ENDD has partnered with Panorama Medicine to repurpose FDA-approved compounds for both STXBP1 and SYNGAP1 disorders. After screening thousands of compounds, we have identified several FDA-approved lead candidates that are blood-brain barrier penetrant and can upregulate target gene expression. We are currently testing them in our animal models for efficacy, phenotype rescue, and toxicology.
The ability to generate human induced pluripotent stem cells (hiPSCs), which are biochemically and morphologically like embryonic stem cells, is a paradigm shift in human disease modeling. Now researchers can generate hiPSCs from non-disease persons as well as from individuals with disease. This is particularly powerful for genetic disorders because hiPSCs – like embryonic stem cells – can differentiate into any cell type in the body. Therefore, we can use neurons derived from patient hiPSCs and compare them to non-disease neurons to identify disease-specific molecular and physiological phenotypes. These phenotypes serve as benchmarks to test the effectiveness of newly developed therapies.
We are developing a panel of STXBP1 and SYNGAP hiPSC lines derived from patients with different pathological variants. We are also using genomic engineering (CRISPR-Cas9) to generate an allelic series of several variants from each gene on a uniform non-disease genetic background, which will enable us to study genotype-phenotype relationships in a more controlled manner. Both types of hiPSC models will permit critically important proof-of-concept studies in human, clinically relevant, cell types.
Dr Deborah French is the lead for Human Stem Cell Model Development at ENDD.
Species differences between rodents and humans have likely played a large role decreased predictive value of rodentmodels for therapeutic development. Many of the therapies we are developing at ENDD require the use of human gene/DNA sequences. Therefore, we have developed humanized mouse of models of STXBP1 and SYNGAP1 where the mouse gene and flanking sequences have been replaced with the human gene and its flanking regulatory sequences. These models will enable the testing of human gene-targeted therapies in an in vivo context. We have confirmed appropriate gene expression from the human alleles relative to the mouse for the STXBP1 and SYNGAP1 humanized heterozygotes and homozygotes. Both fully humanized models have been deposited at the Jackson Labs – prepublication – to make them immediately available to the research community. They are also available upon request (bolandmi@pennmedicine.upenn.edu).
Crossing the humanized mice to heterozygous knock-out mice for each gene will generate humanized mouse models of STXBP1 and SYNGAP1 haploinsufficiency – the primary indication for both disorders – for the assessment of phenotypic rescue after therapeutic intervention. We are also developing humanized mouse lines with specific missense variants (together with corresponding hiPSC lines) for both STXBP1 and SYNGAP1. Once completed, these will also be made available.
Dr Elizabeth Heller, whose niece has SYNGAP1 disorder, leads the CRISPR-related therapeutic development at ENDD. Her laboratory studies the epigenetic mechanisms that regulate gene expression as an underlying etiological factor for neuropsychiatric disorders, such as addiction and depression, as well as neurodevelopmental disorders. At ENDD, her group is developing new tools to upregulate expression of the “good” functioning copy of STXBP1 and SYNGAP1 using an adaption of CRISPR-Cas9 technology called CRISPR activation (CRISPRa).
While at Columbia University, ENDD’s Strategic Director, Dr Michael Boland (whose son has STXBP1 disorder) developed a single AAV-mediated CRISPRa strategy shown to be effective in cortical brain organoids of STXBP1 haploinsufficiency. This and several other types of CRISPR-related therapies developed at ENDD, are being tested in hiPSC and humanized animal models of STXBP1 and SYNGAP1.
Many neurodevelopmental disorders are the result of loss of one functioning copy of a critical gene – a condition referred to as haploinsufficiency. One way to treat haploinsufficiency is to add back a functional copy of the gene in question – a process called gene replacement. Dr Beverly Davidson leads the development of gene replacement therapy at ENDD. The Davidson lab is focused on understanding the molecular etiology of several genetic neurological diseases and developing gene-targeted therapies to treat neurodegenerative disorders (Spinocerebellar Ataxia, Huntington’s disease, Amyotrophic Lateral Sclerosis), lysosomal storage disorders, and neurodevelopmental disorders (STXBP1). Using advanced imaging and sequencing methods their focus is on understanding why certain brain regions are more susceptible than others and improving methods to treat these disorders. A primary therapeutic focus is developing evolved adeno-associated viral (AAV) capsids for enhanced delivery of gene replacement therapies, as well as developing novel tools to regulate the expression of delivered genes.
At ENDD, we are developing and testing gene-targeted therapies for STXBP1 and SYNGAP1 disorders in human induced pluripotent stem cell (hiPSC) models and (humanized) mouse models of STXBP1 and SYNGAP1 haploinsufficiency.
