Role of ACTG2 Variants in Smooth Muscle Determination and Function in Pediatric Intestinal Pseudo-obstruction.

Not Applicable

Not yet recruiting

• Conditions: PIPO

• Registration Number: NCT06687564
• Lead Sponsor: University Hospital, Grenoble

Brief Summary

The primary objective of this study is to describe the transcriptional impact of R178, R257, R40 or A136 variants of the ACTG2 gene on iPS differentiation mechanisms up to organoids derived from PIPO patient samples versus those derived from control / WT patients (generation of IPS from cultured cell lines), at different stages of their experimental ex vivo development.

Detailed Description

Recruited patients will be sampled during a consultation: a blood sample and a biopsy will be taken directly from the patient. Once these samples have been taken, they will be cultured to be reprogrammed into iPS cells, then grown and differentiated into intestinal organoids.

Various experiments will be carried out (as described in the outcomes) to identify at molecular, cellular and tissue level the mechanisms altered in patients with an R178, R257, R40 or A136 variant, assessing their consequence(s) on the development and functionality of the digestive mesenchyme.

Study Timeline

• Study First Submitted: 2024-10-11
• Study First Submitted QC: 2024-11-12
• Study First Posted: 2024-11-13
• Status Verified: 2025-01
• Last Update Submitted: 2025-01-17
• Last Update Posted: 2025-01-22
• Study Start: 2025-02
• Primary Completion: 2030-11
• Study Completion: 2030-11

Recruitment & Eligibility

• Status: NOT_YET_RECRUITING
• Sex: All
• Target Recruitment: 4

Inclusion Criteria

PIPO Population:

1. Minor or adult patient ≥ 4 years of age
2. Patient with PIPO before age 18
3. Male or female
4. Patient with PIPO meeting at least 2 of the ESPGHAN criteria (Thapar et al 2018) and carrying the R178, R257, R40 or A136 mutation of the ACTG2 gene.
5. Patient whose assent has been obtained and whose legal guardians have given their written informed consent
6. Patient affiliated to the French Social Security system or benefiting from an equivalent plan

WT population:

• iPS cell lines MS573 or WT8288 or 202CT or SD378M, from the Nantes University Hospital biological collection and generated from samples from control patients without POIC who have consented to donate their samples.

Exclusion Criteria

PIPO population :

1. Patients with a history of radiotherapy treatment
2. Patient with lymphocyte lineage damage

Study & Design

• Study Type: INTERVENTIONAL
• Study Design: FACTORIAL

Primary Outcome Measures

Name	Time	Method
Description of the transcriptional impact of R178, R257, R40 or A136 variants of the ACTG2 gene	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) through study completion, an average of 5 years	Difference in transcript expression (%) in RNASeq transcriptomic analysis between samples carrying the R178, R257, R40 or A136 variants of the ACTG2 gene on the differentiation mechanisms of iPS up to organoids derived from PIPO patient samples versus those derived from control patients

Secondary Outcome Measures

Name	Time	Method
Evaluate the impact of R178, R257, R40 or A136 variants of the ACTG2 gene on gastrointestinal contractile function between mutant versus WT organoids.	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) an average of 5 years	Difference (%) in isometric contraction force between mutant versus WT organoids by electrical stimulation and specific agonist.
Evaluate the immunofluorescence labeling differential between mutant and WT cells from IPS and organoids	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) an average of 5 years	Difference in fluorescence (%) between mutant and WT cells as cell stages from iPS to organoids
Evaluate the impact of R178, R257, R40 or A136 mutations of the ACTG2 gene on the actin network of fibroblasts derived from skin samples of diseased patients versus those of WTpatients.	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) an average of 5 years	Difference in actin network labeling by immunofluorescence (%) between skin fibroblasts from PIPO and WT patients.
Evaluate the effect of reversion of the R178, R257, R40 or A136 mutation versus WT on functionality and contractility during differentiation of cells into organoids.	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) an average of 5 years	Difference in isometric contraction strength (%) between mutant versus WT versus revertant organoids via specific agonist and electrical stimulation.
Assessing the potential of a chemical library to correct the phenotype	At different stages of their experimental ex vivo development (mesenchymal progenitors, determined smooth muscle cells, differentiated smooth muscle cells and 3D organization of smooth muscle) an average of 5 years	Identification of molecules inducing a significant difference in isometric contraction force (%) between treated mutant organoids, untreated mutant organoids and treated and untreated WT

Trial Locations

Locations (3)

Phymedexp Inserm U1046 - Cnrs Umr 9214; 🇫🇷 Montpellier, France

Tens - Inserm Un Umr 1235; 🇫🇷 Nantes, France

AP-HP Hôpital Robert Debré; 🇫🇷 Paris, France