Parallel Sequencing of Fetal Genome and RNA in the Presence of Ultrasound Warning Signs: a Complementary Approach for the Prenatal Diagnosis of Rare Diseases. (PrenOmics)

Parallel Sequencing of the Fetal Genome and RNA in the Presence of Ultrasound Warning Signs: a Complementary Approach for the Prenatal Diagnosis of Rare Diseases.

Prenatal exome sequencing (ES) is increasingly used for fetuses with ultrasound-detected anomalies but yields 10-15% variants of uncertain significance (VUS), limiting diagnostic performance, particularly in prenatal settings with incomplete phenotypes.

This study aims to evaluate the added value of combined prenatal genome sequencing (GS) and RNA sequencing (RNA-Seq), which are not currently part of routine care. Conducted at AP-HP, it will compare the diagnostic yield of GS + RNA-Seq with the current standard approach (chromosomal microarray analysis + ES), according to variant type (coding, non-coding, and structural). The contribution of systematic RNA-Seq to rapid VUS resolution will be specifically assessed.

Overall, this project will assess the feasibility, diagnostic performance, and clinical utility of implementing GS + RNA-Seq in prenatal diagnosis, supporting future integration into routine care in France.

Study Overview

• Status: Not yet recruiting
• Conditions: Couples Eligible for Prenatal Exome Sequencing
• Intervention / Treatment: Diagnostic test: Genome Sequencing (GS) + RNA-Sequencing (RNA-Seq)

Detailed Description

Couples will be enrolled during a "pre-test" genetic consultation by a physician or a research-trained collaborator (genetic counselor or another investigator).

As part of routine clinical care: chromosomal microarray analysis (CMA) and exome sequencing (ES) will be performed locally, with interpretation by a biologist from the site providing follow-up to the couple.

For research purposes, after obtaining written consent from both the pregnant woman and her partner: samples will be prepared for pre-analytical processing at Pitié-Salpêtrière and sequenced at SeqOIA. Interpretation will be conducted by a biologist at a site different from the one performing the exome analysis, using the Gleaves-P interface dedicated to the project (MOABI).

Transcriptomic analysis (RNA-Seq) will be performed on RNA extracted from amniotic fluid cultures, with and without emetine (a NMD inhibitor). Sequencing will be carried out after Agilent capture at the Genetics Laboratory of Necker-Enfants Malades Hospital (AP-HP). Bioinformatics analysis will be performed by the Imagine platform. Study biologists will interpret results using the PolyRNASEQ interface (qualitative splice junction analysis) and IGV (semi-quantitative analysis and expression).

All results from CMA + ES and GS + RNA-Seq will be reviewed during weekly meetings. In cases of discordant or unexpected results (anticipated for ~10 couples), a reference method will be applied to provide clinically validated results through an accredited prenatal diagnostic center, ensuring no loss of care opportunity.

Results will be discussed in a multidisciplinary prenatal diagnosis meeting (RCP) and integrated into the prognostic discussion. Couples will be seen during a "post-test" genetic consultation for communication of results.

For couples included at Necker-Enfants Malades Hospital only: two additional maternal blood tubes will be collected during routine pregnancy follow-up before amniocentesis. Circulating DNA will be extracted from maternal plasma and subjected to genome sequencing under the same conditions as the amniotic DNA. The generated data will be analysed using bioinformatic approaches integrating artificial intelligence methods to improve the distinction between fetal- and maternal-derived DNA fragments. Interpretation will be performed via the Gleaves-P interface. Non-invasive prenatal genome sequencing results are strictly for research purposes: they will not be communicated to couples and will not be used for clinical decision-making. Their objective is to evaluate the feasibility, accuracy, and performance of non-invasive genome sequencing.

• Study Type: Observational
• Enrollment (Estimated): 100

Contacts and Locations

• Study Contact: Name: Sarah BOUCHARD, Project Manager; Phone Number: +33 01 42 19 28 79; Email: sarah.bouchard@aphp.fr
• Study Contact Backup: Name: Lucile BOUTAUD, Pharm. D PhD; Phone Number: +33 01 44 38 17 47; Email: lucile.boutaud@aphp.fr

Study Locations

• France
  • Île-de-France Region
    • Paris, Île-de-France Region, France, 75015
      • Hopital Necker Enfants Malades
      • Contact: Sarah BOUCHARD, Project Manager; Phone Number: +33 01 42 19 28 79; Email: sarah.bouchard@aphp.fr
      • Contact: Lucile BOUTAUD, Pharm. D PhD; Phone Number: +33 01 44 38 17 47; Email: lucile.boutaud@aphp.fr

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Adult; Older Adult
• Accepts Healthy Volunteers: No

• Sampling Method: Non-Probability Sample

Study Population

Couples whose indication for prenatal exome sequencing has been reviewed and approved by a multidisciplinary prenatal diagnostic center.

Description

Inclusion Criteria:

• Participating couple aged ≥ 18 years
• Ongoing pregnancy for which the indication for exome sequencing has been discussed and validated by a Multidisciplinary Prenatal Diagnosis Center (CPDPN).

While a strict list of indications is not appropriate in the prenatal setting, examples include multiple anomalies not related to a malformation sequence, persistent increased nuchal translucency, hydrops fetalis (anasarca), cleft palate, multiple contractures/arthrogryposis, skeletal dysplasia, bowed femurs, or brain anomalies.

• Sequencing performed on an amniotic fluid sample (chorionic villus sampling is excluded)
• Attendance at a genetic counseling consultation
• Written informed consent obtained for study participation

Exclusion Criteria:

• Couple not covered by the social security system
• Couple deprived of liberty or under legal guardianship or curatorship
• Monoparental pregnancies

Study Plan

How is the study designed?

Number of groups / cohorts

1

Cohorts and Interventions

Group / Cohort

Intervention / Treatment

Genome Sequencing (GS) + RNA-Sequencing (RNA-Seq); Couples (pregnant woman and partner) with a fetal indication requiring prenatal exome sequencing, reviewed and approved by a multidisciplinary prenatal diagnosis center, will be included. Fetal samples (amniotic fluid, including cell cultures) and parental blood samples collected during routine care will be used for standard genetic analyses (chromosomal microarray and exome sequencing) and for research analyses, including genome sequencing (GS), RNA sequencing (RNA-Seq), and bioinformatic evaluations. Non-invasive genome sequencing performed from maternal blood samples (for couples included at Necker-Enfants Malades Hospital only) will be conducted for research purposes only; results will not be returned to couples and will not influence clinical management.

Diagnostic test: Genome Sequencing (GS) + RNA-Sequencing (RNA-Seq); Couples whose indication for trio exome sequencing is approved by a multidisciplinary prenatal diagnostic center are enrolled during a pre-test genetic consultation, during which written informed consent is obtained from both partners. As part of routine care, fetal samples are collected by amniocentesis and parental blood samples are obtained for chromosomal microarray analysis (CMA) and trio exome sequencing (ES), analyzed locally. Amniotic fluid cell cultures are systematically prepared and stored. For research purposes, portions of fetal and parental samples are processed at Pitié-Salpêtrière and sequenced at SeqOIA for trio genome sequencing (GS). Bioinformatics analysis is performed using the MOABI platform, with interpretation via the Gleaves-P interface. RNA sequencing (RNA-Seq) is performed on RNA extracted from amniotic fluid cell cultures at Necker Hospital. For couples included at Necker only, maternal plasma DNA is also sequenced. Non-invasive GS results are for rese

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Evaluation of the diagnostic contribution of RNA sequencing (RNA-Seq) performed in parallel with genome sequencing (GS) in prenatal diagnosis, compared with chromosomal microarray analysis (CMA) and exome sequencing (ES).	Comparison of variants identified by each strategy and the number of additional diagnoses achieved using genome sequencing (GS) combined with RNA sequencing (RNA-Seq) compared with chromosomal microarray analysis (CMA) and exome sequencing (ES) in the prenatal setting. Diagnostic yield will be analyzed according to variant type (single-nucleotide and structural variants) and genomic location (coding vs non-coding regions).	18 months

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Comparison of turnaround times for result delivery between genome sequencing combined with RNA sequencing (GS + RNA-Seq) and the current standard approach of chromosomal microarray analysis and exome sequencing (CMA + ES) in prenatal diagnosis.	Time interval between test prescription and receipt of results for GS + RNA-Seq compared to CMA + ES in prenatal diagnosis	18 months
Comparison of the costs associated with GS + RNA-Seq versus CMA + ES in prenatal diagnostic settings.	Evaluation and comparison of the costs associated with GS + RNA-Seq and CMA + ES approaches.	18 months
Evaluation of the feasibility and analytical performance of non-invasive prenatal genome sequencing performed on circulating cell-free fetal DNA extracted from maternal blood.	Correlation of diagnostic results between invasive foetal genome sequencing performed on DNA extracted from amniotic fluid and non-invasive genome sequencing performed on circulating cell-free foetal DNA extracted from maternal plasma.	18 months
Identification and characterization of potential challenges and obstacles to the implementation of these diagnostic methods in clinical practice.	List of difficulties encountered at each stage, from pre- to post-analytical.	18 months

Collaborators and Investigators

• Sponsor: Assistance Publique - Hôpitaux de Paris
• Collaborators: URC-CIC Paris Descartes Necker Cochin; Laboratoire SeqOIA; Imagine; MOABI
• Investigators: Principal Investigator: Lucile BOUTAUD, Pharm.D PhD, Hôpital Necker Enfants Malades AP-HP

Study record dates

Study Major Dates

• Study Start (Estimated): September 1, 2026
• Primary Completion (Estimated): March 1, 2028
• Study Completion (Estimated): March 1, 2028

Study Registration Dates

• First Submitted: August 6, 2026
• First Submitted That Met QC Criteria: August 25, 2026
• First Posted (Actual): August 27, 2026

Study Record Updates

• Last Update Posted (Actual): August 27, 2026
• Last Update Submitted That Met QC Criteria: August 25, 2026
• Last Verified: August 1, 2026

More Information

Terms related to this study

• Keywords: RNA sequencing; Prenatal diagnosis (PND); Invasive / Non-invasive prenatal diagnosis (NIPD); Exome sequencing (ES); Genome Sequencing (GS); Circulating cell-free fetal DNA (cffDNA)
• Additional Relevant MeSH Terms: Genetic Structures; Genetic Phenomena; Molecular Structure; Biochemical Phenomena; Chemical Phenomena; Base Sequence
• Other Study ID Numbers: APHP251082; N° IDRCB (Other Identifier: 2025-A01594-45)

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: NO

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No