A novel homozygous splicing variant in FREM1 expands the phenotypic spectrum of BNAR syndrome: functional validation and successful PGT-M.

backgroundBifid nose with or without anorectal and renal anomalies (BNAR) is a rare autosomal recessive genetic congenital disorder characterized by bifid nose and renal agenesis, with or without anorectal malformations. Our research identified the genetic factors associated with BNAR in a Chinese pedigree.

methodsMuscle tissue specimens from the fetus and peripheral blood specimens from the parents were collected, and genomic DNA was extracted for whole-exome sequencing. Sanger sequencing was used to verify the emergence of variants in the pedigree. Minigene assays and structural modeling were performed to confirm the influence of intronic variants on mRNA splicing.

resultsWe identified a homozygous splicing variant, c.3274 + 4 A > G, in the FREM1 gene in the fetus. Sanger sequencing revealed that the parents were heterozygous. In vitro and in intro minigene assays revealed exon 18 skipping, causing a nonframeshift, 62 amino acid deletion in FREM1. Structural predictions indicate the deletion severely disrupts the FREM1 CSPG6 domain, replacing its stable interwoven β-sheet architecture with a single β-sheet and an aberrant α-helix. This profound conformational shift likely abolishes the interaction interfaces required for FRAS1/FREM multiprotein complex assembly, impairing its overall function. Based on these findings, the variant was categorized as likely pathogenic according to the American College of Medical Genetics and Genomics guidelines. Accordingly, after being informed about the hazards of the pathogenic variant, the couple selected preimplantation genetic tests for monogenic disorders (PGT-M) and had a healthy baby.

conclusionThis study presents a Chinese fetus with FREM1-related pathology. Moreover, this study extends the phenotypic spectrum of BNAR, while laying a foundation for PGT-M.