A deep intronic CPS1 variant causing pseudo-exon activation identified in an adult with molecularly unconfirmed urea...

Diagnosing proximal urea cycle disorders (UCDs) remains challenging due to the lack of definitive diagnostic biochemical markers, which can lead to delayed or missed diagnosis. Although molecular genetic testing has improved diagnostic accuracy, some patients still harbor only a single detectable pathogenic variant or no identifiable variants in known disease genes. Here, we report a late-onset adult Japanese patient who remained undiagnosed despite strong clinical suspicion of a UCD.

Targeted gene panel sequencing for UCD-associated genes was performed using genomic DNA from the proband. To evaluate potential splicing abnormalities, reverse transcription PCR was performed using blood-derived cDNA to analyze CPS1 transcripts. Functional validation of the candidate splicing variant was conducted using a minigene splicing assay in cultured HEK293T cells.

Targeted gene panel analysis identified a heterozygous CPS1 variant, c.840G>C (p.Val278_Lys280del), inherited from his father. cDNA analysis revealed an aberrant transcript containing a 121-bp pseudo-exon between exons 3 and 4 in CPS1. Subsequent genomic analysis identified a deep intronic variant, c.381+178A>C, located 15 bp upstream of the pseudo-exon acceptor site. A minigene splicing assay confirmed that this variant induces pseudo-exon inclusion.

We identified a novel deep intronic CPS1 variant that causes aberrant splicing through pseudo-exon activation. Partial splicing defects associated with this variant may contribute to the relatively mild clinical phenotype, highlighting the importance of transcript-level analyses for achieving accurate molecular diagnosis of UCD.