Whole-genome sequencing identified a deep intronic COL4A5 variant causing aberrant splicing in a female patient with...

Alport syndrome is an inherited kidney disease caused by pathogenic variants in COL4A3, COL4A4, or COL4A5, encoding type IV collagen chains of the glomerular basement membrane. Genetic confirmation is essential for its diagnosis, prognosis, and donor selection; however, conventional sequencing may fail to detect pathogenic variants, leaving a subset of clinically suspected patients without a genetic diagnosis. We report a 16-year-old girl with persistent hematuria and proteinuria, initially diagnosed with Alport syndrome at 2 years of age based on kidney biopsy findings of a basket-weave glomerular basement membrane and mosaic α5(IV) staining. Despite a family history compatible with this diagnosis, targeted exome sequencing at 8 years of age did not identify pathogenic variants. Whole-genome sequencing at 16 years revealed a deep intronic variant in COL4A5 (NM_000495.5) (c.276 + 1306G > A). In silico analysis with SpliceAI predicted the creation of de novo acceptor and donor splice sites, leading to the inclusion of a 103-bp cryptic exon within intron 4. A minigene assay confirmed aberrant splicing and demonstrated that the inserted exon resulted in a frameshift and premature stop codon (p.Gly93Phefs*99). This case highlights the diagnostic utility of whole-genome sequencing in combination with splicing prediction and functional validation to resolve suspected cases of Alport syndrome in which the genetic cause has remained unclear. As sequencing technologies advance, such integrative approaches may become increasingly important in clinical practice, thereby improving diagnostic yield and supporting the development of RNA-based therapies.