Abstract
Additional file 1: Supplementary Table 1. Quality control (QC) information on pre-processing of DNA methylation samples and probes. Supplementary Table 2. Estimates of inflation and bias in the analysis CpG to CSF biomarker associations in Alzheimer's disease (AD) and cognitively normal (CN) samples. The bacon approach (PMID: 28129774) was implemented using the R package bacon. Conventional approach for inflation estimate is based on the method described in Devlin and Roeder (PMID: 11315092). Supplementary Table 3. Summary of number of significant CpGs and DMRs associated with CSF AD biomarkers. Supplementary Table 4. Significant CpGs associated with CSF total tau in cognitive normal (CN) and Alzheimer's disease (AD) subjects. Highlighted in red are significant DNAm to CSF biomarker associations with P-value < 10-5or FDR < 0.05, or disease by DNAm interaction with P-value < 0.05. Also highlighted in red are gene promoter regions mapped with significant CpGs. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplementary Table 5. Significant CpGs associated with CSF Aβ42 in cognitive normal (CN) and Alzheimer's disease (AD) subjects. Highlighted in red are significant associations with P-value < 10-5, FDR < 0.05, or disease by DNAm interaction with P-value < 0.05. Highlights in yellow indicates overlap with significant DNAm in previous literature. Also highlighted in red are gene promoter regions mapped with significant CpGs. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplementary Table 6. Significant CpGs associated with CSF phosphorylated tau181 in cognitive normal (CN) and Alzheimer's disease (AD) subjects. Values highlighted in red are significant associations with P-value < 10-5, FDR < 0.05, or disease by DNAm interaction with P-value < 0.05. Also highlighted in red are genes with significant CpGs in the promoter regions. Highlights in yellow indicates overlap with significant DNAm in previous literature. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplementary Table 7. Significant DMRs at 5% Sidak adjusted P-value (z_sidak_p) associated with CSF Aβ42 in cognitively normal (CN) subjects and Alzheimer's disease (AD) subjects. For each DMR, annotations include location of the DMR based on hg19/GRCh37 genomic annotation (chr, start, end), nearby genes based on GREAT (GREAT_annotation), and Illumina gene annotations (UCSC_RefGene_Name), location with respect to CpG islands (Relation_to_Island), and overlap with enhancers described in Nasser et al. (2021) study (PMID: 33828297). Direction indicates positive or negative association between DNA methylation at a CpG located within the DMR and CSF biomarker. Highlights in yellow indicates overlap with significant DNAm in previous literature. Highlights in red indicate gene promoter regions mapped with significant DMRs. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplementary Table 8. Significant DMRs at 5% Sidak adjusted P-value (z_sidak_p) associated with CSF pTau in cognitively normal (CN) subjects and Alzheimer's disease (AD) subjects. For each DMR, annotations include location of the DMR based on hg19/GRCh37 genomic annotation (chr, start, end), nearby genes based on GREAT (GREAT_annotation) and Illumina gene annotations (UCSC_RefGene_Name), location with respect to CpG islands (Relation_to_Island), and overlap with enhancers described in Nasser et al. (2021) study (PMID: 33828297). Direction indicates positive or negative association between DNA methylation at a CpG located within the DMR and CSF biomarker. Highlights in yellow indicates overlap with significant DNA methylation loci in previous literature. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplementary Table 9. Significant DMRs at 5% Sidak adjusted P-value (z_sidak_p) associated with CSF total Tau in cogntively normal (CN) subjects and Alzheimer's disease (AD) subjects. For each DMR, annotations include location of the DMR based on hg19/GRCh37 genomic annotation (chr, start, end), nearby genes based on GREAT (GREAT_annotation) and Illumina gene annotations (UCSC_RefGene_Name), location with respect to CpG islands (Relation_to_Island), and overlap with enhancers described in Nasser et al. (2021) study (PMID: 33828297). Direction indicates positive or negative association between DNA methylation at a CpG located within the DMR and CSF biomarker. Highlights in yellow indicates overlap with significant DNA methylation loci in previous literature. The significant DNAm were compared to analysis results of brain samples in Zhang et al. (2020) (PMID: 33257653) and Shireby et al. (2022) (PMID: 36153390). Supplmentary Table 10. Results of pathway analysis using methylGSA (PMID: 30346483). In cognitively normal (CN) subjects, at 25% FDR, a total of 13 KEGG pathways and 76 Reactome pathways were enriched with CSF biomarker-associated CpGs, among which 3 KEGG pathways also reached 5% FDR. In AD samples, at 25% FDR, 10 KEGG pathways and 3 Reactome pathways were enriched with CSF biomarker-associated CpGs, among which 2 Reactome pathways also reached 5% FDR. Shown in red are FDR values less than 0.25, values of FDR < 0.05 are additionally highlighted in bold. Description of pathways that reached FDR < 0.25 in both CN and AD samples are also highlighted in red. Supplementary Table 11. Significant associations between CSF biomarker-associated CpGs and DMRs with target genes in blood samples of cognitively normal (CN) subjects and Alzheimer's disease (AD) subjects. We analyzed CpGs located in the promoter regions and distal regions separately. More specifically, for CpGs located in the promoter region (within ± 2 kb around the transcription start sites (TSS)), we tested the association between CpG methylation with expression levels of the target genes; for CpGs in the distal regions (> 2 kb from TSS), we tested associations between CpG methylation with expression levels of ten nearest genes upstream and downstream from the CpG.Supplementary 12. A total of 30127 CpG - mQTL pairs, associated with 16 unique CpGs, were significant in both cognitively normal (CN) and Alzheimer's disease (AD) sample analyses. The blood mQTLs were obtained from the GoDMC database. Definitions for columns under "Blood mQTLs" can be obtained from README file at http://mqtldb.godmc.org.uk/downloads.Supplementary Table 13. In cognitively normal (CN) samples, a total of 1518 mQTLs overlapped with the 24 GWAS nominated LD blocks in Kunkle et al. (2019) (PMID: 30820047). The mQTLs in blood were obtained from the GoDMC database. Annotations for CpGs include location of the CpG based on hg19/GRCh37 genomic annotation (Chr, Position), Illumina gene annotation (UCSC_RefGene_Name), the type of associated genomic feature (UCSC_RefGene_Group), and location with respect to CpG islands (Relation_to_Island).Supplementary Table 14. In Alzheimer's disease (AD) samples, a total of 41 mQTLs overlapped with the 24 GWAS nominated LD blocks in Kunkle et al. (2019) (PMID: 30820047). The mQTLs in blood were obtained from the GoDMC database. Annotations for CpGs include location of the CpG based on hg19/GRCh37 genomic annotation (Chr, Position), Illumina gene annotation (UCSC_RefGene_Name), the type of associated genomic feature (UCSC_RefGene_Group), and location with respect to CpG islands (Relation_to_Island).Supplementary Table 15. Sensitivity analysis for model that adjust cell type proportions estimated by the IDOL algorithm (PMID: 29843789), as implemented by estimateCellCounts2 function in R package FlowSorted.Blood.EPIC. All Aβ42-associated CpGs remained highly significant, with P-values ranging from 1.10 x 10-10 to 1.81 x 10-4. Supplementary Table 16. Sensitivity analysis for model that adjust cell type proportions estimated by the IDOL algorithm (PMID: 29843789), as implemented by estimateCellCounts2 function in R package FlowSorted.Blood.EPIC. All pTau181-associated CpGs remained highly significant, with P-values ranging from 1.39 x 10-8 to 2.92 x 10-3. Supplementary Table 17. CpGs with significant associations to both CSF AD biomarkers (in ADNI dataset) and Braak stage (in London dataset). Highlighted in red are associations that reached P < 10-5 in ADNI dataset analysis and P < 0.05 in London dataset. Supplementary Table 18. CSF biomarker-associated DMRs with significant associations to both CSF AD biomarkers (in ADNI dataset) and brain pathology (Braak stage in the independent London dataset).