DNA QUANTIFICATION TECHNIQUE AS A INTERPRETATION TOOL IN MITOCHONDRIAL DISEASES

Mitochondrial Diseases Clinical Trial

Official title:

IMPLEMENTING A SINGLE MUSCLE FIBER DNA QUANTIFICATION TECHNIQUE AS A INTERPRETATION TOOL FOR THE VARIANTS OF UNKNOWN SERVICE IN MITOCHONDRIAL DISEASES

Clinical Trial Details — Status: Completed

Administrative data

• NCT number: NCT03216252
• Other study ID #: 16-AOIP-04
• Status: Completed
• Phase: N/A
• Start date: February 2, 2018
• Est. completion date: September 30, 2020

Study information

• Verified date: November 2023
• Source: Centre Hospitalier Universitaire de Nice
• Contact: n/a
• Is FDA regulated: No
• Study type: Interventional

Clinical Trial Summary

2622/5000 Mitochondrial diseases (MM) are the most common metabolic diseases. Since these pathologies are very heterogeneous in clinical terms, only the identification of mutations in nuclear genes or mitochondrial DNA confirms the diagnosis.

The full-scale study of mtDNA by high-throughput sequencing (NGS) is a first step in the diagnostic approach. The recent introduction of this revolutionary new technology has greatly increased the efficiency of mutation identification. However, in addition to known pathogenic mutations, NGS reveals numerous variants whose significance is currently unknown. A major challenge to obtain a reliable diagnosis is therefore the interpretation of the clinical impact of these new rare variants which proves to be very difficult.

Pathogenicity criteria allow the classification of variants from benign to pathogenic. One of the major pathogenicity criteria is a good correlation of heteroplasmic level with tissue or cellular involvement. Indeed, mtDNA mutations are generally heteroplasmic, which corresponds to the coexistence of normal and mutated molecules in the same cell or tissue, the most affected tissues having a high rate of mutation. On a muscle biopsy of an affected patient, the fibers often present an enzyme deficiency in cytochrome c oxidase (COX-negative) which can be demonstrated in immunohistochemistry. The single fiber study allows to isolate the deficient fibers and to quantify the heteroplasmic rate of a variant. The presence of a high level of heteroplasm in the COX-negative fibers, unlike fibers without deficit, is a strong argument in favor of the pathogenicity of this variant. Currently, this technique is not used routinely in diagnostic laboratories but only occasionally in a research framework in some laboratories. It is a heavy technique that consists of a first stage of laser microdissection of the various muscle fibers followed by a second step of quantification of the variant from each fiber. This second step requires a specific focus for each identified variant.

The aim of this pilot study is to develop a new technique for quantification of single-fiber heteroplasmics isolated by NGS laser microdissection. This, independent of the type of variant, will avoid the long and costly adjustments required for each new variant identified and thus facilitate its use

Recruitment information / eligibility

• Status: Completed
• Enrollment: 10
• Est. completion date: September 30, 2020
• Est. primary completion date: September 30, 2020
• Accepts healthy volunteers: No
• Gender: All
• Age group: 0 Years and older

Eligibility

Inclusion Criteria:

• general criteria: major or minor patients, sporadic or isolated cases

• criteria related to pathology:

• Suspected mitochondrial pathology which will be evaluated according to the following criteria (expertise of the clinician of the MM reference center):

• Clinical picture suggestive of a mitochondrial pathology ("illegitimate association" of symptoms, specific syndrome of MELAS type for example, muscular deficit, ptosis ...) AND / OR

• Metabolic assessment suggestive of respiratory tract involvement AND / OR

• Identification of a deficiency involving one or more complexes of the respiratory chain from a muscular specimen

• Presence on the histological analysis of the muscle biopsy of COX-negative fibers

• signing of informed consent for minor patients signed by at least one of the parents or the representative of the parental authority

Exclusion Criteria:

• Persons deprived of their liberty by a judicial or administrative decision;

• Persons hospitalized without consent;

• Persons admitted to a health or social institution for purposes other than research;

• Persons of legal age who are under protection or who are unable to express their consent. Inability to co-operate.

Related Conditions & MeSH terms

• Mitochondrial Diseases

Intervention

Biological:
• samples
Blood samples, oral smear and urine

Locations

Country	Name	City	State
France	CHU de Nice	Nice

Sponsors (1)

Lead Sponsor	Collaborator
Centre Hospitalier Universitaire de Nice

Country where clinical trial is conducted

France, 

Outcome

Type	Measure	Description	Time frame	Safety issue
Primary	number of amplification with the NGS technique		36 months
Secondary	comparaison of the number of amplification between NGS technique and the PCR-RFLP technique		36 months