Our objective is to develop a community-based cohort and novel genomic science resource for defining the biological significance of ancestry-related genomic variation in African-Americans within the GENE-FORECAST :GENomics, Environmental FactORs and the Social DEterminants of Cardiovascular Disease in African Americans STudy. This resource will enable our team to test the working hypothesis that race-ancestry differences in the burden of cardiovascular disease (CVD) reflects the influence of a unique interplay between the distinct genomic variation characteristic of African-Americans (AA) and the exposome of social determinants and environmental factors that influence the pathogenesis of CVD in AA. The specific aims are:

AIM I. To examine the associations between common or ancestry-related DNA variants and CVD risk factors (e.g. hypertension) and phenotypes (e.g. coronary artery calcification) in African-Americans (AA).

AIM II. To examine the associations between health behaviors or social-environmental factors and CVD risk factors and phenotypes in AA.

The study is designed to create a cohort amenable to nested case-control analyses based on a community-based sampling frame with a target size of approximately 1800 self-identified, US -born, African-American (AA) men and women (ages 21-65) to be recruited over the next 5-6 years from the metropolitan Washington DC, Montgomery County (MC) and Prince George s County (PG) areas to be recruited to the NIH Clinical Center. The initial participant recruitment strategy involved two approaches: 1) a random-digit telephone screening survey targeting study-eligible AA that will be consented and invited to an evaluation visit in the NIH Clinical Center which we contracted with a well-established survey group (Southern Research Group \[SRG\]); and 2) a community outreach effort to recruit participants into the Clinical Center by leveraging marketing and the engagement of community-based leaders, organizations and faith-based institutions in the area. We are no longer contracting with SRG but rather focusing on community outreach and marketing for recruitment to the Clinical Center. The contract with SRG was terminated after the first two years of the protocol due to low yield of recruitment to the Clinical Center compared to community outreach.

Given the high burden of CVD among AA, this approach will yield a sample with normal individuals as well as a high proportion of AA with CVD risk factors such as obesity and hypertension that predispose to the eventual clinical signs and symptoms of CVD (e.g. heart attack and stroke). Based on previous epidemiology studies, this protocol s participant ascertainment approach and the target demographic profile; it is anticipated that the prevalence of clinically manifest CVD (history of angina, heart attack or stroke) will be less than 10-15% of the sample. All participants will undergo extensive evaluation in the Clinical Center that includes: medical evaluation (e.g. anthropometrics, blood pressure), laboratory tests (e.g. lipid levels, kidney function), social determinants profiles (e.g. socioeconomic status (SES), perceived stress, discrimination, depression, perceived neighborhood characteristics), blood/urine collection for deep-sequencing based omic analyses (e.g. whole exome sequencing, and RNA-Seq), as well as testing for pre-clinical , biomarkers of the pathobiological processes of CVD or CVD phenotypes (e.g. coronary artery calcification, microalbuminuria, leukocyte telomeres, or vascular dysfunction). It is anticipated that these deep sequencing efforts will yield novel ancestry-related DNA variants associated with the CVD phenotypes; yet with unclear biological significance in elucidating racial disparities in CVD. Accordingly, our protocol also includes a Genotype-to-Phenotype (G2P) component that re- contacts subsets of the cohort based on their genotype (e.g. APOL1 chronic kidney disease risk alleles) for a call-back visit for more in-depth phenotyping and characterization of the potential effect of the DNA variant of interest on human systems biology. In some cases family members of the proband may also be invited to participate in these G2P studies to further characterize the biological significance of these putative functional DNA variants of interest.

The primary outcome variables involve well established CVD phenotypes: 1) CVD risk factors (e.g. hypertension, dyslipidemia), 2) markers of pre-clinical CVD (i.e. coronary artery calcification, coronary plaque burden by cardiac CT angiography (CTA), carotid plaque burden by 3D ultrasound, vascular dysfunction, microalbuminuria, C-reactive protein, Vitamin D levels). The protocol will assess exposures associated with CVD and relevant covariates including: 1) social determinants (e.g. socioeconomic status (SES), perceived stress, discrimination, and depression); 2) environmental factors such as neighborhood characteristics (geospatial features of healthy lifestyles \[e.g. walkability\]) and 3) behavioral factors (e.g. diet, physical activity). The G2P callback visit protocol will involve additional measures of in-depth phenotyping that include: 1) peripheral immune cell phenotyping (e.g. T-cell, monocyte subsets); 2) blood/immune cell RNAseq; 3) iPSC cell line generation (endothelial; vascular smooth muscle cells) and analysis of cardiovascular cell systems biology; 4) HDL proteome analysis, 5) FDG PET/CT and/or PET/MRI scan (vascular inflammation) 6) echocardiography, 7) bisulfite sequencing for identifying sites of DNA methylation, and 8) chromatin immunoprecipitation followed by sequencing (Chip-Seq) for identifying sites of histone modification. It is anticipated that this multi-level, multi-dimensional analysis of genomic and phenotypic characteristics of AA will advance our understanding of the bio-social determinants of the intragroup variance and the increased overall burden of CVD observed among AA.