Association between CSF alpha-synuclein seeding activity and genetic status in Parkinson's disease and dementia with Lewy bodies

Abstract

The clinicopathological heterogeneity in Lewy-body diseases (LBD) highlights the need for pathology-driven biomarkers in-vivo. Misfolded alpha-synuclein (α-Syn) is a lead candidate based on its crucial role in disease pathophysiology. Real-time quaking-induced conversion (RT-QuIC) analysis of CSF has recently shown high sensitivity and specificity for the detection of misfolded α-Syn in patients with Parkinson's disease (PD) and dementia with Lewy bodies (DLB). In this study we performed the CSF RT-QuIC assay in 236 PD and 49 DLB patients enriched for different genetic forms with mutations in GBA, parkin, PINK1, DJ1, and LRRK2. A subgroup of 100 PD patients was also analysed longitudinally. We correlated kinetic seeding parameters of RT-QuIC with genetic status and CSF protein levels of molecular pathways linked to α-Syn proteostasis. Overall, 85% of PD and 86% of DLB patients showed positive RT-QuIC α-Syn seeding activity. Seeding profiles were significantly associated with mutation status across the spectrum of genetic LBD. In PD patients, we detected positive α-Syn seeding in 93% of patients carrying severe GBA mutations, in 78% with LRRK2 mutations, in 59% carrying heterozygous mutations in recessive genes, and in none of those with bi-allelic mutations in recessive genes. Among PD patients, those with severe GBA mutations showed the highest seeding activity based on RT-QuIC kinetic parameters and the highest proportion of samples with 4 out of 4 positive replicates. In DLB patients, 100% with GBA mutations showed positive α-Syn seeding compared to 79% of wildtype DLB. Moreover, we found an association between α-Syn seeding activity and reduced CSF levels of proteins linked to α-Syn proteostasis, specifically lysosome-associated membrane glycoprotein 2 and neurosecretory protein VGF.These findings highlight the value of α-Syn seeding activity as an in-vivo marker of Lewy-body pathology and support its use for patient stratification in clinical trials targeting α-Syn.

Keywords: CSF; GBA; PD; Parkin; RT-QuIC; α-Syn seeding.

Fig. 1

RT-QuIC positive replicates relative fluorescence curves at 30 h stratified by diagnosis and mutation status. A Remarkable differences in RT-QuIC α-Syn seeding profiles were detected in the PD patients’ group when stratifying by PD-associated mutations. While 93% of PD_{GBA_severe} showed a positive RT-QuIC α-Syn seeding profile, 78% of PD_LRRK2 and only 59% of PD_{recessive _heterozygous} gave a positive reaction by RT-QuIC. Strikingly, none of the PD_{recessive_bi-allelic} showed a positive RT-QuIC α-Syn seeding profile (overall p ≤ 0.001) Numbers (n) included: PD_wildtype = 107, PD_{GBA_risk} = 53, PD_{GBA_mild} = 17, PD_{GBA_severe} = 29, PD_LRRK2 = 9, PD_{recessive_heterozygous} = 17, PD_{recessive_biallelic} = 3, DLB_wildtype = 33, DLB_GBA = 16. B While PD_{GBA_severe} patients showed the strongest RT-QuIC seeding kinetics measured by the mean relative fluorescence (RFU) of positive curves (see comparison with PD_wildtype), PD_{recessive_bi-allelic} did not seed at all.

Fig. 2

Longitudinal trajectories of RT-QuIC seeding profiles. Out of the 100 PD patients included in this longitudinal analysis, 86 were RT-QuIC seeding positive while 14 did not show seeding. A Linear mixed model analysis in the longitudinal PD cohort with repeated lumbar puncture and RT-QuIC assay measurements revealed that time had no significant impact on the number of positive RT-QuIC seeding replicates over the course of the disease, neither in those patients who were seeding positive nor in those who were seeding negative (p = 0.670). B, C Similarly, RT-QuIC seeding positives showed stable results of Imax (p = 0.558) and LAG phase (p = 0.324) over time