CLEAR is a prospective longitudinal birth cohort study that will follow a target sample of 1075 pregnant patients in the Detroit area from pregnancy through delivery. Recruitment for CLEAR began in November 2023 and is ongoing. Through June 2025, participants were enrolled into both the CLEAR birth cohort protocol and a birth cohort focused on asthma and allergy [34-36] under a combined protocol. To reduce participant burden and focus on pregnant patients in Detroit, CLEAR-only enrollment began in July 2025, which reduced data and sample collection requirements. The CLEAR protocol is identical for those enrolled both before and after June 2025.

To be eligible for the CLEAR study, participants must be aged ≥18 years, English-speaking, either receiving prenatal care at a Detroit-based Henry Ford Health (HFH) Women’s Health Services (WHS) clinic or residing in Detroit and receiving care at any HFH WHS clinic, and planning to deliver at an HFH hospital. Pregnant patients are eligible to join the study through 28 weeks of gestation. Because the baseline risk of PTB is higher in multiple gestation pregnancies [37], only singleton pregnancies are eligible for CLEAR. However, not all pregnant patients know they are expecting multiples at the time of study recruitment; participants who consent to the study but are later determined to have multiple gestations are removed from the study.

Potentially eligible pregnant patients are identified through the HFH electronic medical record (EMR) system, Epic (Epic Systems Corp). Eligible patients are entered into a study-specific web-based database. This database is an internally developed HFH platform used for recruitment tracking, study communications, appointment scheduling, sample tracking, and incentive management.

CLEAR recruitment uses multiple approaches in an effort to be broadly appealing and accessible to all eligible patients. The primary points of introduction to the study are the following: (1) in-person recruitment at a prenatal care appointment, (2) a recruitment message via the HFH online patient portal (MyChart; Epic Systems Corp), and (3) a mailed postal letter. Eligible patients may be contacted up to 8 additional times using a combination of texting, telephone calls, and email over a 6-week period or until they exit the eligibility window. Patients who decline participation or cannot be reached are assigned a final disposition code (eg, declined to participate, with a brief reason if available, or no contact, respectively) in the CLEAR database, after which no further contact is made.

Patients who express interest and are deemed eligible are sent a consent and Health Insurance Portability and Accountability Act (HIPAA) authorization form by email through the CLEAR Research Electronic Data Capture (REDCap) database. Study data are collected and managed using REDCap electronic data capture tools hosted at HFH [38,39]. REDCap is a secure, web-based software platform designed to support data capture for research studies, providing (1) an intuitive interface for validated data capture, (2) audit trails for tracking data manipulation and export procedures, (3) automated export procedures for seamless data downloads to common statistical packages, and (4) procedures for data integration and interoperability with external sources. Each participant is assigned the same study ID in both the CLEAR REDCap and the CLEAR web-based databases. Upon electronic signature of the CLEAR consent and HIPAA form, participants are automatically emailed a copy of the consent and HIPAA authorization form. CLEAR staff is simultaneously notified to review the form for completeness and update the participant’s status in the CLEAR web database to indicate enrollment.

This study was approved by the HFH Institutional Review Board (13198 and 13306). Eligible pregnant participants provide written informed consent and HIPAA authorization for both their and their baby’s participation. The consent is completed electronically via REDCap, which has previously been shown to be feasible and compliant in clinical research [40]. Participants are provided incentives (in the form of a gift card) for the study activities that they complete. Appropriate data use agreements and material transfer agreements have been obtained to share data and biospecimens across CLEAR projects and cores.

As part of the consenting process, participants are asked to opt in or opt out of several optional study activities. Although the participant does have to make a choice to opt in or opt out for each optional study activity, their specific choice does not impact their overall participation in CLEAR. The optional study activities include the following:

EMR data will be used to determine GA at delivery and define PTB. The “minimal” criteria described by Pennell et al [60] for determining PTB status will be used. GA at delivery will be assigned in accordance with the American College of Obstetricians and Gynecologists guidelines [61]. Our primary analysis will focus on cases defined as PTB if GA at delivery <37 weeks. Secondary analyses will consider the following: (1) continuous GA at delivery, (2) PTB subtypes (eg, extremely preterm if <28 weeks, very preterm if 28 to 32 weeks, and moderate-to-late preterm if 32 to 37 weeks), and (3) whether the PTB was spontaneous (preterm premature rupture of membranes or preterm labor) or indicated for medical reasons.

The CLEAR birth cohort was designed to examine the association of VOCs with PTB, to examine if VOCs impact risk of PTB via maternal inflammation and placental alterations, and to explore potential sources of VOC exposures. A nested case-control approach was chosen to weigh the high cost of laboratory analyses against the need to recruit patients early enough in pregnancy to accrue repeated and multiple biospecimens; nested case-control designs reduce some costs while having minimal impact on statistical efficiency [62]. Given birth rates at HFH and past birth cohort recruitment [63], we anticipate recruiting 1075 pregnant participants into CLEAR. The PTB rate in Detroit in 2018 was 15.2% [64]. We assume that approximately 15% of the CLEAR cohort will experience PTB; thus, 164 PTB cases are expected. Controls with GA at delivery ≥37 weeks at delivery will be 1:1 frequency matched [65] to cases based on infant sex and maternal race. Frequency matching maintains power [66] yet allows us to still adjust for these variables in the analysis. This sample provides sufficient power for the proposed analyses. For example, based on data from pregnant women in the National Children’s Study [56], SD of VOC metabolites on the log₁₀ scale is expected to range from 0.1 to 0.6. With 164 per group for the nested case-control, α=.05, 2-tailed tests, and 80% power, an odds ratio of 1.3, 1.6, or 3.5 for a VOC metabolite with an SD of 0.6, 0.3, or 0.1, respectively, could be detected for associations of a VOC metabolite with PTB. For examining the association of VOCs with placental function, with a false discovery rate of 0.05, there is sufficient power (>80%) to detect correlations of ≥0.3 and fold changes of 2.5.

A combined directed acyclic graph and change-in-estimate approach will guide selection of confounders and effect modifiers [67-70]. Although a large number of potential confounders will be measured, observational studies are subject to unmeasured confounding: e-values will be used to estimate the effect of unmeasured confounding [71,72]. We will consider methods to account for potential bias or misclassification of PTB case or control status by including the method of assigning GA at delivery (first-trimester ultrasound, second-trimester ultrasound, or date of last menstrual period) in models.

VOC results below the limit of quantification will be imputed as in Wei et al [73]. Multiple testing adjustment for high-dimensional data, such as false discovery rate, will be used [74]. With up to 6 VOCs measured, to reduce the multiple-testing burden, chemical mixture modeling will be used. Potential chemical mixture approaches include Bayesian kernel machine regression, weighted quantile sum regression, and principal components analysis [75]; each has distinct strengths and weaknesses. If more than one method is potentially appropriate, robustness of findings will be examined across different methods and all findings will be reported as in Vuong et al [76].

To examine associations between VOC metabolites and PTB, generalized linear mixed regression models will be used, where the repeated VOC metabolites are the covariates and PTB status (yes or no) is the outcome of interest. Generalized linear mixed model elastic net (glmmlasso), which accounts for mixtures and correlation between repeated measures, will be used to examine associations between VOC metabolites and maternal inflammation [77]. The association of VOC mixtures with DNA methylation will be tested using generalized network-based elastic-net linear mixed model (GELMMnet) with a logit link for β values (equivalent to M values), and associations with gene expression (RNAseq) will be tested using GELMMnet with a Poisson distribution [78]. If significant associations are identified above, we will then test if inflammation and/or placental function mediates associations between VOC metabolites and PTB using a mediation approach for dichotomous outcomes that estimates the natural direct and indirect effects for each individual marker [79], which has been extended to account for repeated measures and high-dimensional mediation effects [80]. To explore neighborhood-level or related factors, clustering or hot spot analysis and geographically weighted regression, a spatial analysis technique that uses nonstationary data (eg, demographic factors and environmental characteristics) to model the local relationship between predictors and an outcome (ie, VOC level), will be used [81,82].

Multiple strategies have been implemented to promote participant engagement and retention throughout the study. Participants receive personalized communications, including birthday and welcome baby cards, to acknowledge important milestones and maintain an ongoing connection with the study. CLEAR’s partnerships with clinical providers support trust-building and continuity, while study staff are trained to create positive, supportive encounters that help participants feel valued and respected. To help reduce barriers to continued participation, logistical support is offered, including Lyft (Lyft Inc) rides to and from study appointments, parking reimbursement, and a resource flyer tailored to participant needs (eg, mental health and postpartum assistance). Participants also receive prenatal gifts (eg, diaper bag and vanity kit) and age-appropriate gifts for their baby as tokens of appreciation. Collectively, these strategies are designed to foster a supportive study environment, reduce participant burden, and enhance long-term retention.

The consent process incorporates language specific to follow-up, including permission to use publicly available directories, databases (eg, WhitePages), or social media (eg, Facebook; Meta Platforms Inc) to validate contact information if participants are difficult to reach. This ensures that participants are informed of, and comfortable with, strategies to support continued engagement over time. In addition, participants are asked to provide the name and contact information of a family member or close friend that the CLEAR study can reach out to if the participant is lost to follow-up.

Establishment and study of the CLEAR birth cohort will provide needed data on the role of VOCs in PTB, particularly in older, urban areas such as Detroit, Michigan. Although previous studies have suggested potential associations between VOCs and PTB or other adverse birth outcomes, nearly all have relied on VOC exposures estimated based on residential address, with a single study examining personal exposure to only a single VOC (ie, benzene) [5]. CLEAR will address this limitation via measurement of VOC metabolites in maternal urine across pregnancy. Additionally, better understanding of potential mechanistic pathways, such as impacts on maternal inflammation and on placental function, may reveal areas for interventions to prevent PTB. To date, limited studies have examined these relationships with direct measurements of VOCs and/or in placental tissue; thus, CLEAR will provide unique data on these potential associations. With detailed address information, the CLEAR birth cohort will also allow for the study of potential VOC exposure sources, which are important to identify ways to either decrease exposure or to remediate contaminated areas. Previous studies have identified sources of indoor VOC levels in the homes of children with asthma in Detroit [83]; CLEAR will expand on these previous findings by better understanding sources of direct VOC measurements in pregnant women.

The CLEAR birth cohort addresses limitations of prior studies, importantly by directly measuring VOC exposure via urinary metabolites across pregnancy. Potential limitations, inherent to observational studies, remain. Although CLEAR captures numerous measures of potential confounders, residual confounding remains a potential limitation. This is somewhat mitigated by the proposed use of e-values. Similarly, recruitment bias is a potential issue impacting clinical research studies; because we are recruiting through a health system, we can describe those who do and do not agree to participate in the CLEAR birth cohort (in terms of basic demographic characteristics) which will enable us to address potential bias. Although we propose to measure urinary metabolites and circulating inflammatory biomarkers across pregnancy, not all women enter prenatal care in the first trimester, nor do they complete all study visits, thus missing data are anticipated. Similarly, women who do not deliver at a study hospital or who wish to retain their placenta for personal reasons may not have a placenta for functional measurements. Imputation and/or methods that account for loss-to-follow-up such as inverse probability weighting will be used when feasible. As a postindustrial rust belt city, results will be most generalizable to other similar urban cities, most of which have similarly high PTB rates as Detroit [1,2].

Establishment of the CLEAR birth cohort, currently focused on PTB, provides opportunities for additional future maternal and child research studies. Although outside the scope of the primary aims of this study, additional maternal and infant data collections are planned, including collection of maternal hair, toenails, and stool for assessment of stress biomarkers, additional environmental exposures, and microbiota, respectively, and neonatal blood spots, urine, and stool for measures of inflammation, metabolomics, and microbiota, respectively. Additional surveys over infancy are also planned to measure other exposures, child health and neurodevelopment, and other lifestyle factors.