Autism spectrum disorders (ASDs) are a major public health concern in the United States affecting more than 1% of the nation's children in all racial, ethnic, and socioeconomic groups. The male-to-female prevalence ratio of roughly 4:1 in ASD is a well-recognized, but poorly-understood, phenomenon. An explicit focus on potential etiologic pathways consistent with this gender difference should be a priority in attempts to elucidate ASD causal mechanisms, including those amenable to environmental influence. Androgens, in particular testosterone, produced during pregnancy act on the brain to produce permanent gender differences in structure and function. Some researchers have hypothesized that ASD is an extreme presentation of "male brain," with fetal testosterone as the possible determining factor. Few endocrine disrupting chemicals are known to act on androgens; however, triclosans and triclocarbans (TCS/TCC) have been shown to have androgenic potential and are now widely used in liquid soap, toothpaste, mouth rinse, and other personal care products. Capitalizing on the availability of stored biosamples from a prospective cohort study of 213 mothers of children with ASD at the start of a subsequent pregnancy, we propose to assess prenatal TCS/TCC exposure in maternal prenatal urine samples and fetal testosterone levels in both cord blood and meconium and then and correlate these measures with ASD-related outcomes evaluated at 12 mos and 36 mos of age in the children born into the cohort. Fetal testosterone level will be explored as a potential mediating and/or moderating factor in associations between TCS/TCC and early ASD-related outcomes. The study will investigate a potentially avoidable environmental ASD risk factor, provide evidence related to an etiologic mechanism that comports with the observed ASD gender ratio, and generate i data on a fetal testosterone levels as assessed in meconium.

This project seeks to determine if prenatal air pollution exposures increase the risk of cognitive delays and autistic traits. Emerging evidence suggests that air pollutant exposure may increase risk of neurodevelopmental disorders and autism spectrum disorder (ASD). Our research demonstrates that exposure during gestation and early life to traffic related air pollution (TRP) and regional particulate matter (PM) were associated with risk of autism and neurodevelopmental delays. Other studies indicate neurodevelopmental effects based on prenatal biomarker measures of polycyclic aromatic hydrocarbon (PAH) exposure. However, we do not know if these associations persist over time, nor has a critical period of development been pinpointed. In this study we propose to examine the relationship between prenatal air pollution exposure and early longitudinal measures of cognitive ability and ASD in two prospective samples - Markers of Autism Risk in Babies, Learning Early Signs (MARBLES) and Early Autism Risk Longitudinal Investigation (EARLI). MARBLES and EARLI are both longitudinal studies of pregnant mothers, who have had one child with ASD, increasing the risk for abnormal development and ASD in the subsequent child. Approximately 40% of the infant children from MARBLES and EARLI will have a DD and 1/7 will be diagnosed with an ASD, which enriches our ability to study a broad range of neurodevelopmental outcomes. We will leverage the valuable phenotype and biospecimen resources of MARBLES and EARLI for the first prospective study of prenatal air pollution effects on autistic traits and the trajectory of cognitive development over the first thirty-six months of life. Specific aims are: (1) to assign air pollutant exposure using state of the art modeling techniques for multi- site studies estimating TRP and PM as well as measure novel biomarker measurements of nitro-PAH exposure to freeway-based diesel exhaust and gaseous nitro-PAH pollutants from vehicular and other sources; (2) examine the effect of these exposures on the trajectory of cognitive development using repeated administrations of the Mullen Scales of Early Learning (MSEL); and (3) evaluate the effect of these exposures on autistic traits and ASD diagnoses. Because air pollutant exposure is common and can be mitigated, the potential public health impact of this study is large. Additionally, identification of a risk factor in very young children provides the opportunity for early intervention where reduction of risk for disordered development is still possible.

Autism spectrum disorders (ASD) and related developmental phenotypes are among the most devastating of childhood disorders in terms of lifetime challenges and costs to families. We propose to test the hypothesis that autism and related disorders have an epigenetic basis. We challenge the standard genetic paradigm for autism as incomplete and argue that environmental factors during pregnancy play a critical role in the disorder and that these are mediated by epigenetic mechanisms. We propose a substantially new approach to the etiology of autism and related disorders that integrates genetic, epigenetic, and environmental information through a series of progressive epidemiologic analyses of prospective data beginning at the onset of pregnancy, through the first years of the newborn's life. We have partnered two complementary pregnancy cohorts: 1) the Early Autism Risk Longitudinal Investigation (EARLI) Network, which is recruiting pregnant women at high risk of having a new child with ASD because they already have an autistic child, 2) the Johns Hopkins University National Children's Study site, which is recruiting representative pregnancies in two Maryland counties. Together, these cohorts an extraordinary wealth of data across pregnancy and postnatally in 800 mothers, fathers, and children including: biosamples from mothers at least twice during pregnancy, from children at birth and 12 months, from fathers during the pregnancy, and from 300 placenta; multiple pre-conception and in utero exposures documented and/or directly measured; assessment of child development features associated with ASDs measured at birth (gestational age, birth weight and head circumference) and at 12 months (language, social, cognitive skills). We will perform genome-wide methylation and allele-specific expression analyses on these biosamples to address the following questions: 1) Are there regions of the epigenome that are susceptible to environmental insults occurring before and during pregnancy?; 2) Are there regions of the epigenome that correlate with quantitative newborn and infant developmental phenotypes related to ASD?; 3) How does genetic variation influence these epigenetic findings? The major features of our approach include a) novel genome-wide epigenetic array and statistical methods, b) two complementary pregnancy cohorts, c) longitudinal epigenome analysis through pregnancy and early life, d) exposure measurements through pregnancy, e) quantitative developmental traits at birth and in early life, and f) integration of GWAS with epigenome data. This work will serve as a foundation for a new field of "Epigenetic Epidemiology" and represents an extraordinary opportunity to test the idea that genetics, epigenetics and environment interact before and through pregnancy to modulate the risk of a devastating and common disease, using a state-of-the-art epidemiological and epigenomic design. It will provide the first rigorous analysis of the relationship between nature and nurture in the human epigenome.

The Early Autism Risk Longitudinal Investigation (EARLI) Network will establish and prospectively follow a cohort of mothers of children with an autism spectrum disorder (ASD) at the start of a subsequent pregnancy (with select preconception data collection). This enriched-risk cohort will have the potential to serve as a study population for a variety of observational studies on the etiology and natural history of ASDs. The Network includes an Administrative Center at the Drexel University School of Public Health, a Data Coordinating Center at University of California Davis, a Central Laboratory and Repository at the Johns Hopkins School of Public Health, and four field sites: 1) Drexel University School of Public Health / Children's Hospital of Philadelphia; 2) University of California Davis / MIND Institute; 3) Johns Hopkins University School of Public Health / Kennedy Krieger Institute; and 4) Northern California Kaiser Permanente. The EARLI Network will implement a core epidemiologic data collection protocol focusing on: 1) prospective documentation of exogenous exposure during pregnancy and early life; 2) collection and banking of biologic samples during pregnancy and early-life, allowing measurement of exposure biomarkers, endogenous risk biomarkers, genotypes, and gene-expression profiles; and 3) follow-up of the newborn sibling through 36 months of age to gather data on ASD diagnoses, continuous ASD behavioral domains, and other potential behavioral endophenotypes. The goal is to have data on 1,000 siblings of ASD probands from fetal life through 36 months of age. This will allow sufficiently-powered estimation of associations among exposures and biomarkers measured during the pre-, peri-, and neonatal periods and ASD risk as well as candidate gene, gene-environment interaction, epigenetic, and maternally-mediated genetic effects. Full data collection in the target sample size will require additional five-years beyond the initial award period; however, by the end of the award the Network will have followed 875 pregnancies and completed 250 24-month follow-up visits. The attached application describes a plan for establishing the cohort and presents a set of exemplary specific aims to be partially addressed during the initial award period and more completely addressed once data collection is completed. Other types of specific aims that could be addressed through additional analyses of EARLI Network cohort data are highlighted.