Targeted bisulfite sequencing analysis of candidate genes associated with Alzheimer’s disease.

Abstract Background Recent epigenome‐wide association studies (EWAS) have identified a number of loci in specific genes that show robust and reproducible alterations in DNA methylation in Alzheimer’s disease brain samples. The standard method to assess methylation in EWAS is via microarrays, however, these only target a limited number of methylation sites in each gene. Therefore, further analysis of methylation changes across the entire gene are required to determine the exact extent and pattern of methylation changes in disease. In this study we have performed targeted bisulfite sequencing for candidate genes in the Brains for Dementia Research (BDR) tissue sample resource, which is a highly characterized cohort containing tissue with a high degree of standardized pathological, clinical and administrative data available to allow comparative studies. Method Prefrontal cortex brain samples from 60 individuals were selected from the BDR cohort and grouped by Braak stage (Control 0‐II; mild cognitive impairment III‐IV; AD V‐VI). The DNA was extracted, before 30 genomic regions of interest, identified from previous AD EWAS, were captured using Agilent Sure Select target baits. The DNA was next‐generation bisulfite sequenced, before the sequence reads were aligned and the methylation status of cytosine residues were called using the Bismark Bisulfite Mapper program. Differentially methylated positions (DMPs) were then analyzed across the three groups. Result The degree and extent of methylation within the targeted genomic regions were identified and quantified for each group before being analyzed using linear regression models. Conclusion This study builds on previous work that identified differential methylation in several genomic regions that were associated with Braak stage. By identifying the exact positions that are subjected to differential methylation this work provides further evidence that dysregulation of methylation is associated with pathological changes in AD prefrontal cortex.