JPND - Multicellular organoids: modeling, mechanisms and therapy development for C9ORF72-associated neurodegeneration.

Lead Research Organisation: University of Sheffield
Department Name: Neurosciences

Abstract

Amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) are devastating disorders. The most genetic cause of ALS and FTD is linked to mutations in C9ORF72 gene. While significant discoveries have recently been made in the genetics of ALS/FTD, patients still have no real therapeutic treatments. The lack of any cure for ALS-linked C9ORF72 is attributable primarily to: (i) Poor understanding of the molecular pathogenesis of neurodegeneration; (ii) Lack of reliable animal models mimicking the multicellular and multi-mechanism complexity of the human disease; (iii) Delivery of therapeutically attractive molecules has been hampered by inefficient delivery methods including factors like the blood-brain barrier; (iv) Ineffective targeting of therapeutic agents specifically to the diseased sites of the brain and spinal cord. Research efforts aimed at understanding how mutations in ALS causative genes leads to motor neuron injury are of the utmost importance to enable therapeutic development for these disorders. The limitations associated with existing C9orf72 models can be overcome by using specific patient cells - derived 3D models named organoids. Thus, organoids provide unique opportunities as a system for the development of pharmacological or tailored gene therapies for C9ORF72-linked neuronal injury. Here, we assembled a multidisciplinary research team with complimentary expertise to examine strategies to overcome some of these challenges. The overall aims of our research programme are: (1) Generate 3D in vitro multicellular organoids models from iPSCs derived from healthy and patients with C9ORF72 ALS/FTD; (2) Fully characterise the newly generated models using established assays in the consortium (e.g. molecular markers and electrophysiology); (3) Explore how neuronal injury happen in ALS; (4) develop treatment for C9orf72 linked ALS..

Technical Summary

The delivery of therapeutic agents to the central nervous system remains a major challenge. This is especially the case for the devastating disorders at the focus of the current proposal [amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD)]. The most common genetic form of ALS/FTD is caused by expansion of a hexanucleotide repeat (G4C2) in the non-coding region of chromosome 9 open reading frame 72 (C9ORF72). The lack of any cure for ALS-linked C9ORF72 is attributable primarily to: i) Poor understanding of the molecular pathogenesis of neurodegeneration, the contribution of each cell type to the disease and the underlying causes of the variability that characterises the patient population; ii) Lack of reliable animal models mimicking the multicellular and multi-mechanism complexity of the human disease; iii) Delivery of therapeutically attractive molecules has been hampered by inefficient delivery methods including factors like the blood-brain barrier; iv) Ineffective targeting of therapeutic agents specifically to the diseased CNS site and/or cell type. Research efforts aimed at understanding how mutations in ALS causative genes affect various cell types and pathways, thus leading to motor neuron degeneration are of the utmost importance to enable therapeutic development for these disorders. The limitations associated with existing C9orf72 animal models can be overcome by using specific patient induced pluripotent stem cells (iPSC)- derived 3D models. Here, we assembled a multidisciplinary team with complimentary expertise to overcome some of these challenges. The overall aims of our research programme are: 1) Generate 3D in vitro multicellular organoids models from iPSCs derived from healthy and patients with C9ORF72 ALS/FTD; 2) Fully characterise the newly generated models using established assays in the consortium; 3) Explore mechanistic pathways (e.g. pro-inflammatory, anti-oxidant); 4) Therapeutic screening in collaboration with AstraZeneca.

Planned Impact

The current proposal will use multidisciplinary approaches to achieve 3D modeling, investigate mechanistic pathways and therapy development for C9ORF72-linked ALS/FTD. The expected outcomes at completion are:

Short- and mid-term
Generation of the 3D organoid system would provide a great toll for:
1) Drug screening: the 3D model generated from healthy and C9ORF72 patients will be used to test a range of compounds. This will allow establishment of partnerships with other pharma companies [in addition to AZ (see MICA)]. The promising targets will be tested in animal models of disease. We are already in conversation with our patent office to discuss IP on the use of this 3D tool for drug screening.
2) The 3D system will be useful for 3Rs: Replacement, Reduction and Refinement. A Pubmed search using the terms: spinal cord, neurodegeneration and either mouse or rat returned 703 studies in 2017 alone. Considering each study reports data from an average of 44 animals, we estimate that >30,100 rodents were used in 2017 alone to study the neurodegeneration process in the spinal cord. Once proven functional and relevant to complex CNS studies, we expect other groups to set up this system. To facilitate the process, we will deposit our cells, protocols and data in public repositories. If this system will replace even only 1% of the animals used in 1 year, this would correspond to replacement of about 3,000 animals/year. In addition, the spinal cord organoids (SCO) generated here will be more relevant to study disease mechanisms and achieve patient stratification, one of the major challenges in the treatment of neurodegenerative diseases.
3) Data from WP3 will allow identification of new pathways and therapeutic targets

Long term and implementation
Successful generation of efficacy proof-of-concepts based on drug and gene therapies (WP4 and WP5) will likely lead to clinical development of the best targets. This will be done as follows:
1) Data generated from WP4 and WP5 will allow initiation of clinical development: a) presentation of the preclinical package to regulatory bodies as part of scientific advisory meetings (EMA, MHRA); b) GLP regulatory safety studies in rodents; c) GMP manufacturing of clinical products; d) Preparation and submission of Clinical Trial Application (CTA), e) Recruitment and initiation of clinical trials in C9ORF72 patients.
2) IP generation and potential commercialisation: commercialisation of our potential products can be achieved: a) establishment of start-up to take the product forward; b) or partnership with biotech or pharma companies; c) or licencing the product to a private entity.

Publications

10 25 50
 
Description Gene Therapy Innovation and Manufacturing Centre
Amount £500,000 (GBP)
Organisation University of Sheffield 
Sector Academic/University
Country United Kingdom
Start 12/2021 
End 07/2022
 
Description Non-pay BRC funding
Amount £500 (GBP)
Organisation Sheffield Teaching Hospitals NHS Foundation Trust 
Sector Public
Country United Kingdom
Start 11/2021 
End 12/2021
 
Title 3D spinal cord model 
Description Cells forming the spinal cord have been bio printed in gelatine to study their interaction in 3D 
Type Of Material Technology assay or reagent 
Year Produced 2021 
Provided To Others? No  
Impact There are no existing spinal cord organoids available, hence this is one of the first multi-cellular models that will be generated to study complex interactions between cells and the effect of therapeutic agents 
 
Description Gene Therapy Innovation and Manufacturing Centre (GTIMC) 
Organisation Cell and Gene Therapy Catapult
Country United Kingdom 
Sector Private 
PI Contribution Project funded by the MRC and LifeArc to establish a Gene Therapy Innovation & Manufacturing Centre (GTIMC) in Sheffield. Several partners across the North of England, Midlands and Wales are participating in this initiative. Lead: Mimoun Azzouz
Collaborator Contribution Cell & Gene Therapy Catapult: - Tech transfer AAV platform. - Assay standardisation. - Training through ATAC. - Regulatory engagement University of Sheffield: - GTIMC management - GMP manufacturing. - Gene therapies delivery to patients - Process innovation - Training and skills - Regulatory engagement MW- ATTC: - Clinical pathways for gene therapy delivery - Training Genomic Laboratory Hub: - Access to clinicians, patients and research groups in rare disease - Networking links to other Genomic Laboratory Hubs Lonza AG - Downstream manufacturing AAV platforms for commercialisation - Training and placements Birmingham Women's and Children's Hospital - Gene therapy trials in haematological, metabolic, and neuromuscular disorders Leeds Teaching Hospitals - Delivery of AAV therapies to patients with rare diseases - Training and skills University of Liverpool: - AAV safety and immunogenicity - Genomic Laboratory Hub North-West: Rare diseases NIBSC: - Reference materials - Standardising analytical methods - Training in regulatory sciences. Universities of Bradford & York: - Rare diseases - Training and skills University of Leicester - Data sciences - EU rare diseases network CPI: - Process development and scale-up for AAV. - Scalable manufacturing solutions transfer to ShefVec. Cobra Biologics: - GMP grade plasmid and vector for late phase and commercialisation. - Training: manufacturing, analytics, regulatory affairs. ATAC: - Upskill apprentices. - Use GTIMC MSc Course to develop Levels 6/7 apprenticeship programmes. Northern Health Science Alliance - Source partners through NHSA member organisations. - Secure investment into the Hubs. LifeArc Translational Support: - Regulatory engagement - IP and commercialisation
Impact The award will start in July 2021
Start Year 2021
 
Description IMI 
Organisation University of Cambridge
Department Department of Physiology, Development and Neuroscience
Country United Kingdom 
Sector Academic/University 
PI Contribution This is a large European collaboration aiming at the identification of mechanisms involved in the immune response to AAV during clinical trials. My team will be modelling the immune response in vitro using immunocompetent CNS organoids
Collaborator Contribution Other partners in the consortium will be studying immune response in vivo or in other cell types outside the CNS
Impact The collaboration has just started. Present outputs are formed by databases, animal models and cell resources.
Start Year 2020
 
Description JPND-Multicellular organoids: modeling, mechanisms and therapy development for C9ORF72-associated neurodegeneration 
Organisation JPND Research
Country Global 
Sector Academic/University 
PI Contribution Lead applicant on this JPND Award. Project is scheduled to start May 2020
Collaborator Contribution Coordinator UK, Mimoun Azzouz, University of Sheffield (SITraN) Partner 2: Sweden, Elena Kozlova, University of Uppsala (UU) Partner 3: UK, Nicola Hamilton, King's College London (KCL) Partner 4: Turkey, Esra Cagavi, Istanbul Medipol University (IMU) Partner 5: Finland, Tarja Malm, University Eastern Finland (UEF) Partner 6: France, Guilhem Velvé-Casquillas, Elvesys.
Impact Project is starting in May 2020
Start Year 2020
 
Description Visit to Uppsala 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Postgraduate students
Results and Impact The post-doc employed on the project gave an overview of ALS and the JPND project to students and staff at the University of Uppsala
Year(s) Of Engagement Activity 2021