Genome-wide Epistasis for Cardiovascular Severity in Marfan Study

NCT ID: NCT06257004

Last Updated: 2024-02-13

Basic Information

• Recruitment Status: RECRUITING
• Total Enrollment: 200 participants
• Study Classification: OBSERVATIONAL
• Study Start Date: 2020-11-30
• Study Completion Date: 2025-10-31

Brief Summary

Marfan syndrome (MFS) is an autosomal dominant connective tissue disorder with pleiotropic manifestations in the ocular, skeletal and cardiovascular systems. Morbidity and mortality are mostly determined by aortic root aneurysm dissection and rupture. Although mutations in FBN1, the gene coding for the extracellular matrix protein fibrillin-1, are the well-established genetic cause of this condition, there is a very poor correlation between the nature or location of the causal FBN1 mutation and the phenotypical outcome. Indeed, wide intra- and interfamilial phenotypical variability is observed. So, even with an identical primary mutation in all family members, the clinical spectrum varies widely, from completely asymptomatic to sudden death due to aortic dissection at a young age. The precise mechanisms underlying this variability remain largely elusive.

Consequently, a better understanding of the functional effects of the primary mutation is highly needed and the identification of genetic variation that modifies these effects is becoming increasingly important. In this project, we have carefully selected different innovative strategies to discover mother nature's own modifying capabilities with respect to Marfan syndrome aortopathy.

Detailed Description

In this project we will focus on the cardiovascular, or more specific, the TAAD (Thoracal Aorta Aneurysma Dissection) expressivity of the Marfan syndrome. The most frequent mutations in FBN1 (fibrilin-1 ), with significant aortopathy expressivity is p.Ile2585Thr; c.7754T>C and p.Ala882Val; c.2645C>T). We will limit the used population to p.Ile2585Thr; c.7754T>C mutation since this is the biggest population.

Marfan syndrome subjects carrying an identical FBN1 mutation show a variable aortopathy expressivity, even within one family. We hypothesize that the cardiovascular phenotypical variability is under control of genetic modifiers.

The first approach strategy involves ranking of carriers of the specific FBN1 mutation that present with significant variable aortopathy expressivity according to the severity of aortic aneurysma disease (based on Z-score, timing of surgery and manual expert curation). We will stratify these mutation carrying individuals in three groups: mild or no aortic disease (UMC, unaffected mutation carrier)), severely affected (AMC, affected mutation carrier), and participants with indeterminate data.

The second approach is the molecular characterisation of the 25% extreme cohort (AMC and UMC) using WGS (Whole Genome Sequencing) and linkage analysis.

Finally subjects peripheral blood mononuclear cells (PBMCs) of 10 severely affected mutation carrier (AMC) and 10 unaffected mutation carriers (UMC) as well as 2 controls will be reprogrammed to iPSCs (induced Pluripotential Stem Cells). These cells will finally be differentiated into VSMC's (VasculairSmoth Muscle Cells). The genomic integrity and identity of the iPSCs and the VSMCs will be validated using RT-PCR and immunocytochemistry.

Transcriptomic (i.e. RNA-sequencing) data will be acquired from these specific induced pluripotent stem cell-derived vascular smooth muscle cells (iPSC-VSMCs).

We will be able to filter the WGS data based on variant quality and location in genes that are differentially expressed when comparing the AMC and UMC iPSC-VSMCs, via the synchronization of both data types. This approach will allow us to identify the modifier gene. Once candidate modifier genes (and hence candidate modifier variants) have been identified, their modifying capacity will be functionally checked in relevant cell- or animal models. The choice of the model system will be determined based on the nature of the identified modifier. In an animal model, we will prove its effect by crossing an animal carrying the variant of interest with a MFS model, which should significantly alter the cardiovascular phenotype. Depending on the function and evolutionary conservation of the identified modifier gene, zebrafish or mouse models will be used.

Alternatively, the identified modifier will be functionally validated using the cutting-edge CRISPR/Cas9 genome editing technology in the available and thoroughly functionally characterized iPSC-VSMC lines.

Further evidence for a modifying role of the most interesting candidate genes will be obtained by performing targeted re-sequencing of these genes' coding and regulatory sequences in, again, the 25% most and least severely cardiovascular affected MFS cases of a large replication cohort consisting of more than 3000 clinically and molecularly (FBN1 mutation-positive) characterized index cases.

Whenever possible, segregation of the remaining candidate modifier variants with protection from TAAD will be investigated in available gDNA samples of the probands' relatives carrying the FBN1 mutation.

Conditions

Marfan Syndrome; FBN1 Mutation

Study Design

• Observational Model Type: CASE_CONTROL
• Study Time Perspective: PROSPECTIVE

Study Groups

AMC (Affected Mutation Carrier)

FBN1 mutation (Marfan Syndrome) - Phenotype cardiovascular severe outcome

Saliva collection (screening of all participants)

Intervention Type: GENETIC

All participants will give a salive sample (self-sampling kit) - selection of 25% extremes (UMC and AMC) participants for WGS and linkage analysis

Bloodsampling

Intervention Type: GENETIC

Based on the results of the WGS, a selection will be made of the 5% most extremes (UMC and AMC) participants for the iPSC-VSMC generation

UMC (Unaffected Mutation Carrier)

FBN1 mutation (Marfan Syndrome) - Phenotype cardiovascular mild outcome

Saliva collection (screening of all participants)

Intervention Type: GENETIC

All participants will give a salive sample (self-sampling kit) - selection of 25% extremes (UMC and AMC) participants for WGS and linkage analysis

Bloodsampling

Intervention Type: GENETIC

Based on the results of the WGS, a selection will be made of the 5% most extremes (UMC and AMC) participants for the iPSC-VSMC generation

Interventions

Saliva collection (screening of all participants)

All participants will give a salive sample (self-sampling kit) - selection of 25% extremes (UMC and AMC) participants for WGS and linkage analysis

Intervention Type: GENETIC

Bloodsampling

Based on the results of the WGS, a selection will be made of the 5% most extremes (UMC and AMC) participants for the iPSC-VSMC generation

Intervention Type: GENETIC

Eligibility Criteria

Inclusion Criteria

• Participants with proven mutation (p.Ile2585Thr;c.7754C>T) in the FBN1 gene

Exclusion Criteria

-

• Eligible Sex: ALL
• Accepts Healthy Volunteers: No

Sponsors

University Hospital, Ghent

OTHER

Sponsor Role: collaborator

University Hospital, Antwerp

OTHER

Sponsor Role: lead

Responsible Party

Responsibility Role: SPONSOR

Principal Investigators

Bart Loeys, Prof,MD,PhD

Role: PRINCIPAL_INVESTIGATOR

University Hospital, Antwerp

Paul Coucke, Prof,MD,Ing

Role: STUDY_CHAIR

University Hospital, Ghent

Locations

University Hospital Antwerp

Edegem, Prins Boudewijnlaan 43/6, Belgium

Site Status: RECRUITING

Countries

Belgium

Central Contacts

Bart Loeys, Prof,MD,PhD

Role: CONTACT

bart.loeys@uantwerpen.be

++32-3-2759768

Paul Coucke, Prof,PhD,Ing

Role: CONTACT

paul.coucke@ugent.be

++32-9-3323634

Facility Contacts

Bart Loeys

Role: primary

bart.loeys@uantwerpen.be

+3232759768

Paul Coucke

Role: backup

paul.coucke@ugent.be

+3293323634

References

Verstraeten A, Luyckx I, Loeys B. Aetiology and management of hereditary aortopathy. Nat Rev Cardiol. 2017 Apr;14(4):197-208. doi: 10.1038/nrcardio.2016.211. Epub 2017 Jan 19.

Reference Type: BACKGROUND

PMID: 28102232 (View on PubMed)

von Kodolitsch Y, De Backer J, Schuler H, Bannas P, Behzadi C, Bernhardt AM, Hillebrand M, Fuisting B, Sheikhzadeh S, Rybczynski M, Kolbel T, Puschel K, Blankenberg S, Robinson PN. Perspectives on the revised Ghent criteria for the diagnosis of Marfan syndrome. Appl Clin Genet. 2015 Jun 16;8:137-55. doi: 10.2147/TACG.S60472. eCollection 2015.

Reference Type: BACKGROUND

PMID: 26124674 (View on PubMed)

Groth KA, Gaustadnes M, Thorsen K, Ostergaard JR, Jensen UB, Gravholt CH, Andersen NH. Difficulties in diagnosing Marfan syndrome using current FBN1 databases. Genet Med. 2016 Jan;18(1):98-102. doi: 10.1038/gim.2015.32. Epub 2015 Mar 26.

Reference Type: BACKGROUND

PMID: 25812041 (View on PubMed)

Loeys BL, Dietz HC, Braverman AC, Callewaert BL, De Backer J, Devereux RB, Hilhorst-Hofstee Y, Jondeau G, Faivre L, Milewicz DM, Pyeritz RE, Sponseller PD, Wordsworth P, De Paepe AM. The revised Ghent nosology for the Marfan syndrome. J Med Genet. 2010 Jul;47(7):476-85. doi: 10.1136/jmg.2009.072785.

Reference Type: BACKGROUND

PMID: 20591885 (View on PubMed)

Braverman AC. Medical management of thoracic aortic aneurysm disease. J Thorac Cardiovasc Surg. 2013 Mar;145(3 Suppl):S2-6. doi: 10.1016/j.jtcvs.2012.11.062. Epub 2012 Dec 20.

Reference Type: BACKGROUND

PMID: 23260459 (View on PubMed)

De Backer J, Loeys B, Leroy B, Coucke P, Dietz H, De Paepe A. Utility of molecular analyses in the exploration of extreme intrafamilial variability in the Marfan syndrome. Clin Genet. 2007 Sep;72(3):188-98. doi: 10.1111/j.1399-0004.2007.00845.x.

Reference Type: BACKGROUND

PMID: 17718856 (View on PubMed)

Franken R, Teixido-Tura G, Brion M, Forteza A, Rodriguez-Palomares J, Gutierrez L, Garcia Dorado D, Pals G, Mulder BJ, Evangelista A. Relationship between fibrillin-1 genotype and severity of cardiovascular involvement in Marfan syndrome. Heart. 2017 Nov;103(22):1795-1799. doi: 10.1136/heartjnl-2016-310631. Epub 2017 May 3.

Reference Type: BACKGROUND

PMID: 28468757 (View on PubMed)

Renard M, Muino-Mosquera L, Manalo EC, Tufa S, Carlson EJ, Keene DR, De Backer J, Sakai LY. Sex, pregnancy and aortic disease in Marfan syndrome. PLoS One. 2017 Jul 14;12(7):e0181166. doi: 10.1371/journal.pone.0181166. eCollection 2017.

Reference Type: BACKGROUND

PMID: 28708846 (View on PubMed)

Granata A, Serrano F, Bernard WG, McNamara M, Low L, Sastry P, Sinha S. An iPSC-derived vascular model of Marfan syndrome identifies key mediators of smooth muscle cell death. Nat Genet. 2017 Jan;49(1):97-109. doi: 10.1038/ng.3723. Epub 2016 Nov 28.

Reference Type: BACKGROUND

PMID: 27893734 (View on PubMed)

Other Identifiers

1S81820N

Identifier Type: OTHER_GRANT

Identifier Source: secondary_id

B3002021000292

Identifier Type: OTHER

Identifier Source: secondary_id

20/08/087

Identifier Type: -

Identifier Source: org_study_id