How can you effectively treat a disease or disorder when you don’t know what causes it? To develop novel therapeutics, researchers need a firm understanding of the biologic factors that underpin dysfunction — so that they can come up with solutions for how to fix or stop the problem. The most severe class of epilepsy disorders, the developmental and epileptic encephalopathies (DEE), have been difficult to treat. DEE are characterized by significant global developmental delays and early-onset seizures often resistant to available therapies. Children with DEE can also have behavioral challenges and autism. DEE were regarded for many years as non-genetic, but thanks to next-generation technology, researchers have gained novel insights into how genetics impact DEE – and are now using that knowledge to create new therapies.
DEE occurs sporadically in families, so there was no obvious genetic link at first, and physicians believed that developmental issues would improve if they could treat the seizures. However, thanks to rapid advances in next-generation sequencing technologies, genetic discovery in DEE has exploded, revealing previously unappreciated pathogenic genetic variants as the most common cause of dysfunction.
These discoveries have significantly impacted diagnosis for epilepsy and other rare diseases, with up to 50% of individuals with DEE who undergo genetic testing now receiving a precise genetic diagnosis. However, there remains a gap between our ability to provide a genetic diagnosis and our ability to use that diagnosis for precision therapy.
At St. Jude, the laboratory of Heather Mefford, MD, PhD, Center for Pediatric Neurological Disease Research, is focused on identifying mutations that cause DEE in patients and determining their biological implications to help inform the development of precision therapies.
Given that only half of the individuals with DEE receive a precise genetic diagnosis using targeted and exome sequencing, scientists in the Mefford lab are expanding efforts to make genetic testing more robust. These efforts include adding to analyses short-read and long-read whole genome sequencing in trios (the affected individual and both parents). This testing allows them to search for causative abnormalities, including noncoding mutations, structural or copy number variants and repeat expansions. Moreover, they are decoding the epigenome to identify rare differentially (hyper- or hypo-) methylated regions in affected individuals.
To date, the Mefford lab has genetically screened approximately 1,000 unsolved (no known genetic cause) epilepsy cases using targeted gene and exome sequencing and approximately 150 trios using genome sequencing. They are actively growing their cohort through collaborations and establishing a repository of patient samples for functional studies and targeted therapy development.
Conducting laboratory research is only possible when there are reliable models to study. Biopsy specimens from patients are rarely available for studying DEE, so the Mefford lab works with induced pluripotent stem cells (iPSCs). The researchers engineer these cells by collecting fibroblasts from patients with unsolved DEE and, after reprogramming, transforming them into 2D neuronal models and 3D brain organoids. With these models, they can perform functional studies in disease-relevant cell types that are patient-specific.
Patient-derived brain organoids are specialized models that have demonstrated certain characteristics seen in DEE, such as aberrant neuronal firing, further highlighting their utility. Working closely with collaborators and families involved in patient advocacy groups, the researchers collect samples from individuals with and without a genetic diagnosis to generate iPSCs. With other members of the St. Jude Pediatric Translational Neuroscience Initiative, the Mefford lab is establishing a biorepository specifically for individuals with neurological conditions with onset in childhood as a departmental, institutional and external rare disease resource.
By studying these models, Mefford lab scientists have generated multi-omic data sets that allow them to identify changes in the transcriptome and proteome. This work reveals affected cellular pathways and highlights therapeutic targets. Moreover, patient-derived organoids will enable them to examine unique aspects of neuronal development and electrophysiology and to test potential therapies.
Targeted therapy for rare diseases using an N-of-1 approach (treating a single genetic variant found in a single patient) has, in some cases, demonstrated early success. However, approaching therapy development one patient or one variant at a time is not sustainable. Furthermore, the approach is restricted to patients with specific genetic variants that find willing scientific partners.
A proactive therapy design that directly addresses disease mechanisms is likely a better approach to benefit more patients. The Mefford lab is establishing a robust pipeline from gene discovery and disease and variant modeling to therapeutic testing to accelerate the implementation of targeted therapy for a broader group of patients.
Numerous approaches for gene-targeting therapies have emerged and continue to evolve, so the time is right to approach rare diseases with hope for treatment. Examples include RNA-based therapies (e.g., antisense oligonucleotides (ASOs), SINEUPs, mRNA tethering), CRISPR-based approaches (CRISPRa, CRISPRi) and gene replacement strategies.
Developing novel therapeutics starts with understanding dysfunction at the genomic level. By gaining insights into the mechanisms that cause, drive and control DEE in patients, the Mefford lab, alongside their collaborators and partners, is poised to help transition potential new therapeutics from an idea in the lab to treatments in the clinic.