C9orf72 dipeptide repeat proteins: Molecules and models

Frontotemporal lobar degeneration (FTLD) is the second most common cause of dementia after Alzheimer's disease and is related to motor neuron disease. Around 40% of patients with FTLD have a family history of disease indicating that genetics plays a large role in the development of the condition. Scientists have identified several genes that cause FTLD and one of these is called C9orf72. Currently we don't really know how this gene causes disease. This programme of work is aimed at trying to address this. With FTLD caused by C9orf72 five different repetitive proteins are abnormally produced in brain cells. We want to genetically modify mice so that they make two of these repetitive proteins to try and mimic disease in these. This will tell us if these repetitive proteins contribute to the process that leads brain cells to die. We will look at the effects on mouse behaviour and whether they cause the mice brain cells to die. We will study the levels of soluble and insoluble repetitive proteins in these mice over time. In addition, with the help of the Manchester Brain Bank and the Brains for Dementia Research Network we will measure levels of these proteins in the brains of people with the C9orf72 gene to see if levels affect disease presentation or pathology. In addition, we have brain tissue from three patients with FTLD that died of other causes early in their dementia diagnosis. We will also look at the levels of the soluble forms of the repetitive proteins in these to see if they are different early in the disease course.

The repeat expansion mutation of C9orf72 is the most common genetic cause of Frontotemporal lobar degeneration (FTLD). The expansion is translated producing 5 different di-peptide repeat proteins (DPRs). However, there is controversy over the role of the DPRs in disease. In various cell and animal models of C9orf72 disease DPRs are clearly neurotoxic. However, DPR pathology in patients does not correlate very well with neurodegeneration. It is possible this is due to DPR pathology being lost in areas susceptible to the DPR neurotoxicity leaving resistant areas with intact DPR pathology to be observed at post mortem. To address this we will make powerful inducible mouse models of DPRs and study the effects of DPR expression on the brain over time. We have produced expression constructs using alternative codons to those in the hexanucleotide repeat which produce inclusions without the repetitive RNA generating RNA foci.The inclusions we model are ubiquitin, p62 and ubiquilin-2 positive and recapitulate the pathology that is observed in patients. We will use these to generate state of the art inducible, single copy, Rosa26 integration site, transgenic mice. We will make one line producing cytoplasmic inclusions (pro-ala DPR) and one with intranuclear inclusions (pro-arg DPR). We will characterise the behaviour and pathology of these mice over a 18 month period. In addition, we will compare their brain transcriptome to that from a patient with the expansion. Also, to investigate the effect of DPRs on phenotype we will measure soluble and insoluble DPRs using novel western blotting and ELISA techniques in a large series of human brains. These data will be compared to clinical and pathological data. Collectively, we will generate data on the precise role of DPRs in the neurodegenerative process arising from the repeat expansion mutation in C9orf72.

Patients, their families and those others providing care and support will benefit most from this research. Given that frontotemporal lobar degeneration generally affects younger people of a working age its socioeconomic impact is huge due to loss of income and associated cost of care that is required. To alleviate this impact it is vital that an effective therapy is developed.

These people will benefit on several fronts. Much of this project is focused on understanding the key biological pathways involved in the neurodegenerative process using effective models to recapitulate the features seen in FTLD. The aim of this is to hopefully highlight potential therapeutic targets and/or strategies within these pathways.

Furthermore, as this project will establish the role of DPRs in the neurodegenerative process in FTLD and ALS this work will also be of benefit to academics working within this area.

In the longer term this work has the potential to benefit pharmaceutical companies and biomarker development programmes by identifying targets, this obviously also has potential accompanying economic benefits to the economy of the United Kingdom.

If these models do eventually aid in the development of effective therapeutics for FTLD and ALS then this would also benefit the general public. A reduction in the emotional, practical and financial impact placed on carers and patients would increase the quality of life of many thousands of people each year.