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[Preprint]. 2025 Aug 14:2025.08.12.25333453.

doi: 10.1101/2025.08.12.25333453.

Epigenetic clocks and longitudinal plasma biomarkers of Alzheimer's disease

Bowei Zhang¹, Linda K McEvoy², Steve Nguyen¹, Mark A Espeland³, Stephen R Rapp⁴, Steve Horvath⁵, Ake Lu⁵, Andrea Z LaCroix¹, Caroline M Nievergelt⁶, Adam X Maihofer⁶, Susan M Resnick⁷, Michelle M Mielke⁸, Kenneth Beckman⁹, Danni Li¹⁰, Brian Silver¹¹, JoAnn E Manson^{12

13}, Luigi Ferrucci¹⁴, Aladdin H Shadyab^{1

15}

Affiliations

¹ Herbert Wertheim School of Public Health and Human Longevity Science, University of California San Diego, La Jolla, CA, USA.
² Kaiser Permanente Washington Health Research Institute, Seattle, WA, USA.
³ Division of Gerontology and Geriatric Medicine, School of Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁴ Department of Psychiatry and Behavioral Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁵ Altos Labs, San Diego, CA, USA.
⁶ Department of Psychiatry, School of Medicine, University of California San Diego, La Jolla, CA, USA.
⁷ Laboratory of Behavioral Neuroscience, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
⁸ Department of Epidemiology and Prevention, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁹ Genomics Center, University of Minnesota, Minneapolis, MN, USA.
¹⁰ Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, MN, USA.
¹¹ Department of Neurology, UMass Chan Medical School, Worcester, MA, USA.
¹² Division of Preventive Medicine, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
¹³ Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA, USA.
¹⁴ Longitudinal Studies Section, Translational Gerontology Branch, National Institute on Aging, Baltimore, MD, USA.
¹⁵ Division of Geriatrics, Gerontology, and Palliative Care, Department of Medicine, University of California San Diego, La Jolla, CA, USA.

PMID: 40832399
PMCID: PMC12363716
DOI: 10.1101/2025.08.12.25333453

Epigenetic clocks and longitudinal plasma biomarkers of Alzheimer's disease

Bowei Zhang et al. medRxiv. 2025.

[Preprint]. 2025 Aug 14:2025.08.12.25333453.

doi: 10.1101/2025.08.12.25333453.

Authors

Affiliations

¹ Herbert Wertheim School of Public Health and Human Longevity Science, University of California San Diego, La Jolla, CA, USA.
² Kaiser Permanente Washington Health Research Institute, Seattle, WA, USA.
³ Division of Gerontology and Geriatric Medicine, School of Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁴ Department of Psychiatry and Behavioral Medicine, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁵ Altos Labs, San Diego, CA, USA.
⁶ Department of Psychiatry, School of Medicine, University of California San Diego, La Jolla, CA, USA.
⁷ Laboratory of Behavioral Neuroscience, National Institute on Aging, National Institutes of Health, Baltimore, MD, USA.
⁸ Department of Epidemiology and Prevention, Wake Forest University School of Medicine, Winston-Salem, NC, USA.
⁹ Genomics Center, University of Minnesota, Minneapolis, MN, USA.
¹⁰ Department of Laboratory Medicine and Pathology, University of Minnesota, Minneapolis, MN, USA.
¹¹ Department of Neurology, UMass Chan Medical School, Worcester, MA, USA.
¹² Division of Preventive Medicine, Department of Medicine, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA.
¹³ Department of Epidemiology, Harvard T.H. Chan School of Public Health, Boston, MA, USA.
¹⁴ Longitudinal Studies Section, Translational Gerontology Branch, National Institute on Aging, Baltimore, MD, USA.
¹⁵ Division of Geriatrics, Gerontology, and Palliative Care, Department of Medicine, University of California San Diego, La Jolla, CA, USA.

PMID: 40832399
PMCID: PMC12363716
DOI: 10.1101/2025.08.12.25333453

Abstract

Introduction: Chronological age is the strongest risk factor for Alzheimer's disease and related dementias (ADRD). However, the association of accelerated biological aging relative to chronological age with ADRD pathology is unclear.

Methods: In a cohort of 2,366 cognitively unimpaired older women from the Women's Health Initiative Memory Study, we examined associations of five baseline measures of epigenetic age acceleration (EAA) with 15-year changes in plasma ADRD biomarkers.

Results: At baseline, higher AgeAccelPheno was associated with lower amyloid-β42 to amyloid-β40 (Aβ42:Aβ40) ratio, and higher AgeAccelGrim2 was associated with elevated neurofilament light (NfL). Longitudinally, higher DunedinPACE - which measures the pace of biological aging - was associated with faster increases in phosphorylated tau at threonine 181 (p-tau181), p-tau217, NfL, and glial fibrillary acidic protein (GFAP) over 15 years.

Discussion: Accelerated biological aging, particularly as indicated by DunedinPACE, was associated with increasing levels of plasma ADRD biomarkers over time.

Keywords: Alzheimer’s disease; GFAP, NfL; amyloid beta; biological aging; blood biomarkers; dementia; epigenetic clocks; p-tau181; p-tau217, plasma; women’s health.

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Conflict of interest statement

The Regents of the University of California are the sole owner of patents and patent applications directed at epigenetic biomarkers for which Steve Horvath is a named inventor; SH is a founder and paid consultant of the non-profit Epigenetic Clock Development Foundation that licenses these patents. SH is a Principal Investigator at Altos Labs. Brian Silver discloses the following relationships: the National Heart, Lung, and Blood Institute (NHLBI) grant funding R01 HL164485 (Madsen TE, PI); Women’s Health Initiative Steering Committee and Outcome Adjudications Chair (NHLBI); NIH study section member (StrokeNet, NeuroNext, Special Emphasis Studies); Medicolegal malpractice review (consultant); Manager: Magnapeutics, LLC. Michelle M. Mielke has served on scientific advisory boards and/or has consulted for Acadia, Althira, Biogen, Cognito Therapeutics, Eisai, Lilly, Merck, Novo Nordisk, Neurogen Biomarking, and Roche; received speaking honorariums from Novo Nordisk, PeerView Institute, and Roche; and receives grant support from the National Institute of Health, Department of Defense, Alzheimer’s Association, and Davos Alzheimer’s Collaborative. B Zhang, LK McEvoy, S Nguyen, MA Espeland, SR Rapp, A Lu, AZ LaCroix, CM Nievergelt, AX Maihofer, SM Resnick, K Beckman, D Li, JE Manson, L Ferrucci, and AH Shadyab declare no conflicts of interest.

Figures

**Figure 1.. Associations of Epigenetic Clocks with Plasma Biomarkers of ADRD at Baseline**
Abbreviations: EAA = Epigenetic Age Acceleration; ADRD = Alzheimer’s diseases and related dementias; CI = confidence interval; SD = standard deviation; Aβ = amyloid-β; p-tau181 = tau phosphorylated at threonine 181; p-tau217 = tau phosphorylated at threonine 217; GFAP = glial fibrillary acidic protein; NfL = neurofilament light. Associations between epigenetic age acceleration (EAA) measures and plasma biomarkers were based on linear regression models. Standardized log2-transformed biomarkers levels are shown with 95% confidence intervals per 1-SD increase in EAA. EAA include AgeAccelHorvath (SD 5.38 years), AgeAccelHannum (SD 5.01 years), AgeAccelPheno (SD 6.81 years), AgeAccelGrim2 (SD 4.36 years), and DunedinPACE (SD 0.11 years of physiologic decline). Biomarkers include Aβ42:Aβ40, GFAP, NfL, p-tau181, and p-tau217. Models were adjusted for age, hormone therapy treatment arm, education, smoking status, race, ethnicity, physical activity, body mass index, diabetes, cardiovascular disease, hypertension, total cholesterol, HDL cholesterol, estimated glomerular filtration rate, and blood cell composition.

**Figure 2.. Changes in Plasma Biomarkers over an Average of 15 years**
Abbreviations: SD = standard deviation; Aβ = amyloid-β; p-tau181 = tau phosphorylated at threonine 181; p-tau217 = tau phosphorylated at threonine 217; GFAP = glial fibrillary acidic protein; NfL = neurofilament light. Distribution of plasma biomarker levels at baseline and a second time point an average of 15 years later among participants with both measurements (N= 873). Biomarkers include plasma Aβ42:Aβ40, p-tau181, p-tau217, GFAP, NfL. Violin plots display the distribution density. Boxplots indicate median and interquartile ranges. Means and SDs are presented above each group. Values above the 99^th percentile were truncated to reduce the influence of extreme values.

**Figure 3.. Associations of Baseline Epigenetic Clocks with Changes in Plasma Biomarkers of ADRD Over an Average of 15 Years**
Abbreviations: EAA = Epigenetic Age Acceleration; ADRD = Alzheimer’s diseases and related dementias; CI = confidence intervals; SD = standard deviation; Aβ = amyloid-β; p-tau181 = tau phosphorylated at threonine 181; p-tau217 = tau phosphorylated at threonine 217; GFAP = glial fibrillary acidic protein; NfL = neurofilament light. Associations between baseline epigenetic age acceleration (EAA) measures and rates of changes per decade in plasma biomarkers based on linear mixed effects regression models. Standardized log2-transformed rate of change in plasma biomarkers are shown with 95% confidence intervals per 1-SD increase in EAA at baseline. EAA include AgeAccelHorvath (SD 5.35 years), AgeAccelHannum (SD 4.92 years), AgeAccelPheno (SD 6.73 years), AgeAccelGrim2 (SD 4.28 years), and DunedinPACE (SD 0.11 years of physiologic decline). Biomarkers include Aβ42:Aβ40, GFAP, NfL, p-tau181, and p-tau217. Models were adjusted for age, time since baseline in decades, hormone therapy treatment arm, education, smoking status, race, ethnicity, physical activity, body mass index, diabetes, cardiovascular disease, hypertension, total cholesterol, HDL cholesterol, estimated glomerular filtration rate, and blood cell composition.

**Figure 4.. Estimated Changes in Plasma Biomarkers of ADRD over an Average of 15 Years across 3 Levels of DunedinPACE from a Fully Adjusted Linear Mixed-Effects Model**
Abbreviations: ADRD = Alzheimer’s disease and related dementias; SD = standard deviation; p-tau181 = tau phosphorylated at threonine 181; p-tau217 = tau phosphorylated at threonine 217; GFAP = glial fibrillary acidic protein; NfL = neurofilament light. Linear mixed-effects models contained random intercepts and slopes adjusted for age, time since baseline, hormone therapy treatment arm, education, smoking status, race, ethnicity, physical activity, body mass index, diabetes, cardiovascular disease, hypertension, total cholesterol, HDL cholesterol, estimated glomerular filtration rate, and blood cell composition. An interaction term between DunedinPACE and time was included. Results are Z-transformed DunedinPACE set to the mean (1.05) or 1-SD below (0.94) and above the mean (1.16). Biomarkers include GFAP, NfL, p-tau181, and p-tau217.

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References

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This is a preprint.

Epigenetic clocks and longitudinal plasma biomarkers of Alzheimer's disease

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Epigenetic clocks and longitudinal plasma biomarkers of Alzheimer's disease

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Conflict of interest statement

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