A molecular biofluid signature of multiple system atrophy (MSA): CSF neurofilament light chain and α-synuclein seeding as complementary biomarkers allow to distinguish MSA from sporadic adult-onset ataxia

BackgroundAccurate diagnosis of multiple system atrophy (MSA) is critical for clinical management and efficient trial designs, yet remains challenging, particularly distinguishing MSA (especially cerebellar-subtype [MSA-C]) from sporadic adult-onset ataxia (SAOA). Combining a marker of neuroaxonal degeneration, neurofilament light chain (NfL), with a marker of the pathogenic MSA hallmark, -synuclein seeding activity, may define a mechanistically-informed CSF signature of MSA, enabling sensitive and specific differentiation from SAOA even in early disease. MethodsWe analyzed 60 cross-sectional patient CSF samples (n=32 clinically diagnosed MSA [MSAclin] 22/32 MSA-C; n=28 SAOA) for NfL (Simoa) and -synuclein seeding activity (seed amplification assay [synSAA], Piperazine-N,N-bis(2-ethanesulfonic acid)-based), and assessed diagnostic accuracy, disease-duration correlations, and trial power using biomarker-based stratification. ResultsAge-adjusted NfL was higher in MSAclin than SAOA (3859 vs. 997pg/mL), yielding 96.9% sensitivity and 85.7% specificity. SynSAA was concordant with clinical diagnosis (25/32 MSAclin synSAA-positive; 23/28 SAOA synSAA-negative), with 78.1% sensitivity and 85.2% specificity (all confirmed in MSA-C subgroup). Both biomarkers displayed divergent trajectories with disease duration: NfL peaked early before declining (r=-0.45, p=0.01); whereas synSAA maximum fluorescence intensity increased (r=0.42, p=0.016), suggesting greater synSAA signal with accumulating MSA burden. Integrating both biomarkers in MSA treatment trials allows sample-size reduction by 20% versus NfL alone. ConclusionsCSF NfL and synSAA capture complementary aspects of MSA biology: while NfL provides high diagnostic accuracy for MSAclin, peaking early, synSAA adds mechanistic specificity for -synuclein seeding activity and might allow target engagement assessment. Combined, they might enable biological diagnostic frameworks, molecular trial stratification, and treatment monitoring in MSA. Key messagesO_ST_ABSWhat is already known on this topicC_ST_ABSWhile highly warranted for clinical management and efficient treatment trial design, accurate diagnosis of multiple system atrophy (MSA) against overlapping and reciprocally mimicking conditions such as sporadic adult-onset ataxia (SAOA) remains clinically challenging, especially in early disease stages. A mechanistically informed biofluid signature of MSA might enable sensitive and specific differentiation from SAOA, even in early disease stage. Recently merging molecular markers reflecting neuroaxonal damage (NfL) and -synuclein seeding activity (measured by the seed amplification assay; synSAA) might here show particular promise. What this study addsThis is the first study to systematically assess the ability of both CSF NfL and CSF -synuclein seeding activity to distinguish clinically diagnosed MSA (MSAclin) from SAOA, thereby offering a window into underlying MSA biology in patients in vivo. Our findings suggest that the rate of axonal degeneration is most pronounced in early MSA disease stages but decreases with longer disease duration; whereas -synuclein seeding signal activity increases as MSA-related disease burden accumulates. Finally, it demonstrates the impact of a combined molecular fluid signature of MSA for improving trial design: a biomarker-based stratification of MSA subjects in future MSA treatment trials combining NfL plus -synuclein seeding activity allows to reduce sample sizes by 20% compared to NfL alone. How this study might affect research, practice or policyThe findings from this study may help to molecularly diagnose patients with MSA against overlapping and reciprocally mimicking conditions such as SAOA, in particular and even in early disease stages. Moreover, they might lay the foundation for a future biologically-informed diagnostic framework of MSA; support trial stratification for more efficient upcoming MSA treatment trials; and might facilitate molecular treatment effect monitoring in MSA, in particular in synuclein-targeted treatment trials.