Case Report: Genetic and functional analysis of a novel homozygous intronic CD46 variant in a patient with atypical hemolytic uremic syndrome

Atypical hemolytic uremic syndrome (aHUS) is a complement-mediated thrombotic microangiopathy driven by excessive complement activation. Genetic abnormalities in complement-related genes are detected in approximately half of the patients with aHUS. CD46 (membrane cofactor protein, MCP) is a transmembrane cofactor for factor I that facilitates inactivation of C3b and C4b on the cell surface. Among CD46 variants associated with aHUS, intronic variants can cause aberrant splicing and loss of cell-surface CD46. However, it is challenging to predict the pathogenicity of noncanonical splice-site variants, even with machine-learning-based algorithms. In this study, we investigated the potential pathogenicity of a novel intronic CD46 variant identified in a patient with aHUS. A 27-year-old woman presented with recurrent aHUS triggered by COVID-19 infection and was treated with eculizumab. She had no family history of aHUS. Following treatment with eculizumab and methylprednisolone pulse therapy, her laboratory findings improved, and she was discharged on day 15 of hospitalization. Genetic analysis revealed a homozygous intronic variant, c.287-5T>C, in the CD46 gene. Although in silico analysis predicted a low likelihood of pathogenicity, we performed further genetic and expression analysis of CD46 in the patient and her asymptomatic heterozygous mother. Flow cytometry showed markedly reduced CD46 expression on peripheral blood mononuclear cells, whereas her heterozygous mother exhibited decreased but detectable expression. RT-PCR demonstrated exon 3 skipping in the patient’s transcript, resulting in a premature termination codon. A minigene splicing assay reproduced the same exon-skipping event observed in the peripheral blood cDNA. These findings indicate that the novel homozygous CD46 intronic variant c.287-5T>C, whose pathogenicity could not be established by in silico prediction alone, causes aHUS via aberrant splicing and loss of cell-surface CD46 expression. This study highlights the crucial role of functional validation in interpreting noncanonical splice-site variants in complement-mediated diseases.