Abstract
OBJECTIVE: Biallelic intronic AAGGG repeat expansions in RFC1 cause Cerebellar Ataxia with Neuropathy and Vestibular Areflexia Syndrome and may also contribute to isolated sensory neuropathy. The clinical significance of both heterozygous and biallelic RFC1 expansions in more diverse patient populations remains unclear-partly due to the absence of accurate, user-friendly computational tools specifically tailored for tandem repeat analysis. METHODS: To discern the relationship between RFC1 expansions and idiopathic peripheral neuropathy (iPN), we performed whole-genome sequencing (WGS) followed by PCR-based confirmation in a large, well-characterized U.S. cohort consisting of 788 iPN patients (369 pure small fiber neuropathy (SFN), 266 sensorimotor, 144 pure sensory, and 9 pure motor). We developed an integrative pipeline combining ExpansionHunter Denovo and Expansion Hunter coupled with unsupervised clustering to reliably detect and genotype RFC1 expansions from short-read WGS data, achieving 98.2% concordance with repeat-primed PCR based validation. RESULTS: Biallelic RFC1 expansions were absent in 879 controls but present in 2.8% of iPN patients (Fisher's exact p = 5.9×10 (-8) ), including 6.2% of pure sensory, 2.2% of SFN, and 1.5% of sensorimotor neuropathy, indicating that motor nerve involvement should not exclude patients from RFC1 repeat screening. We also observed a markedly increased frequency of monoallelic expansions in iPN compared to controls (13.2% versus 2.5%; Fisher's exact p = 3.4×10 (-17) ), without evidence of secondary mutations or expansions on the other allele. INTERPRETATION: Our approach provides a robust, cost-effective method for detecting RFC1 expansions from WGS data. Our findings indicate that both heterozygous and homozygous AAGGG repeat expansions in RFC1 can contribute to development of iPN.