16-mer hypocretin-1/orexin-A in cerebrospinal fluid to diagnose narcolepsy

Abstract

Study Objectives Cerebrospinal fluid (CSF) hypocretin-1/orexin-A quantification via radioimmunoassay (RIA) is used for diagnosing narcolepsy type 1(NT1), but its limitations require alternative methods. Using liquid chromatography–mass spectrometry (LC–MS), this study aimed to 1) fully characterize CSF orexin-A fragments detected by the gold standard RIA method, and 2) assess diagnostic relevance of measuring the most prevalent fragment, a 16-mer, in patients with NT1, narcolepsy type 2 (NT2), idiopathic hypersomnia (IH), and nonspecified hypersomnolence (NSH). Methods CSF samples were analyzed using RIA and LC–MS in patients with hypersomnolence disorders evaluated at the French Narcolepsy National Reference Center. Fractionation techniques isolated orexin-A and its fragments, which were identified via targeted MS. Statistical analysis compared LC–MS performance against RIA. Results CSF samples from 115 patients (54.8% females, mean age 30.4±15.8 years) including 52 with NT1, 6 NT2, 13 IH, and 44 NSH were analyzed by RIA and LC–MS. A 16-mer orexin-A fragment was identified and quantified by LC–MS, with lower levels in NT1. This fragment correlated strongly with RIA-measured orexin-A (r=0.83, p