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Impact of Mutations in Aminoacyl tRNA Synthetases on Protein Translation and Cellular Stress (FIBROMARS)

Primary Purpose

Interstitial Lung and Liver Disease, Infantile Liver Failure Syndrome 1, Neurologic, Endocrine and Pancreatic Disease, Multisystem, Infantile-Onset 2

Status
Not yet recruiting
Phase
Not Applicable
Locations
France
Study Type
Interventional
Intervention
Skin biopsy
Sponsored by
Assistance Publique - Hôpitaux de Paris
About
Eligibility
Locations
Arms
Outcomes
Full info

About this trial

This is an interventional basic science trial for Interstitial Lung and Liver Disease focused on measuring Cytosolic aminoacyl-tRNA synthetase, Mutations in the MARS1 gene, Mutations in the LARS1 gene, Mutations in the YARS1 gene, Mutations in the FARSA gene

Eligibility Criteria

undefined - undefined (Child, Adult, Older Adult)All SexesDoes not accept healthy volunteers

Inclusion Criteria:

  • Patients carrying mutations in the MARS1, LARS1, YARS1 or FARSA gene responsible for a multi-systemic phenotype
  • Information and consent of the patient if an adult and of the holders of parental authority if a minor patient and of the minor patient

Exclusion Criteria:

- Non-consent of one of the holders of parental authority or of the minor patient or of adult patient

Sites / Locations

  • Hôpital Necker-Enfants Malades

Arms of the Study

Arm 1

Arm Type

Experimental

Arm Label

Patients

Arm Description

Patients with mutations in the MARS1, LARS1, YARS1 or FARSA genes and cared at Necker Hospital.

Outcomes

Primary Outcome Measures

Determination of total protein content
Determination of total protein content by Bicinchoninic acid assay.
Incorporation of d-methionine and d-phenylalanine into proteins
Incorporation of methionine and phenylalanine by labelled amino-acid fluorescent assays using ready-to-use kits.
Study of polysomes profiling
Study of polysome profils by differential sedimentation on sucrose gradients.
Study of the assembly of the ribosomal 43S pre-initiation complex
Study of the assembly of the ribosomal 43S pre-initiation complex by co-immunoprecipitation experiments.
Phosphorylation of eIF2α and 4EBP and the expression of ATF4
Phosphorylation of eIF2α and 4EBP and the expression of ATF4 by western blot.
Ribosome profiling
Ribosome profiling by high throughput sequencing.
Transfer RNA (tRNA) sequencing
Transfer RNA (tRNA) sequencing by high throughput sequencing.
Production of reactive oxygen species (ROS)
Production of reactive oxygen species (ROS) by fluorescent measurement after cells' incubation with 2',7'- dichlorodihydrofluorescein diacetate (H2DCFDA).

Secondary Outcome Measures

Full Information

First Posted
June 17, 2022
Last Updated
January 16, 2023
Sponsor
Assistance Publique - Hôpitaux de Paris
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1. Study Identification

Unique Protocol Identification Number
NCT05514470
Brief Title
Impact of Mutations in Aminoacyl tRNA Synthetases on Protein Translation and Cellular Stress
Acronym
FIBROMARS
Official Title
Impact of Loss-of-function Mutations of Genes Encoding Cytosolic Aminoacyl-tRNA Synthetases on Protein Translation and Responses to Cellular Stress
Study Type
Interventional

2. Study Status

Record Verification Date
January 2023
Overall Recruitment Status
Not yet recruiting
Study Start Date
April 2023 (Anticipated)
Primary Completion Date
April 2026 (Anticipated)
Study Completion Date
April 2026 (Anticipated)

3. Sponsor/Collaborators

Responsible Party, by Official Title
Sponsor
Name of the Sponsor
Assistance Publique - Hôpitaux de Paris

4. Oversight

Studies a U.S. FDA-regulated Drug Product
No
Studies a U.S. FDA-regulated Device Product
No
Data Monitoring Committee
No

5. Study Description

Brief Summary
Mutations in the genes encoding cytosolic aminoacyl-tRNA synthetases are responsible for early-onset multisystemic diseases including to varying degrees interstitial lung disease, liver damage, neurological and digestive disorders, and systemic inflammation. These are rare and severe diseases whose pathophysiology is poorly understood. The investigative team hypothesizes that mutations within these genes are responsible for a decrease in protein translation and lead to a cellular stress response similar to that induced by amino acid deprivation. The investigative team also hypothesizes that these alterations could be corrected by high-dose supplementation in the culture medium of the corresponding amino acid. The main objective of the study is to precisely determine the consequences of cytosolic aminoacyl-tRNA synthetase mutations at the cell level on protein translation.
Detailed Description
Mutations in the genes encoding cytosolic aminoacyl-tRNA synthetases are responsible for early-onset multisystemic diseases including to varying degrees interstitial lung disease, liver damage, neurological and digestive disorders, and systemic inflammation. These are rare and severe diseases whose pathophysiology is poorly understood. The investigative team hypothesizes that mutations within these genes are responsible for a decrease in protein translation and lead to a cellular stress response similar to that induced by amino acid deprivation. The investigative team also hypothesizes that these alterations could be corrected by high-dose supplementation in the culture medium of the corresponding amino acid. The main objective of the study is to precisely determine the consequences of cytosolic aminoacyl-tRNA synthetase mutations at the cell level on protein translation. The parameters below will be studied in vitro in cell culture from skin biopsies of patients and commercially available human dermal fibroblasts from newborns and adults: Determination of total protein content The incorporation of d-methionine, leucine, tyrosine or phenylalanine into proteins The study of polysomes profiling The study of the assembly of the ribosomal 43S pre-initiation complex The phosphorylation of eIF2α and 4EBP and the expression of ATF4 Ribosome profiling Transfer RNA (tRNA) sequencing The production of reactive oxygen species (ROS) The results of these studies will be compared: Between patient cells and control cells Between genetically corrected patient cells, by stable transfection of the wild-type cDNA of the concerned genes and uncorrected cells Between patient cells cultured in medium enriched with methionine, phenylalanine, tyrosine or leucine and cells cultured in standard medium.

6. Conditions and Keywords

Primary Disease or Condition Being Studied in the Trial, or the Focus of the Study
Interstitial Lung and Liver Disease, Infantile Liver Failure Syndrome 1, Neurologic, Endocrine and Pancreatic Disease, Multisystem, Infantile-Onset 2, Rajab Interstitial Lung Disease With Brain Calcifications 2
Keywords
Cytosolic aminoacyl-tRNA synthetase, Mutations in the MARS1 gene, Mutations in the LARS1 gene, Mutations in the YARS1 gene, Mutations in the FARSA gene

7. Study Design

Primary Purpose
Basic Science
Study Phase
Not Applicable
Interventional Study Model
Single Group Assignment
Masking
None (Open Label)
Allocation
N/A
Enrollment
22 (Anticipated)

8. Arms, Groups, and Interventions

Arm Title
Patients
Arm Type
Experimental
Arm Description
Patients with mutations in the MARS1, LARS1, YARS1 or FARSA genes and cared at Necker Hospital.
Intervention Type
Other
Intervention Name(s)
Skin biopsy
Intervention Description
A skin biopsy performed on the forearm or thigh depending on the patient's age and wishes, with a biopsy punch with a diameter of 3 to 4 mm depending on the child's age (3 for children under 3 years, 4 beyond). Culture of fibroblasts and immortalization.
Primary Outcome Measure Information:
Title
Determination of total protein content
Description
Determination of total protein content by Bicinchoninic acid assay.
Time Frame
Day 0
Title
Incorporation of d-methionine and d-phenylalanine into proteins
Description
Incorporation of methionine and phenylalanine by labelled amino-acid fluorescent assays using ready-to-use kits.
Time Frame
Day 0
Title
Study of polysomes profiling
Description
Study of polysome profils by differential sedimentation on sucrose gradients.
Time Frame
Day 0
Title
Study of the assembly of the ribosomal 43S pre-initiation complex
Description
Study of the assembly of the ribosomal 43S pre-initiation complex by co-immunoprecipitation experiments.
Time Frame
Day 0
Title
Phosphorylation of eIF2α and 4EBP and the expression of ATF4
Description
Phosphorylation of eIF2α and 4EBP and the expression of ATF4 by western blot.
Time Frame
Day 0
Title
Ribosome profiling
Description
Ribosome profiling by high throughput sequencing.
Time Frame
Day 0
Title
Transfer RNA (tRNA) sequencing
Description
Transfer RNA (tRNA) sequencing by high throughput sequencing.
Time Frame
Day 0
Title
Production of reactive oxygen species (ROS)
Description
Production of reactive oxygen species (ROS) by fluorescent measurement after cells' incubation with 2',7'- dichlorodihydrofluorescein diacetate (H2DCFDA).
Time Frame
Day 0

10. Eligibility

Sex
All
Accepts Healthy Volunteers
No
Eligibility Criteria
Inclusion Criteria: Patients carrying mutations in the MARS1, LARS1, YARS1 or FARSA gene responsible for a multi-systemic phenotype Information and consent of the patient if an adult and of the holders of parental authority if a minor patient and of the minor patient Exclusion Criteria: - Non-consent of one of the holders of parental authority or of the minor patient or of adult patient
Central Contact Person:
First Name & Middle Initial & Last Name or Official Title & Degree
Alice HADCHOUEL, MD, PhD
Phone
1 44 49 48 47
Ext
+33
Email
alice.hadchouel-duverge@aphp.fr
First Name & Middle Initial & Last Name or Official Title & Degree
Hélène MOREL
Phone
1 71 19 63 46
Ext
+33
Email
helene.morel@aphp.fr
Overall Study Officials:
First Name & Middle Initial & Last Name & Degree
Alice HARCHOUEL, MD, PhD
Organizational Affiliation
Assistance Publique - Hôpitaux de Paris
Official's Role
Principal Investigator
First Name & Middle Initial & Last Name & Degree
Isabelle SERMET-GAUDELUS, MD, PhD
Organizational Affiliation
Assistance Publique - Hôpitaux de Paris
Official's Role
Study Director
Facility Information:
Facility Name
Hôpital Necker-Enfants Malades
City
Paris
ZIP/Postal Code
75015
Country
France
Facility Contact:
First Name & Middle Initial & Last Name & Degree
Alice HADCHOUEL, MD, PhD
Phone
1 44 49 48 47
Ext
+33
Email
alice.hadchouel-duverge@aphp.fr

12. IPD Sharing Statement

Plan to Share IPD
No

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Impact of Mutations in Aminoacyl tRNA Synthetases on Protein Translation and Cellular Stress

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