Snowball, Albert; (2018) Gene therapy for focal neocortical epilepsy: Optimising viral vectors for clinical translation. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Approximately 0.5% of the world’s population suffers from focal epilepsy. While traditional pharmacotherapy provides essential relief for millions of patients, almost half continue to experience uncontrolled seizures. Of these, only a select few are suitable for surgical intervention. There is therefore a pressing need to develop alternative treatments for refractory focal epilepsy. One particularly promising approach is gene therapy. In previous work by our group, pathological epileptiform activity evoked by injecting tetanus neurotoxin into the rat motor cortex was suppressed by lentiviral delivery of a KCNA1 gene encoding the voltage-gated potassium channel Kv1.1. Similar therapeutic efficacy was achieved using a combined chemical-genetic approach involving adeno-associated viral (AAV) delivery of the modified muscarinic receptor hM4D(Gi), activated by its selective ligand clozapine N-oxide. Despite the pre-clinical success of these therapies, several features of the viral vectors made them unsuitable for human administration. Moreover, it was unclear if the reduction of pathological epileptiform activity in the motor cortex could be extrapolated to more common, longer-lasting seizures arising from other brain regions. Here we present two novel gene therapy vectors designed to maximise their translational potential. As before, the vectors comprise a KCNA1-encoding lentivirus and an hM4D(Gi)-encoding AAV. Translational optimisation of the vectors included: sequence engineering to boost transgene expression (and reduce Kv1.1 inactivation); a custom CAMK2A promoter to restrict expression to excitatory neurons; and, for the KCNA1 lentivirus, packaging into an integration-deficient vector to reduce the risk of insertional mutagenesis. The ability of a novel ligand to activate hM4D(Gi) receptors was also investigated. Our translationally optimised gene therapies were tested for efficacy in a rat model of focal epilepsy characterised by long-lasting visual cortex seizures. In randomised, blinded pre-clinical trials designed to closely mimic clinical trial conditions, the KCNA1 therapy rapidly and persistently suppressed seizures, but the hM4D(Gi) therapy failed to do so.

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