Expanding the pipeline of therapeutic targets rooted in a strong biological rationale is key to maximizing the chance of success in future clinical trials. In 2022, the Target ALS Independent Review Committee (IRC) identified seven collaborative projects for funding. In just two years, these projects have achieved significant milestones. Target ALS is proud to highlight these below:
Project title: Probing RNA-binding protein aggregates at the nanoscale using in-silico-designed aptamers.
Researchers: Mathew Horrocks (University of Edinburgh, Project Leader), Jenna Gregory (University of Aberdeen), Gian Tartaglia and Elsa Zacco (Italian Institute of Technology), and Neil Shneider (Columbia University).
Key accomplishments: Through innovative in silico development and validation in post-mortem human tissue, these researchers successfully engineered FUS and TDP-43 aptamers that selectively recognize pathological protein aggregates without interfering with the aggregation process. These aptamers have been instrumental in advancing the understanding of ALS pathology, contributing to multiple publications that uncovered key discoveries—including novel nucleolar FUS and nuclear TDP-43 pathology, TDP-43 pathology within the amygdala, TDP-43/NEK1 colocalization, and correlations between FUS pathology and cognitive impairment. Building on these achievements, ongoing research aims to leverage the TDP-43 aptamer as a potential biomarker to distinguish ALS cases from healthy controls in cerebrospinal fluid (CSF), further expanding its clinical relevance.
Project title: Exploring the landscape of ALS genetics with machine learning and optical pooled screens.
Researchers: Johnathan Cooper-Knock (University of Sheffield, Project Leader), Michael Snyder (Stanford University), Ophir Shalem (University of Pennsylvania), and Eran Hornstein (Weizmann Institute of Science).
Key accomplishments: Through large-scale genetic analysis, this research team identified hundreds of novel ALS-associated genes that influence motor neuron function, paving the way for targeted investigations using CRISPR-based optical pooled screens and organelle phenotyping. Among these discoveries, two genes emerged as critical drivers of ALS: WDR49, expressed in astrocytes, and CCDC146, expressed in neurons. Dysfunctional WDR49+ astrocytes may play a key role in ALS progression, as mutations in WDR49 are linked to familial ALS (fALS), and these astrocytes are found near neurons exhibiting TDP-43 pathology. Meanwhile, CCDC146 was first identified in a screen for non-coding variants influencing ALS risk. By using antisense oligonucleotides (ASOs) in both in vitro and in vivo models, the team demonstrated that CCDC146 knockdown (KD) could provide therapeutic benefits for C9-driven and sporadic ALS cases. In an aggressive mouse model driven by TDP-43 mislocalization, ASO knockdown of CCDC146 extended survival by more than 50%, with some animals living five-times longer than any untreated animal. Overcoming significant technological hurdles, the researchers successfully conducted CRISPR-based optical pooled screening in iPSC-derived neurons. They also assessed how their hits influenced the “organellome” of patient-derived disease-relevant cells. These efforts revealed that several key genes influence TDP-43 localization, and that TDP-43 mislocalization profoundly disrupts the neuronal organellome, shedding new light on ALS pathophysiology.
Targeting the CCDC146 Gene for Therapeutic Benefit – A Consortia Approach
“If we find a genetic change, we know it’s a driver of the disease. It’s not a consequence of disease. And that’s really exciting because if we can correct that upstream driver, then potentially we’ve got a therapy.” – Dr. Johnathan Cooper-Knock, Clinician Scientist at The University of Sheffield.
Through the Target ALS Basic Biology Consortia Program, Dr. Johnathan Cooper-Knock (University of Sheffield, UK), Eran Hornstein (Weizmann Institute, Israel), Ophir Shalem (University of Pennsylvania, USA), and Mike Snyder (Stanford University, USA) have been collaborating for the past two years to enable the study of multiple genes or genetic mutations simultaneously using optical pooled screens. By using a unique combination of high-powered genetic analyses with cutting-edge lab techniques, the team expects to identify multiple candidate drug targets for ALS. Our Basic Biology programs encourage exploratory research like this to better understand disease biology and generate new targets for potential treatments. Through this work, the team has already identified that increased expression of the CCDC146 gene can cause ALS phenotypes in iPSC-derived motor neurons. The group then created an antisense oligonucleotide (ASO) that targets CCDC146. Importantly, this ASO isn’t limited to providing benefit for one form of ALS – the team’s preliminary work indicates the CCDC146 ASO reverses ALS phenotypes in iPSC-derived neurons from both C9orf72 ALS patients and from sporadic ALS patients. Work presented by the team at International ALS/MND in December showed that the ASO also extends lifespan in a TDP-43 mouse model. These data suggest the CCDC146 ASO has the potential to become a therapy for multiple forms of ALS. Due to their outstanding achievements and the high impact of their work, the Target ALS Independent Review Committee (IRC) unanimously awarded the team a third year of funding to allow them to dive deeper into the biology of CCDC146 in ALS and validate potential new treatment avenues and targets from their original screens.
Project title: Scalable phenotyping of ALS-associated point mutations, using base editor pooled genomic screens and a single cell transcriptomic readout.
Researchers: Eran Hornstein (Weizmann Institute of Science, Project Leader), Hemali Phatnani (New York Genome Center), Pietro Fratta (University College, London), and Michael Ward (NIH/NINDS).
Key accomplishments: This research team conducted large-scale genetic screens in HT29 cells and optimized prime editing screening in iPSCs, uncovering key insights into ALS-associated gene function. Base editing screens in HT29 cells revealed widespread transcriptional and pathway alterations following the editing of ALS-related genes, highlighting common molecular changes that may contribute to disease progression. Additionally, spatial transcriptomics analyses of ALS and FTD brain tissue offer a unique opportunity to identify shared disease mechanisms across these neurodegenerative disorders, with data now available in the Data Engine for further exploration. A major technological advancement from the Fratta lab includes the development of cutting-edge tools to track TDP-43 loss of function (LOF). These include detailed analysis of CE emergence at varying levels of TDP-43 depletion, fluorescent reporters to visualize TDP-43 LOF, and high-throughput CE detection using multiplex PCR. Contributions from the Ward lab include developing fluorescence in situ hybridization (FISH) assays for cryptic exons (CEs), massive improvements in the efficiency of pooled gene editing in iPSCs using Prime Editing, and identification of morphological fingerprints of TDP-43 loss of function in iPSC neurons that enable high content microscopy screens. Together, these innovations provide a powerful framework for studying TDP-43 dysfunction and its role in ALS and related diseases.
Project Title: Investigation of Ms4a genes as potential novel therapeutic targets for ALS.
Researchers: Paul Greer (Project Leader), Robert Brown, and Dorothy Schafer (UMass Chan Medical School).
Key accomplishments: This research team made a significant breakthrough in ALS by identifying Ms4a6c as a key player in motor neuron degeneration. SOD1^G93A mice with deletions of all Ms4a family members exhibited improved motor function and extended survival. Deleting just Ms4a6c in these mice not only prevented motor neuron death but also restored muscle innervation, demonstrating its potential as a therapeutic target and underscoring its role as the critical Ms4a family member that could modulate ALS symptoms and progression.
Previous literature indicated that Ms4a genes are primarily active in microglia, and functional studies by the University of Massachusetts team provided key mechanistic insights in ALS-relevant models. Specifically, Ms4a6c knockout microglia exhibited reduced synaptic material phagocytosis, suggesting that excessive synaptic pruning by microglia may contribute to ALS progression. These findings offer new avenues for understanding microglial involvement in ALS and highlight the Ms4a gene family as a promising target for therapeutic intervention.
Project Title: Evaluation of Novel Strategies to Ameliorate Aberrant Central Motor Synapse Elimination in ALS.
Researchers: Brian McCabe and Bernard Schneider (Swiss Federal Institute of Technology, Project Co-Leaders), and Sabine Liebscher (Ludwig-Maximilians University).
Key accomplishments: This research team uncovered a critical mechanism of synaptic loss in ALS, demonstrating that the selective depletion of inhibitory central motor system synapses occurs across multiple disease models, including Drosophila TDP-43 null flies , humanized Drosophila TDP-43 models, and SOD1^G93A mice. This synaptic loss was linked to the aberrant expression of phosphatidylserine (PtdSer), a lipid recognized by phagocytic cells, suggesting an overactive process of synapse elimination in ALS.
To counteract this pathological process, the team developed AAV-PtdSer-MASK, a gene therapy designed to mask aberrant PtdSer signaling and prevent excessive synapse clearance. When expressed from astrocytes, AAV-PtdSer-MASK selectively engaged with premotor synapses, increasing synapse density in an ALS mouse model (SOD1^G93A). Furthermore, intrathecal administration of AAV-PtdSer-MASK in adult SOD1^G93A mice led to significant motor function improvements, as quantified through an advanced machine learning-driven swimming assay. These findings highlight the therapeutic potential of targeting synapse loss in ALS and provide a promising avenue for future interventions.
Read more about the McCabe Consortia’s research in our feature on them: Unveiling the Path to ALS Therapies: A Journey from Flies to Humans.
Project Title: Does the accumulation of disease-associated forms of TDP-43 in platelets parallel ALS pathophysiology in the nervous system?
Researchers: Ruth Luthi-Carter (AC Immune SA, Project Co-Leader), Robert Bowser (Barrow Neurological Institute, Project Co-Leader), Abdulbaki Agbas (Kansas City University), and Emanuele Buratti (International Centre for Genetic Engineering and Biotechnology).
Key accomplishments: This research team made a groundbreaking discovery that platelets serve as a rich source of TDP-43 in the bloodstream, offering a new avenue for studying ALS-related biomarkers. Their work revealed distinct molecular forms of TDP-43 in platelets within patients and healthy individuals, identifying a high-molecular-weight TDP-43 species in ALS patients, while controls predominantly exhibited a dimeric TDP-43 species in platelets. Additionally, ALS patients showed significantly lower levels of full-length TDP-43 in their platelets compared to healthy controls. These findings provide critical insights into TDP-43 biology in peripheral blood and may open new possibilities for developing blood-based biomarkers for ALS diagnosis and progression monitoring.
Project Title: Targeting hypermetabolism for ALS treatment.
Researchers: Francesco Roselli and Johannes Dorst (Ulm University, Project Co-Leaders), Luc Dupuis (Université de Strasbourg), and Liang Li (University of Alberta).
Key accomplishments: This research team generated large-scale lipidomic and metabolomic datasets from ALS patients, healthy controls, and ALS mouse models, providing a comprehensive view of metabolic alterations in the disease. Their findings identified decreased levels of ANGPTL3 and ANGPTL4 in serum as a peripheral endophenotype specific to SOD1-related ALS. In sporadic ALS, ANGPTL3 and ANGPTL4 levels correlated with hypothalamic atrophy, establishing a critical link between central and peripheral metabolic dysfunction in ALS.
Lipidomic profiling further revealed distinct differences between ALS patients and healthy controls, with genetic and non-genetic ALS cases sharing similar lipidome signatures while also exhibiting unique subtype-specific variations. Murine ALS models mirrored these human metabolic phenotypes. Notably, ANGPTL alterations in SOD1^G93A mice were strongly associated with neuroinflammation in the hypothalamus, highlighting a potential mechanistic connection between metabolic dysregulation and neurodegeneration. These findings provide valuable insights into ALS pathophysiology and may inform future biomarker and therapeutic development.
Target ALS is exceptionally proud to support these significant strides in research as we work towards a world where Everyone Lives. If you have any questions about this research or would like to learn more, reach out to Ellen Guss at Ellen.Guss@targetals.org.
