GWAS of plasma GFAP in East Asian cohort

Exploratory Score: 0.950 Price: $0.50 Alzheimer's disease human patients - East Asian cohort (K-ROAD) Status: proposed

What This Experiment Tests

Exploratory experiment designed to discover new patterns targeting SLC10A7, APOE in human patients - East Asian cohort (K-ROAD). Primary outcome: plasma GFAP levels

Description

Genome-wide association study of plasma glial fibrillary acidic protein (GFAP) levels, a biomarker of astrocytic activation and neuroinflammation. GFAP has been included in previous biomarker GWAS but this provides ancestry-specific findings in an East Asian population. The study identified genome-wide significant associations at the SLC10A7 locus and strong associations at the APOE locus.

TARGET GENE

SLC10A7, APOE

MODEL SYSTEM

human patients - East Asian cohort (K-ROAD)

ESTIMATED COST

TIMELINE

0 months

PATHWAY

astrocytic activation, neuroinflammation, glial response

SOURCE

extracted_from_pmid_41804841

PRIMARY OUTCOME

plasma GFAP levels

Scoring Dimensions

📖 Wiki Pages

APOE — Apolipoprotein Egene APOE - Apolipoprotein Escidex_docs APOE contributes to Alzheimer's disease by regulathypothesis APOE Genotyping for Neurodegenerative Disease Riskdiagnostic APOE Lipid Metabolism Pathway in Alzheimer's Diseamechanism APOE-Expressing Astrocytescell APOE contributes to Alzheimer's disease by regulathypothesis APOE Lipid Dysregulation Causal Chain in ADmechanism gfap-biomarker-adbiomarker GFAP (Glial Fibrillary Acidic Protein Gene)gene GFAP in Alzheimer's Diseasebiomarker GFAP (Glial Fibrillary Acidic Protein) - Biomarkerbiomarker GFAP (Glial Fibrillary Acidic Protein) - Diagnostidiagnostic GFAP-Guided Astrocyte Modulation Therapyidea GFAP (Redirect)redirect

Protocol

Phase 0: Study Design & Population Recruitment (Weeks 1-8)

Timepoints: Week 1-2 (design), Week 3-8 (recruitment)

Cohort: 2,000 East Asian participants from K-ROAD (Korean R&D database), aged 60-85 years, both sexes
Stratification: 1,000 AD cases (NINCDS-ADRDA criteria), 1,000 cognitively normal controls
Inclusion: East Asian ancestry (self-reported + PCA validation), available plasma, informed consent
Exclusion: Other neurological diseases, recent stroke (<6 months), autoimmune conditions

Phase 1: Sample Collection & Processing (Weeks 6-12)

Timepoints: Fasting blood collected at 8:00-10:00 AM

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Phase 0: Study Design & Population Recruitment (Weeks 1-8)

Timepoints: Week 1-2 (design), Week 3-8 (recruitment)

Cohort: 2,000 East Asian participants from K-ROAD (Korean R&D database), aged 60-85 years, both sexes
Stratification: 1,000 AD cases (NINCDS-ADRDA criteria), 1,000 cognitively normal controls
Inclusion: East Asian ancestry (self-reported + PCA validation), available plasma, informed consent
Exclusion: Other neurological diseases, recent stroke (<6 months), autoimmune conditions

Phase 1: Sample Collection & Processing (Weeks 6-12)

Timepoints: Fasting blood collected at 8:00-10:00 AM

Blood collection: 10 mL EDTA plasma + 5 mL serum per participant
Centrifugation: 1,500 × g, 15 min, 4°C within 30 min of collection
Aliquoting: 4 × 250 μL plasma aliquots, 2 × 250 μL serum aliquots, stored at -80°C
DNA extraction: QIAamp DNA Blood Kit (Qiagen, cat# 51106) from buffy coat
DNA QC: NanoDrop 260/280 ratio >1.8, Qubit fluorometry >50 ng/μL

Phase 2: Genotyping (Weeks 10-18)

Timepoints: Week 10 (DNA QC), Week 12-16 (genotyping), Week 17-18 (QC)

Platform: Illumina Asian Screening Array (ASA) + custom Content (SLC10A7, APOE flanking regions)
Genotyping: 730,000 markers + 5,000 custom SNPs
APOE genotyping: rs429358 (C>T), rs7412 (T>C) for ε2/ε3/ε4 determination
SLC10A7 SNP selection: 1000 Genomes East Asian LD tagging (r² > 0.8) + known missense variants
Quality control:
Sample call rate >98%, SNP call rate >99%
HWE p > 1×10⁻⁶
Remove ancestry outliers (>4 SD from East Asian cluster in PCA)

Phase 3: Plasma GFAP Measurement (Weeks 8-20)

Timepoints: Single measurement per participant at baseline

Assay: Simoa Human GFAP Advantage Kit (Quanterix, cat# 103346)
Sample preparation: Thaw plasma on ice, centrifuge 10,000 × g, 5 min, 4°C
Dilution: 1:4 in sample diluent
Instrument: Simoa HD-X Analyzer (Quanterix)
Detection range: 8.22-4,000 pg/mL
Intra-assay CV: <5%, Inter-assay CV: <10%
Batch randomization: Blind duplicates (10% of samples) randomly distributed across plates

Phase 4: GWAS Analysis (Weeks 18-28)

Timepoints: Week 18 (imputation), Week 20-24 (association), Week 25-28 (validation)

Imputation: Michigan Imputation Server (Eagle v2.4, minimac4), TOPMed r2>0.3
Association model: Linear regression (quantitative trait) in PLINK 2.0
Covariates: Age, sex, APOE ε4 status, PC1-PC10, batch effect
Genomic inflation: λGC < 1.05
Significance threshold: Genome-wide (p < 5×10⁻⁸) + suggestive (p < 1×10⁻⁵)
Conditional analysis: GCTA-COJO for independent signals
SLC10A7 expression QTL: eQTL analysis in DICE database + Genotype-Tissue Expression (GTEx) v8

Phase 5: Replication & Functional Validation (Weeks 26-40)

Timepoints: Week 26-30 (independent cohort), Week 30-36 (cell model), Week 36-40 (bioinformatics)

Replication cohort: 1,500 East Asian participants from BioBank Japan
Cell model: iPSC-derived astrocytes from 3 donors per genotype (SLC10A7 rs1145016 variants)
siRNA knockdowns: Lipofectamine RNAiMAX (Thermo Fisher), 30 nM siSLC10A7 (Santa Cruz, sc-90178)
GFAP ELISA: Cell supernatant collected at 72h post-transfection, R&D Systems Duoset (cat# DYC2468)
Western blot: Anti-GFAP (Cell Signaling, cat# 8245), anti-β-actin loading control

Expected Outcomes

Primary signal at APOE locus: rs429358 associated with plasma GFAP at p = 1.2×10⁻¹², ε4 carriers showing 1.4-fold higher GFAP vs ε3/ε3 (95% CI: 1.28-1.54)

Novel signal at SLC10A7 locus: rs1145016 (missense, Ala91Thr) associated at p = 3.7×10⁻⁸, T allele associated with 1.18-fold lower GFAP (95% CI: 1.10-1.27)

Heritability estimate: SNP-based heritability (h²SNP) of plasma GFAP = 0.22 (SE 0.04), consistent with prior European studies despite population differences

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Success Criteria

Primary GWAS threshold: At least one SNP at APOE or SLC10A7 reaches p < 5×10⁻⁸ in discovery cohort (n=2,000); if APOE signal fails, task is considered failed regardless of other signals
Effect size magnitude: Detected SNP effects must exceed 0.12 SD change in plasma GFAP per allele; smaller effects (<0.10 SD) indicate insufficient power or population stratification
Replication requirement: Lead SNPs must replicate with p < 0.05/3 = 0.017 (Bonferroni for 3 primary hypotheses) in BioBank Japan cohort with directionally consistent effect

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Related Hypotheses (5)

Selective APOE4 Degradation via Proteolysis Targeting Chimeras (PROTACs)0.795

Competitive APOE4 Domain Stabilization Peptides0.784

APOE4-Specific Proteolytic Fragment Inhibition Therapy0.777

APOE4 Allosteric Rescue via Small Molecule Chaperones0.765

APOE Isoform Expression Across Glial Subtypes0.743

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