Children born in the context of poverty have 10 years lower life expectancy and an increased likelihood of having compromised health and development, highlighting the urgent need for effective interventions to improve child developmental trajectories. Supportive perinatal interventions reduce health risk rooted in early experience, but to evaluate interventions’ effects on offspring development, researchers must either wait years for health outcomes to emerge in offspring, or use high-cost, labor intensive assessments such as MRI. Causal evidence of perinatal interventions’ effects on a biological level could provide mechanistic insight into interventions’ influence, contributing non-invasive, proximal indicators to evaluate, compare, and refine interventions for specific populations. However, existing studies on perinatal biological embedding are based primarily on observational data and do not provide the requisite experimental leverage for causal inference. The proposed project capitalizes on a cluster randomized trial (cRCT) of a structured, multi-component perinatal intervention, the Learning Clubs to a) experimentally test the hypothesis of perinatal biological embedding via the mechanism of DNA methylation (DNAm) in offspring and b) identify non-invasive biological indicators that could be used to evaluate perinatal interventions’ early effects on child development. Studies in rodents causally implicate DNAm in the biological embedding of perinatal adversity. Novel potential biomarkers of perinatal biological embedding in humans include 1) a cross-tissue DNAm signature of glucocorticoid (GC) exposure, developed using hippocampal progenitor cells, and validated using neonatal blood samples and 2) the Pediatric Buccal Epigenetic clock (PedBE), derived from buccal cells, which provides a measure of epigenetic aging, which is associated with compromised infant brain growth and neurodevelopment. Leveraging these two methodological advances and a cRCT of Learning Clubs (n=1245), we will generate DNAm biomarkers across infancy and paired genetic data using low pass whole genome sequencing. We propose that maternal Learning Clubs participation, versus care as usual, will alter the biological embedding of perinatal adversity, as reflected by DNAm biomarkers of GC exposure and pediatric epigenetic aging over infancy, which will correlate with infant development at 24 months. Aim 1: Test whether the Learning Clubs causes variation in DNAm at GC-sensitive sites in infants over the first two years of life. Aim 2: Determine whether Learning Clubs causes differences in epigenetic age in infants over the first two years of life. Aim 3: Identify DNAm signatures as predictors of infant development at 24 months of age. The results would provide causal evidence for perinatal biological embedding of supportive intervention via DNAm, and contribute to direct, proximal indicators of perinatal interventions’ salutary effects against developmental risk in the cont...