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Quantifying subpercent nuclear TDP‐43 loss in cells and ALS cortex using junction‐specific cryptic exon RT‐qPCR.

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Title: Quantifying subpercent nuclear TDP‐43 loss in cells and ALS cortex using junction‐specific cryptic exon RT‐qPCR.
Authors: Koide, Shingo1 (AUTHOR), Ikegami, Ichiko1 (AUTHOR), Hanyu, Ryutaro1 (AUTHOR), Koike, Yuka Mitsuhashi2 (AUTHOR), Yamagishi, Takuma1 (AUTHOR), Toyama, Genri1 (AUTHOR), Washida, Aya1 (AUTHOR), Tada, Mari3 (AUTHOR), Kakita, Akiyoshi3 (AUTHOR), Onodera, Osamu1,2 (AUTHOR), Sugai, Akihiro1 (AUTHOR) akihiro.sugai@bri.niigata-u.ac.jp
Source: FEBS Letters. Jan2026, Vol. 600 Issue 1, p119-130. 12p.
Subjects: Amyotrophic lateral sclerosis, TDP-43 proteinopathies, Drug monitoring, Frontotemporal lobar degeneration, Reverse transcriptase polymerase chain reaction, Neurodegeneration, Biomarkers, Exons (Genetics)
Abstract: Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are progressive neurodegenerative diseases characterised by nuclear TDP‐43 loss. Its hallmark, cryptic exon (CE) splicing, is often masked in bulk tissue analyses by the low abundance of affected neurons. We developed an ultrasensitive RT‐qPCR assay targeting STMN2 CE using one exon–CE junction‐spanning primer and the other within the CE. The design expands the dynamic range sevenfold: TDP‐43 knockdown boosted STMN2 CE levels 1395‐fold in differentiated SH‐SY5Y neurons. Spike‐in tests set detection at 0.16% deficient cells. Crucially, the assay revealed a 42‐fold CE increase in ALS motor cortex, previously missed by conventional primers. This streamlined tool enables precise quantification of TDP‐43 dysfunction and sensitive pharmacodynamic monitoring for future ALS‐FTD therapeutic studies. Impact statementBecause cryptic‐exon signals are diluted in bulk tissue, we developed a junction‐spanning STMN2 RT‐qPCR with sub‐percent sensitivity. This deployable biomarker will aid ALS/FTD researchers and drug developers by standardizing measurements and enabling sensitive pharmacodynamic monitoring of therapies targeting nuclear TDP‐43 dysfunction. [ABSTRACT FROM AUTHOR]
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Database: Engineering Source
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Abstract:Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are progressive neurodegenerative diseases characterised by nuclear TDP‐43 loss. Its hallmark, cryptic exon (CE) splicing, is often masked in bulk tissue analyses by the low abundance of affected neurons. We developed an ultrasensitive RT‐qPCR assay targeting STMN2 CE using one exon–CE junction‐spanning primer and the other within the CE. The design expands the dynamic range sevenfold: TDP‐43 knockdown boosted STMN2 CE levels 1395‐fold in differentiated SH‐SY5Y neurons. Spike‐in tests set detection at 0.16% deficient cells. Crucially, the assay revealed a 42‐fold CE increase in ALS motor cortex, previously missed by conventional primers. This streamlined tool enables precise quantification of TDP‐43 dysfunction and sensitive pharmacodynamic monitoring for future ALS‐FTD therapeutic studies. Impact statementBecause cryptic‐exon signals are diluted in bulk tissue, we developed a junction‐spanning STMN2 RT‐qPCR with sub‐percent sensitivity. This deployable biomarker will aid ALS/FTD researchers and drug developers by standardizing measurements and enabling sensitive pharmacodynamic monitoring of therapies targeting nuclear TDP‐43 dysfunction. [ABSTRACT FROM AUTHOR]
ISSN:00145793
DOI:10.1002/1873-3468.70198