Establishment of an in vitro platform for the development of n-of-1 antisense oligonucleotide treatments for ultra-rare metabolic disorders of the CNS

• Applicant: Dr. Marlen Lauffer
• Subject Area: Molecular and Cellular Neurology and Neuropathology ; Human Genetics
• Term: since 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 521414448

Antisense oligonucleotides (ASOs) are chemically-modified pieces of RNA, that can be designed to specifically hybridize to any target transcript, and can be applied to modulate pre-mRNA splicing to restore mRNA and protein function, thus serving as genetic therapies. As seen for spinal muscular atrophy, ASOs can be employed as disease-modifying therapies for severe genetic disorders of the central nervous system (CNS). Since these compounds are target-specific, they can be directed toward different genetic variants and even be used as customized therapies for individual patients. Hence, ASOs provide a possibility to close the treatment gap for patients with rare diseases for whom efficacious treatments are largely missing.Every ASO has to be designed and developed individually, which is cost- and time-consuming, and there are no standardized platforms available that could accelerate the efforts. Therefore, this project aims at establishing a standardized in vitro platform for the development of n-of-1 ASO treatments.The platform will be based on cellular models that allow to investigate each compound in a human environment to ensure target-specificty, avoid the use of animal models, and enable the identification of off-target effects. The toolbox will include studies on patient-derived fibroblasts as well as human induced pluripotent stem cells differentiated into 2D and 3D neuronal systems. With these modeling systems, it will be possible to study the effect of each ASO on transcript and protein levels and the overall functionality of the cells. It will further allow for the investigation of the neurotoxicity of each compound on the 2D and 3D neuronal cultures. There are no reliable neurotoxity platforms available yet for the investigation of ASOs and this project will provide a new toolkit for the study of neurotoxicity in 2D and 3D systems.To test and validate the platform, the rare disease group of lysosomal storage disorders (LSDs) will be used. This is a heterogeneous group of diseases and patients present with variable ages of symptom onset as well as a variety of severe symptoms affecting multiple organs. Most patients exhibit neurological symptoms like seizures and developmental delay, with some LSDs demonstrating a sole or predominantly neurological phenotype. Effective therapies and treatment management are missing for most forms of LSDs. There is proof of concept showing that patients with LSDs can benefit from ASO treatments. One individualized ASO treatment for a patient with Batten’s disease has been developed by Tim Yu and colleagues and further pre-clinical studies are ongoing for example Alexander disease and different forms of neuronal ceroid lipofuscionsis, all subforms of LSDs. Taken together, this project will establish a pre-clinical in vitro platform for the development and testing of ASO therapies for individual patients suffering from LSDs.

• DFG Programme: WBP Fellowship
• International Connection: Netherlands
• Hosts: Professorin Annemieke Aartsma-Rus, Ph.D.; Professorin Willeke van Roon-Mom, Ph.D.