A growing body of data indicates that the propagation of pathogenic protein aggregates across neural systems could be mediated by misfolded protein seeds that are released and taken up by anatomically connected neurons causing disruption of their function. Therefore, blocking this process may help arrest the progression of Parkinson’s (PD) or Alzheimer’s (AD) disease. The Consortium IMPRiND, funded by the Innovative Medicine Initiative (IMI), is a group of European academic laboratories and members of the European Federation of Pharmaceutical Industries and Associations (EFPIA) that aims to delineate and target critical steps in the propagation of α-synuclein (α-syn) and tau assemblies between neurons. Our programme is collaborative and mobilizes diverse expertise in order to deliver disease-relevant phenotypes suitable for high throughput screening and validation models using iPSC, organotypic cultures and animal models.
Over the fifth and final year, we have made significant progress in delivering on our objectives. This has been facilitated by regular teleconferences and one virtual Consortium meeting.
The main research efforts of IMPRIND are focused on the following areas: (a) Generation of brain-derived or amplified assemblies and their characterisation using biochemical techniques, (b) Cryo-EM structure of tau and α-syn assemblies, (c) Conduction of CRISPR screens in cell lines followed by cross-validation in primary neuronal screens, (d) bioinformatic integration and analysis of screens based on druggability and gene expression data, (e) development of advanced cell-based models or animals for validation of selected targets and (f) Data dissemination. Over the duration of the project we have achieved the following objectives:
(i) LMB, Lilly and Janssen have optimised and cross-validated protocols to isolate and quality control brain-derived tau assemblies from Alzheimer’s disease brain. CNRS has established methodologies for amplification of proteopathic assemblies from brain homogenates of -synucleinopathies.
(ii) LMB has published the Cryo-EM structures of tau filaments isolated from Alzheimer’s disease, Pick’s disease and chronic traumatic encephalopathy (Nature, 2017, 2018, 2019, 2021) as well as Cryo-EM structures of MSA filaments (Nature 2020). CNRS has published the Cryo-EM structure of de novo generated (eLife 2019) and brain amplified -syn fibrils (BioRxiv 2021).
(iii) Development of assays suitable for high through-put screening and completion of primary screens were completed by UOXF, Novartis and HLU. These include focused CRISPR/Cas9 proteostasis screens for syn and tau, genome-wide CRISPR screens for tau, primary neuronal screens using siRNA for 300 targets for tau and the DUB screen for -syn.
(iv) Hits from these screens were further analysed in an integrated fashion by Lilly to prioritise targets for further validation in primary neurons.
(v) Validation of prioritised hits was performed by Novartis and HLU in primary neurons using siRNA or shRNA for Tau and siRNA for ⍺-syn. Janssen also assessed selected hits using CRISPR/Cas9 knockout in primary neurons. Further assessment of selected hits and additional targets for Tau was performed at Janssen using CRISPR/Cas9 knockout in primary neurons and for -synuclein at Cellectricon using a shRNA library prepared at DZNE.
(vi) A number of advanced models have been fully characterised and their suitability for target validation has been established. These include the iPSC-based dopaminergic neuronal model (UOXF, Nature Comm 2021), organotypic cultures (DZNE, Mol Neurodegener. 2021) and in vivo mouse (BRFAA and HLU) models for -syn. Tau seeding mouse models have been established by Lilly and Janssen. The transplantation of iPSC-derived neurons for in vivo assessment of Tau propagation and the tau drosophila model (both VIB) have also been completed. iPSC-based tau organoids (UCAM) and hNP-based tau neuronal model (DZNE) have also undergone initial characterisation. UBx has developed a model of tau propagation in primates using AD brain derived fibrils
(vii) Different AAV mediated silencing approaches were tested and characterized in vivo. Janssen, BRFAA and HLU have optimised the delivery method for effective knockdown of hits from the genetic screens using SNCA and MAPT RNAi in proof of principle studies. Knockdown of targets is on-going.
(viii) We generated an on-line registry for critical reagents and initiated the deposition of data in public repositories to enable sharing of datasets or dissemination following publications. Website updates and news, social media messages, and a number of conference presentations and posters have completed the dissemination activities. We have also organised two international symposia during the biennial ADPD meetings in 2019 and 2021.
