). We establish that direct delivery of the recombinant protease to the CNS is required for improvement of neuropathology and lifespan extension. Together these data support the continuation of the pre-clinical studies for the application of rhCTSD in the treatment of NCL.Abbreviations: AIF1/IBA1: allograft inflammatory factor 1; BBB: blood brain barrier; CNS: central nervous system; CTSB: cathepsin B; CTSD: cathepsin D; CTSL: cathepsin L; ERT: enzyme replacement therapy; GFAP: glial fibrillary acidic protein; INL: inner nuclear layer; LAMP1: lysosomal-associated membrane protein 1; LAMP2: lysosomal-associated membrane protein 2; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; LDL: low-density lipoprotein; LRP1: low density lipoprotein receptor-related protein 1; LSD: lysosomal storage disorder; MEFs: mouse embryonic fibroblasts; M6P: mannose 6-phosphate; mCTSD: mature CTSD; NCL: neuronal ceroid lipofuscinosis; ONL: outer nuclear layer; PB: phosphate buffer; proCTSD: pro-cathepsin D; LRPAP1: low density lipoprotein receptor-related protein associated protein 1; rhCTSD: human recombinant CTSD; SAPC: saposin C; SAPD: saposin D; ATP5G1: ATP synthase, H+ transporting, mitochondrial F0 complex, subunit C1 (subunit 9); SQSTM1/p62: sequestosome 1; TPP1: tripeptidyl peptidase I.