S55. ANXA11 mutations prevail in Chinese ALS patients with and without cognitive dementia

Abstract

Introduction Amyotrophic lateral sclerosis (ALS) is a fatal neurological disease characterized by progressive paralysis and ultimately respiratory failure within 5 years of symptom onset. Approximately 5–10% of ALS cases exhibit familial inheritance (FALS) and causative gene mutations can be found in 60% of FALS patients. The remaining 90–95% of ALS cases show sporadic manner (SALS) and mutations in the same genes are responsible for 10% of SALS patients. To date, rare variants in more than 30 genes have been reported to cause or be associated with ALS. Recently, Smith and et al. identified mutations in ANXA11 in ALS patients of European ancestry but pathogenicity of ANXA11 in other ALS cohorts remained unproved. In the current study, we investigated ANXA11 mutations in Chinese ALS patients with or without cognitive decline. Methods: Study population A total of 383 Chinese ALS patients were recruited at ALS clinic of Neurology Department, Peking Union Medical College Hospital (PUMCH) from January 2016 to August 2017. Mutation screening All coding exons and at least 100 bp of flanking introns of ANXA11(NM_145869) were sequenced using ABI 3730 automated DNA-sequencing system. Rare variants (MAF Transcripts investigation of the splicing mutation Total RNA was extracted using TRIzol (Invitrogen) from peripheral white blood cells. Then 2.5 μ g RNA were used to perform Reverse transcriptase-PCR. cDNA was amplified using primer pair ANXA11-RT-F(5′-CCATGAGCTACCCTGGCTAT-3′) and ANXA11-RT-R(5′-GACTCCCCAGGCAGTCAAT-3′) locating at ANXA11 exon 4 and exon 8 (shown in Fig. 3), respectively. The PCR products were isolated on a 1.5% agarose gel. DNA fragments of interest were gel-purified and sequenced. Results We identified 6 nonsynonymous heterozygous mutations (c.107C>G, p.P36R; c.119A>G, p.D40G; c.382G>A, p.V128M; c.687T>A, p.S229R; c.904C>T, p.R302C; c.1471G>A, p.G491R) and 1 splice-site mutation(c.174-2A>G, p.A58_Q187del) in 10 unrelated patients, accounting for a mutant frequency of 5.6% (1/18) in FALS, 2.3% (8/353) in SALS and 8.3% (1/12) in ALS-FTD. A novel p. P36R mutation was identified in one FALS index, one SALS patient and one ALS-FTD. The splicing mutation(c.174-2A>G) caused in-frame skipping of the entire exon 6 and further supported the loss-of-function mechanism. Conclusion ANXA11 gene is the most frequently mutated genes in Chinese patients with SALS. The study findings further expand the ANXA11 phenotype, firstly highlighting its pathogenic role in ALS-FTD.