Diagnostic potential of the amniotic fluid cells transcriptome in deciphering mendelian disease: a proof-of-concept

Abstract

RNA sequencing (RNA-seq) is emerging in genetic diagnoses as it provides functional support for the interpretation of variants of uncertain significance. However, the use of amniotic fluid (AF) cells for RNA-seq has not yet been explored. Here, we examined the expression of clinically relevant genes in AF cells (n = 48) compared with whole blood and fibroblasts. The number of well-expressed genes in AF cells was comparable to that in fibroblasts and much higher than that in blood across different disease categories. We found AF cells RNA-seq feasible and beneficial in prenatal diagnosis (n = 4) as transcriptomic data elucidated the molecular consequence leading to the pathogenicity upgrade of variants in CHD7 and COL1A2 and revising the in silico prediction of a variant in MYRF. AF cells RNA-seq could become a reasonable choice for postnatal patients with advantages over fibroblasts and blood as it prevents invasive procedures.

Incidental Genetic Finding in a Fetus

“…In another study, individuals harboring low-penetrance RB1 PSVs (eg, splice-site and missense SVs) developed bilateral retinoblastoma later than patients with null PSVs . Molecular screening methods (eg, RNA sequencing of amniotic fluid cells) have enabled clinicians to assess whether a specific RB1 SV is likely to be associated with any phenotypic manifestations . However, there currently are no clinically acceptable predictive parameters to better define the phenotypic expression of RB1 PSV carriers.…”

“…4 Molecular screening methods (eg, RNA sequencing of amniotic fluid cells) have enabled clinicians to assess whether a specific RB1 SV is likely to be associated with any phenotypic manifestations. 10 However, there currently are no clinically acceptable predictive parameters to better define the phenotypic expression of RB1 PSV carriers. The case presented herein reports an incidental RB1 PSV detected on unrelated genetic testing, posing a clinical and ethical dilemma to both clinician and family.…”

Integrative omics approaches to advance rare disease diagnostics

“…Therefore, non‐invasive biospecimens for transcriptomic approach recently got more attention. Lee et al examined the potential of RNA‐seq from amniotic fluid cells for prenatal diagnostics 29 . They identified a high similarity of gene expression profiles to fibroblasts and provided a diagnosis to 4 patients (8%).…”

“…It was successfully applied as a complementary approach to WES and WGS in more than 18 studies (Table S2). There are multiple case reports in which RNA‐sequencing (RNA‐seq) provided the necessary functional evidence to support a molecular diagnosis 24–39 . Transcriptomics can be used for VUS reclassification and for prioritisation of overlooked likely pathogenic variants.…”

Improved detection of aberrant splicing using the Intron Jaccard Index

“…Identifying splicing aberrations on RNA sequencing (RNA-seq) data provides more direct evidence of the presence of splicing defects and reveals the resulting transcript isoforms. This approach has been successfully used to diagnose patients with rare genetic disorders in large cohorts (14)(15)(16)(17)(18)(19)(20). Following this initial success, computational tools specialized on detecting aberrant splicing from RNA-seq data have been developed including LeafcutterMD (21), SPOT (22), and FRASER (23).…”