Alpha-synuclein (aS) amyloid formation is involved in Parkinson’s disease (PD); therefore, small molecules that target aS and affect its aggregation are of interest as future drug candidates. We recently reported modified ring-fused 2-pyridones that modulate aS amyloid formation in vitro. Here, we describe the effects of such molecules on behavioral parameters of a Drosophila model of PD (i.e., flies expressing human aS), using a new approach (implemented in a commercially available FlyTracker system) to quantify fly mobility. FlyTracker allows for automated analysis of walking and climbing locomotor behavior, as it collects large sequences of data over time in an unbiased manner. We found that the molecules per se have no toxic or kinetic effects on normal flies. Feeding aS-expressing flies with the amyloid-promoting molecule FN075, remarkably, resulted in increased fly mobility at early time points; however, this effect switched to reduced mobility at later time points, and flies had shorter life spans than controls. In contrast, an amyloid inhibitor increased both fly kinetics and life span. In agreement with increased aS amyloid formation, the FN075-fed flies had less soluble aS, and in vitro aS-FN075 interactions stimulated aS amyloid formation. In addition to a new quantitative approach to probe mobility (available in FlyTracker), our results imply that aS regulates brain activity such that initial removal (here, by FN075-triggered assembly of aS) allows for increased fly mobility.

Transthyretin amyloidosis (ATTR) is a fatal disease caused by the systemic aggregation and deposition of transthyretin (TTR), a blood transporter that is mainly produced in the liver. TTR deposits are made of elongated amyloid fibrils that interfere with normal tissue function leading to organ failure. The current standard care for hereditary neuropathic ATTR is liver transplantation or stabilization of the native form of TTR by tafamidis. In our previous work, we explored an additional strategy to halt protein aggregation by capping pre-existing TTR fibrils with structure-based designed peptide inhibitors. Our best peptide inhibitor TabFH2 has shown to be effective at inhibiting not only TTR aggregation but also amyloid seeding driven by fibrils extracted from ATTR patients. Here we evaluate the effects of peptide inhibitors in two Drosophila models of neuropathic ATTR and compared their efficacy with diflunisal, a protein stabilizer currently used off-label for the treatment of ATTR. Our peptide inhibitor TabFH2 was found the most effective treatment, which resulted in motor improvement and the reduction of TTR deposition. Our in vivo study shows that inhibiting TTR deposition by peptide inhibitors may represent a therapeutic strategy for halting the progression of ATTR.

Transthyretin amyloidosis (ATTR) is a fatal disease caused by the systemic aggregation and deposition of transthyretin (TTR), a blood transporter that is mainly produced in the liver. TTR deposits are made of elongated amyloid fibrils that interfere with normal tissue function leading to organ failure. The current standard care for hereditary neuropathic ATTR is liver transplantation or stabilization of the native form of TTR by tafamidis. In our previous work, we explored an additional strategy to halt protein aggregation by capping pre-existing TTR fibrils with structure-based designed peptide inhibitors. Our best peptide inhibitor TabFH2 has shown to be effective at inhibiting not only TTR aggregation but also amyloid seeding driven by fibrils extracted from ATTR patients. Here we evaluate the effects of peptide inhibitors in two Drosophila models of neuropathic ATTR and compared their efficacy with diflunisal, a protein stabilizer currently used off-label for the treatment of ATTR. Our peptide inhibitor TabFH2 was found the most effective treatment, which resulted in motor improvement and the reduction of TTR deposition. Our in vivo study shows that inhibiting TTR deposition by peptide inhibitors may represent a therapeutic strategy for halting the progression of ATTR.SIGNIFICANCE STATEMENTFamilial Amyloid Polyneuropathy (FAP) is a hereditary condition caused by the deposition of transthyretin (TTR) in nerves. Marked by progressive deficit and disability, FAP has no cure and limited therapeutic options. The replacement of the production source of mutant TTR by liver transplantation and the stabilization of native TTR by compounds, current lines of treatment, often fail to halt disease progression. Previously, we discovered that two segments of TTR drive amyloid deposition, and designed structure-based peptide inhibitors. Here we evaluate these peptide inhibitors in FAP models of Drosophila. The most efficient inhibitor resulted in an improvement of locomotor abilities and a reduction of TTR deposition. This study points to peptide inhibitors as a potential therapeutic strategy for FAP.