In-vivo characterisation of an autoregulatory gene expression system to regulate miHD or miHD-BDNF as a treatment for Huntington's disease

Abstract

Huntington's disease (HD) is a fatal autosomal dominant inherited neurodegenerative disorder caused by a polyglutamine expansion in the huntingtin protein (HTT). There are currently no treatments that can halt or reverse the course of HD. Reducing the expression of either total or mutant HTT expression is hypothesised to have a therapeutic benefit. This can be achieved by the transduction of a miRNA against HTT by an adeno-associated viral (AAV) vector into an HD brain. Promising research also suggests increasing the expression of brain-derived neurotrophic factor (BDNF) could provide protective effects against HD. In this thesis, the therapeutic potential of AAV9 vector delivered miHD and miHD-BDNF under the expression of a CBA promoter was addressed in an acute rat model of mutant HTT overexpression. Our data demonstrated attenuation of HTT aggregate formation by miHD; interestingly this was counteracted in rats coexpressing BDNF. miHD-BDNF caused an increase in reactive astrocytes and neuronal degradation in the claustrum and septal nuclei. This toxicity caused by BDNF has not been detected in previous studies and is likely due to the use of prepro-BDNF in the miHD-BDNF treatment construct. Overexpression of miLuc also increased HTT aggregate formation, however, it did not affect soluble HTT expression, posing potential issues with treatment of HD by unregulated expression of miRNA. To make these treatments more clinically viable their expression needs to be more tightly regulated; this can be achieved by placing them under a gene regulation system. In our lab, a novel gene regulation system is being developed which relies on an increased protease expression (a Huntington's disease-specific signal) to express the therapeutic transgene. The second aim of this thesis was to validate the therapeutic potential of these same treatment groups under the expression of a STOP-expression system, representing the maximum potential output of the novel gene regulation system. When expressed under the STOP-expression system the increase in HTT aggregate formation by miLuc was lost, as well as the ability of miHD to effectively attenuate HTT expression. The loss of HTT expression attenuation by the STOP-miHD could be counteracted by increasing the dose given to the rats, increasing the ratio of miHD to HTT. miHD-BDNF expression by the STOP-expression system did not remove the toxic effects of BDNF. It may be less toxic to express a co-infusion of prepro-BDNF with enzyme tissue plasminogen activator (which cleaves pro- BDNF into its mature form) or directly treat with mature-BDNF.