Towards personalised medicine for rare inherited brain diseases

Project title 
Towards personalised medicine for rare inherited brain diseases by identifying and manipulating variant RNA transcripts

Supervisors names
Prof Sara Mole
Dr Chris Minnis

Background:
The greatest abundance of variation in transcription occurs in the brain1, the site of significant inherited disease pathology and likely functional consequences2. We developed a bioinformatic and experimental pipeline for the most common Batten disease3 gene, CLN3 (MRC award MR/V033956), manuscript in preparation), and have collected fresh blood and brain bank samples for other diseases which await analyses. Analysing public (healthy control) long-read RNAseq databases4 for several inherited diseases of interest has revealed much diversity in usage of transcripts, with usage of the main transcript for specific disease genes varying from 21% (i.e. no dominant transcript) to >90%, and variation in transcript distribution across organs.
 

Aims/Objectives:
The project will: (1) apply state-of-the-art deep RNA sequencing technology to comprehensively identify disease transcript isoforms, including those missing exons or gaining new sequence; (2) investigate interesting templates at a cell level to understand their effect on protein function to answer questions such as how much disease pathogenesis associated with a particular mutation is due to partial loss of activity or the acquisition of new characteristics; whether disease severity correlates with the prevalence of particular variant transcripts; and whether the concentration of variant transcripts can be manipulated to reduce or exacerbate the disease. 

Methods:
1.    RNA is purified from blood or brain samples and the PacBio Sequel system (<1% error rate) is used for long read sequencing with targeted capture of disease gene cDNA to provide true transcript diversity and proportionality, revealing rare transcripts not accessible by traditional sequencing methods.
2.    For conserved genes the fission yeast S. pombe as well as human cells are used as model cell systems to study the functionality of peptides encoded by individual variant transcripts.
3.    Develop more accurate cell models to study transcripts arising from particular gene mutations to better understand their cellular consequences. e.g. brain organoid models can be developed through collaboration.
4.    Enhance disease pathology and pathogenesis of disease progression from mechanistic cascades.
5.    Design new therapeutic strategies based on enhancing the concentration of templates that are beneficial and reducing the concentration of templates that are deleterious. 
 

Timeline:
 Identification of variation in transcripts produced in inherited diseases of current interest, 1-12 mo; functional study of selected key transcripts in cell models, 6-30 mo; therapeutic design and development, 18-33 mo; thesis writing and submission 33-36 mo.
 

References:
1.    Zhang D, et al., Incomplete annotation has a disproportionate impact on our understanding of Mendelian and complex neurogenetic disorders. Sci Adv, 2020. 6: eaay8299. DOI: 10.1126/sciadv.aay8299.
2.    Clark MB, et al., Long-read sequencing reveals the complex splicing profile of the psychiatric risk gene CACNA1C in human brain. Mol Psychiatry, 2020. 25: 37-47. DOI: 10.1038/s41380-019-0583-1.
3.    Mole SE, et al., Clinical challenges and future therapeutic approaches for neuronal ceroid lipofuscinosis. Lancet Neurol, 2019. 18: 107-16. DOI: 10.1016/S1474-4422(18)30368-5.
4.    ENCODE Poject Consortium, et al., Expanded encyclopaedias of DNA elements in the human and mouse genomes. Nature, 2020. 583: 699-710. DOI: 10.1038/s41586-020-2493-4.

Contact
Sara Mole, s.mole@ucl.ac.uk