Abstract title: Effects of lentiviral shRNA-mediated targeting of selected genes on alpha-synuclein aggregation in cortical neurons

Poster board number (onsite): 721
Poster number: P0805 / #624

Authors: Linnea Strid Orrhult1 , Emmy Rannikko2 , Anne Vuorenpää2 , Johan Pihl3 , Paul Karila1 , Andrii Domanskyi2 , Ibrahim Malik2
Affiliations: 1Cellectricon AB, Discovery, Mölndal, Sweden, 2Orion Corporation, Discovery, Turku, Finland, 3Cellectricon AB, R&d, Mölndal, Sweden

Aims: Insoluble aggregates (Lewy bodies) consisting of the misfolded protein alpha-synuclein (αSyn) progressively accumulate in the nervous system of most Parkinson’s disease (PD) patients. In the current study, the aim was to identify targets involved in modulation of αSyn aggregation using an in vitro model.
Methods: Based on literature, 20 targets were chosen for their potential role in modulating αSyn aggregation. Lentiviral shRNAs were added to primary mouse (E18) cortical cultures and mouse pre-formed αSyn fibrils (PFFs) were added 6 days later. An unbiased, automated image analysis workflow was used to assess the effect of selected mRNAs on αSyn aggregation in NeuN-positive neurons and MAP2-positive neurites one and two weeks after addition of PFFs.
Results: Lentiviral shRNA-mediated downregulation of some targets caused effects on neuronal survival. However, one week after PFF addition, targeting Zscan21, Vps35 and Fyn resulted in an increase, whereas targeting Ppp1r15a, USP14 and Bach1 resulted in a decrease in αSyn aggregation without effects on neuronal health. Two weeks after PFF addition, targeting Atp13a2, Fyn and Aimp2 resulted in an increase, and targeting Ppp1r15a, Rhot1, Ttbk1 and Bach1 resulted in a decrease in αSyn aggregation without effects on neuronal health. For some of the targets, the effect on αSyn aggregation was thus time-dependent.
Conclusions: Our in vitro model is useful to test potential targets affecting the αSyn aggregation process over time. We provide a list of potential targets for modulation of αSyn aggregation, which should next be tested in αSyn aggregation models in human neurons and in vivo aiming to develop disease-modifying therapies for PD.