Models, modifiers and novel treatments of Joubert syndrome

Cystic kidney disease accounts for 10% of the 40,000 UK patients requiring renal replacement therapy (dialysis and transplantation). Cystic kidney disease is part of a group of disorders referred to as the "ciliopathies" that cause various combinations of cystic kidney disease, retinal degeneration and brain abnormalities in patients. There are no current disease modifying treatments for these conditions.

The term "ciliopathy" covers a broad spectrum of disorders and patients typically can show a wide range of symptoms. For example, a mutation within the same gene can cause different symptoms in different individual patients. Clearly this complicates efforts to understand the cause of the disease, make accurate diagnoses, and develop specific treatments.
The archetypal ciliopathy is known as Joubert Syndrome (JS) which is predominantly caused by mutations in the CEP290 gene. We have created a mouse model of JS that more closely resembles the human condition than any other and have identified a previously unrecognized abnormality in the kidney that responds positively to drug treatment when we test kidney cells that have been isolated from diseased kidneys.
Our proposal focuses on identifying the factors involved in the complex presentation of ciliopathies to answer the question of why patients carrying mutations in the same gene show different symptoms and if they require different treatments?
To do this, we will breed our JS mutant mice with a different strain of mouse to mimic the genetic complexity of humans (note laboratory mice are inbred, whilst humans are outbred) to give mice with a range of disease severity. In this relatively simple system, it will be possible to identify the factors that affect disease severity.
In addition to the specific benefits to the field of ciliopathies, this proposal will provide a demonstration of the potential of using mouse genetics to understand complex human diseases in general.
This project is likely to provide long-term benefit to patients with inherited ciliopathies as it brings forward in a tangible way the prospect of personalized medicine and individualized treatments for patients. Specifically, the findings of this research proposal will help to categorize and precisely diagnose specific forms of cliliopathy. In parallel, we will assess novel potential treatments tailored to these specific forms. The fact that we have already identified one drug that can restore normal function to JS kidney cells suggests that this next round of research will make a significant step towards a treatment to reduce the morbidity and mortality associated with ciliopathies and reduce the need for dialysis and transplantation in affected patients.

We have created a Cep290 mutant mouse model of Joubert Syndrome (JS) that accurately recapitulates human JS, including the variability of phenotype dependent upon genetic background. Furthermore, we have demonstrated that Hh signalling is disrupted in developing renal collecting duct (CDC) cells in these mice and that treatment with a Hh agonist (purmorphamine) can restore normal collecting duct cell function in mutant primary cells ex vivo.
The aim of this proposal is to understand the complexity of JS with a view to identifying potential novel therapeutic targets and compounds. This will be achieved by:
The identification of genetic modifiers of the murine phenotype. C57BL/6 (severe) mice will be crossed with 129/Ola (mild) mutant mice to generate F2 mice that will be screened (Sequenom) to broadly identify loci associated with severity of cystic kidney disease. Affymetrix gene expression arrays will be used to compare kidneys from the parent strains and from a panel of F2 mutant mice to highlight potential modifier genes within these loci.
The identification of novel disease alleles mutated in JS patients. Synentic regions in JS patients having undergone whole exome sequending will be compared with the corresponding to the candidate loci identified in mice. This approach will also serve to validate candidate genes found in mice.
Functional assessment (loss/gain of function) of modifier genes in primary CDCs from JS mice and patients. The effects of loss-of-function (siRNA knockdown) and/or gain of function (lentiviral overexpression) will be evaluated in primary cells from our mouse model and ultimately in JS patient CDCs. The effects of these modifier genes will be assessed in terms of Hh agonist rescue.
This project will address the issue of phenotypic heterogeneity in JS and determine the effects of this heterogeneity upon drug efficacy.

This project will have impact in the following areas:
Health and well -being of ciliopathy patients: An improvement of day to day health and increase quality of life of patients with ciliopathies is a future benefit of this study. We anticipate this project will hasten the identification of novel treatments to for ciliopathies and cystic kidney disease. We expect our work to become translational within 5 years of this study. The impact of avoiding/postponing renal replacement therapy from an early age are self evident, in terms of health improvement and the complications from dialysis and transplantation.
General reduction in healthcare consumption: Renal replacement therapy costs the NHS 30K per patient per year. Cystic kidney disease accounts for 10% of the 40,000 UK residents on renal replacement therapy. Novel therapies that delay the progression of this disease are likely to reduce healthcare consumption overall.
Researchers within the field of ciliopathies: Identification of novel molecular pathways and disease (modifying) genes in Joubert syndrome will allow insights into related ciliopathies. Following the end of the grant cycle and after publication in peer reviewed discovery journals we would make microarray data and resources (including murine tissue samples) available to the scientific community. The research is of important interest to Rare Diseases consortia in the UK as well as the European Renal Association committee for inherited cystic kidney diseases. We anticipate new collaborations to be formed on the strength of our data and the creation of such novel and unique resources. Our studies will inform the approach to many other ciliopathy diseases as we will elucidate the important mechanistic details involved. Staff working with us on this project will develop skills in personalized medicine which will be applicable to many avenues of basic science research and current and future NHS treatments.
Mouse geneticists and human geneticists: This project will exploit the use of mouse models to probe finer aspects of human disease heterogeneity.

MRC and Genome England 100K Genomes Project: This work will link potential treatments to genetic variants affecting severity of disease and generate a reference framework of genes involved in ciliopathies to help annotate the human 100K genomes project.