Antisense Oligonucleotide (ASO) Development for Rare and Neglected Diseases

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We have engaged in productive partnerships across various therapeutic areas in collaboration with NIH and external academic researchers and companies. During the current fiscal year, progress was made in meeting the objectives of these various collaborations. The team is further refining the high-content ASO screening assays utilizing organoids. Complementing this work, algorithms to analyze the data from these assays are under active development in collaboration with the informatics team and confirm the ability of these data to predict toxicity accurately. The animal toxicity experiments of the ASOs were initiated to benchmark the high-content screening assays. In collaboration with clinical groups at the NIH Clinical Center, the design of the clinical trial is underway and will shortly be submitted for consideration. ASO Toxicity Assay Development: We continue refining the high-content screening assays using cell-based platforms that will substitute for low-throughput animal toxicity experiments. These screening platforms rely on organoids to predict the toxic side effects of ASOs and improve the safety pharmacology of this drug class. In parallel, we are constructing and benchmarking algorithms that predict toxicity, allowing bad ASOs to be eliminated early in the design phase. In this way, we are speeding up the process of ASO development and making it cheaper and faster to produce disease-modifying therapies for rare diseases. We collaborate with pharmaceutical companies and academic researchers to achieve these ambitious targets. For example, we continue to work with Ionis Pharmaceuticals and Roche, two of the leaders in the ASO field, on our in vitro and in vivo screening platform development. Our approach will make the domain knowledge accessible to all stakeholders. We are collaborating on developing and validating 2D and 3D human cell-based ASO toxicity assays using in vitro and nonclinical in vivo toxicity and safety data our collaborators have previously generated. We have incorporated iPSC-derived human neuronal cells into the development of these assays. The approach supports using a patient's cells in future IND-directed safety and toxicity studies for N=1 or a few diseases. Development of a Predictive Model of Safety and Toxicity of Candidate ASO Therapies: In collaboration with Charles River laboratories and our pharmaceutical partners, we continue to develop a platform to evaluate the safety and acute toxicity of ASOs in mice. These animal data have been compared to the results of cell-based screening assays and have validated the ability of the in vitro assays to predict the toxicity properties of ASOs in animals and to rely on the in vitro cell-based studies for future safety readouts. The project's initial focus is on acute and chronic dose-limiting toxicities encountered following intrathecal administration of ASOs targeting CNS neurological disorders. Our screening assay reduces the need for future animal experimental testing and predicts the safety of prospective candidate ASOs intended for clinical use. This project aims to provide sufficient and confirmatory in vivo data to the point that the FDA accepts cell-based in vitro data instead of conventional animal toxicology studies currently required for advancing new ASOs to clinical trials. N=1 Gene Identification in Neurodegenerative and Systemic Rare Diseases: Individualized N=1 therapies target a minimal number of people, even as few as one. We continue to work with the N=1 Collaborative, academia, foundations, and biotech companies to bring customizable treatments targeting the underlying genetic defect as safely and quickly as possible in rare diseases. We are focused on using antisense oligonucleotides (ASOs), as they are rapidly customizable, cost-efficient to manufacture, straightforward to administer and have a growing safety and efficacy record. Through these efforts, we are establishing a standardized framework for individualized medicine that will extend to other customizable platform technologies such as siRNAs, RNA therapeutics, and CRISPR. To test our drug development pipeline, we are working on two genes, KIF5A and CHCHD10, that cause amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD) as demonstration projects. ALS is a rare neurodegenerative disease characterized by progressive loss of motor neurons leading to respiratory failure and death. Approximately 7,000 Americans die of ALS yearly, which will increase dramatically in the future because of population aging. In collaboration with Dr. Derek Narendra (NINDS) and Dr. Justin Kwan, the director of the NINDS ALS clinic at the NIH Clinical Center, we work on developing ASOs and screening assays to identify lead ASOs for both genes and perform the IND-enabling studies with the aim of clinical trials. ASO Delivery Development: Most rare diseases affect the central nervous system, meaning gene therapies such as ASOs must be delivered intrathecally. While manageable, this route of administration adds to the complexity when scaling up treatments within the general rare disease population. We explore advanced delivery approaches, such as lipid nanoparticles, chemical structure, and conjugation with targeting moieties. Each system shows promise, but the goal of oral delivery of gene therapy agents requires considerable ongoing effort. The resources at NCATS bring key competencies to speed up research on delivery approaches. For example, we collaborate with Optimeos Life Sciences to apply their tunable lipid nanoparticle technology to deliver ASOs. We are working on in vitro and in vivo experiments to explore additional aspects of this innovative delivery system to neurons and other tissues. Together, we are exploring this payload delivery method to decrease the dose and mitigate the toxicity of ASOs. If successful, this novel modality will empower the ASO field to find a safe way to distribute this new class of therapeutics to target tissues. Initial experiments demonstrate the uptake of the nanoparticles into neurons and other cell types, highlighting the potential value of this new delivery approach. ASO Development for Infectious Diseases: In addition to treating rare diseases, we also deploy ASO technologies to treat emerging viruses. In particular, we are using ASOs to knock down the viral replication of SARS-CoV-2. This project collaborates with colleagues in the Early Translation Branch (ETB) at NCATS and Dr. Avindra Nath at NINDS. This effort maps out regions of the viral genome that are amenable to RNA-directed therapies. The underlying notion is that this approach can be more broadly applied to emerging pathogens once their genomic sequence has been isolated. In collaboration with biotech companies, we are also working on ASO liver and systemic toxicity discovery and assay development to test the candidate ASOs for infectious diseases.