Development of Targeted RNA-Seq for Amyotrophic Lateral Sclerosis Diagnosis

NCT ID: NCT06083584

Last Updated: 2025-05-20

Study Results

Results pending

The study team has not published outcome measurements, participant flow, or safety data for this trial yet. Check back later for updates.

Basic Information

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Recruitment Status

RECRUITING

Total Enrollment

192 participants

Study Classification

OBSERVATIONAL

Study Start Date

2023-11-22

Study Completion Date

2027-05-31

Brief Summary

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Genetic diagnosis of Amyotrophic Lateral Sclerosis (ALS) could identify the origin of the disease, potentially allowing the patient to pursue targeted/gene therapy. However, many familial forms of ALS are genetically undiagnosed, either because no variant has been detected in the genes of interest, or because the detected variant(s) have uncertain significance. Currently, molecular diagnosis takes place in two stages: 1) Search for the GGGGCC expansion in the C9ORF72 gene by RP-PCR; 2) Analysis of the coding regions by high-throughput sequencing of a panel of 30 genes involved in ALS.

Many of these variants of uncertain significance affect splicing. Their impact can be predicted using in silico tools, but only an analysis of the patient's RNA can confirm their pathogenic nature. Currently, the analysis of transcripts is only done a posteriori, when a variant predicted to impact splicing is detected on the patient's DNA. RT-PCR followed by Sanger sequencing then verifies the impact of the splice variants. This method confirmed the impact of certain splice variants in patients. However, this method is time-consuming and requires custom development, and is mutation/gene/patient-dependent. In contrast, high-throughput RNA sequencing (RNA-Seq) simultaneously analyzes the splicing of numerous genes, with a global approach, applicable to all patients. This approach avoids the custom design of primers, which can be biased by the interpretation of splicing predictions, while RNA-Seq systematically captures and sequences all the transcripts. Finally, RNA-Seq provides additional information compared to DNA sequencing such as the detection of exon skipping, intron inclusion, and the creation of fusion transcripts.

In the GTEx project (GTEx Consortium, 2013), expression levels of human genome transcripts were quantified by RNA-Seq. Using these results, the study investigators measured expression of transcripts of known ALS genes in whole blood. Applying a threshold value of 0.5 transcripts per million reads (TPM), 25 of the 30 ALS genes currently analyzed by NGS in routine diagnostics at Nîmes University Hospital could be eligible for a complete analysis by RNA-Seq. None of the French laboratories carrying out genetic analyzes of ALS has yet developed RNA-Seq as a routine diagnostic tool. The study laboratory receives more than 600 requests for genetic diagnosis of ALS patients per year. The aim of this study is therefore to develop a global method for analyzing RNA transcripts of ALS genes to categorize the mutations to improve the diagnostic management of patients.

Detailed Description

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Conditions

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Amyotrophic Lateral Sclerosis

Study Design

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Observational Model Type

COHORT

Study Time Perspective

PROSPECTIVE

Study Groups

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Positive controls

6 patients already in database. The 6 confirmed splicing mutations are: DCTN1 (NM\_004082.5): c.3209G\>T, OPTN (NM\_001008211.1) : c.1613-7T\>G, FUS (NM\_004960.4) : c.764+8T\>A, GRN (NM\_002087.4): c.835+1G\>A, GRN (NM\_002087.4): c.709-3C\>G, SPG11 (NM\_025137.4): c.3039-5T\>G

RNA sequencing

Intervention Type OTHER

RNA-Seq (Sureselect XT HS2 RNA) from patient blood sample

Negative controls

30 patients with familial hypercholesterolemia. The absence of splicing anomalies in the SLA genes after confirmation by RT-PCR followed by Sanger sequencing of the absence of anomalies for the 6 variants listed above for each of the 30 individuals.

RNA sequencing

Intervention Type OTHER

RNA-Seq (Sureselect XT HS2 RNA) from patient blood sample

Exploratory cohort

156 ALS: 20 ALS patients with splice variants predicted to be deleterious by in silico prediction software; 136 panel-analysis-negative ALS patients (priority will be given to familial ALS)

RNA sequencing

Intervention Type OTHER

RNA-Seq (Sureselect XT HS2 RNA) from patient blood sample

Interventions

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RNA sequencing

RNA-Seq (Sureselect XT HS2 RNA) from patient blood sample

Intervention Type OTHER

Eligibility Criteria

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Inclusion Criteria

* Have a prescription for a genetic diagnosis of ALS (or familial hypercholesterolemia for the control cohort)
* Have given their informed consent for the genetic study and the biobank
* The patient must be a member or beneficiary of a health insurance plan

Exclusion Criteria

* The patient is under safeguard of justice or state guardianship
Minimum Eligible Age

18 Years

Eligible Sex

ALL

Accepts Healthy Volunteers

No

Sponsors

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Association pour la Recherche sur la Sclérose Latérale Amyotrophique et autres maladies du motoneurones

UNKNOWN

Sponsor Role collaborator

Centre Hospitalier Universitaire de Nīmes

OTHER

Sponsor Role lead

Responsible Party

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Responsibility Role SPONSOR

Principal Investigators

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Claire Guissart

Role: PRINCIPAL_INVESTIGATOR

CHU de Nimes

Locations

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CHU de Bordeaux

Bordeaux, , France

Site Status RECRUITING

CHU de Clermont-Ferrand

Clermont-Ferrand, , France

Site Status RECRUITING

CHU de Lyon

Lyon, , France

Site Status RECRUITING

La Timone

Marseille, , France

Site Status RECRUITING

CHU de Montpellier

Montpellier, , France

Site Status RECRUITING

CHU de Nîmes

Nîmes, , France

Site Status RECRUITING

CHU de Toulouse

Toulouse, , France

Site Status RECRUITING

Countries

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France

Central Contacts

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Claire Guissart

Role: CONTACT

04 66 68 32 07

Facility Contacts

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Gwendal Le Masson

Role: primary

Nathalie Guy

Role: primary

Emilien Bernard

Role: primary

Annie Verschueren

Role: primary

Elisa De la Cruz

Role: primary

Anissa Megzari

Role: primary

04.66.68.42.36

Blandine Acket

Role: primary

Other Identifiers

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NIMAO/LOCAL/2023/CG-01

Identifier Type: -

Identifier Source: org_study_id

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