OSTEOPONTIN AS A PROGNOSTIC BIOMARKER IN AMYOTROPHIC LATERAL SCLEROSIS

Background: Ostepontin is a crucial regulator of microglia activity, and has been implicated in gray matter atrophy in multiple sclerosis (MS) and Alzheimer’s disease (AD). In Amyotrophic Lateral Sclerosis (ALS), an invariably fatal neurodegenerative disorder affecting upper and lower motor neurons (MN), osteopontin (SPP1) was found to be upregulated in perivascular fibroblasts before any microglia activation in the presymptomatic phase in sporadic ALS patients. Objectives: Osteopontin could be a novel prognostic biomarker, reflecting vascular non-cell autonomous processes modulating disease progression in ALS. We aimed to correlate osteopontin (SPP1) levels with other known biomarkers of neurodegeneration and neuroinflammation, and to explore its relation with clinical variables of disease progression in ALS. Methods: Serum and CSF samples from ALS patients (n=76) and age and sex-matched controls (n=36), followed up from the 1st September 2008 to the 1st May 2025 in the Neurology Unit of A.O.U. Modena, were employed for biomarker analysis by Ella semi-automated immunoassay. Neurofilament Light (NfL) and Heavy Chain (NfH), Chitinase-3-like protein 1 (CHI3L1), SerpinA1, Triggering Receptor Expressed On Myeloid cells 2 (TREM2) were quantified together with SPP1. Correlations with clinical and demographic variables were run with Spearman correlation analysis and comparisons were performed with Mann-Whitney U tests or Dunn test with pairwise comparisons and Bonferroni correction. Cox regression analysis was used to confirm SPP and other clinical and biomarker variables prognostic value. Results: SPP1 concentrations in serum and CSF significantly correlate (rho=0.26, P=0.006). In the CSF, and to a minor extent, also in serum, SPP1 levels associate to TREM2 (rho=0.39, P=0.004 and rho=0.21, P=0.054, respectively), but did not show any association with the other biomarkers. SPP1 in serum rose according to King’s staging (Stage 0-1: median: 36.15, IQR:26.60-54.83; stage 2: median: 58.63, IQR: 39.94- 76.63, stage 3: median: 57.67, IQR: 43.24-114.71; P=0.006). Though CSF SPP1 did not correlate with ALSFRS-R or disease progression rate at sampling (rho=0.013, P=0.91), it associated with the monthly decline in ALSFRS-R from the time of sampling to the last observation (rho=0.25, P=0.029). In Cox univariate analysis corrected for age at sampling, CSF SPP1 non-significantly predicted worse prognosis (HR= 1.002, 95%CI 1-1.004, P=0.16), while in multivariate adjusted analysis with all the biomarkers, it confirmed as a negative independent prognostic factor (HR=1.006, 95%CI 1.003 -1.009, P<0.0001), together with serum NfL and CSF CHI3L1, while CSF TREM2 appeared as protective factor (HR=0.95, 95%CI: 0.92- 0.98, P<0.0001). Discussion: SPP1 confirmed as negative prognostic factor for ALS, independent of other well-known markers of neurodegeneration, as neurofilaments, or glial activation, as CHI3L1. Interestingly, SPP1 levels in CSF correlate with another marker of microglial regulation, TREM2, though the latter showed an opposite effect on ALS progression. This might suggest a complex interplay between non-motor neuron cells inside the central nervous system after the onset of ALS, which deserves further research in neuropathology or in vivo studies.