Exome sequencing reanalysis identifies a novel likely pathogenic CFAP54 variant and expands the phenotypic and...

Primary ciliary dyskinesia (PCD) is a rare genetic disorder caused by structural or functional abnormalities of motile cilia, characterized by considerable clinical and genetic heterogeneity. Although exome sequencing (ES) can improve the diagnostic rate of PCD, more than 30% of patients with clinically suspected PCD remain undiagnosed by initial ES. The American College of Medical Genetics and Genomics (ACMG) recommends periodic reanalysis of ES data to increase the diagnostic yield.

We investigated a 35-year-old female patient with bronchiectasis and a strong clinical suspicion of PCD from Peking Union Medical College Hospital. The patient's ES data, which had initially yielded negative results in August 2024, was reanalyzed in early 2025 employing a "genotype-phenotype-inheritance pattern" strategy. A minigene assay was conducted to validate the pathogenicity of the identified CFAP54 splicing variant. Variant pathogenicity was classified according to the ACMG/AMP guidelines.

The patient had a history of rhinitis and neonatal pneumonia. Pulmonary function tests revealed moderate obstructive ventilatory dysfunction. ES reanalysis identified a homozygous variant, CFAP54 (NM_001306084.2):c.6965 + 5G >A, which was initially classified as a variant of uncertain significance. Minigene assays confirmed that this variant induced exon 50 skipping, resulting in a frameshift and a premature termination codon (loss-of-function). This variant was subsequently reclassified as "Likely Pathogenic".

This study is the first to describe CFAP54:c.6965 + 5G >A and confirm its pathogenicity. This finding brings the total number of CFAP54-associated PCD patients to eight and the number of distinct CFAP54 mutations to twelve, thereby enriching the phenotypic and genotypic spectrum of this gene. Furthermore, this effective strategy of "ES reanalysis + minigene verification" resolved the diagnostic dilemma in initially ES-negative PCD cases, providing a replicable molecular diagnostic framework for similar scenarios.