Laura Helou

Introduction: Autism spectrum disorder (ASD) is a neurodevelopmental condition with substantial genetic and phenotypic heterogeneity. However, populations of African ancestry remain underrepresented in genomic studies, limiting understanding of ASD genetic architecture. This study aimed to characterize rare, clinically relevant genetic variants in a Rwandan pediatric ASD cohort using trio‐based whole‐exome sequencing (WES). Methods: Trio‐based WES was performed in 31 Rwandan pediatric patients with ASD (aged 2–18 years) and their parents. Variants were analyzed using a trio‐based workflow and classified according to American College of Medical Genetics and Genomics/Association for Molecular Pathology (ACMG/AMP) guidelines. Results: Eleven candidate variants were identified in 9 of 31 patients, including four likely pathogenic variants and seven variants of uncertain significance. This resulted in a diagnostic yield of 12.9% (4/31), expanded to 29.0% when phenotypically concordant variants of uncertain significance were considered. Most likely pathogenic variants were identified in individuals with syndromic ASD who presented with intellectual disability, epilepsy, and global developmental delay. Likely pathogenic findings included two single nucleotide variants in GABRB3 , SYNGAP1 , and two copy‐number variants involving the GNAS locus and chromosome 1p35.3‐p35.2. Conclusions: The diagnostic yield observed in this cohort is consistent with previous trio‐based WES studies of ASD. The findings support the clinical utility of WES for the genetic evaluation of ASD and underscore the need for expanded genomic studies in African populations.

Les éléments transposables sont des éléments génétiques mobiles présents dans presque tous les génomes eucaryotes et procaryotes. Ils co-évoluent avec leurs hôtes et ont participé au façonnage de leur environnement génomique au cours de leur évolution. Ils sont en outre à l’origine de nouveaux gènes dérivés de séquences codantes de transposon par domestication.La transposase domestiquée piggyBac, PGBD5, est très conservée depuis l’origine des vertébrés. C’est une transposase fonctionnelle et capable de transposer un élément piggyBac d’insecte, Ifp2. Elle s’exprime spécifiquement dans le système nerveux central et plus précisément dans les neurones pyramidaux de l’hippocampe, de l’amygdale et du bulbe olfactif qui sont respectivement impliqués dans les mémoires spatiale, contextuelle, émotionnelle et olfactive. La conservation élevée de PGBD5 chez les vertébrés suggère que sa fonction et ses cibles chromosomiques le seraient également.Mon travail de thèse traite de l’origine et de l’activité de la transposase domestiquée PGBD5 dans le génome des vertébrés.Dans un premier temps, nous avons étudier les capacités de transposition d’une transposase piggyBac sans le domaine CRD. En effet, PGBD5 peut être caractérisé comme étant une transposase ayant perdu son domaine CRD au cours de la domestication. Nous avons vérifié la nécessité de la présence d’un domaine C-terminal pour l’activité de transposase-recombinase.Dans un second temps, nous avons étudier l’histoire évolutive de pgbd5 et des pgbd en général. Nous avons identifié le pble phylogénétiquement le plus proche de pgbd5 et ses capacités à être transposé par PGBD5.Enfin, nous avons étudier les capacités de PGBD5 à exciser et réintégrer dans de nouveaux locus des pble humains afin d’évaluer la capacité de PGBD5 à mobiliser ces éléments présents dans le génome humain et d’affiner l’identification des cibles de la transposase.

Genomic first-tier approaches are increasingly explored as a strategy for newborn screening, raising major challenges for variant interpretation in asymptomatic populations. Within the BabyDetect project, we evaluated the clinical relevance of variants detected in three key one-carbon metabolism genes – MTHFR , CBS , and MAT1A – by comparing next-generation sequencing with biochemical data. Variants of uncertain significance (VUS) in those three genes were observed in approximately 4% of newborns. All likely pathogenic or pathogenic variants were present in the heterozygous state. Methionine levels, measured prospectively as part of routine newborn screening, and total homocysteine levels, assessed retrospectively, remained within neonatal reference ranges for both VUS and likely pathogenic/pathogenic variant carriers, with no significant differences between variant groups. These findings indicate that neither VUS nor heterozygous likely pathogenic/pathogenic variants in these genes exert a measurable biochemical impact in the neonatal period. Our results demonstrate the critical value of integrating biochemical data to contextualize genomic findings. This combined approach supports cautious interpretation of genomic results in population-based newborn screening and argues against the necessity to consider VUS in the absence of biochemical or clinical abnormalities.

Newborn screening (NBS) has evolved significantly since its inception, yet many treatable rare diseases remain unscreened due to technical limitations. The BabyDetect study used gene panel sequencing to expand NBS to treatable conditions not covered by conventional biochemical screening. We present here the analytical validation of this workflow, assessing sensitivity, precision, and reproducibility using dried blood spots from newborns. We implemented strict quality control thresholds for sequencing, coverage, and contamination, ensuring high reliability. Longitudinal monitoring confirmed consistent performance across more than 5900 samples. Automation of DNA extraction improved scalability, and a panel redesign enhanced the coverage and selection of targeted regions. By focusing on known pathogenic/likely pathogenic variants, we minimized false positives and maintained clinical actionability. Our findings demonstrate that gene panel sequencing-based NBS is feasible, accurate, and scalable, addressing critical gaps in current screening programs.

Baby Detect Project, started in September 2022, aimed to create a newborn screening test using targeted next-generation sequencing for all early-onset, treatable, and serious conditions. The elaborated gene panel covers 405 genes, associated with 165 genetic conditions, and includes RB1, linked to retinoblastoma, the only oncological disease tested for. Germline RB1 mutations concern around 50% of all retinoblastoma cases and 100% of the most severe, bilateral cases. Ninety percent of them occur de novo, which delays the diagnosis by about a year with subsequent loss of vision and sometimes the eye itself. Detecting children with germline RB1 mutation at birth would greatly improve functional and anatomic outcomes, limiting invasive treatments and general anesthesias through early childhood. We discuss herein the novel approach of population screening, the rationale for newborn testing for RB1 mutations, the incidence of expected cases, the reliability of the test and its costs. The next step is to move to a nation-scale population; this initiative marks a landmark in retinoblastoma patients’ care.

PGBD5 is encoded by a gene domesticated at the chordate origin from a DNA transposon of the piggyBac family. During its evolution, PGBD5’s sequence has been under strong purifying selection among vertebrate genomes. This suggests PGBD5 functions in the development and physiology of chordates, as recently demonstrated in mouse and human brain development, where it was implicated in double strand DNA breaks on neurons. However, biochemical PGBD5 activities remain undefined due to lack of appropriate in vitro model systems. Furthermore, coevolution of PGBD5 with hosts has likely reshaped some of its functions, resulting in differences between vertebrate PGBD5 and the insect piggybac transposase (PB). Recent studies have found that PGBD5 can interact with two different “species” of piggyBac -like transposon element ( pble ). Here, we show that human PGBD5 can interact with four “species” human pbles and to promote their chromosomal integration in cells, a property it shares with insect PB. Human PGBD5 can also bind to distinct chromosomal copies of human pble in cell type-specific manner, and to genomic loci containing inverted repeats in human cells akin to those found in subterminal insect pble ends. These findings expand the scope of potential biological activities of PGBD5 and other domesticated DNA transposases.

Newborn Screening (NBS) has evolved significantly since its inception, yet many treatable rare diseases remain unscreened due to technical limitations. The BabyDetect study used a targeted next-generation sequencing (tNGS) panel to expand NBS to treatable conditions not covered by conventional screening. We present here the analytical validation of this workflow, assessing sensitivity, precision, and reproducibility using dried blood spots from newborns. We implemented strict quality control thresholds for sequencing, coverage, and contamination, ensuring high reliability. Longitudinal monitoring confirmed consistent performance across more than 5,900 samples. Automation of DNA extraction improved scalability, and a panel redesign enhanced the coverage and selection of targeted regions. By focusing on known pathogenic/likely pathogenic variants, we minimized false positives and maintained clinical actionability. Our findings demonstrate that tNGS-based NBS is feasible, accurate, and scalable, addressing critical gaps in current screening programs.

The rapid development of therapies for severe and rare genetic conditions underlines the need to incorporate first-tier genetic testing into newborn screening (NBS) programs. A workflow was developed to screen newborns for 165 treatable pediatric disorders by deep sequencing of regions of interest in 405 genes. The prospective observational BabyDetect pilot project was launched in September 2022 in a maternity ward of a public hospital in the Liege area, Belgium. In this ongoing observational study, 4,260 families have been informed of the project, and 3,847 consented to participate. To date, 71 disease cases have been identified, 30 of which were not detected by conventional NBS. Glucose-6-phosphate dehydrogenase deficiency was the most frequent disorder detected, with 44 positive individuals. Of the remaining 27 cases, 17 were recessive disorders. We also identified one false-positive case in a newborn in whom two variants in the AGXT gene were identified, which were subsequently shown to be located on the maternal allele. Nine heterozygous variants were identified in genes associated with dominant conditions. Results from the BabyDetect project demonstrate the importance of integrating biochemical and genomic methods in NBS programs. Challenges must be addressed in variant interpretation within a presymptomatic population and in result reporting and diagnostic confirmation.

Purpose: Genomic newborn screening programs are emerging worldwide. With the support of the local pediatric team of Liege, Belgium, we developed a panel of 405 genes that are associated with 165 early-onset, treatable diseases with the goal of creating a newborn screening test using targeted next-generation sequencing for all early-onset, treatable, and serious conditions. Methods: A process was developed that informed the future parents about the project and collected their consent during a face-to-face discussion with a trained investigator. The first baby was screened on 1 September 2022. The main objective of the study was to test the feasibility and the acceptability of targeted sequencing at birth as a first-tier newborn screening approach to detect treatable genetic conditions or genetic conditions for which a pre-symptomatic or early symptomatic clinical trial is available. Results: As of 20 June 2024, the parents of 4425 children had been offered the test; 4005 accepted (90.5%) and 420 refused (9.5%). The main reasons for refusal were the research nature of the project and the misunderstanding of what constitutes genetic conditions. Conclusions: These data demonstrate the high acceptability of genomic newborn screening in a properly informed population.