The pathological hallmark of a range of neurodegenerative diseases, including Motor Neurone Disease (MND), is the presence of protein aggregates in affected neurons. Protein aggregates progress continually from the site of onset to disturb cellular processes, ultimately resulting in neuron degeneration [1]. The only intracellular degradative pathway that can purge the cells of these aggregates and dysfunctional organelles is autophagy. Since impaired/reduced autophagy may contribute to MND pathogenesis, upregulating autophagy offers a potential therapeutic option.
This study aimed to design and synthesise novel pharmacological agents with non-toxic autophagy-inducing activity in motor neuronal NSC-34 cells. A series of peptide analogues of the evolutionarily conserved domain of Beclin-1 protein, the master regulator of autophagy, were synthesised. To enhance the cellular uptake and the cytosolic bioavailability of peptides, they were cyclised through disulphide, thioether and lactam bond formation and conjugated to a cell-penetrating peptide (CPP) domain (a fragment of apolipoprotein E) and an endosomal escape (EE) domain (a fragment of hemagglutinin-2 protein). The mRFP-GFP-LC3 tandem fluorescent protein was used to determine the autophagy-inducing effect of the peptides. We have shown that the peptides can induce autophagy at 5 micromolar concentration. We have also demonstrated that cyclisation and the addition of a CPP and EE domain to the peptides further increase their autophagy-inducing potency. More importantly, the most potent peptide analogue significantly decreased the levels of aggregate-prone C9orf72 dipeptide repeat protein and mutant SOD1 protein associated with MND. Subsequently, the Beclin peptide reduced the toxicity associated with the expression of these aggregate-prone proteins by reducing cleaved caspase-3 activation. Moreover, the autophagy effect of the peptide on motor neurons in autophagy reporter mice was analysed. In conclusion, this study provides evidence for a novel peptide-based approach to enhance autophagy as a unique and safe treatment modality with potential therapeutic benefits in MND and other proteinopathies.