Misfolded TAR DNA-binding protein 43 (TDP-43) is the primary pathological hallmark of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). While seed amplification assays (SAAs), such as real-time quaking-induced conversion (RT-QuIC), have shown promise in detecting misfolded TDP-43 in cerebrospinal fluid (CSF) and olfactory mucosa, technically accessible methodologies are urgently required for widespread clinical application. We developed a streamlined, non-immunoprecipitation-based TDP-43 RT-QuIC assay to assess seeding activity in brain tissue and CSF. We evaluated its diagnostic performance using CSF from patients with TDP-43 proteinopathies and control subjects, and further examined its association with neurofilament light chain (NfL) and tau-related biomarkers. In CSF analysis, the assay demonstrated positive seeding activity in 70% (21/30) of patients with ALS and dementia, 50% (5/10) of patients with FTLD, and 40% (8/20) of patients with ALS alone. The assay exhibited excellent specificity, yielding negative results in >99% (199/200) of control samples, including those with autoimmune or electrophysiological abnormalities. Furthermore, CSF analysis demonstrated significantly higher NfL levels in TDP-43 SAA-positive cases compared to SAA-negative cases (p < 0.0008). The highest NfL concentrations were observed in the SAA-positive ALS with dementia and ALS cohorts, contrasting with lower levels in FTLD. Tau-related biomarkers exhibited no significant differences between the groups. Our streamlined, non-immunoprecipitation TDP-43 RT-QuIC assay provides highly specific detection of pathological TDP-43 seeding activity. While the assay detects the underlying TDP-43 proteinopathy rather than distinguishing between ALS and FTLD clinical phenotypes, its technical simplicity and combined utility with NfL measurements offer a robust, scalable framework for biomarker development. This approach provides a practical foundation for future multi-center validation and international standardization efforts.
K. Satoh, M. Shimamura, Takeshi Fujimoto et al.· Current Issues in Molecular...· 0 citations
TDP-43 proteinopathies, encompassing amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration (FTLD), and limbic-predominant age-related TDP-43 encephalopathy (LATE), represent a heterogeneous spectrum of devastating neurodegenerative disorders. For decades, the diverse clinical presentations of these diseases have complicated antemortem diagnosis and hindered the development of disease-modifying therapies. However, recent breakthroughs in basic science are beginning to address these clinical barriers, although substantial hurdles to practical clinical application remain. Structural elucidation via cryo-electron microscopy (Cryo-EM) has shattered the single-protein amyloid dogma by revealing that TDP-43 can form hetero-amyloid filaments with ANXA11, thereby providing a molecular basis for pathological strain diversity. Concurrently, the pathogenic focus has shifted toward nuclear loss of function, which triggers a systemic “RNA crisis” characterized by aberrant alternative polyadenylation (APA) and cryptic exon inclusion (e.g., STMN2, UNC13A). Crucially, this metabolic collapse is profoundly exacerbated by patient-specific genetic risk factors, acting synergistically in a “two-hit” model of neurodegeneration. To translate these findings to the clinic, next-generation diagnostic tools are emerging. Integrating neuron-derived extracellular vesicle (EV) isolation with Seed Amplification Assays (SAAs) holds promise to help overcome the structural camouflage that limits current PET imaging, potentially offering ultra-sensitive, functional strain identification in biofluids. While these structural and diagnostic milestones provide a strong foundation for precision medicine, major challenges in assay standardization and clinical validation must be addressed. Advanced therapeutic strategies—namely, splice-switching antisense oligonucleotides (ASOs) that directly restore RNA metabolism, combined with the targeted suppression of neuronal hyperexcitability—are now entering clinical trials. This review synthesizes how decoding the structural and RNA-metabolic complexities of TDP-43 is paving a promising pathway from bench to bedside, while critically discussing current translational limitations.
M. Shimamura, K. Satoh· Pathogens· 0 citations
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