Preclinical gene therapy for genetic urinary bladder disease

In this project we will test cures for a devastating bladder disease.

This disease runs in families and makes children and adults unable to fully empty their bladders. Instead, urine builds up in the bladder and becomes infected, causing kidney failure.

At present there is no cure for this disease and affected people need to put a catheter into their bladder several times a day to drain urine.

Our Manchester research team have identified two faulty genes in people who have this disease. We discovered that these genes control how nerves grow into the bladder. These same nerves then control how our bladders fill up and store urine, and also control how our bladders change shape to let us pass water.

We also study mice with the same genetic condition. Like people with the disease, the mice have trouble emptying their bladders.

In this project we will use 'gene therapy' to cure these mice that have genetic urinary bladder disease. The therapy is given to mice just after they are born to give the best chance of treating the disease before it has done too much damage.

The successful outcome of our project will be a first step towards curing people with severe bladder disease beginning in childhood. This would mean that these people no longer have life-long incontinence of urine.

In future, this new type of treatment could be used in other life-threatening inherited genetic diseases that affect the kidney and urinary bladder. Such diseases are important causes of kidney failure in children and young adults and yet they currently have no definitive cures.

Heterozygous Hpse2 or Lrig2 parent mice are mated to generate neonatal litters with wild-type, heterozygous and homozygous mutants in a 1:2:1 ratio.

Neonatal litters will be intravenously administered AAV9 viral vectors and followed for six weeks, covering the period of bladder functional maturation to young adulthood.

Our Objectives are:

1. To significantly improve urination defects in mutant mice. If successful, this outcome will be most relevant to treating the crippling symptoms of this disease. Here, we will use the voided stain on paper method to assess urination frequency and volumes, respectively increased and decreased in UFS compared with wild-type mice.

2. To document effects on bladder physiology, bladder nerves, and expression of genes in pelvic ganglia. These results will help us better understand the pathobiology of UFS. Cystometry under terminal anaesthesia will be used to measure pressures inside bladders (increased in UFS) and seek vesicoureteric reflux of urine (present in UFS). We will quantify bladder nerves using peripherin, tyrosine hydroxylase and nNOS immunostaining because we reported aberrant patterns in untreated UFS mice. We will quantify Hpse2 and Lrig2 by in situ hybridisation, and neuronal associated genes (e.g. Sox10, Cntnap2, Capza1 and Wnt4) that we found were deregulated in untreated UFS mutant mouse bladders

3. To compare the efficacy of a neural-specific (SYN1) promoter with the broad action CAG (compound CMV/ACTB) promoter, as being used in human gene transfer studies. We reason that the neural-specific promoter will be most relevant to UFS, given that it appears to have a neurogenic basis.

4. To provide safety information, in addition to urinary tract analyses, we will measure blood levels of liver enzymes and full blood count and undertake histology of extra-renal organs. As well as assessing mutants, we will compare virus administered with vehicle-only administered wild-type mice.

1. The applicants will benefit from the research because it will advance, and bring together, their established and fruitful lines of study in urinary tract disorders and preclinical therapies for models of early onset genetic disease. The named research assistant (Lopes) has undertaken a PhD in growth factors and urinary tract development, and this will be her first post after that, applying her basic knowledge to treating a disease of the urinary tract.

2. The wider scientific community will benefit from the advance in biological knowledge about UFS. Over the last few years, our research has discovered that this is a disease with abnormal bladder innervation. How bladder becomes innervated has been little studied in molecular terms, despite the bladder being an important organ. Our study will further this area and be of great interest to developmental biologists and physiologists.

3. Although UFS is rare, it causes life-long morbidity, so it is potentially devastating for affected individuals. The successful outcome of our project will be a first step towards curing people with a severe bladder disease. This would mean that these people no longer have life-long incontinence of urine.

4. What is more, in future, our new treatment strategy could become a template to be used in mouse models of other genetic diseases that affect the urinary tract. Such diseases are important causes of kidney failure in children and young adults, and several are known to have monogenic causes. For example, these diseases include prune belly syndrome, and we showed a subset of families carry mutations of CHRM3 that codes for the acetylcholine receptor on bladder smooth muscle. This will be the next, and already available, mouse model we treat with AAV9.

5. Around 2% of the NHS health budget is used to treat severe renal disease. Urofacial syndrome has no definite cure but treating symptoms is still expensive. Chronic intermittent bladder catheterisation costs £4,000/year and dialysis costs £30,000/year. If early onset urinary tract diseases could be treated with a 'one shot' viral-mediated therapy (as being done for spinal muscular atrophy and haemophilia) there is the long term prospect of significant savings for the health service.

6. We use University of Manchester Intellectual Property Limited (UMIP), a subsidiary appointed to advise on and facilitate the protection and commercialisation of intellectual property. Each case is assigned a business manager with expertise in industry and commercialisation. Although our current discussions with UMIP do not point to an immediate patent, they will steer our path towards patient application over the coming decade, and facilitate links with industry, where appropriate.