Small molecule modulators of alternative splicing as a novel therapy for amyotrophic lateral sclerosis (ALS)

Amyotrophic lateral sclerosis (ALS) is a debilitating neurodegenerative disease caused by motor neuron loss that currently has no cure or a disease-modifying treatment. Pathological changes to a protein called TDP-43 are nearly ubiquitous in ALS and are believed to be one of the main causative mechanisms in this disorder. In ALS, TDP-43 becomes mislocalised from the nucleus where it normally resides, to the cytoplasm of motor neurons, which leads to loss of its nuclear functions. Recent transcriptomic studies demonstrated that although TDP-43 regulates a multitude of transcripts, one of the most marked effects of TDP-43 depletion is on a transcript involved in axonal maintenance, stathmin2. TDP-43 depletion drives premature polyadenylation and aberrant splicing in stathmin2 pre-mRNA, leading to the production of a non-functional, truncated mRNA and suppression of stathmin2 protein production. Crucially, impairment of neurite and axonal growth in TDP-43 depleted cells can be rescued by restoring stathmin2 levels. Overall, stathmin2 loss of function may be a final common pathway of motor neuron degeneration across different ALS subtypes. Stathmin2 protein is almost exclusively expressed in the CNS therefore its therapeutic targeting is not expected to cause on-target toxicity in other organs/tissues. The ability to restore normal stathmin2 RNA splicing and hence normal stathmin2 protein levels in cells with depleted TDP-43 can be used as a means to efficiently rescue axonal degeneration in ALS motor neurons. Thus stathmin2 represents a universal and extremely attractive therapeutic target for ALS.

Traditional drug discovery efforts have focussed on the protein but it has recently become possible to identify drugs which address the altered processing of RNA. Two small molecule splicing modulators branaplam and risdiplam were developed for a fatal childhood disease spinal muscular atrophy (SMA). Small molecules with their superior pharmacokinetic profile provide a good alternative to antisense oligonucleotides. The discovery of biologically active small molecule compounds acting on RNA and RNA-protein complexes can be supported by phenotypic assays as exemplified by the discovery of both splicing modulators for SMA in cellular reporter-based phenotypic screens.

This project is aimed at the identification and validation of small molecules capable of correcting abnormal stathmin2 RNA splicing, downstream of TDP-43 dysfunction, for ALS research and drug discovery. The student will generate and characterise reporter cell lines (using CRISPR editing and minigene construction); perform imaging-based high-content screening of compound libraries; develop secondary assays and carry out compound validation studies. Validation studies will involve analysis of axonal phenotypes and transcriptome-wide changes in patient-derived ALS cell models treated with hit compounds. Training in relevant techniques will be provided at the laboratories of Dr Tatyana Shelkovnikova and Prof Kurt De Vos at the Sheffield Institute of Translational Neuroscience (SITraN), where the project will be performed.

How to apply:

Please complete a University Postgraduate Research Application form available here: www.shef.ac.uk/postgraduate/research/apply

Please clearly state the prospective main supervisor in the respective box and select 'Neuroscience' as the department.

Enquiries:

Interested candidates should in the first instance contact Dr Tatyana Shelkovnikova ([Email Address Removed])