To fight neurodegenerative diseases, several therapeutic strategies have been proposed that, to date, are either ineffective or at the early preclinical stages. Intracellular protein aggregates represent the cause of about 70% of neurodegenerative disorders, such as Alzheimer’s disease. Thus, autophagy, i.e., lysosomal degradation of macromolecules, could be employed in this context as a therapeutic strategy. Searching for a compound that stimulates this process led us to the identification of a 37/67kDa laminin receptor inhibitor, NSC48478. We have analysed the effects of this small molecule on the autophagic process in mouse neuronal cells and found that NSC48478 induces the conversion of microtubule-associated protein 1A/1B-light chain 3 (LC3-I) into the LC3-phosphatidylethanolamine conjugate (LC3-II). Interestingly, upon NSC48478 treatment, the contribution of membranes to the autophagic process derived mainly from the non-canonical m-TOR-independent endocytic pathway, involving the Rab proteins that control endocytosis and vesicle recycling. Finally, qRT-PCR analysis suggests that, while the expression of key genes linked to canonical autophagy was unchanged, the main genes related to the positive regulation of endocytosis (pinocytosis and receptor mediated), along with genes regulating vesicle fusion and autolysosomal maturation, were upregulated under NSC48478 conditions. These results strongly suggest that 37/67 kDa inhibitor could be a useful tool for future studies in pathological conditions.

Mitochondrial dysfunction is a core cellular abnormality in Down syndrome (DS), yet the mechanisms underlying impaired mitochondrial quality control remain poorly defined. Here, we provide an integrated characterization of the autophagy–mitophagy–lysosome axis in human trisomic fetal fibroblasts (DS-HFFs). We show that trisomic cells display a paradoxical combination of elevated basal autophagy/mitophagy-related markers and defective mitochondrial clearance, indicative of a saturated and inefficient quality control system. DS-HFFs exhibit increased steady-state levels of PINK1, PARKIN, OPTN, NDP52, LC3-II, and p62, accumulation of autophagic vacuoles, reduced autophagic flux, and impaired delivery of mitochondria to lysosomes. We further demonstrate that metformin alleviates the saturation of the autophagy–mitophagy pathway by modulating multiple components of the mitochondrial quality control pathway, including attenuation of mTOR-associated signaling, enhancement of autophagic flux and normalization of mitophagy-related protein levels. Crucially, metformin increases mitochondria–lysosome association and rescues lysosomal degradative competence as shown by increased Dq-BSA activity, and increased maturation of Cathepsin D. Together, our findings support mitophagy inefficiency due to pathway saturation as a key pathogenetic mechanism in trisomic cells and identify metformin as a pharmacological tool capable of relieving the proteostatic overload intrinsic to trisomy 21. More broadly, these results align with a model in which aneuploidy imposes a chronic proteostatic burden that compromises organelle quality control.

Protein-misfolding neuropathic diseases are characterized by the accumulation in the central nervous system of misfolded proteins, whose irreversible change into beta-sheets sticky conformation facilitates the protein to form aggregates, which represent the main commonality shared by these disorders, including Prion diseases, Alzheimer’s disease, Parkinson's disease, Amyotrophic lateral sclerosis and Frontotemporal dementia. To explain the mechanisms through which protein misfolding and aggregation lead to the development of these diseases, at least three models have been proposed. Besides the widely accepted gain- or loss-of-function model for misfolded proteins, the inflammation model proposes that abnormal protein aggregates act as culprits causing chronic inflammatory response that leads to cell death. Historically, inflammatory events were viewed as result of toxicity from the inability of the cell to clear misfolded proteins. However, how misfolded proteins cause inflammation remains still an open issue. Given the current lack of an overarching view describing the connection between protein misfolding-related neurodegenerative diseases and the inflammasome, we aim to outline this relationship in the present review. We integrate recent insights into the mechanistic links between protein misfolding and NLRP3 inflammasome activation, highlighting the impact of the recently described “prionoid-activity” of ASC specks on the progression of different neurodegenerative misfolding diseases.