VGF: a novel and predictive biomarker of Parkinson’s disease

Parkinson’s disease (PD) is a progressive neurodegenerative disorder characterized by a prodromal phase in which non-motor symptoms precede the onset of classical motor impairments. Identifying reliable biomarkers capable of detecting PD during these early stages remains a major clinical challenge. VGF is a neurosecretory precursor protein that undergoes extensive processing to generate multiple neuronal products (peptides/proteins) with neuromodulatory functions. Accumulating evidence indicates that proVGF and several of its derived products—particularly those encompassing the proVGF C-terminus, NAPP, and TLQP amino acid sequences—are dysregulated at both central and peripheral levels in PD animal models that recapitulate early and advanced stages of the disease. Importantly, alterations in proVGF C-terminal-derived products have also been reported in drug-naïve PD patients; however, no evidence is currently available regarding changes in the NAPP and TLQP regions of the proVGF in these patients. Furthermore, the temporal- and region-specific dynamics of proVGF and its processed products throughout PD progression remain poorly understood. The present study aimed to quantify specific VGF-derived products—involving the proVGF C-terminus, NAPP, TLQP, and AQEE amino acid sequences—and to identify those exhibiting early alterations in prodromal PD by integrating data from a prodromal/progressive PD animal model and human plasma samples. Material and methods: A PD mouse model based on chronic intranasal administration of rotenone was employed (n = 26 PD vs. n = 26 control mice). Animals were evaluated at multiple time points (2, 3, and 6 months post-treatment) to capture prodromal and progressive stages of the disease. Behavioral assessments were performed at each point including both motor and non-motor evaluations, such as olfactory testing. Molecular analyses comprised the quantification of mRNA levels of VGF, α-synuclein, and tyrosine hydroxylase, alongside parallel protein-level analyses performed by immunofluorescence and/or ELISA in brain regions relevant to PD pathology (olfactory bulb and substantia nigra) and in plasma. To investigate the temporal dynamics of VGF processing during disease progression, specific antibodies targeting distinct VGF-derived products were used and selected on the basis of previous findings. In addition, an antibody against AQEE-related products—previously found to be implicated in other diseases—was employed to assess the specificity of VGF sequence alterations in PD. To assess the translational relevance of the experimental findings, human plasma samples were analyzed using competitive ELISA with the corresponding human VGF antibodies to determine whether alterations observed in the animal model were recapitulated in patients. The human cohort included drug-naïve PD patients (n = 20), treated PD patients (n = 50; equally divided between short-term–treated and advanced-stage cases), and age-matched healthy controls (n = 39). Additionally, individuals diagnosed with REM sleep behavior disorder (RBD), a condition widely regarded as prodromal to PD (n = 20), along with age-matched controls, were included. To evaluate disease specificity, plasma samples from individuals with other neurodegenerative, neurological, and autoimmune disorders were also analyzed (total n = 122, including patients and age-matched controls).Results: In rats, non-motor alterations—specifically olfactory dysfunction—were detected at 3 months, preceding the onset of motor deficits and gastrointestinal dysfunction, which became evident at 6 months. Olfactory impairment at 3 months was associated with α-synuclein accumulation and dopaminergic dysfunction in both the olfactory bulb and substantia nigra (persisting also at 6 months), whereas corresponding mRNA levels remained unchanged until 6 months. Importantly, at 3 months, concomitant with these early pathological changes, several VGF-derived prod