Tuberous sclerosis, drug development, seizures, tumours, cognition
Basic research, Translational and applied research, Maintenance of colonies of genetically altered animals
Tuberous sclerosis complex (TSC) is a genetic disorder caused by mutation of one of two genes: Tsc1 and Tsc2. TSC is a prevalent disorder affecting one in every 6000 newborns, and is characterised by the growth of tumours throughout the organs of the body, epilepsy, mental retardation and autism. There is no cure for TSC and current treatments are limited to removal of tumours, minimising complications that arise from tumours, and relief of symptoms- mainly antiepileptic medications to treat the unrelenting seizures suffered by patients with TSC.
We will establish and maintain a colony of mutant TSC rats, the Eker rat, to support ongoing clinical development of new drugs for the treatment of this condition. We will assess whether our drugs may be beneficial for the treatment of seizures in TSC, reduce the number of tumours and whether they can alleviate the cognitive deficits and autistic traits that are commonly seen. If efficacious, we will use tissue from these and additional rats for in vitro experiments to determine the mechanism(s) by which our drugs exert their beneficial effects which will support clinical licensing applications.
Objective 1: Determine the anti-epileptic efficacy of a given study drug using behavioural monitoring methods.
Objective 2: Determine the neuropsychological tolerability profile of a given test compound using cognitive, emotional and motor behavioural tasks.
Objective 3: Determine the anti-tumoural actions of a given study drug using ex vivo methods in Tsc tissue.
Objective 4: Determine the underlying mechanism of action by which the study drug exerts its effects using in vitro methods in Tsc2 tissue.
Knowing whether our drugs work in this rat model of TSC will help regulators, industry, clinicians and patients benefit by informing and, if results are positive, speeding up the process of clinical licensing of these drugs.
Knowing how these drugs work will help us better understand why the disease occurs and will allow us to develop better medicines to treat the disorder.
Tsc2 mutant (Eker) rats; ~2500 over 5 years
Pairs of animals containing the Tsc2 mutation will be bred together. 33% of animals will exhibit no disease symptoms, 66% of animals will exhibit kidney tumours, cerebral hamartomas and mild cognitive impairments. Therefore, this work is considered to be of severe severity.
We will need to take tissue samples from these animals in order to determine which have the disease and which do not. This can cause some pain but will be minimised by pre-treatment with a local anaesthetic.
We will assess the anti-epileptic activity of our study drugs in these animals. In order to do this, we will use animal models that induce either a single seizure or full-blown recurrent epilepsy. Seizures can cause suffering and may be fatal. However, whilst these symptoms can appear distressing, seizures are preceded by a loss of conscious awareness by the animal such that it is unaware of the induced state (as is the case with humans epilepsy patients).
We will also treat animals with our study drugs over a number of months to assess any changes in tumour development and/or cognitive and autistic traits. Whilst these drugs have been well tolerated in our long experience we cannot rule out rare reactions in a tiny proportion of animals.
All animals will be killed at the end of the study unless asymptomatic animals are transferred to other authorised users to establish colonies for their own research.
TSC is a disorder that affects the whole body (brain and periphery) producing tumours, epilepsy, cognitive deficits and autism and, as such, it is too complex for current in vitro approaches to properly model it. Regulatory requirements for new drugs demand testing in whole animal models.
The number of animals will be minimised via the following:
i) Experimental group sizes will be determined by statistical power calculations.
ii) A professional statistician will be consulted, when required, to ensure that our statistical designs are appropriate and that the number of animals required is minimised, yet adequate precision and power are maintained.
iii) We will not breed more animals than we will need for our experiments.
iv) Animals, or animal tissue, may be used for multiple experiments, if appropriate.
v) Use established protocols to maintain high data quality and reduce variability in experimental results, leading to increased precision and the need for fewer animals in our experiments.
We will use rats which are the least sentient species possible that can be used. There are a number of different models of TSC, however none of them exactly mimic the human form of the disease. For this program of work we have chosen to utilise the Eker rat, the most widely used animal model to study TSC and the model that exhibits the greatest number of TSC symptoms of interest. Similarly to human TSC, the Eker rat produces a subtle cognitive phenotype and autistic-like behaviour, exhibits tumours and harmatomas primarily in liver, spleen and kidneys and, although it does exhibit spontaneous seizures (unlike the human form of the disease), it does show higher propensity for seizures after induction of kindling.
We will only breed sufficient animals to meet our planned experimental needs and we will use animals as early as it is scientifically possible for our experiments to minimise the time during which tumours manifest. Although we plan to utilise our animals early in their life cycle (6 months max), previously it has been shown that tumour burden itself is not lethal and Eker rats have been shown to survive to ~24 months old despite developing kidney tumours by 4 months old.
We have extensive experience (>8 years) of using whole animal models of epilepsy and seizure, including working with two severe severity. In addition, we will use our strong links with Dr Robert Waltereit from the University of Dresden who has significant experience of working with Eker rats to assist with our project. Our group is, therefore, one of the best placed in the UK to ensure that the welfare of these animals is maintained to the highest standard possible given their disease status.