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Alzheimer's disease (AD) is a multifactorial neurodegenerative disease characterised by dysregulation of various cellular and molecular processes. Apart from environmental and lifestyle effects, genetic variations of the apolipoprotein E (APOE) gene plays a significant role in AD risk and progression, but these factors are poorly understood from a mechanistic perspective. In chapter 2, a meta-analysis of blood and CSF biomarkers of AD was performed, noting that the range of biomarkers studied has been restricted to a handful of classical proteins (Aβ and tau) and heavily focused on CSF. Still, more research is needed to establish robust blood tests to complement CSF or imaging tests for non-invasive testing options. Mass spectrometry significantly outperforms conventional antibody-based approaches such as ELISA and western blotting in specificity and quantification of low abundant proteins. Plasma proteomics has historically been limited by the lack of throughput and sensitivity, owing mainly to the complexity of the plasma samples. In chapter 3, I have developed a method for determining the fractionation strategy that provides in-depth plasma proteome coverage identifying 4,385 total proteins. This work demonstrates that simpler and faster approaches can provide substantial proteome coverage in conventional biochemistry laboratories. In chapter 4, I performed label-free proteomics analysis on plasma samples from clinical cohorts, using the newly developed fractionation method. Longitudinal and cross-sectional analyses of normal ageing and ageing with progression to MCI and AD were performed, based on plasma proteomic changes in the Sydney Memory and Ageing Study cohort. A replication cohort was used in chapter 5, the Australian Imaging, Biomarkers and Lifestyle study, which also included APOE e3 and e4 allele carriers. This additional information facilitated plasma proteome profiling to understand the impact of APOE e3 and APOE e4 carriers on AD dementia. Apart from comparing the effect of APOE genotypes on the AD proteome, I have confirmed a panel of reliable AD biomarkers that are consistently changing in both cohorts. In conclusion, I have successfully developed and applied MS-based fractionation methods for in-depth plasma proteome coverage of age, cognition and disease-related changes. Finally, a list of 44 plasma biomarkers consistently dysregulated in both AD cohorts presents a promising foundation for future clinical studies. |
