Genetic and pharmacological targeting of Heat shock protein 72 (Hsp72) in the 5xFAD*Tg30 mouse model of Alzheimer’s disease

Abstract

The impact of Alzheimer’s disease (AD) is profound. In Australia, around 460,000 patients have dementia, AD being the most common form. An estimated 1.2 million people are involved in their care and it is the second leading cause of death. It is difficult to provide statistics purely on AD due to the difficulty in diagnosis, but an estimated 70% of dementia cases are AD (322,000 patients) (Dementia Australia 2018). Overexpression of the cytoprotective Heat Shock Protein 72 (Hsp72) is currently under investigation as a potential therapeutic option for prevention and treatment of AD due to its many protective mechanisms. The series of projects in this thesis aim to better understand the effects of upregulating Hsp72 in relation to behaviour, cognition and metabolism in a mouse model of AD.

The aim of the first study was to replicate and extend on the characterisation of a recently developed mouse model of AD. The mutant 5xFAD (overexpressing human APP and Presenilin-1) was crossed with Tg30 (overexpressing tau) to produce 5xFAD*Tg30 mice and investigate the effects of tau accumulation in the presence of amyloid pathology, which closely resembles human AD. A comprehensive battery of behavioural and physiological tests were performed to observe cognitive and physical decline over time, between 3-8 months of age. In agreement with one previous report (Heraud et.al 2014), we observed a decline in Rotarod performance from 6 months onwards compared with wildtype control (WT). While we noted a decrease in spatial awareness and memory (Morris water maze) and significant genotype differences in anxiety-like behaviour (large open field) by 8 months, memory function (Y-maze) and novel object recognition were not significantly affected. The 5xFAD*Tg30 mice had a 40% decrease in survival by 10 months of age. Additionally, 5xFAD*Tg30 mice were smaller, with a significant difference in body weight, tibialis anterior skeletal muscle weight and tibia length. Our results successfully reproduced aspects of the previously published description of this model (Heraud et.al 2014), while adding further additional characterisation of the model. This initial characterisation study demonstrated that the model develops many aspects associated with AD including frailty, early death and initiation of cognitive decline to complement the previously described AD-like brain pathology, specifically the presence of amyloid-beta pathology with tau accumulation and neurofibrillary tangle (NFT) development, found in this model (Heraud et.al 2014). Hsp72 has been shown to play a cytoprotective role in AD- related research by inhibiting amyloid-beta oligomerization, enhancing its clearance, restoring tau homeostasis and inhibiting neuronal apoptosis. Hence, the aim of the second study was to genetically overexpress Hsp72 and study its effects on the 5xFAD*Tg30 mouse model of AD. As in the first study, we crossed 5xFAD and Tg30 mice, to create the double transgenic, 5xFAD*Tg30, then crossed the double transgenic mice with Hsp72 overexpressing transgenic mice (Hsp72 Tg) to create the triple transgenic, 5xFAD*Tg30*Hsp72Tg. BGP -15 is a compound described to be a co-inducer of Hsp72, and therefore, our third study hypothesized that pharmacologically activating Hsp72 with this compound may be advantageous in preventing or delaying AD progression and associated pathology. 5xFAD*Tg30 mice were treated with BGP-15 or vehicle in their drinking water in a randomised and blinded study in both male and female mice from the ages of 2-10months. A comprehensive battery of behavioural and metabolic tests was conducted for both the second and third studies, and results compared to littermate WT mice. We observed significant declines in Rotarod, Y-Maze and Novel Object performance and increased seizure activity in the 5xFAD*Tg30 mice compared to wildtype, however neither Hsp72 overexpression (study 2) nor BGP-15 treatment (study 3) rescued this decline. Hsp72 overexpression was partially effective in maintaining lean mass in male mice and improved performance on the elevated plus maze. In males there was a 38% decrease in survival rates by 10months of age in the 5xFAD*Tg30 mice, which was improved by BGP-15 treatment to only a 14% loss (p=0.07). In conclusion, BGP-15 may improve survival rates in a gender specific manner (male), while overexpression of Hsp72 leads to maintenance of lean mass in male 5xFAD*Tg30 mice. Both avenues used to target Hsp72 were insufficient, however, to protect against the observed cognitive deficits in the model.