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

Overview

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.

Interventions

• Procedure Biopsy
  A skin biopsy and a blood sample will be taken to culture the iPS cells and intestinal organoids.

Primary outcome measures

• Description of the transcriptional impact of R178, R257, R40 or A136 variants of the ACTG2 gene [Time frame: 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]

Secondary outcome measures (5)

• Evaluate the impact of R178, R257, R40 or A136 variants of the ACTG2 gene on gastrointestinal contractile function between mutant versus WT organoids. [Time frame: 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]
• Evaluate the immunofluorescence labeling differential between mutant and WT cells from IPS and organoids [Time frame: 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]
• 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. [Time frame: 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]
• 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. [Time frame: 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]
• Assessing the potential of a chemical library to correct the phenotype [Time frame: 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]

Eligibility criteria

Inclusion criteria

PIPO Population:

• Minor or adult patient ≥ 4 years of age
• Patient with PIPO before age 18
• Male or female
• 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.
• Patient whose assent has been obtained and whose legal guardians have given their written informed consent
• 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 :

• Patients with a history of radiotherapy treatment
• Patient with lymphocyte lineage damage

Criteria are shown verbatim from the registry (in English). Final eligibility is always assessed by the study center.

Healthy volunteers: No

Study design

Allocation

Non-randomized

Model

Factorial

Masking

Open label

Primary purpose

Basic science

Study locations

France · 3 centers

• Phymedexp Inserm U1046 - Cnrs Umr 9214 — Montpellier
• Tens - Inserm Un Umr 1235 — Nantes
• AP-HP Hôpital Robert Debré — Paris

Publications

• Zenzeri L, Tambucci R, Quitadamo P, Giorgio V, De Giorgio R, Di Nardo G. Update on chronic intestinal pseudo-obstruction. Curr Opin Gastroenterol. 2020 May;36(3):230-237. doi: 10.1097/MOG.0000000000000630. PMID 32073506
• Fournier N, Fabre A. Smooth muscle motility disorder phenotypes: A systematic review of cases associated with seven pathogenic genes (ACTG2, MYH11, FLNA, MYLK, RAD21, MYL9 and LMOD1). Intractable Rare Dis Res. 2022 Aug;11(3):113-119. doi: 10.5582/irdr.2022.01060. PMID 36200034
• Assia Batzir N, Kishor Bhagwat P, Larson A, Coban Akdemir Z, Baglaj M, Bofferding L, Bosanko KB, Bouassida S, Callewaert B, Cannon A, Enchautegui Colon Y, Garnica AD, Harr MH, Heck S, Hurst ACE, Jhangiani SN, Isidor B, Littlejohn RO, Liu P, Magoulas P, Mar Fan H, Marom R, McLean S, Nezarati MM, Nugent KM, Petersen MB, Rocha ML, Roeder E, Smigiel R, Tully I, Weisfeld-Adams J, Wells KO; Baylor-Hopki PMID 31769566
• de Santa Barbara P, van den Brink GR, Roberts DJ. Development and differentiation of the intestinal epithelium. Cell Mol Life Sci. 2003 Jul;60(7):1322-32. doi: 10.1007/s00018-003-2289-3. PMID 12943221
• Mahe MM, Brown NE, Poling HM, Helmrath MA. In Vivo Model of Small Intestine. Methods Mol Biol. 2017;1597:229-245. doi: 10.1007/978-1-4939-6949-4_17. PMID 28361322

Identifiers

NCT: NCT06687564 · 38RC23.0368 · 2024-A01516-41

Primary sources (government registries)

• ClinicalTrials.gov (US NIH)

View this study on ClinicalTrials.gov ↗