Amyloid-β1-43 cerebrospinal fluid levels and the interpretation of APP, PSEN1 and PSEN2 mutations

Abstract

Background: Alzheimer's disease (AD) mutations in amyloid precursor protein (APP) and presenilins (PSENs) could potentially lead to the production of longer amyloidogenic Aβ peptides. Amongst these, Aβ_1-43 is more prone to aggregation and has higher toxic properties than the long-known Aβ_1-42. However, a direct effect on Aβ_1-43 in biomaterials of individuals carrying genetic mutations in the known AD genes is yet to be determined.

Methods: N = 1431 AD patients (n = 280 early-onset (EO) and n = 1151 late-onset (LO) AD) and 809 control individuals were genetically screened for APP and PSENs. For the first time, Aβ_1-43 levels were analysed in cerebrospinal fluid (CSF) of 38 individuals carrying pathogenic or unclear rare mutations or the common PSEN1 p.E318G variant and compared with Aβ_1-42 and Aβ_1-40 CSF levels. The soluble sAPPα and sAPPβ species were also measured for the first time in mutation carriers.

Results: A known pathogenic mutation was identified in 5.7% of EOAD patients (4.6% PSEN1, 1.07% APP) and in 0.3% of LOAD patients. Furthermore, 12 known variants with unclear pathogenicity and 11 novel were identified. Pathogenic and unclear mutation carriers showed a significant reduction in CSF Aβ_1-43 levels compared to controls (p = 0.037; < 0.001). CSF Aβ_1-43 levels positively correlated with CSF Aβ_1-42 in both pathogenic and unclear carriers and controls (all p < 0.001). The p.E318G carriers showed reduced Aβ_1-43 levels (p < 0.001), though genetic association with AD was not detected. sAPPα and sAPPβ CSF levels were significantly reduced in the group of unclear (p = 0.006; 0.005) and p.E318G carriers (p = 0.004; 0.039), suggesting their possible involvement in AD. Finally, using Aβ_1-43 and Aβ_1-42 levels, we could re-classify as "likely pathogenic" 3 of the unclear mutations.

Conclusion: This is the first time that Aβ_1-43 levels were analysed in CSF of AD patients with genetic mutations in the AD causal genes. The observed reduction of Aβ_1-43 in APP and PSENs carriers highlights the pathogenic role of longer Aβ peptides in AD pathogenesis. Alterations in Aβ_1-43 could prove useful in understanding the pathogenicity of unclear APP and PSENs variants, a critical step towards a more efficient genetic counselling.

Keywords: Alzheimer mutations; Alzheimer’s disease (AD); Amyloid-β 1–43 (Aβ1–43); Cerebrospinal fluid (CSF); Oxford Nanopore Technologies (ONT) long-read sequencing.

Fig. 1

CSF levels of Aβ_1–43, Aβ_1–43/Aβ_1–40 and Aβ_1–43/Aβ_1–42 ratios in the three mutation carrier groups compared to controls. Scatter plots show Aβ_1–43 (a), Aβ_1–43/Aβ_1–40 (b) and Aβ_1–43/Aβ_1–42 (c) CSF levels in controls and in carriers of known pathogenic mutations, of VUS and of PSEN1 p.E318G. Values of mean ± SD are given. p value indicators correspond to the values assessed with Kruskal-Wallis: *p < 0.05, ***p < 0.0001

Fig. 2

Aβ_1–43 CSF levels in carriers of known pathogenic mutations or VUS compared with the control group. The CSF levels of Aβ_1–43 for each carrier of a known pathogenic mutation (in stripes) or a VUS (in gray) are shown in the bar plots together with the control group (in black). Error bars indicate the SD of the duplicate measurements for the mutation carriers and the average of the values for the controls. The asterisks (*) indicate the carriers of one APOE ε4 allele

Fig. 3

CSF levels of Aβ_1–42 and Aβ_1–42/Aβ_1–40 ratio in the three mutation carrier groups compared to controls. Scatter plots show Aβ_1–42 (a) and Aβ_1–42/Aβ_1–40 (b) CSF levels in controls and in carriers of known pathogenic mutations, of VUS and of PSEN1 p.E318G. Values of mean ± SD are given. p value indicators correspond to the values assessed with Kruskal-Wallis: *p < 0.05, **p < 0.01, ***p < 0.0001

Fig. 4

CSF levels of sAPPα, sAPPβ, Aβ_1–43/sAPPα and Aβ_1–43/sAPPβ ratios in the three mutation carrier groups compared to controls. Scatter plots show sAPPα (a), sAPPβ (b), Aβ_1–43/sAPPα (c) and Aβ_1–43/sAPPβ (d) CSF levels in controls and in carriers of known pathogenic mutations, of VUS and of PSEN1 p.E318G. Values of mean ± SD are given. p value indicators correspond to the values assessed with Kruskal-Wallis: *p < 0.05, ***p < 0.0001

Fig. 5

Transcript analysis of PSEN1 in patient 16. The bar graph shows the relative quantifications of exon 6 in the double carrier (patient 16, PSEN1 p.G183V, p.P49L), single carrier (PSEN1 p.G183V) and non-carrier lymphoblast cells CHX treated (CHX) and untreated (UNT). Relative quantifications of splice junctions were calculated by dividing the number of junction-supporting reads by the total number of reads spanning the PSEN1 transcript. The quantifications for both CHX and UNT of the non-carriers are reported as averages (values of SD for CHX ± 0.001052869 and for UNT ± 0.000671837) (a). Visualization of the PSEN1 exon 6 cDNA MinION reads from Integrative Genomic Viewer software (IGV). Sequencing reads of PSEN1 cDNA of the double and single carriers confirm exon 6 skipping due to the PSEN1 p.G183V mutation (b)