C9FTD/ALS is a rapidly progressive and debilitating neurological disease caused by expansion of a simple tandem repeat sequence in the gene C9ORF72. C9FTD/ALS is the number one inherited cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). There are no effective treatments for C9FTD/ALS, nor the other neurological repeat expansion disorders In C9FTD/ALS and most other repeat expansion disorders, of which over two dozen exist, the repeat expansion is transcribed into expanded tandem repeat-containing RNA, or xtrRNA, which mediates the molecular mechanisms of disease. Although the molecular chain of events that lead to C9FTD/ALS pathology are still unclear, it is widely accepted that reducing or blocking production of the xtrRNA will lead to effective therapeutic treatments that can halt disease progression. The key will lie in identifying treatments that can selectively inhibit transcription of large repeat expansions without affecting normal gene expression. A protein called Supt4h1 (Spt4 in yeast) acts as a processivity factor to the core RNA polymerase II enzyme to improve transcription across repetitive, structurally complex, or large regions of the genome. Previous studies have demonstrated that Supt4h1 is largely dispensable in yeast and its knock-down in C9FTD/ALS model organisms and patient-derived cells significantly mitigates disease pathology. Supt4h1 interacts with RNA polymerase II through dimerization with Sup5h, a core transcription factor. A crystal structure of this dimer reveals precise molecular contacts. Thus, we will develop an Supt4h1- Supt5h dimerization assay that can rapidly report dimerization status and is amenable to high throughput drug screening. The assay is colorimetric and quick, making high throughput chemical library screening accessible and cost-effective. We will search for small molecules that inhibit dimerization. After counter- screening and titration of the best inhibitors, we will further characterize the ability of top molecules to block C9ORF72 xtrRNA transcription and downstream molecular markers of disease pathology in patient- derived cells. This project will potentially deliver lead small molecules for further development as drug candidates for C9FTD/ALS. This project may also help fill an urgent gap in therapeutics for a number of diseases comprising an entire class of neurological disorders. The Supt4h1-Supt5h dimerization assay developed here will be useful for screening other libraries of small molecules or active biologics.
The number one genetic cause of frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS) is a mutation in the gene C9ORF72 that results in a massive expansion of repetitive nucleotide sequence. This repeat expansion is made into mutant RNA, which causes a molecular cascade of disease. This project will develop and use a high-throughput screen of over 130,000 compounds in search of small molecules that can block production of the toxic RNA and lead to drug candidates for FTD and ALS.