RICERCA DI BIOMARCATORI NELLE MALATTIE NEUROLOGICHE RARE

Spinal Muscular Atrophy (SMA) is an autosomal recessive disorder due to progressive degeneration of lower motor neurons (LMN) resulting in muscle atrophy and possible respiratory and bulbar involvement. Pathogenic hallmark is the loss of function of Survival Motor Neuron Protein (SMN), encoded by mutated or deleted SMN1 gene. SMN2, the centromeric paralogous gene of SMN1, produces only a small amount of functional SMN (Levin, 2019). Nusinersen is the only disease modifying therapy approved for SMA patients. Neurofilaments light chain (NfL) are intermediate filaments exclusively expressed in neurons. As NfL levels raise following axonal damage, they might be promising diagnostic and prognostic biomarkers in motor neuron diseases (Gaetani, et al., 2019). A different intermediate filament, peripherin (PRPH) assembles with neurofilaments mostly in peripheral neurons and LMN (Zhao, 2016). Human Profilin-1 (PFN-1) is a small actin-binding protein that promotes actin polymerization and has a role in regulating cytoskeletal architecture and dynamics of neurons (Witke, 2004). This study aims to investigate the role of NfL as disease and treatment-response biomarker in a cohort of adult SMA type 2 and 3 patients. 33 SMA type 2 and 3 patients were recruited at the Neuromuscular Center of Padua Hospital, where nusinersen treatment was administered from February 2018 to September 2019 in a loading phase (L1 baseline, L2 day 14, L3 day 28, and L4 day 63) and a maintenance phase (M1-M6, every four months). Cerebrospinal fluid (CSF) samples were collected at each administration; NfL was tested at each time point, additional neurodegeneration biomarkers total tau (t-Tau) and phosphorylated tau (p-Tau) proteins were tested at time points L1 and L3. PFN-1 was tested at each time point as an exploratory muscular biomarker. Further testing of PRPH was conducted at baseline on 20 type 3 patients. NfL concentration was determined with a commercial enzyme-linked immunosorbent assay (ELISA) kit (UmanDiagnostics, Umea, Sweden). T-Tau and pTau were measured with an automated Chemiluminescent Enzyme Immunoassay (CLEIA) analyzer (LUMIPULSE G600 II by Fujirebio, Japan); PFN-1 was measured in serum with a commercial ELISA kit (Cusabio, China); PRPH was tested in serum with a commercial ELISA kit (Abbexa, UK). Both PFN-1 and PRPH results were compared each to a control group of healthy subjects (HC). Neuromuscular outcomes were tested at L1, L4, M1, M2 and M3 with appropriated validated motor scales. Baseline CSF NfL, t-Tau and p-Tau levels were overall included in the reference ranges for healthy donors. Mean NfL was 211.97 ± 180.9 ng/l in SMA patients, compared to 809.53 ± 1,065.26 ng/l in controls. Correlation was found between baseline log[NfL] and age both in SMA patients and control group. Also log[t-Tau] and log[p-Tau] correlated with log[NfL] at L1, but not at L3, although a slight significant increase was found in t-Tau and p-Tau at L3. NfL significantly increased in loading phase until L3 (mean increase 285.45 ng/l). From L4 NfL started to decrease and no significant difference was found with baseline at M1, M2 and M3. PFN-1 at baseline was higher in SMA than in healthy controls (mean 989 vs 608 ng/l, p=0.0018). PFN-1 showed a complex dynamic during loading phase, with a significant reduction at L4. PRPH was significantly higher compared to HC (median 5.653 µg/l vs 3.168 µg/l, p=0.02). No correlation was found between NfL and motor scores at each time point. Our study partially reinforces recently published results in similar patients (Wurster, et al., 2019) (Wurster, et al., 2020) (Faravelli, et al., 2020), adding insights on NfL dynamic during the first month of treatment. Neurodegenerative biomarkers might inadequately relate to long disease duration. Serum PFN-1 and PRPH might be more sensitive to reveal LMN damage, therefore subsequent studies are needed to clarify their potential prognostic value.