Secular trends in breast cancer incidence in younger women suggest environmental factors, like exposure to environmental chemicals, may play a role in rising incidence. One of the strongest risk factors for developing breast cancer, next to family history, is high mammographic breast density, which is defined as the proportion of fibroglandular breast tissue relative to fat as seen on a mammogram. Phthalates, a ubiquitous endocrine disrupting chemical, have the potential to interfere with endogenous hormones like estrogen and androgens. There is growing evidence from animal and epidemiologic studies indicating distinct periods of heightened susceptibility to endocrine disrupting chemicals throughout the life course, particularly during critical windows of breast development. Exposure to hormonally active environmental chemicals like phthalates may be a modifiable risk factor for breast cancer, therefore reducing or eliminating exposure could have substantial public health benefits. The overarching goal of this dissertation was to assess the relationship between exposure to phthalates during two critical windows of susceptibility, the prenatal and pregnancy periods, and its effect on breast tissue composition in adolescence and adulthood. First, a comprehensive review of epidemiologic studies summarized the body of evidence for the association between phthalate exposure and intermediate markers known to be in the causal pathway of breast cancer risk (age at breast development, menarche, and breast tissue composition). This systematic review of the literature aimed to identify potential patterns of evidence by outcome and timing of exposure. Evidence from this review suggested that phthalate exposure during the prenatal and childhood periods may play a role in altering menarche. Findings for phthalate exposure and age at breast development were inconclusive. There was a considerable lack of epidemiologic data on phthalate exposure and breast tissue composition throughout the life course. Based on one study, there is a potential association between phthalate exposure during pre-puberty and altered breast tissue density in adolescent girls. No study assessed the relationship between phthalate exposure during the prenatal or pregnancy period and subsequent breast tissue composition. Second, an examination for the association between prenatal phthalate exposure and breast tissue composition measured in adolescence (Chapter 3) and the association between phthalate exposure during pregnancy and breast tissue composition measured during or after the postpartum transient period (Chapter 4) aimed to address this major gap identified from the comprehensive review. The empirical chapters of this dissertation used data from an ongoing longitudinal birth cohort study of mothers and their children conducted by the New York City Columbia Center for Children's Environmental Health and the Breast Cancer and the Environment Research Project (CCCEH-BCERP). The CCCEH-BCERP study cohort has prospective data on nine phthalate metabolite concentrations measured during the third trimester of pregnancy and breast tissue composition measured in a subsample of mother-daughter dyads. Notably, we used novel non-invasive methods (optical breast spectroscopy) in this younger cohort of mothers and daughters to objectively measure specific components of the bulk breast composition before mammography screening age. There was significant evidence of altered breast tissue composition in both mothers and daughters. For daughters (n=127, mean age 15.2 ± 1.9 years), prenatal exposures to select low molecular weight (LMW) and high molecular weight (HMW) phthalate metabolites altered overall breast density in opposing directions, which appears to be driven by significant altered percent breast water. There was a significant association between higher prenatal levels of a LMW phthalate metabolite (monobutyl phthalate) and lower levels of overall breast density (adjusted β = -0.32; 95% CI: -0.51, -0.13) and significant association between sum of di(2-ethylhexyl) phthalate (∑DEHP), a HMW phthalate metabolite, and higher levels of overall breast density in girls (adjusted β = 0.20; 95% CI: 0.05, 0.34). For mothers (n=133, mean age 41 ± 5.3 years at follow-up), there was a significant association between two LMW phthalate metabolites and lower levels of percent breast collagen. Additionally, there was a significant inverse relationship between levels of mono-(3-carboxypropyl), a HMW phthalate metabolite, and percent total hemoglobin of the breast (adjusted β =-0.03; 95% CI: -0.06, 0.00, p=0.05). Overall, this dissertation increased our understanding of the impact that exposure to phthalates during critical windows of susceptibility may have on specific components of the breast. Reducing exposure to both HMW and LMW phthalates may have an impact in reducing breast cancer risk, particularly for girls prenatally exposed, as there was stronger evidence of higher overall breast density and percent water from exposure to select HMW phthalates. Future prospective studies should confirm these results as findings might provide an opportunity for modifying potential breast cancer risk.

Introduction Endocrine-disrupting chemicals (EDCs) interfere with the endocrine system, disrupting hormone production and potentially causing adverse effects on exposed individuals and their offspring. EDCs are often found in mixtures, increasing the complexity of exposure assessment and health risk evaluation. EDCs are hypothesized to disrupt hormonal pathways essential for healthy pregnancy progression, with potential implications for gestational weight gain (GWG) and fetal growth. Phthalates are a widespread class of EDCs found in various consumer products, from personal care items to plastic materials, resulting in their pervasive presence in the environment. In this dissertation, I investigated associations between prenatal exposure to phthalate metabolites (PthM), both individually and in mixtures, and two critical maternal-fetal health outcomes: GWG and fetal growth, including birthweight. I addressed three primary aims across five chapters. First, I introduce PthM and outline the specific research objectives. Second, I systematically reviewed the literature on prenatal phthalate exposure and its relationship with GWG, identifying consistencies, limitations, and methodological gaps. Third, I conducted an empirical study identifying PthM trajectories throughout pregnancy and their associations with GWG and fetal growth. Fourth, I assessed the PthM mixture effect on GWG and birthweight and quantified individual contributions of each component within the mixture. Each aim contributed a distinct perspective: the systematic review synthesized the existing body of knowledge and highlighted research gaps; the trajectory analysis captured dynamic exposure patterns associated to maternal and fetal outcomes; and the mixture analysis assessed the combined PthM mixture effect, reflecting real-world exposure scenarios. Finally, I conclude with a discussion of the overall findings including strengths and limitations, and implications for EDC research and public health. Together, these chapters provide a comprehensive understanding of prenatal PthM exposure’s effect on pregnancy and birth outcomes by integrating current evidence, examining longitudinal exposure patterns, and evaluating mixture effects. Material and Methods In the systematic review, I included peer-reviewed studies from major databases, each examining associations between prenatal exposure to PthM, measured in blood and/or urine, and GWG. For the empirical analyses, I used data from the control group of a nested case-control study within the Nulliparous Mothers To Be (nuMoM2b) Study, a large and diverse cohort of nulliparous pregnant women in the United States, with repeated measures of maternal phthalate exposure including replacement phthalates, weight, and fetal growth (using estimated fetal weight). Eleven PthM were measured in maternal urine samples collected at up to three points across pregnancy. I used latent class growth analysis (LCGA) to explore trajectories of PthM over pregnancy and its associations with GWG and birthweight. Using Bayesian Weighted Quantile Sum (BWQS) regression, I assessed associations between PthM mixture concentrations and maternal-fetal outcomes, specifically GWG and birthweight, while generalized linear models (GLM) were used to evaluate associations with individual metabolites. I further explored these associations through stratified analyses across key subgroups: first trimester BMI categories, fetal-sex, and race/ethnicity, to identify vulnerable populations and potential effect modifications. Results In the systematic review, I found that prenatal exposure to certain PthM were negatively associated with both total and period-specific weight gain, although no single PthM demonstrated consistent associations across all studies. Evidence regarding prenatal exposure to mixtures of PthM and GWG varied across studies. There was indication of BMI-specific associations, particularly more pronounced associations among overweight and obese women. Significant differences were observed across studies regarding exposure and timing of exposure assessment, outcome assessment, and the methods used to analyze these associations. In the first empirical study, I identified stable trajectory classes for high (HMWP) and low (LMWP) molecular weight PthM. Replacement PthM demonstrated patterns of temporal variability. Specific trajectories of HMWP and LMWP were associated with lower GWG, with obese women, non-Hispanic Black/African American and Hispanic women, and those delivering female infants being particularly affected. Period-specific analyses highlighted early pregnancy as a sensitive period, with specific PthM significantly associated with lower GWG. Similarly, certain HMWP, LMWP, and replacement PthM trajectories were associated with lower infant birthweight, particularly among obese, underweight/normal-weight women, non-Hispanic White and Hispanic women, and those delivering male infants. In the second empirical study, I found negative associations between one-quartile increase in the PthM mixture on both GWG and infant birthweight in the crude and adjusted BWQS models. MBZP and MiBP contributed the most weights to the outcome-specific mixtures. Stratified analysis showed that the negative association with GWG was more pronounced among women delivering female infants, those with higher first trimester BMI, and Non-Hispanic Black women. For infant birthweight, the negative associations were observed in women delivering females, underweight and normal weight women and Non-Hispanic White women. Notably, the ordering of the contributions of PthM such as MBZP and MiBP within the mixture analysis was consistent across stratified groups. The mixture analysis corroborated the individual PthM analysis, showing consistent negative associations of MBZP and MiBP with GWG and infant birthweight. Conclusions Overall, there were consistent patterns of modest negative associations between PthM, including replacement phthalates, and both GWG and fetal growth, although most infants in this sample were born within a healthy birthweight range. These findings emerged following triangulation across multiple analytical approaches—trajectory analysis, individual metabolite analysis, and mixture analysis—highlighting that prenatal exposure to PthM is associated with lower GWG and birthweight.