Pandemrix-induced narcolepsy is associated with genes related to immunity and neuronal survival

Abstract

Background: The incidence of narcolepsy rose sharply after the swine influenza A (H1N1) vaccination campaign with Pandemrix. Narcolepsy is an immune-related disorder with excessive daytime sleepiness. The most frequent form is strongly associated with HLA-DQB1*06:02, but only a minority of carriers develop narcolepsy. We aimed to identify genetic markers that predispose to Pandemrix-induced narcolepsy.

Methods: We tested for genome-wide and candidate gene associations in 42 narcolepsy cases and 4981 controls. Genotyping was performed on Illumina arrays, HLA alleles were imputed using SNP2HLA, and single nucleotide polymorphisms were imputed using the haplotype reference consortium panel. The genome-wide significance threshold was p < 5 × 10^-8, and the nominal threshold was p < 0.05. Results were replicated in 32 cases and 7125 controls. Chromatin data was used for functional annotation.

Findings: Carrying HLA-DQB1*06:02 was significantly associated with narcolepsy, odds ratio (OR) 39.4 [95% confidence interval (CI) 11.3, 137], p = 7.9 × 10^-9. After adjustment for HLA, GDNF-AS1 (rs62360233) was significantly associated, OR = 8.7 [95% CI 4.2, 17.5], p = 2.6 × 10^-9, and this was replicated, OR = 3.4 [95% CI 1.2-9.6], p = 0.022. Functional analysis revealed variants in high LD with rs62360233 that might explain the detected association. The candidate immune-gene locus TRAJ (rs1154155) was nominally associated in both the discovery and replication cohorts, meta-analysis OR = 2.0 [95% CI 1.4, 2.8], p = 0.0002.

Interpretation: We found a novel association between Pandemrix-induced narcolepsy and the non-coding RNA gene GDNF-AS1, which has been shown to regulate expression of the essential neurotrophic factor GDNF. Changes in regulation of GDNF have been associated with neurodegenerative diseases. This finding may increase the understanding of disease mechanisms underlying narcolepsy. Associations between Pandemrix-induced narcolepsy and immune-related genes were replicated.

Keywords: (MeSH); Autoimmune diseases; Drug-related side effects and adverse reactions; Genetic variation; Genome-wide association study; Glial cell line-derived neurotrophic factor; H1N1 subtype; Influenza A virus; Influenza vaccines; Narcolepsy; Pharmacogenetics; RNA, long noncoding.

Fig. 1

Analysis of principal components 1 and 2 (PC 1 and PC2) for cases (n = 42) and controls (n = 4891) in the discovery cohort. Comparison is made with Utah residents with Northern and Western European ancestry from the CEPH collection (CEU), Han Chinese in Beijing, China (CHB), Japanese in Tokyo, Japan (JPT), and Yoruba people in Ibadan, Nigeria (YRI).

Fig. 2

Manhattan plot of the genome-wide association analysis. All analyses were made on 42 cases of Pandemrix-associated narcolepsy vs 4891 population controls with 8.6 million SNPs after imputation, adjusted by sex and genetic principal components 1–4. The red line shows the threshold for genome-wide significance of 5 × 10⁻⁸. A) Main analysis. The top SNP is located in the human leukocyte antigen (HLA) region on chromosome 6 position 32,213,150 according to Genome Reference Consortium human assembly GRCh37. B) Adjustment for HLA-DQB1*06:02. The top SNP was rs62360233 on chromosome 5, located near glial cell line-derived neurotrophic factor (GDNF) anti-sense 1 (AS1), GDNF-AS1 (OR = 8.6 [95% CI 4.2, 17.5], p = 2.6 × 10⁻⁹). (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Fig. 3

Meta-analysis of T-cell receptor alpha joining (TRAJ). Results for the TRAJ variant rs1154155 from the discovery and replication cohorts, and meta-analysis using a fixed effects model.