Introduction
In prenatal diagnosis and genetic counseling, "synonymous variants" are often classified as "Variants of Uncertain Significance" (VUS), creating a bottleneck for a definitive diagnosis. However, a study on a family with Meckel syndrome (MKS) demonstrates how the RDDC RNA Splicer tool can crack this challenge. In this research, RDDC accurately predicted that a "synonymous" VUS in the RPGRIP1L gene would cause two distinct aberrant splicing patterns. This prediction was subsequently confirmed perfectly by a minigene assay, successfully upgrading the VUS to a pathogenic variant and guiding the family to a healthy birth via Preimplantation Genetic Testing (PGT-M).
Research Challenge: A "Synonymous" VUS Halting PGT-M
This study centered on a couple with a history of four adverse pregnancies, including fetal encephalocele and polydactyly. Whole Exome Sequencing (WES) of the fourth fetus revealed compound heterozygous variants in the RPGRIP1L gene: one known pathogenic variant (c.1351-11A>G) inherited from the mother, and another variant, c.1581G>A (p.Gln527=), inherited from the father. This paternal variant was "synonymous" (it doesn't change the amino acid) and thus classified as a VUS by ACMG. To provide the family with accurate PGT-M, it was imperative to first determine if this synonymous variant was truly pathogenic.
RDDC's Precise Prediction: Unveiling Two Aberrant Splicing Patterns
To evaluate the variant's function before investing in experimental resources, the research team utilized the RDDC RNA Splicer tool. The AI model's analysis provided critical and specific pathogenic hypotheses. It predicted that this "silent" c.1581G>A variant would actually devastate the splicing motif, leading to two primary aberrant splicing outcomes:
Aberrant Splicing Patterns
- A 26 bp retention of intron 13: This would cause a frameshift and a premature termination codon (PTC).
- Skipping of exon 13: This would result in a 174 bp deletion, also disrupting the protein.
This prediction clearly indicated that the synonymous variant was pathogenic via a loss-of-function mechanism.
Experimental Validation and Clinical Success: From Prediction to Healthy Neonate
Based on RDDC's clear predictions, the team performed a minigene assay. The experimental results robustly confirmed RDDC's forecast: the vector carrying the c.1581G>A mutation indeed produced two aberrant transcripts—one retaining 26 bp of intron 13, and the other completely skipping exon 13. The perfect concordance between the in silico prediction and the in vitro results provided the critical pathogenic evidence (fulfilling ACMG criteria PS3) needed to upgrade the VUS to "Likely Pathogenic" (LP).
Clinical Success
This definitive molecular diagnosis was the cornerstone for the subsequent clinical intervention. It allowed the team to confidently design a PGT-M strategy, screen embryos, and select a single wild-type, chromosomally normal embryo for transfer. The family ultimately welcomed a healthy baby. This case powerfully demonstrates that RDDC RNA Splicer is a robust tool for deciphering the pathogenicity of "synonymous" VUS, providing precise, testable splicing predictions that bridge the gap between basic research and clinical applications like PGT-M.
Content Source and Disclaimer
This article is a compilation and interpretation of the scientific study cited below, intended to highlight the application of RDDC bioinformatics tools. All research data and conclusions belong to the original authors and publication.
Original Article
Xu H, Pu J, Wu Z, et al. Case report: Successful PGT-M based on the identification of a spliceogenic variant in the RPGRIP1L gene through Minigene assay. Frontiers in Genetics. 2024 Feb 20;15:1354366.
Article Link: https://pubmed.ncbi.nlm.nih.gov/39479399/
