Dysregulated WNT signaling as a driver of motor neuron loss in Amyotrophic Lateral Sclerosis

Amyotrophic Lateral Sclerosis (ALS) (also known as motor neuron disease), is caused by a loss of motor nerves or neurons. These nerves carry signals from the brain and spinal cord to the muscles for movement and breathing. Loss of motor nerves in these patients leads to progressive muscle paralysis and eventually death due to breathing problems. About ~10% of ALS cases have a family history with mutations identified in several genes with diverse functions. However, the molecular mechanisms driving motor neuron loss in ALS are still poorly understood. Understanding these mechanisms is paramount in developing therapies that can halt or even reverse the relentless neuronal loss.

With the advent of a new stem cell technology called reprogramming, skin or blood cells from patients can be converted into an embryonic state. These reprogrammed cells are called induced pluripotent stem cells (iPSC) and have the potential to be converted to any cell type in the body. Consequently, iPSC's from ALS patients can now be coaxed to become motor neurons i.e. the same kind of cells that are lost in ALS patients. This technology enables the development of human models of ALS in a dish, allowing scientists to interrogate what goes wrong within these cells to cause them to die in ALS. Using such models, I have uncovered that an important signalling pathway called WNT may be responsible for driving MN death in ALS.

Signalling pathways within a cell are similar to specific applications on a smart phone. Each pathway is designed to perform a specific task. Signalling pathways can communicate with other pathways to execute complex functions within a cell. Inappropriate increase or decrease in the activity of a pathway can result in serious consequences for a cell. One such pathway is the WNT signalling pathway. This pathway functions in most cells in the human body, including the nervous system. Within the nervous system, it regulates important functions such as the structure of neuron and how neurons communicate with each other. Given its important, WNT signalling is tightly regulated.

Data generated in my lab indicates that this pathway is aberrantly activated in motor nerves in ALS patients. Additionally, we found that if this pathway is inhibited using drugs, ALS motor nerves live longer in a dish. However, we do not understand how activation of this pathway causes motor nerves to die and whether inhibiting this pathway will have other unintended consequences on motor nerves. This proposal will use the iPSC technology to generate motor nerves from ALS patients and use these nerves to understand the details of how WNT functions in ALS. The results of this study will generate deeper insights into why ALS motor nerves die and highlight whether WNT can be targeted as a therapy for ALS.

Amyotrophic Lateral Sclerosis (ALS) (also known as motor neuron disease), is a fatal neurodegenerative disease characterized by a loss of motor neurons (MNs). Understanding why these neurons die is critical if we hope to develop therapeutics to halt or reverse the relentless neurodegeneration in ALS patients. My preliminary work using in vitro ALS models have uncovered a role of WNT signalling in causing ALS MN death. This proposal will detail the mechanism of how WNT dysregulation in ALS MNs leads to MN death. I will also explore the role of WNT in MN homeostasis and investigate whether targeting WNT is a viable approach to improve MN function in ALS.

Although, most ALS cases are sporadic, ~10% of cases have a genetic basis with mutations found in genes with diverse functions including SOD1(free radical scavenger) and FUS(RNA processing). SOD1 is the second most common cause of genetic ALS, while recent evidence has uncovered shared pathological links between FUS ALS and sporadic ALS. I have developed in vitro models of ALS-associated MN dysfunction using induced pluripotent stem cells (iPSCs) derived from ALS patients with mutations in SOD1 and FUS. My preliminary work utilizing these models uncovered that canonical WNT signalling is activated in both SOD1 and FUS MNs. Using bioinformatics analysis of independent gene expression datasets from familial FUS and TDP43 ALS MNs as well as sporadic ALS MNs, I uncovered that WNT activation may be a common theme in ALS. I will use the iPSC-based models I have developed to gain a mechanistic understanding into how WNT activation disrupts MN homeostasis and leads to MN dysfunction or death in ALS. These investigations will advance our understanding of the underlying cause of MN death in ALS and highlight WNT as a potential target to develop ALS therapeutics.

The immediate beneficiaries of this research will be scientists trying to decipher the molecular networks underlying motor neuron degeneration in ALS. In addition to this scientific community, the project findings are expected to be beneficial to the following communities in the short and long term as follows:
1. Academic community: The results from this study will highlight WNT, an important signalling pathway as a potential driver of neurodegeneration in ALS and generate mechanistic data into the role of WNT in motor neurons. WNT has also been implicated in other late-onset neurodegenerative disorders such as Alzheimer's disease, Parkinson' disease and Huntington's disease, and will be of interest to the community of scientists studying late-onset neurodegeneration. Additionally, these results will also prove valuable to researchers investigating the role of WNT in neuronal development and homeostasis. All the phenotypic characterizations will be made available through publications and conference presentations. The genomics datasets will be deposited in public repositories as described below.

2. The genomics data generated in this project will be made publicly available by depositing the raw files in ArrayExpress or the Gene Expression Omnibus. These datasets will outlive the grant period and enable future meta-analysis to identify new pathways to target in ALS. The data will also prove valuable to data mining companies such as Verge Genomics and Human Longevity that aim to develop in silico models to understand late-onset human diseases. Researchers developing software tools to analyse single cell data will find these datasets very useful to benchmark their algorithms.

3. The results of the proposed study will be of great interest to pharmaceutical companies that are trying to develop therapeutics against ALS. The WNT pathway and its downstream genes identified as potential drivers of motor neuron loss in this proposal would be attractive targets for drug development. Drug development typically requires human relevant models of disease that can treated with thousands of drugs and phenotypes assayed to measure drug response and toxicity. The in vitro models implemented in this project would immediately benefit such efforts and would greatly accelerate the drug-discovery process for ALS. Working closely with the University of Exeter's Innovation, Impact and Business (IIB) directorate, we will actively seek collaborative ventures with the pharmaceutical industry to further develop our models for drug discovery or the identified candidates for targeted drug development. This will result in industrial investment in the form of direct research funding, access to high-tech equipment or creation of research positions.

4. In the long term, the proposed study would contribute to the development of new therapeutics for ALS thereby benefiting patients directly.

5. Finally, the proposed research will enable cross-disciplinary training of the PDRA and the Research Technician across fields as diverse as single cell genomics, bioinformatics, stem cell biology and high-content microscopy, equipping them with skills for a career not only in academia but also in industry.